Microdacyn Wound Care

Transcript

Microdacyn Wound Care www.sdomedical.com Composición Microdacyn Wound Care® Solución Compuestos Fórmula química Concentración (%) Agua H2O 99,9 Cloro libre disponible Ácido hipocloroso HOCl Hipoclorito de sodio NaOCl Cloruro de sodio NaCl Carbonato sódico Na2CO3 Hidróxido sódico NaOH Peróxido de hidrógeno H2O2 Dióxido de cloro Ozono PH ClO2 O3 6,2-7,8 Composición Microdacyn Hidrogel® Sodium Magnesium Lithium Fluorsilicate Polydimethylsiloxane, trimethylsiloxy Sodium Phosfhate Sodium Chloride Hipochlorous Acid Sodium Hypochlorite Elecrolysed Water Ph Gelling Film forming NaH2PO4 NaCL HOCL NaOCL Vehicle 6,2-7,8 TABLA DE ACCION ANTIMICROBIANA DE MICRODACYN MICROORGANISMO Estafilococos aureus Meticilino Resistente (MRSA) Enterococos faecium resistente a vancomicina(VRE-MDR) Enterococos faecalis resistente a vancomicina (VRE) Estafilococos epidermis Estafilococos haemolyticus Estafilococos hominis Estafilococos saprophyticus Estreptococos pyogenes Micrococo luteus Bacteria Escherichia coli GramAcinetobacter baummannii Negativa Bacteroides fragilis Enterobacter aerogenes Haemophilius influenzae Klebsiella oxytoca resistente a MDR Klebsiella pneumonia Proteus mirabilis Pseudomonas aeruginosa Serratia marcescens Levadura Candida albicans TrichophytuonMentagrophytes Bacteria GramPositiva TIEMPO DE % DE ELIMINACIÓN DISMINUCIÓN 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos 30 segundos MDR: Fármaco multiresistente MRSA: resistente a la meticilina Estafilococos aureus VRE Enterococo resistente a la vancomicina. 99.999 99.999 99.999 99.999 99.999 99.996 99.998 99.999 99.996 99.999 99.999 99.999 99.997 99.999 99.999 99.999 99.993 99.999 99.999 99.999 99.998 99.999 99.999 99.999 Virus Testing Summary of all strains tested with Microcyn. All the virus listed below had complete inactivation by Microcyn Name Oculus Source HIV-1 ATS- TR31061-02 Influenza A Apptec - TR32003 Microcyn Viral Inactivation >3.75 log 10min EPA carrier test >5.09 log 1 min Type Enveloped Capsid Group IV +ssRNA Yes Yes Yes Helical No icosahedral Yes Helical No icosahedral Retroviridae Group V –ssRNA Orthomyxoviridae Parvovirus Human Coronavirus Rhinovirus type 37 ATS -TR31061-04 ATS – 31285-01 ATS- TR31286-01 >5 log 15 min EPA carrier test >3 log 15 sec, 30 sec >3.25 log 1 min 1% soil 2.75 log 15 sec 3.25 log 30 sec 2.75 log 1 min >4.75 log 5 min 1% soil Group II ssDNA Parvoviridae Group IV +ssRNA Coronaviridae Group IV +ssRNA Picornaviridae Name Oculus Source FVC – ATCC 2280 Feline calcivirus – surrogate for Norwalk Virus. Influenza A H1N1 Johannesburg/82/96 Flu A H3N1 Sydney/5/97 HSV1 MacIntyre-92 HSV2 clinical-87 Polio 1 lab standard Rhino 1A clinical-75 RSV Human respiratory syncytial virus clinical; no date available WNV – West Nile Virus NY-99 Hepatitis B Taketa-Grahm, Gutierrez, Thatcher. Microcyn Viral Log Reductions /Inactivation 5 log 1 min 6 log 5 min Taketa-Grahm, Gutierrez, Thatcher. Taketa-Grahm, Gutierrez, Thatcher. Taketa-Grahm, Gutierrez, Thatcher. Taketa-Grahm, Gutierrez, Thatcher. Taketa-Grahm, Gutierrez, Thatcher. Taketa-Grahm, Gutierrez, Thatcher. Taketa-Grahm, Gutierrez, Thatcher. 7 log 1 min 7 log 5 min 7 log 1 min 7 log 5 min 6 log 1 min 6 log 5 min 6 log 1 min 6 log 5 min 4-6 log 1 min - TBD Taketa-Grahm, Gutierrez, Thatcher. NA 7 log 1 min 8 log 5 min Not Tested. Type Enveloped Capsid Group IV +ssRNA No Spherical Yes Helical Yes Helical Yes icosahedral Yes icosahedral No icosahedral No icosahedral Yes Yes Yes spherical Yes icosahedral Caliciviridae Group V –ssRNA Orthomyxoviridae Group V –ssRNA Orthomyxoviridae Group I dsDNA Herpesviridae Group I dsDNA Herpesviridae Group IV +ssRNA Picornaviridae 8 log 1 min 8 log 5 min 4 log 1 min 4 log 5 min Group IV +ssRNA Picornaviridae Group V –ssRNA Paramyxoviridae Group IV +ssRNA Flaviviridae Group VII dsDNA-RT Hepadnaviridae Filoviridae viral family of RNA viruse Eficacia , seguridad y compatibilidad de la tecnología Microcyn ® Introducción Este documento proporciona una visión general del rendimiento, seguridad y compatibilidad de los resultados de un estudio relacionado con la tecnología Microcyn™ y enumera las características generales de la tecnología Microcyn™. Esta tecnología es la base de aplicaciones de productos individuales, como Dermacyn™ Wound Care, Desinfectante Oculus Microcyn 60 etc. Descargo de responsabilidad: La siguiente información no debe interpretarse como una guía de aplicación para los distintos productos. Las normativas son específicas para la aplicación de cada producto y y se requiere una aprobación normativa individual (por ejemplo, la certificación CE según la Directiva de Dispositivos Médicos 93/42/CEE). Por favor, consulte las instrucciones de uso del producto correspondientes para las aplicaciones específicas del producto. Tecnología Patentada Eficacia Bactericida (ensayo en portador) Las características bactericidas de Microcyn™ se ensayaron utilizando el método de Dilución de Uso AOAC, un ensayo en portador, según los requisitos de las directrices de Estados Unidos EPA DIS/TS5-01. Los organismos analizados fueron Pseudomonas aeruginosa (ATCC nº 15442), Staphylococcus aureus (ATCC nº 6538) y Salmonella choleraesuis (ATCC nº 10708). Se utilizaron 60 muestras por organismo para probar las propiedades bactericidas de Microcyn™. Cada muestra consistió en un cilindro de acero inoxidable con una película de las bacterias específicas secada sobre el mismo. Los cilindros fueron expuestos durante 10 minutos a Microcyn™ y luego transferidos a recipientes con un medio de subcultivo. Tras una incubación de 48 horas a 35 °C ± 2 °C, las muestras fueron observadas en busca de bacterias supervivientes. Conclusión: para cada organismo, el número de bacterias que sobrevivieron fue 1 sobre 60 muestras o menos, cumpliendo así con los requisitos EPA. [ATS Labs., Estados Unidos] Bactericida (Prueba en suspensión) Las propiedades bactericidas de Microcyn™ se evaluaron de acuerdo con el borrador de norma BS EN 13727:1999, un ensayo en suspensión. Para cada organismo, se prepararon suspensiones bacterianas con una concentración entre 1,5 x 108 y 10 x 5,08 /ml para Pseudomonas aeruginosa (ATCC nº 15442), Staphylococcus aureus (ATCC nº 6538) y Enterococcus hirae (ATCC nº 10541). Se añadió 1 ml de cada una de las suspensiones bacterianas a 8 ml de Microcyn™ junto con 1 ml de albúmina bovina (0,03 g/l) como sustancia interferente. Las mezclas de ensayo se mantuvieron a 20 °C durante 15 minutos y luego se cultivaron en placas de agar tripticasa-soja (TSA). Después de 48 horas de incubación a 36 °C ± 1 °C, se observó el crecimiento de las placas. Conclusión: para los tres organismos ensayados se consiguió una reducción de la carga bacteriana de más de 105, en de un tiempo de contacto de 15 minutos con Microcyn™ en cinco réplicas realizadas en dúplex. [Micromed Laboratories lnc., Estados Unidos] Cepas resistentes a bactericidas (MRSA) En este ensayo se probaron las características bactericidas de Microcyn™ contra Staphylococcus aureus resistentes a la Meticilina (ATCC nº 33592; MRSA), según las directrices DIS/TSS-05. Utilizando el método de dilución de uso AOAC, 20 penicilindros con una película bacteriana secada sobre ellos se sumergieron durante 10 minutos en Microcyn™. Tras la exposición, los cilindros se transfirieron a un medio de cultivo neutro y se incubaron durante 48 horas a 35 - 37 °C. Después de la incubación, las muestras fueron observadas en busca de bacterias supervivientes. Conclusión: en las condiciones de esta investigación, ninguna de las 20 muestras mostró crecimiento, lo que demuestra el cumplimiento de los requisitos EPA. [ATS Labs., Estados Unidos] Cepas resistentes a bactericidas (VRE) En esta ensayo se probaron las características bactericidas de Microcyn™ contra Enterococcus faecalis resistentes a la vancomicina (ATCC nº 51299; VRE), según las directrices DIS/TSS-05. Utilizando el método de dilución de uso AOAC, 20 penicilindros con una película bacteriana secada sobre ellos se sumergieron durante 15 minutos en Microcyn™. Tras la exposición, los cilindros se transfirieron a un medio de cultivo neutro y se incubaron durante 48 horas a 35 - 37 °C. Después de la incubación, las muestras fueron observadas en busca de bacterias supervivientes. Conclusión: en las condiciones de esta investigación, ninguna de las 20 muestras mostró crecimiento, lo que demuestra el cumplimiento de los requisitos EPA. [ATS Labs., Estados Unidos] Tuberculicida (Mycobacterium) En este ensayo se probaron las propiedades bactericidas de Microcyn™ contra Mycobacterium bovis (O'T nº 105401) según las directrices EPA DIS/TSS-06 para pruebas de tuberculicidas. Se prepararon suspensiones bacterianas con una carga de un 5% de materia orgánica (suero fetal bovino). Se inoculó un solo tubo con la suspensión bacteriana, seguido de exposición a Microcyn™ 5 minutos. Conclusión: durante la incubación durante 20 días a 35 °C ± 2 °C no se observó crecimiento y se concluyó que Microcyn™ cumple con los requisitos tuberculicidas de esta investigación. [ATS Labs., Estados Unidos] Virucida Microcyn™ fue ensayado para determinar sus características virucidas contra el virus de inmunodeficiencia humana tipo I (HN-D, cepa HTLV-IIIB, según las directrices EPA DIS/TS5-07 de EE.UU. El virus se aplicó como una película seca sobre placas de Petri de vidrio estériles, seguido por exposición a Microcyn™ durante 10 minutos. Posteriormente, el virus se separó de la sustancia de ensayo por filtración en gel y se tituló mediante diluciones en serie para el ensayo de inactivación. Conclusión: por grupos de dilución (7 grupos: de 10-1 "a 10-7), se observaron 8 muestras y no se detectó ninguna actividad infecciosa. La reducción en el título viral fue 3,75 log 10 para todas las muestras. En las condiciones de esta investigación, Microcyn ™ demostró la inactivación completa del HIV-I tras un tiempo de exposición de 10 minutos, de acuerdo con los requisitos para virucidas definidos por la EPA. [ATS Labs., Estados Unidos] Fungicida Microcyn™ fue ensayado para determinar sus características fungicidas contra el hongo Tichophyton mentagrophytes (ATCC nº 9533). El ensayo se realizó de acuerdo con las directrices EPA DIS/TS5-06, utilizando el método de dilución de uso AOAC. Las células fúngicas se aplicaron en forma de película seca en soportes de acero inoxidable y se expusieron durante 10 minutos a Microcyn™, con una carga orgánica de un 5% (suero fetal bovino). Después de la exposición los portadores se transfirieron a un medio de subcultivo neutralizante y se ensayaron en busca de supervivientes después de la incubación. Conclusión: En las condiciones de este experimento, Microcyn™ resultó fungicida frente a Tichophyton mentagrophytes tras una exposición de diez minutos a 20 °C. [ATS Labs., Estados Unidos] Esporicida Microcyn™ fue ensayado para determinar sus características esporicidas contra esporas Bacillus atrophaeus (ATCC nº 6633). El ensayo se realizó de conformidad con el borrador de la norma BS EN 14347:2002. Se preparó una solución de esporas con una concentración de esporas de 108-109 / ml a partir de una suspensión de esporas comercial. Las muestras de ensayo contenían 1 ml de la solución de esporas, 1 rnl de agua y 8 ml de Microcyn™. La solución de esporas se expuso durante 15 minutos a Microcyn™, después de la exposición las muestras se cultivaron y se incubaron durante cuatro días a 36 ± 1 ºC. Conclusión: En las condiciones de este experimento Microcyn ™ resultó esporicida contra esporas de Bacillus subtilis. La reducción media de las esporas fue de log 6.5, cumpliendo de este modo los requisitos del método de ensayo aplicado. [Micromed Laboratories lnc., Estados Unidos] Pioneros en innovación en ciencias de la vida y tecnologías disruptivas que están revolucionando la industria Seguridad / Biocompatibilidad Genotoxicidad Microcyn "fue evaluado en cuanto a su potencial genotóxico según la directiva 474 de la OCDE (Ensayo de micronúcleos en eritrocitos de mamíferos), según la norma ISO 10993-3:2003. La sustancia ensayada se inyectó por vía intraperitoneal en una población de ensayo de 5 machos y 5 hembras de ratón. La dosis aplicada fue 12,5 ml de Microcyn™ / kg de peso corporal durante 2 días consecutivos. Después de la inyección, los ratones fueron observados en cuanto a su salud en general y reacciones adversas. Al tercer día, los ratones finalizaron el tratamiento y se determinó la relación entre eritrocitos policromáticos y normocromáticos. Además, se examinó la incidencia de micronucleación en los eritrocitos policromáticos. Conclusión: las observaciones clínicas no revelaron signos de toxicidad durante el estudio. El examen microscópico de frotis de médula ósea de los animales utilizados mostró ausencia de toxicidad en los animales tratados comparable a la de los resultados encontrados en el ensayo negativo, en el que se utilizó un 0,9% de solución de NaCl. La misma comparación se hizo para la micronucleación de los eritrocitos policromáticos. En los animales tratados y en los animales del ensayo negativo, este número no aumentó, mostrando la ausencia de efectos mutagénicos. En base a este estudio, se concluyó que Microcyn™ no causa efectos genotóxicos. [Biomatech SAS, Francia] Citotoxicidad Esta prueba se ejecutó de acuerdo con la norma ISO 10993-5:1999 para determinar el potencial de Microcyn™ de causar citotoxicidad. Se colocó un disco de filtro con 0,1 ml de Microcyn™ sobre una superficie agarosa, superponiendo directamente una monocapa de fibroblastos de ratón (L-929). Las muestras preparadas se observaron en busca de daños citotóxicos al cabo de 24 horas de incubación a 37 ºC en presencia de un 5% de CO 2. Las observaciones se compararon con muestras de control positivas y negativas. Conclusión: las muestras que contenían Microcyn™ no presentaron ninguna evidencia de lisis celular o toxicidad, mientras que los controles positivos y negativos fueron según lo previsto. En base a este estudio se concluyó que Microcyn™ no genera efectos citotóxicos. [Namsa, Estados Unidos] Sensibilización de la piel Se realizó un estudio de sensibilización de la piel en 15 machos y 15 hembras de cobaya albino de pelo corto para determinar si Microcyn™ produce alguna reacción de sensibilización. La prueba se realizó según las directrices de EE.UU. EPA OPPTS 870.2600 y es necesaria para el cumplimiento de la norma ISO 10993-10:2002. En un grupo de control se incluyeron 5 animales de cada sexo. Estos animales permanecieron sin tratar durante cinco semanas, y posteriormente se les administró una dosis única de 0,4 ml de Microcyn™. Los animales restantes fueron tratados una vez por semana durante tres semanas con 0,4 ml de Microcyn™, descansaron durante 2 semanas y en la quinta semana del experimento se les administró, una vez más, 0,4 ml de Microcyn™. Conclusión: no se observó irritación de la piel en los animales del grupo de control. Tampoco se observó ninguna irritación en el otro grupo. Debido a la ausencia de respuesta cutánea, se llegó a la conclusión de que Microcyn no es sensibilizante de la piel. [Stillmeadow, Inc., Estados Unidos] Irritación cutánea Se realizó un ensayo para cumplir con la norma ISO 10993-10:2002 a fin de determinar el potencial de irritación de la piel de Microcyn™. En esta prueba, en tres conejos blancos de Nueva Zelanda se aplicó una dosis de 0,5 ml de Microcyn™ en los puntos de ensayo especificados en el animal. La sustancia de ensayo se administró en una gasa y se fijó a los puntos de ensayo para producir 4 horas de contacto con la piel. La piel tratada se observó durante 72 horas a intervalos de tiempo regulares y se comparó con puntos de piel no tratada en el mismo animal. Los animales fueron observados en cuanto a formación de edema, formación de eritema o escaras. Conclusión: todos los animales fueron encontrados sanos durante todo el estudio. No se detectó irritación de la piel durante todo el periodo de estudio. En base a este estudio, se concluye que la irritación cutánea de Microcyn™ es despreciable. [Northview Pacífic Laboratories Inc., Estados Unidos] Irritación ocular Se efectuó un ensayo de irritación ocular para cumplir con la norma ISO 1099310:2002 a fin de determinar el potencial de irritación ocular de Microcyn™. En esta prueba, en tres conejos blancos de Nueva Zelanda se aplicó una dosis de 0,1 ml de Microcyn™ en el ojo derecho de cada animal. El ojo izquierdo se dejó sin tratar y sirvió como control. Tras la aplicación de la sustancia de ensayo, los ojos se examinaron a intervalos de tiempo regulares para ver si había irritación. Conclusión: todos los animales fueron encontrados sanos durante todo el estudio. No se detectó irritación ocular durante todo el período de estudio. En base a este estudio, se concluye que Microcyn™ no causa irritación ocular. [Northview Pacífic Laboratories Inc., Estados Unidos] Toxicidad oral aguda Se evaluó el potencial de toxicidad oral aguda de Microcyn™ cuando se administra como una dosis por sonda a un nivel de 5000 mg / kg de peso corporal para una población de ensayo consistente en 3 ratas albinas hembras. El ensayo se realizó según el reglamento EPAOPPTS 870.1100 de EE.UU. a fin de cumplir con la norma ISO 10993-11:1996. Los animales tratados recibieron 4,98 ml de Microcyn™/kg de peso corporal y se observaron durante 14 días. Conclusión: no se detectó mortalidad ni signos de toxicidad clínicos o de conducta. La necropsia realizada al final del estudio no reveló anomalías observables. En base a este estudio, se concluye que Microcyn™ no causa toxicidad oral. [Stillmeadow, Inc., Estados Unidos] Toxicidad dérmica aguda La prueba fue ejecutada de acuerdo con el reglamento EPAOPPTS 870.1200 de EE.UU. para determinar la toxicidad dérmica aguda y la irritación cutánea relativa de Microcyn™, según se requiere para el cumplimiento de la norma ISO 10993-11:1996. Microcyn™ se aplicó a la piel intacta de 5 machos y 5 hembras de conejo albino. La dosis de Microcyn™ fue de 5050 mg/kg de peso corporal, aplicada sobre la zona dorsal del tronco. El área de ensayo posteriormente se cubrió con una gasa. Después 24 horas, el área se enjuagó con agua para eliminar todo resto de sustancia de ensayo. Conclusión: las observaciones realizadas durante 14 días después de la aplicación de la sustancia de ensayo no mostraron signos de irritación dérmica en los animales en ningún momento. Todos los animales tenían aspecto normal durante el tiempo del estudio y la necropsia realizada a la finalización del estudio no reveló anomalías observables. En base a este estudio, se concluye que Microcyn™ no genera toxicidad dérmica. [Stillmeadow, Inc., Estados Unidos] Toxicidad por inhalación aguda El ensayo se realizó según el reglamento EPAOPPTS 870.1300 para determinar la toxicidad aguda por inhalación de Microcyn™, según se requiere para el cumplimiento de la norma ISO 10993-11:1996. Microcyn™ se aplicó en forma de aerosol a 5 machos y 5 hembras de rata albina. La dosis de Microcyn™ fue de 2,16 mg / l de aire, aplicada por inhalación nasal durante 4 horas consecutivas. Conclusión: las observaciones realizadas durante 14 días después de la aplicación de la sustancia ensayada no revelaron signos de actividad farmacológica o tóxica. Todos los animales sometidos al ensayo parecían normales durante el periodo del estudio, y la necropsia realizada a la finalización del estudio no revelaron anomalías observables. En base a este estudio, se concluye que Microcyn™ no causa efectos tóxicos por inhalación. [Stillmeadow, Inc., Estados Unidos] Estudio de seguridad y eficacia de tratamiento para heridas cutáneas Se realizó un estudio con 16 ratas para evaluar la tolerabilidad local de Microcyn™ y sus efectos sobre la histopatología del lecho de las heridas en un modelo de cicatrización de heridas dérmicas de espesor completo. Secciones cutáneas teñidas con tricrómico de Masson y secciones teñidas con colágeno de tipo II de heridas quirúrgicas tratadas con Microcyn™ o con solución salina fueron evaluadas por un patólogo veterinario titulado. Las secciones se evaluaron en cuanto a la cantidad de expresión de colágeno tipo 2 como manifestación de la proliferación del tejido conjuntivo, la morfología de los fibroblastos y la formación de colágeno, la presencia de neoepidermis en la sección transversal, la inflamación y el grado de ulceración cutánea. Conclusión: los resultados sugieren que Microcyn™ fue bien tolerado en ratas administrado en las condiciones de este estudio, no hubo lesiones histopatológicas relacionadas con el tratamiento en las secciones de piel, ni en el lado izquierdo ni en el derecho (tratados con Microcyn™ y solución salina respectivamente). No hubo diferencias histopatológicas relevantes entre las heridas tratadas con solución salina y las tratadas con Microcyn™, lo que indica que el tratamiento con Microcyn™- fue bien tolerado. No hubo diferencias significativas en la expresión de colágeno tipo 2 entre las heridas tratadas con solución salina y las tratadas con Microcyn™, incluyendo que Microcyn™ no tiene efecto adverso sobre los fibroblastos o la elaboración de colágeno bajo las condiciones de este estudio. Compatibilidad con materiales Se realizó un ensayo de corrosión de acuerdo con los requisitos de la norma ASTM G60-01. Los materiales ensayados fueron: PVC, acero inoxidable 303, acero inoxidable 316, aluminio HA, titanio, aluminio, poliéster, teflón, polipropileno, caucho natural, HDPE, neopreno, nylon, silicona, policarbonato, poliuretano, polietileno de baja densidad, polisulfona, UHMWP y acero inoxidable 416. Se prepararon fragmentos representativos a partir de los materiales de ensayo y se pusieron en contacto diario durante 30 minutos con Microcyn™ durante 20 días consecutivos. Durante la prueba y al final de la misma, los fragmentos se examinaron en cuanto a su aspecto visual y el peso. Conclusiones: Los metales industriales como el aluminio sin tratar y el acero inoxidable 416 mostraron signos de corrosión después de haber estado en contacto con Microcyn™ durante 20 días. Todos los demás materiales mantuvieron su aspecto visual y su peso a niveles constantes. En base a los resultados de este estudio se determinó que Microcyn™ es compatible y no corrosivo con todos los materiales ensayados, excepto aluminio y acero inoxidable 416. Con estos dos últimos, se concluye que Microcyn™ tiene una corrosividad similar a la del agua [Micromed Laboratories Inc., EE.UU.] Control de calidad Microcyn™ se fabrica conforme a la norma ISO 13485:2003. Todos los lotes de producto fabricados son ensayados en la planta de producción en cuanto a rendimiento antimicrobiano utilizando el método de suspensión contra esporas de Bacillus subtilis (una reducción mínima de 106 es el requisito aplicable). Además, se realiza un control del producto en cuanto a pH, potencial redox y niveles de cloro libre disponibles como parte del control de calidad en el proceso de producción. Pruebas de estabilidad Se ha demostrado que Microcyn™ se mantiene estable y eficaz cuando envejece. La composición química del producto y la eficacia antimicrobiana tras envejecimiento acelerado han sido ensayadas según las directrices pertinentes. En base a estos datos, el producto tiene una vida útil de un (1) año. ™ AGUA ELECTROLIZADA H2O CLORURO DE SODIO CLNA ACIDO HIPOCLOROSO HOCL HIPOCLORITO DE SODIO NaOCL DATOS CLÍNICOS Indicaciones terapéuticas Microdacyn60™ Wound Care es una solución electrolizada indicada para utilizarse en el desbridamiento, la irrigación y la hidratación de las heridas, úlceras, cortes abrasiones y quemaduras de naturaleza tanto aguda como crónica. Microdacyn60™ presenta también un efecto antiinflamatorio directo sobre la herida. Es aplicable en cavidades como la boca, nariz y oído. El Ácido Hipocloroso y el Hipoclorito Sódico son sustancias auxiliares que pueden tener un efecto antimicrobiano local. Al reducir la cantidad de microorganismos y contribuir a mantener un entorno húmedo, permite al organismo realizar su propio proceso de cicatrización. Microdacyn60 Wound Care puede aplicarse de manera generalizada como parte de un tratamiento integral de heridas. MANIPULACIÓN Microdacyn60™ Wound Care es biocompatible para su uso indicado, no irrita la piel, los ojos ni la garganta humanos, y no es un sensibilizador de la piel. No se requieren precauciones de manipulación especiales. Microdacyn60™ Wound Care está preparado para su uso, no es necesario mezclarlo ni diluirlo. Aplique Microdacyn60™ Wound Care directamente a la zona afectada. Utilice Microdacyn60™ en cada cambio de apósito de la herida. Puede aplicarse de manera generalizada como parte de un tratamiento integral de heridas. CONSERVACION Microdacyn60™ Wound Care debe conservarse en su recipiente hermético original a condiciones ambiente. Conservar a temperatura ambiente y prevenir el congelamiento. No es inflamable. Después del uso, el recipiente debe cerrarse de nuevo antes de guardarlo. ESTABILIDAD Hasta que se abre, la estabilidad y la esterilidad de Microdacyn60 Wound Care está garantizada hasta la fecha de caducidad (el año y el mes de caducidad AAAAMM se indican en el frasco). Cuando lo abra, anote la fecha en el frasco, en el apartado “fecha de apertura”. La estabilidad y la esterilidad de la solución no utilizada está garantizada por un máximo de 30 días después de la apertura. ™ MODO DE EMPLEO: Microdacyn60™ Wound Care está preparado para su uso, no es necesario mezclarlo ni diluirlo. Aplique Microdacyn60™ Wound Care directamente a la zona afectada. Utilice Microdacyn60™ en cada cambio de apósito de la herida. La saturación del apósito con Microdacyn60 Wound Care facilitará la retirada del apósito. TRATAMIENTO Empape/sature el apósito con Microdacyn60 Wound Care en cada cambio de apósito, como y cuando sea necesario. La inmersión se permite un máximo de 15 minutos diarios durante el tratamiento inicial. ADVERTENCIAS Y ASPECTOS QUE HAY QUE TENER EN CUENTA Si el precinto a prueba de manipulación indebida está roto, no utilice el producto. Utilice siempre solución nueva, no reutilice el producto. Uso externo, no apto para inyección o consumo. No utilice este producto si es alérgico a sus ingredientes. No utilice Microdacyn60 Wound Care junto con otras soluciones tópicas sin el asesoramiento de un profesional sanitario. En caso de heridas graves, profundas, o si le preocupa algún otro aspecto del tratamiento consulte con un profesional sanitario. Si presenta signos de infección ( aumento de dolor, hemorragia, pus etc) o si la herida no muestra signos de cicatrización, consulte a un profesional sanitario. Si presenta una reacción adversa consulte con el profesional sanitario. Además de Microdacyn60 Wound Care en algunos casos puede ser necesario el uso de antibióticos sistémicos o locales. Mantenga fuera del alcance y de la vista de los niños. ELIMINACION La solución no es tóxica y es segura para el medio ambiente. No se requieren condiciones de eliminación especiales. ESTERILIDAD La esterilidad depende de los conservantes que contiene el líquido. MICRODACYN60™ WOUND CARE SOLUCION PRESENTACIONES ! ™ AGUA ELECTROLIZADA H2O CLORURO DE SODIO CLNA. SODIO MAGNESIO FLUORSILICATO ACIDO HIPOCLOROSO HOCL HIPOCLORITO DE SODIO NaOCL DATOS CLÍNICOS Indicaciones terapéuticas Microdacyn60™ Wound Care es un hidrogel electrolizado indicado para utilizarse en el desbridamiento y la hidratación de las heridas, úlceras, cortes abrasiones y quemaduras de naturaleza tanto aguda como crónica. Microdacyn60™ presenta también un efecto antiinflamatorio directo sobre la herida. Es aplicable en cavidades como la boca, nariz y oído. El Ácido Hipocloroso y el Hipoclorito Sódico son sustancias auxiliares que pueden tener un efecto antimicrobiano local. Al reducir la cantidad de microorganismos y contribuir a mantener un entorno húmedo, permite al organismo realizar su propio proceso de cicatrización. Microdacyn60 Wound Care puede aplicarse de manera generalizada como parte de un tratamiento integral de heridas. MANIPULACIÓN Microdacyn60™ hidrogel es biocompatible para su uso indicado, no irrita la piel, los ojos ni la garganta humanos, y no es un sensibilizador de la piel. No se requieren precauciones de manipulación especiales. Microdacyn60™ hidrogel está preparado para su uso, no es necesario mezclarlo ni diluirlo. Aplique Microdacyn60™ hidrogel directamente a la zona afectada. Utilice Microdacyn60™ hidrogel en cada cambio de apósito de la herida. Puede aplicarse de manera generalizada como parte de un tratamiento integral de heridas. CONSERVACION Microdacyn60™ hidrogel debe conservarse en su recipiente hermético original a condiciones ambiente. Conservar a temperatura ambiente y prevenir el congelamiento. No es inflamable. Después del uso, el recipiente debe cerrarse de nuevo antes de guardarlo. ESTABILIDAD Hasta que se abre, la estabilidad y la esterilidad de Microdacyn60 hidrogel está garantizada hasta la fecha de caducidad (el año y el mes de caducidad AAAAMM se indican en el frasco). Cuando lo abra, anote la fecha en el frasco, en el apartado “fecha de apertura”. La estabilidad y la esterilidad de la solución no utilizada está garantizada por un máximo de 30 días después de la apertura. ™ C.N.172398.7 MODO DE EMPLEO: Microdacyn60™ hidrogel está preparado para su uso, no es necesario mezclarlo ni diluirlo. Aplique Microdacyn60™ hidrogel directamente a la zona afectada. Utilice Microdacyn60™ en cada cambio de apósito de la herida. La saturación del apósito con Microdacyn60 hidrogel facilitará la retirada del apósito. TRATAMIENTO Limpie y seque el área. Aplique inmediatamente una capa delgada de Microdacyn60 hidrogel en la zona afectada. Utilice Microdacyn60 hidrogel una o dos veces al día y en cada cambio de apósito, como y cuando sea necesario. Cubra con un apósito o vendaje. ADVERTENCIAS Y ASPECTOS QUE HAY QUE TENER EN CUENTA Si el precinto a prueba de manipulación indebida está roto, no utilice el producto. Utilice siempre solución nueva, no reutilice el producto. Uso externo, no apto para inyección o consumo. No utilice este producto si es alérgico a sus ingredientes. No utilice Microdacyn60 hidrogel junto con otras soluciones tópicas sin el asesoramiento de un profesional sanitario. En caso de heridas graves, profundas, o si le preocupa algún otro aspecto del tratamiento consulte con un profesional sanitario. Si presenta signos de infección (aumento de dolor, hemorragia, pus etc) o si la herida no muestra signos de cicatrización, consulte a un profesional sanitario. Además de Microdacyn60 hidrogel en algunos casos puede ser necesario eln uso de antibióticos sistémicos o locales. Si presenta una reacción adversa consulte con el profesional sanitario. Mantenga fuera del alcance y de la vista de los niños. ELIMINACION Microdacyn60 hidrogel no es tóxico y es segura para el medio ambiente. No se requieren condiciones de eliminación especiales. ESTERILIDAD La esterilidad depende de los conservantes que contiene el líquido. Microcyn Technology ¿QUE ES? DISRRUPTIVE TECHNOLOGY La Tecnología Microcyn® se fabrica en Estados Unidos, está registrada y regulada por la compañía Oculus Innovative Sciences, Inc. La tecnología patentada Microcyn® mimetiza la misma composición en oxigeno y cloro que fabrican los neutrófilos del sistema inmunitario del cuerpo humano. Los neutrófilos son las células blancas más abundantes en los humanos y forman parte integral del sistema inmunitario. Los productos basados en la tecnología Microcyn® son tan seguros como la solución salina, biocompatibles, y estables, y han demostrado en una amplia variedad de investigaciones y estudios clínicos su capacidad de matar una amplia gama de patógenos, incluyendo cepas resistentes a algunos antibióticos ( incluyendo MRSA y VRE), virus, hongos y esporas, a la vez que reducen la inflamación y incrementan el aporte sanguíneo a los tejidos, todo ello con un 100% de seguridad y no toxicidad para los tejidos y células humanas. Los productos basados en la tecnología Microcyn® pueden ser utilizados en la nariz, ojos y boca. La tecnología Microcyn® tiene aprobaciones legales gubernamentales globales incluyendo múltiples aprobaciones de la F.D.A. en EEUU, múltiples aprobaciones de la CE, y varias aprobaciones de la E.P. A. en Estados Unidos; además de aprobaciones de los gobiernos de Canadá, México, Emiratos Árabes, Kuwait, Arabia Saudí, Iraq, Jordania, y varios países Norte Africanos. Es esta seguridad, efectividad, y no toxicidad, junto con su amplia eficacia en el mundo de uso médico como desinfectante, germicida, bactericida, tuberculicida, esporicida y virucida la que le ha proporcionado su revolucionario reconocimiento mundial. Microdacyn® es un producto de alto desarrollo científico ampliamente utilizado a nivel hospitalario, y también como desinfectante en cualquier superficie dura inanimada no porosa. Se fabrica en Estados Unidos por una compañía registrada y regulada por del Departamento de Salud y Servicios Humanos Administración de Alimentos y Medicamentos (FDA) , Oculus Innovative Sciences , la cual ha obtenido la certificación ISO 13485:2003 ,y la TNO Certification BV, que otorga un instituto independiente reconocido por el Consejo Holandés de Acreditación. MECANISMO DE ACCIÓN. COMO ACTUA MICRODACYN® En el mundo de la infección, los científicos han desarrollado una tecnología global que mimetiza eficazmente la acción química del mecanismo de defensa del cuerpo humano. La Tecnología Microcyn® es tan segura como la solución salina, sin ninguna toxicidad para el cuerpo humano, pero puede erradicar bacterias y virus de cualquier tipo, en cuestión de segundos. Esta tecnología química basada en la oxidación y la electrólisis es la plataforma básica de eficacia que se encuentra detrás de todos los productos de la gama Microdacyn® . A partir de agua purificada y solución salina, cuando la corriente eléctrica pasa a través del agua purificada, los átomos de hidrógeno y oxígeno de la molécula de agua se dividen en iones de hidrógeno con carga positiva y iones hidróxido de carga negativa. Dado que estos iones ya no son estables, buscan hacia fuera otros electrones que puedan estabilizarlos. La solución resultante, súper oxidada, es una “asesina” potente de los organismos unicelulares como las bacterias, virus y otros microorganismos que son potencialmente letales para los seres humanos y el medio ambiente. Cuando Microdacyn® entra en contacto con microorganismos unicelulares, los iones en alta concentración, corren hacia la pared celular bacteriana, donde la concentración de iones es menor y por ósmosis rompen la pared celular (bacteriana o viral) hidratandola hasta que la célula estalla y explota eliminando así cualquier posibilidad de mutación. El avance tecnológico de Oculus Innovative Sciences es que ha sido capaz de embotellar estos iones inestables, el oxígeno y el cloro que forma de manera natural, manteniendo el ph de la solución neutro, por lo que es inofensivo para los tejidos sanos. Esta es la misma forma de Cloro que los neutrófilos (células blancas de la sangre) crean naturalmente en el cuerpo durante el metabolismo para promover el sistema inmunológico. En nuestro organismo contamos con varios litros de esta forma de cloro, el cual forma parte esencial de nuestro sistema inmunitario y acelera la respuesta de curación ante una infección. De esta forma la tecnología Microcyn® ha adquirido su revolucionario posicionamiento como líder global en el tratamiento de la infección, y el sobrenombre de “El sistema inmunológico humano dentro de una botella”. Esta solución tiene una estabilidad mínima de 24 meses. El diagrama inferior muestra como los iones inestables y altamente concentrados en la solución Microcyn, situada fuera de una bacteria o virus quiere ir dentro de la célula donde hay una concentración inferior de iones. Los iones tratan de re-equilibrarse a si mismos de esta manera. Cuando la membrana o la pared celular de la bacteria o virus es dañada, los iones entran en el microbio y interrumpen su acción. El organismo, básicamente, explota y no hay manera de que se produzca una mutación o desarrolle una resistencia al microorganismo. Esta elegante, sencilla y natural tecnología está ahora a su alcance de la mano de los productos Microdacyn distribuidos en España por SDOMEDICAL. http://www.youtube.com/watch?v=HTK4Lb3mzUY International Immunopharmacology 7 (2007) 1013 – 1024 www.elsevier.com/locate/intimp Super-oxidized solution inhibits IgE-antigen-induced degranulation and cytokine release in mast cells ☆ J. Medina-Tamayo a , E. Sánchez-Miranda a , H. Balleza-Tapia a , X. Ambriz a , M.E. Cid a , D. González-Espinosa b , A.A. Gutiérrez b , C. González-Espinosa a,⁎ a Pharmacobiology Department, Cinvestav, South Campus, Mexico City, Mexico Cell Therapy Unit, National Institute of Rehabilitation, Mexico City, Mexico b Received 13 July 2006; received in revised form 2 March 2007; accepted 19 March 2007 Abstract Activation of the high affinity IgE receptor (FcεRI) through IgE-antigen complexes induces mast cell degranulation, synthesis of lipid mediators and cytokine production. These effects are involved in Type I hypersensitivity reactions and controlling them has been the main objective of many anti-allergic therapies. Here we report that pretreatment of murine bone marrow derived mast cells (BMMC) with super-oxidized solution (SOS) inhibits FcεRI dependent-β hexosaminidase and cytokine release. This effect is exerted without altering total protein tyrosine phosphorylation, MAPK activation, cytokine mRNA accumulation or calcium mobilization after FcεRI triggering. Our data suggest that this neutral pH-SOS acts like a mast cell-membrane stabilizer inhibiting the cell machinery for granule secretion without altering the signal transduction pathways induced by IgE-antigen receptor crosslinking. © 2007 Elsevier B.V. All rights reserved. Keywords: Mast cell; Degranulation; Cytokine production; Allergy; IgE receptor; Super-oxidized solution 1. Introduction Abbreviations: SOS, super-oxidized solution; SCF, stem cell factor; MCs, mast cells; BMMCs, bone marrow-derived mast cells; CCL3/MIP1-α, macrophage inhibitory protein 1 alpha; IgE, Immunoglobulin E; FcεRI, High affinity-IgE receptor; NFAT, nuclear factor of activated T cells; 7-AAD, 7 aminoactinomycin; PMA, phorbol 12 myristate 13 acetate; TNF-α, tumor necrosis factor alpha; DNP-HSA, dinitrophenol-human serum albumin. ☆ AA Gutiérrez conducted the research in Mexico and had become a paid employee of Oculus IS at the time of drafting this paper. ⁎ Corresponding author. Pharmacobiology Department, Cinvestav, South Campus Calzada de los Tenorios 235 Col. Granjas Coapa, 14330 Mexico City 14330 Mexico. Tel.: +52 55 5061 2800x2875; fax: +52 55 5603 3957. E-mail address: [email protected] (C. González-Espinosa). 1567-5769/$ - see front matter © 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.intimp.2007.03.005 Mast cells (MCs) have long been recognized as effector cells of hypersensitivity Type I reactions and protective responses against parasites [1,2]. These cells begin their differentiation in the bone marrow from CD34+ hematopoietic progenitors and migrate to the bloodstream as committed cells. In peripheral tissues [3], they acquire their distinctive phenotype under the influence of stem cell factor (SCF), interleukin (IL)-3 and various other locally-produced mediators [4]. Although MCs do not represent a single homogenous, functional-cell population, there is one characteristic that unites them all with basophils which is the possession of high affinity membrane receptors for immunoglobulin 1014 J. Medina-Tamayo et al. / International Immunopharmacology 7 (2007) 1013–1024 (Ig) E (FcεRI). When these receptors are activated by IgE-antigen interactions, the FcεRI signal transduction pathway can lead to 1) histamine and protease-containing granule release, 2) lipid-derived inflammatory mediator production and 3) cytokine synthesis [5]. The secretion of these mediators is responsible for early and late phases of allergic responses [6]. Depending on the anatomic site where activated mast cells are located, the immediate secreted molecules can play a regulatory role of smooth muscle contraction, immune cell chemotaxis, endothelial cell layer permeability and mucus production. Mast cell activation, therefore, plays an important role in a wide array of inflammatory disorders [7]. Investigation into the mechanisms that control histamine release by mast cells has constituted an important milestone in immunopharmacology [8]. Based on this knowledge, diverse pharmacological agents have been developed to prevent or ameliorate the activation and secretion of mast cells. Among them, β2 adrenoceptor stimulants, methylxantines, glucocorticoids, antihistamines and mast cell stabilizers (chromones) are the main classes of drugs widely used for allergic diseases [9]. These agents are frequently utilized as tools for basic research too, and have helped to better understand mast cell physiology and the FcεRI signal transduction system. Pharmacological stabilization of mast cells is currently used for the therapy of asthma [10,11], rhinitis [12], atopic dermatitis [13], seasonal conjunctivitis [14] and even chronic leg ulcers [15]. This strategy has also been applied to abrogate ocular allergic inflammation [16], prevent adhesion formation after pelvic surgery [17], and diminish inflammation and hyperalgesia induced by nerve injury [18]. Sodium cromoglycate and nedocromyl are the most commonly used mast cell stabilizers for inhibiting cytokine release from bronchial mast cells [19]. When inhaled several times a day, cromoglycate will inhibit both, immediate and late asthmatic responses elicited by antigen or exercise [11]. Given the intimate relationship between mast cells and multiple inflammatory diseases, stabilization of mast cell activity would be expected to be therapeutically effective. Interestingly, there are some reports showing beneficial effects of super-oxidized solutions in disorders related to mast cell activation, suggesting that these solutions might have a direct modulator activity on this type of cells [20–22]. Super-oxidized solutions (also known as superoxidized water, SOW) are produced by the electrolysis of purified water and sodium chloride. Water is broken down into oxygen, ozone and other unstable oxidized species. However, the main active chemical species generated during this process are hypochlorite and hypochlorous species. Despite solutions commercially available are all produced by electrolysis and they differ in the concentration of active components and pH [23]. Therefore, their biocidal activity and toxicity profiles also vary depending on these two characteristics. Microbicidal activity has been shown against bacteria, viruses, fungi and spores in vitro [24,25]. In a rat-burn model, for example, the researchers applied SOS directly to the infected lesion with Pseudomonas and were able to efficiently reduce blood levels of endotoxin, systemic effects of septicemia and mortality rates [21]. Similarly in humans, skin infections and ulcers [26], inflammatory skin disorders (e.g. pemphigus, psoriasis) [27], burns [28] and peritonitis [20,29] have all improved their outcomes with the use of SOS treatment. But whether these better outcomes are related to the antiseptic efficacy of the SOS, or to a synergystic anti-inflammatory effect, is unknown [29,30]. To the best of our knowledge, there is no information on the direct effects that a SOS might exert on the inflammatory process itself or in the process of mast cell activation. It is well established that studies on mast cell physiology are useful to identify possible therapeutic targets for allergic/inflammatory diseases. Murine bone marrowderived mast cells (BMMCs) are one of the most common cells used for this purpose. These cells are generated from murine bone marrow cultured in the presence of IL-3 (for the generation of mucosal-type mast cells) or IL-3 and SCF (for the development of connective tissue-type mast cells) [31]. Mucosal-type BMMCs are particularly useful to analyze distinct molecular events leading to signal transduction, degranulation, leucotriene production, and cytokine mRNA expression and secretion [2,3]. We therefore explored the effects of a pH neutral-SOS on the activation and secretion processes of mucosaltype BMMCs after FcεRI triggering using IgE-antigen. For this purpose, cells were incubated in serial dilutions of a pH neutral-SOS for a short period of time before being activated. The SOS treatment was not toxic but blocked both, IgE/antigen- and calcium ionophoreinduced mast cell degranulation in a concentration dependent manner. SOS pretreatment also significantly inhibited the release of TNF-α, CCL-3, IL-13 and IL-6 from activated mast cells. Yet, the treatment did not interfere with early stages of the signal transduction mechanism of the FcεRI receptor, since protein tyrosine phosphorylation, MAPK activation, calcium mobilization and cytokine mRNA production were observed after receptor crosslinking. Altogether these data suggest that SOS pretreatment is able to partially inhibit the secretion of mediators of both, acute and late induced responses in mast cells. The exact targets of SOS in mast cells are unknown but clearly deserve further evaluation. J. Medina-Tamayo et al. / International Immunopharmacology 7 (2007) 1013–1024 2. Materials and methods Culture media, bovine fetal serum, essential amino acids, and sodium pyruvate solutions for cell growth and differentiation were from Invitrogen (Carlsbad, California). Antibodies used for western blot analysis included anti-p38 (Cat. # SC-535), ERK2 (Cat. # SC-154 ), and p-ERK2 (Cat. # 7976 ) from Santa Cruz Biotechnology (California, USA), p-p38 (Cat. # 9211S) from Cell Signaling (Beverly, MA), and anti-pTyr, clone 4G10, (Cat. # 05-321) from Upstate Biotechnology (Lake Placid, NY). Monoclonal anti-DNP IgE (clone SPE-7), calcium ionophore A23187, PMA and reagents for buffer preparation, electrophoresis and calcium mobilization assays were purchased from Sigma Chemical Co. (St. Louis MO). Antigen stimulation of the cells was performed adding distinct amounts of dinitrophenol-human serum albumin (DNP-HSA, Sigma Chemical Co. Cat # A-6661) to overnight IgE sensitized BMMC. The SOS used for this study was a stable, neutral pH preparation commercially available in Mexico (Microcyn®, Oculus Innovative Sciences, CA, USA). 2.1. Mice and BMMC culture Bone marrow derived mast cells (BMMC) were isolated from the tibia of 4–8 weeks old mice (strain 129S1/SvImJ), from Jackson Laboratories (Maine, USA, stock # 002448). Briefly, total bone marrow was isolated and cultured in RPMI media containing IL-3 (20 ng/mL) and 10% Fetal Bovine Serum (FBS) in order to differentiate mast cells, as previously described [31]. After 4–6 weeks of culture, between 97 to 99% of the cells were positive to FcεRI staining as measured by a standard flow cytometry assay [32]. 2.2. SOS treatments A pH neutral, super-oxidized solution (Microcyn®) that contains a small amount of free available chlorine (b80 ppm) in the form of hypochlorous acid, sodium hypochlorite and sodium chloride was used for this study. This solution is 1015 produced through the electrolysis of purified water containing limited amounts of chloride ion in a unique multi-chamber cell (Oculus Innovative Sciences). This stable, super-oxidized solution was mixed with concentrated Tyrode's/BSA buffer (see degranulation assay section) in order to obtain solutions with different percentages of SOS (v/v) without significantly altering the osmolarity or pH of Tyrode's/BSA buffer. Sensitized cells (see Degranulation section) were preincubated with Tyrode's/BSA buffer alone, or with 10% to 50% SOS containing-Tyrode's/BSA buffers, at 37 °C for distinct periods of time to test viability or during 15 min to test physiological activation. After SOS pretreatments, cells were collected and resuspended in fresh SOS free-Tyrode's/BSA in order to be stimulated with antigen or with a mixture of calcium ionophore and PMA, as a positive control for cell activation. 2.3. Viability assays Cell viability analysis of SOS exposed mast cells was carried out by three independent methodologies. Propidium iodide incorporation, annexin V exposition and 7AAD positive cells were measured as follows. Mature BMMCs (5 × 105) were centrifuged at room temperature during 5 min at 800 ×g. After removal of the supernatant, cells were carefully resuspended in 0.5 mL of the different SOS dilutions in Tyrode's/ BSA (see previous section) and kept at 37 °C, 5% CO2. After different times of exposure, treatment was stopped by addition of 0.5 mL of the complete cell culture media. Cells were then pooled by centrifugation during 5 min at 800 ×g, the resultant pellets were disaggregated and cells were resuspended and stained with 7-aminoactinomycin D (7-AAD) according to the manufacturer's instructions (7-AAD; BD., Palo Alto, CA. USA). For long-term cytotoxicity showed in Table 1, SOS dilutions were made in complete RPMI media (see cell culture section). Osmolarity was not significantly affected by dilution process, since mean osmolarity of complete medium containing 50% SOS of three different lots was 284.9 ± 1.05, 296.34 ± 0.78, 317.82 ± 0.48, determined using an Advanced 3D3 Osmometer Table 1 Effect of 50% SOS or 0.0075% sodium hypochlorite on mast cell viability depending on exposure time Cell viability Treatment determination method t=0 t = 5 min t = 15 min t = 30 min t = 60 min t = 120 min t = 4 h 7AAD 7AAD 7AAD Propidium iodide Propidium iodide Propidium iodide Annexin C Annexin C Annexin C 99 ± 1% 99 ± 2% 99 ± 5% 99 ± 0.6% 98 ± 1.4% 99 ± 2.3% 99 ± 0.03% 99 ± 0.08% 99 ± 0.16% 99 ± 2% 99 ± 2% 99 ± 8% 99 ± 1.4% 99 ± 2.0% 99 ± 3.2% 99 ± 0.01% 99 ± 0.05% 99 ± 0.12% 99 ± 1% 99 ± 3% 99 ± 6% 99 ± 0.3% 99 ± 0.5% 98 ± 0.3% 99 ± 0.07% 99 ± 0.09% 95 ± 1.5% ⁎ 99 ± 4% 99 ± 5% 95 ± 9% 98 ± 1.4% 97 ± 2.1% 88 ± 2.6% ⁎ 99 ± 0.01% 98 ± 0.03% 86 ± 2.6% ⁎ 99 ± 5% 97 ± 5% 90 ± 12% ⁎ 98 ± 0.5% 96 ± 0.5% ⁎ 86 ± 4%⁎ 99 ± 0.02% 98 ± 0.03% 87 ± 3.6% ⁎ 99 ± 5% 95 ± 4% ⁎ 85 ± 12% ⁎ 98 ± 0.7% 95 ± 0.7% ⁎ 89 ± 6% ⁎ 99 ± 0.03% 98 ± 0.09% 80 ± 14% ⁎ Buffer 50% SOW 0.0075% sodium hypochlorite Buffer 50% SOW 0.0075% sodium hypochlorite Buffer 50% SOW 0.0075% sodium hypochlorite 99 ± 6% 95 ± 11% ⁎ 66 ± 12% ⁎ 96 ± 2% ⁎ 93 ± 4% ⁎ 54 ± 19% ⁎ 99 ± 0.10% 99 ± 0.07% 47 ± 16% ⁎ Time-dependent cytotoxicity of 50% SOS or 0.0075% sodium hypochlorite on mature BMMC's. Cell viability was determined using three different reagents as described in Materials and methods section. Results are expressed as percentage of viable cells in each condition. No difference was noticed between buffer treated or non-treated BMMCs. ⁎ denote significant differences from cells exposed to Tyrode's BSA/culture media mixture. 1016 J. Medina-Tamayo et al. / International Immunopharmacology 7 (2007) 1013–1024 from John Morris Scientific. As a control for long-term cytotoxicity, different dilutions of a Voluette Analytical Chlorine Standard (HACH Company, Loveland, CO, USA) were prepared in complete media and cells were pretreated during distinct periods of time in order to test viability. Given the presence of NaCl in complete media, after chlorine addition, sodium hypoclorite (NaOCl) was produced and the amount of NaOCl in the preparation was measured by using a molar extinction coefficient of 350 M− 1 cm− 1 at 290 nm at pH 12 [33]. The ratio of necrosis and apoptosis were determined simultaneously in the cells using annexin V-FITC and propidium iodide according to the manufacturer's protocol (Annexin-VFluos Staining kit, ROCHE, NJ, USA). Positive stain cells were analyzed on a FACScalibur flow cytometer (BD., Palo Alto, CA, USA). All experiments were conducted in triplicate. 2.4. Statistical analysis Statistical analysis was performed by the t-test for Dependent Samples. p b 0.05 was considered significant. The Tukey honest significant difference test was used for the analysis of variance (ANOVA) between groups. The Statistica™ version 6.00 software was used for these analyses. 2.5. Degranulation assays Mature BMMCs were sensitized by overnight incubation in media containing 300 ng/mL of monoclonal anti-DNP IgE (clone SPE7). The following morning, cells were washed and resuspended in SOS-free or SOS-containing Tyrode's/BSA buffer (20 mM HEPES pH7.4, 135 mM NaCl, 5 mM KCl, 1.8 mM CaCl2, 1 mM MgCl2, 5.6 mM glucose, and 0.05% bovine serum albumin) and kept at 37 °C during 15 min. Two million cells in 1 mL buffer were stimulated with different concentrations of DNP-HSA (antigen) or calcium ionophore/ PMA at 37 °C during 30 min for degranulation, or during 1 h for total RNA isolation. After the indicated times, cells were collected by centrifugation and supernatants were used for βhexosaminidase activity determination and pellets for total RNA isolation, as previously described [34]. 2.6. RNA isolation, RT-PCR and RPA assays Pellets from non-stimulated and stimulated cells were used for total RNA isolation. Briefly, the cell pellet obtained in degranulation assays was resuspended in 1 mL of Trizol following manufacturer's instructions (Invitrogen, Carlsbad, CA, USA). RNA concentration was calculated by absorbance at 260 nm and cDNA synthesis was performed using a first strand DNA synthesis kit (Invitrogen, cat # 12371-019). PCR amplification of cDNAs was conducted in the presence of specific primers for distinct cytokines or GAPDH genes, following the manufacturer's instructions (Mouse Inflammatory Cytokine Set2ms, BioSource™; Invitrogen Carlsbad, CA, USA). RT-PCR products were separated in TBE-prepared 2% agarose gels and analyzed in a UVP image analysis system (Upland, CA, USA). Ribonuclease I Protection Assays (RPA) were conducted using specific multiprobe RPA sets labeled with 33P, following the instructions provided in the kits (In vitro transcription and RPA kits, BD Pharmingen, cat. # 556850 and # 556134, respectively). Protected fragments were separated on denaturing polyacrylamide gels, which were dried and exposed to Kodak Biomax film at − 80 °C. Representative results of at least four independent experiments are shown. 2.7. Western blotting Ten million sensitized BMMCs were pretreated with SOS free-or SOS containing-Tyrode's/BSA buffers, during 15 min at 37°. After this time, cells were collected and resuspended in 250 μl of fresh Tyrode's/BSA buffer. BMMCs were then stimulated by the addition of 30 ng/mL DNP-HSA antigen for 10 min at 37 °C and the stimulus further stopped by adding 250 μl of 2× Laemmli buffer to each tube. Samples were boiled during 5 min and 30 μl of the sample was separated on 10% Tris–glycine-SDS polyacrylamide gels. Proteins were transferred to PVDF membranes using standard procedures. Membranes were blocked 1 h at room temperature in 4% nonfat milk. First antibody incubation was performed overnight at 4 °C using recommended dilutions. The following day, antibody excess was eliminated by washing the membrane several times in TTBS buffer (50 mM Tris; 150 mM NaCl; 0.1% Tween 20). Secondary antibody incubations were performed 1 h at RT using 1:10,000 dilutions of HRPconjugated specific antibodies. Chemiluminiscence was measured with a SuperSignal detection kit (Roche, NJ, USA). 2.8. Cytokine secretion The concentration of murine cytokines in supernatants from treated and non-treated, stimulated and naive cells, was evaluated by standard ELISA kits as recommended by the manufacturer (Biosource-Invitrogen, Carlsbad, CA). Evaluated cytokines included TNFα (cat. KMC3011), CCL-3 (MIP1α) (cat. KMC2201), IL-6 (cat. KMC0061) and IL-3 (cat. KMC0031). Supernatants were obtained as follows: 40 million of sensitized BMMC were preincubated with SOS free-or SOS containingTyrode's/BSA buffers for 15 min at 37 °C. After this time, cells were collected and resuspended in 10 mL of fresh cell culture media supplemented with a protease inhibitor cocktail (Roche, NJ, USA). Cells were stimulated with 30 ng/ml of DNP-HSA at 37 °C for different periods of time, at the end of which 1 mL of media was collected from the supernatant and frozen down at −80 °C for cytokine determination. Supernatants of nonstimulated cells were used as negative controls. Between 20 to 60 μl of the supernatant was used for ELISA cytokine detection, depending on the specific cytokine tested. 2.9. Calcium mobilization and NFAT transcription factor activity Calcium mobilization was measured as described elsewhere [35]. In brief, sensitized BMMC were loaded with J. Medina-Tamayo et al. / International Immunopharmacology 7 (2007) 1013–1024 1017 5 mM Fura-2AM dissolved in Tyrode's/BSA at 37 °C for 30 min. After loading, cells were pre-incubated with SOS free or SOS containing Tyrode's/BSA buffers, during 15 min at 37°. Cells were collected and resuspended in cold Tyrode's/ BSA until use. For stimulation, 4 million cells were placed in a 2 mL cuvette of a Jobin Yvon-Fluorolog Spectrofluorometer and the ratio of fluorescence at 340/380 nm was recorded during 100 s before adding antigen (30 ng/mL) or Calcium ionophore/PMA (100 nM/1 μM) to the cells. Ratio of fluorescence was recorded for an additional 300 s and calibration was performed by the addition of Triton X-100 (for maximal fluorescence) and 10 mM EGTA (for minimal fluorescence) to the cuvette. NFAT transcription factor activity was determined using a commercial TransAM Kit for NFATc1 activation, from Active Motif (Cat # 40269). Briefly, 10 million mast cells were sensitized and pre-incubated with buffer, 25% SOS or 50% SOS during 15 min. After this time, cells were washed once and resuspended in Tyrode's/BSA buffer, prior to being activated during 15 min using 9 or 27 ng/ml of DNP-HSA at 37 °C. Total nuclear extracts were performed and NFAT activity was tested following instructions provided by the manufacturer. Data are presented as optical density values at 450 nm in non-treated and treated BMMCs. 3. Results 3.1. SOS pretreatment does not affect BMMC viability The neutral SOS herein tested is a solution with broad antimicrobial activity commonly used in wound care treatment. Yet, its direct effects on cells mediating allergic or inflammatory processes, like mast cells, had never been explored. Thus, the cytotoxicity of this SOS was first evaluated in IgE-sensitized BMMCs treated with Tyrode's/ BSA buffer containing distinct concentrations of SOS (i.e. 1% to 50%, v/v). After incubation for 15 min at 37 °C, cells were processed to simultaneously evaluate cell viability, necrosis and apoptosis by flow cytometry. Fig. 1 shows the concentration-response curves of BMMCs cultures exposed to different dilutions of SOS. Cell viability was analyzed by exclusion of 7AAD. Of all SOS dilutions tested, only 50%-SOS induced a statistically significant reduction in BMMCs' viability in comparison to the control group ( p b 0.05). Despite this, the cell viability of BMMCs pretreated with 50% SOS was ≥ 95%. Apparently, 50% SOS induced cell death through necrosis because ≥ 3% of the cells incorporated propidium iodide in the flow cytometry analysis (Fig. 1B). This percentage, although low, was also significantly higher than the rest of the groups ( p b 0.05). Apoptosis, on the contrary, does not seem to be the mechanism by which 50% SOS induces cell death. In fact, the percentages of cells exposing Annexin-V in the cellular surface ranged from 0.4% to 0.9% in all groups (Fig. 1C). These results show that mild cytotoxicity in BMMCs is only induced by 50%-SOS in vitro. Instead, lower SOS concentrations (i.e. 1% to 25%-SOS) do not seem to affect BMMCs cell membrane permeability as measured in these assays. Fig. 1. Mast cell treatment with SOS (SOW) does not affect cellular viability. BMMCs (5 × 105) were incubated at 37 °C for 15 min in Tyrode's/BSA alone, or in the same buffer containing different dilutions of neutral pH SOS (v/v). After this time, cells were analyzed by flow cytometry looking for 7AAD (A), propidium iodide (B) and Annexin (C) positive cells. SOS solutions were prepared as described in the Materials and methods section. Results are expressed as the mean from three independent experiments ±SD. To establish the sustained effect of SOS on mast cellcytokine release, a time-course experiment was first conducted to show that cell viability was not significantly modified for several hours after a 15 min exposure to 50% SOS. As described in previous sections, cells were collected and incubated for 15 min in complete media containing Tyrode's/BSA buffer, 50% SOS or 0.0075% sodium hypochlorite. Care was taken to keep the osmolarity of the media in the range of 275 to 317 mOsm/L in all testing conditions. At the end of each 1018 J. Medina-Tamayo et al. / International Immunopharmacology 7 (2007) 1013–1024 exposure. These experiments show that hypochlorite is far more toxic than SOS to mast cells, even when both solutions contained a similar concentration of free available chlorine. 3.2. SOS pretreatment of BMMC inhibits FcεRI-induced degranulation Mast cell degranulation initiates the early phase of allergic responses. In order to test the effect of SOS treatment on the degranulation of mast cells, IgE-antigen-induced β-hexosaminidase release was measured in BMMCs pretreated with either, SOS free-or SOS containing-Tyrode's/BSA buffer for 15 min at 37°. Pretreatment of the cells with 25% and 50% SOS inhibited antigen-induced mast cell degranulation by 75–80% (Fig. 2A). This effect was also observed when cell preincubation Fig. 2. SOS (SOW) pretreatment of mast cells inhibits antigen and calcium ionophore/PMA-induced degranulation. (A) Sensitized mature BMMCs were collected from culture media and resuspended in normal Tyrode's/BSA buffer or in the same buffer containing different dilutions of neutral pH SOS (v/v) at 37 °C for 15 min. After this time, cells were collected, washed once and resuspended in SOS-free Tyrode's/BSA (37°). Then they were stimulated with different concentrations of DNP-HSA (antigen) during 30 min. After this time, cells were collected and β-hexosaminidase activity was determined in the supernatant. A pellet of non-stimulated cells was lysed with 0.5% Triton and used as the positive control (i.e. 100% enzymatic activity). Results are expressed as percent of the total βhexosaminidase activity. (B) Degranulation stimulated by antigen or A23187/PMA was inhibited by SOS containing Tyrode's/BSA. Cells were treated as in panel A but β-hexosaminidase release was measured after antigen (30 ng/ml) or calcium ionophore/PMA (100 nM/1 μM) stimulation. The graph shows the mean degranulation values obtained from 5 independent experiments ±SE. incubation time, new fresh RPMI media was added to each tube and cell viability was determined. Propidium iodide (PI) incorporation, annexin V exposition and 7AAD staining at different times of incubation at 37 °C were the techniques used for cell viability determination. As shown in Table 1, viability in 50%SOS-treated cells dropped to 95% after 2 and 4 h post exposure as evaluated by 7AAD. Instead, 0.0075% hypochlorite significantly reduced mast cell viability down to 90%, 85% and 66% at 1, 2 and 4 h post exposure, respectively. Another difference was that hypochlorite induced cell death through both, necrosis and apoptosis, as demonstrated by increasing PI incorporation and annexin V exposition from minute 30 post- Fig. 3. SOS (SOW) pretreatment of mast cells does not prevent antigendependent protein tyrosine phosphorylation and MAPK activation. Mature, sensitized BMMCs were collected from culture media and resuspended in SOS free- or SOS containing-Tyrode's/BSA buffers at 37 °C during 15 min. After this time, cells were washed once in SOS-free Tyrode's/BSA (37 °C) and stimulated with 30 ng/ml DNP-HSA during 3 min. Reaction was stopped by addition of boiling 2× Laemmli buffer and total proteins were analyzed using phospho-tyrosine (A), or phospho ERK and phospho p38 (B) specific antibodies. A representative image of three independent experiments is shown. When SOW was added alone, the final concentration was 50%. J. Medina-Tamayo et al. / International Immunopharmacology 7 (2007) 1013–1024 1019 was conducted in 1% SOS. Of note, the inhibition of antigeninduced mast cell degranulation was not avoided by addition of increasing antigen doses. Furthermore, the blockage of β-hexosaminidase release induced by other stimuli, like calcium ionophore and PMA, was also inhibited by 90 ± 10% in cells pretreated with 50% SOS (Fig. 2B). These results suggest that SOS blockage of β hexosaminidase release is a non-specific effect on the secretion process. 3.3. SOS pretreatment does not affect FcεRI-induced tyrosine phosphorylation and MAPK activation FcεRI receptor crosslinking rapidly induces tyrosine phosphorylation of a variety of proteins inside mast cells leading to degranulation and cytokine production. Since some tyrosine kinase-inhibitors are able to halt β-hexosaminidase release [35], we tested the effect of SOS pretreatment on antigen induced-tyrosine phosphorylation of proteins in mast cells. Neither of the two SOS dilutions tested (i.e. 25–50%) prevented total tyrosine phosphorylation after FcεRI activation, as judged by western blot using an antiphosphotyrosine Fig. 4. SOS (SOW) pretreatment of mast cells does not inhibit calcium mobilization after antigen or calcium ionophore stimulation. Sensitized, mature BMMCs were collected from culture media and resuspended in FURA-2 AM containing Tyrode's/BSA buffer at 37 °C during 30 min. Then, cells were washed once and resuspended in normal Tyrode's/BSA buffer or in 50% SOS-containing buffer at 37 °C during 15 min. After this incubation, cells were washed again and resuspended in SOS-free Tyrode's/BSA buffer before to be transferred to a spectrofluorometer cuvette. Fluorescence 340:380 ratio was recorded during 100 s and after this time (A) 30 ng/ml of DNPHSA (Antigen, Ag) or (B) 100 nM calcium ionophore (A23187) was added to the cells. Traces were followed during 200 s after stimuli addition. Traces shown are representative from three independent experiments with similar results. Fig. 5. SOS (SOW) pretreatment of mast cells does not significantly inhibit antigen-dependent cytokine mRNA accumulation. Mature, sensitized BMMCs were collected from culture media and resuspended in normal Tyrode's/BSA buffer or in SOS-containing buffer at 37 °C during 15 min. After this time, cells were washed once in SOS-free Tyrode's/BSA (37 °C) and stimulated with 30 ng/ml DNP-HSA during 1 h. Total RNA isolation, RT-PCR or RPA was performed as described in the Materials and methods section. mRNA accumulation of GAPDH, IL-2, IL-3, IL-4 and IL-6 was determined by RT-PCR (A), and GAPDH, TNF-α and IL-13 mRNA accumulation was determined by RPA (B). When SOW was added alone, the final concentration was 50%. 1020 J. Medina-Tamayo et al. / International Immunopharmacology 7 (2007) 1013–1024 (4G10) antibody (Fig. 3A). Yet, total tyrosine-phosphorylated proteins decreased after antigen stimulation if mast cells were pretreated with 25% or 50% SOS. Since IgE receptor crosslinking is able to induce MAPK activation, we also tested the effect of the same SOS dilutions on p38 and ERK2 MAP kinase phosphorylation levels in mast cells (Fig. 3B). In the absence of antigen-induced stimulation, the sole exposure of BMMCs to 50% SOS induced ERK2 and p38 phosphorylation. Antigen stimulation for 10 min further elevated SOS induced-MAPK activation in BMMCs. According to these results, it is possible that SOS pretreatment can activate MAPK without impairing the capacity of the FcεRI receptor to further stimulate MAPK activity. 3.4. SOS pretreatment does not inhibit FcεRI-induced calcium mobilization or NFAT-transcription factor activation in mast cells Mast cell degranulation and the synthesis of some cytokines, are absolutely dependent on calcium mobilization [36,37]. Given the fact that SOS pretreatment inhibits the degranulation capabilities of mast cells, we examined the rise of intracellular calcium in 50% SOS pretreated-mast cells during IgE-antigen or calcium ionophore stimulation. Under these experimental conditions, calcium mobilization was not affected in stimulated mast cells with either agent if they were pretreated in 50% SOS for 15 min (Fig. 4A,B). Since NFATtranscription factor is activated after IgE-antigen stimulation and calcium rise in mast cells, we also investigated NFAT binding activity in 50% SOS-treated cells. As it can be observed in Fig. 4C, no statistical difference on NFAT activation was observed between SOS treated- and non-reated-BMMCs after antigen stimulation. Thus, SOS does not seem to interfere with calcium mobilization or calcium activated-effectors in mast cells. 3.5. SOS pretreatment does not prevent IgE-antigen-induced cytokine mRNA accumulation in mast cells Murine BMMCs, through activation of distinct Src family kinases, are able to synthesize many different cytokines and chemotactic factors in response to antigen stimulation, including: IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-10, IL-12, IL-13, TNF-α, TGF-β, GM-CSF, IFN-γ, CCL2, CCL3 and CCL4, among others [38]. It is known that FcεRI crosslinking Fig. 6. SOS (SOW) pretreatment of mast cells significantly inhibits antigen-induced cytokine secretion. A total of 40 million mature, sensitized BMMCs were collected from culture media and resuspended in normal Tyrode's/BSA buffer or in SOS-containing buffer at 37 °C during 15 min. After this time, cells were washed, resuspended in 10 ml of SOS-free, complete media at 37 °C and stimulated with 30 ng/ml DNP-HSA (antigen). Samples of the supernatant were taken at indicated times and cytokine content was determined by cytokinespecific ELISA kits. Results of three independent experiments ±SE are presented. J. Medina-Tamayo et al. / International Immunopharmacology 7 (2007) 1013–1024 activates a signal transduction system that modulates the activity of a number of different transcription factors (i.e. NFAT, NFκB, AP-1). These, in turn, are responsible for de novo synthesis of cytokine mRNA [39]. Once secreted, cytokines induce recruitment and proliferation of T cells, macrophages, eosinophils and other cellular types that initiate the inflammatory response of late phase-allergic reactions. We, therefore, tested the effect of 25% and 50% SOS-preincubation on cytokine mRNA accumulation in BMMCs. For this purpose, RT-PCR or RPA assays were used to measure IL-2, IL-3, IL-4, IL-6, TNF-α and IL-13 specific transcripts in mast cells pretreated with SOS free-and SOS-containing Tyrode's BSA for 15 min at 37 °C (Fig. 5). In the absence of antigenic stimulation, the sole exposure of mast cells to 50% SOS did not induce detectable cytokine mRNA synthesis. Pretreatment of mast cells with 25% or 50% SOS, however, did not prevent FcεRI-induced cytokine mRNA synthesis. 3.6. SOS pretreatment diminishes cytokine secretion in mast cells. Since SOS pretreatment of BMMC had inhibited FcεRIinduced degranulation but not FcεRI-induced cytokine mRNA synthesis, we further evaluated cytokine release in control and SOS-preincubated cells by ELISA. For these experiments, IgEsensitized cells were preincubated in SOS free- or SOS containing-Tyrode's/BSA buffer for 15 min at 37 °C. After this time, cells were washed and resuspended in SOS-free culture media containing 30 ng/ml DNP-HSA, as described in “Materials and methods”. Cells were then incubated at 37 °C and samples taken from the supernatant at different time points to determine the amount of cytokine release after IgE-antigen stimulation. Cytokines detected included TNF α, CCL3 (MIP1α), IL-6 and IL-13 (Fig. 6). The extent of inhibition of cytokine release in SOS-pretreated cells was found to be concentration dependent and molecule specific. In general, 50%-SOS was more effective than 25% SOS pretreatment in the inhibition of antigen-induced cytokine release, except for IL-6 secretion. After pretreatment of mast cells with 50% SOS, the inhibitory effect was more noticeable in TNF-α release (83 ± 10% max. inhibition), followed by IL-13 and MIP-1α (50 ± 7%) and IL-6 (40 ± 6%). Interestingly, the inhibitory effect of SOS-pretreatment was sustained from 2 to 24 h and there were not significant changes in the concentration of all four cytokines tested in the supernatant of BMMCs' cultures, during this period of time. 4. Discussion Inhibition of degranulation of mast cells has been proposed as a feasible therapeutic target for allergic and inflammatory diseases [8,9]. In this study we report that pretreatment of a model of mucosal-type mast cells (BMMCs) with different dilutions of a neutral pH superoxidized solution (SOS), does neither significantly reduce the viability of the cells nor interfere on early 1021 FcεRI signal transduction processes. Yet, SOS pretreatment importantly inhibits the early secretion of granule content and the late secretion of at least four regulatory and chemoattractant cytokines. These results are intriguing and suggest that SOSs might be potent stabilizers of mast cell activity with potential therapeutic applications. We first evaluated the toxicity of SOS on mast cells. As expected, the treatment of BMMCs with different SOS dilutions did not significantly affect cell viability as assayed by three independent methods (i.e. 7AAD and propidium iodide exclusion and annexin V detection) (Fig. 1, Table 1). These results are in accordance with the lack of toxicity in primary irritancy and sensitization studies conducted with this same SOS in skin and mucosas, as well as in in vitro experiments (i.e. agarose overlay using fibroblasts) [30]. SOS pretreatment, however, was able to block the degranulation of mast cells induced by IgE-antigen crosslinking of the FcεRI receptor. This effect was concentration dependent, with reductions of allergen induced-histamine release ranging from 86 ± 8% and 27 ± 5% in pretreated cells with 50% and 1% SOS, respectively. Since calcium ionophore and PMA-induced degranulation were affected to the same extent, we concluded that the effects of SOS pretreatment on histamine release in mast cells are receptor-independent (Fig. 2A). In order to further explore the potential mechanism of action of SOS on degranulation, we evaluated the total tyrosine phosphorylation and MAPK phosphorylation in antigen activated-BMMCs (Fig. 3). It is known that molecular crosslinking of the FcεRI receptor activates a complex series of biochemical reactions involving the activation of distinct Src-family kinases (Fyn and Lyn) [38], activation of Syk kinase, phosphorylation of adapter proteins (like LAT, and Gab2), activation of Phospholipase C and PI3K enzymes and intracellular calcium mobilization [5]. This complex transduction system also involves the activation of MAP kinases, phosphorylation of transcription factors, and synthesis of new cytokine mRNA [5]. We therefore evaluated the global tyrosine phosphorylation level after FcεRI activation, as judged by western blot (using 4G10 antibody) and the phosphorylation levels of ERK2 and p38 MAP kinases. Total tyrosine phopshorylation after FcεRI triggering seems to decrease in mast cells pretreated with 25% or 50% SOS. In contrast, the sole exposure of the cells to 25% or 50% SOS-containing buffers elicited the phosphorylation of ERK2 and p38 proteins (Fig. 2B). These phenomena could be due to the induction of a mild cellular stress by the low concentration of chlorine species found in this SOS 1022 J. Medina-Tamayo et al. / International Immunopharmacology 7 (2007) 1013–1024 (b80 ppm). These species are not themselves free radicals but these oxygen-containing molecules can facilitate free-radical formation [43]. In fact, oxidative damage is known to induce ERK and p38 activation, and even cell death, depending on the time of exposure and concentration of the oxidizing agent [40]. Due to the low concentration of chlorine species found in this particular SOS, the mast cells could have been stressed without compromising the viability. Taken together, these results suggest that SOS pretreatment does not significantly affect the signaling machinery depending on tyrosine phosphorylation of the FcεRI receptor. Another required condition for IgE-antigen inducedmast cell degranulation is intracellular calcium mobilization. Unexpectedly, cells pretreated with 50% SOS did not show inhibition of calcium mobilization nor in NFAT activation after allergen-or calcium ionophorestimulation (Fig. 4). This result supports the hypothesis that SOS pretreatment does not interfere with the signal transduction process required for calcium mobilization and it does not seem to modify the membrane capacity of calcium influx. However, it does suggest that SOS pretreatment can induce uncoupling of calcium flux and granule secretion in this cells, since it has been widely understood that calcium mobilization is an essential requisite for IgE-antigen induced degranulation [41]. Cytokine mRNA expression was also determined in control and SOS-treated cells after 1 h of IgE-antigen stimulation. As measured by RT-PCR and RPA, we did not detect a significant effect of 50% SOS pretreatment on IL-2, IL-3, IL-4, IL-6, TNF-α and IL-13 mRNA accumulation either in basal or IgE-antigen induced conditions (Fig. 5). In contrast, cytokine secretion was severely inhibited by SOS pretreatment in a concentration dependent and molecule specific way. TNF-α secretion, for example, was inhibited by 80%, whereas CCL3 and IL-13 release were blocked by 50%, and IL-6 by only 40%. According to these results, SOS pretreatment seems to be able to inhibit the cytokine secretory pathway(s) in mast cells, without altering the transcription machinery. Altogether, our study suggests that this particular neutral pH super-oxidized solution induces most of the effects of a classical mast cell stabilizer. Cromolyn sodium and nedocromil sodium are typical membrane stabilizers because they produce blockage on mast cell degranulation without affecting other parameters of cell activation [42]. Similarly, the SOS herein used was able to strongly inhibit both antigen and calcium ionophore-induced mast cell degranulation without affecting cell viability or even FcεRI signal transduc- tion processes. The precise mechanism(s) by which this SOS inhibits degranulation without disturbing RNA synthesis and early signal transduction is unknown. It is possible that the components of this SOS partially modify cytoskeleton or plasma membrane targets that uncouple calcium rise from granule secretion. Instead, this SOS does not appear to inhibit intracellular signaling processes depending on tyrosine phosphorylation. At this point, however, it is not possible to make a generalization about the effects of SOS in mast cells because not all super-oxidized solutions have the same properties [23]. The more acidic or alkaline the solutions are, the more corrosive and toxic they become [43, 44]. Another major problem with previous SOSs formulations, had been the lack of stability for long periods of time. Basically, most of SOSs available today are stable for only few hours or days. But, apparently, neither one of these two problems seem to be of major concern with the neutral pH-SOS herein tested. In aging studies, for example, it was demonstrated that this SOS was stable for up to 2 years. Preclinical data also showed that this SOS does not irritate or sensitize skin and mucosas (e.g. ocular, peritoneal, nasal, oral) [30]. Finally, pilot human studies have been successfully conducted in various clinical conditions, including those associated with mast cell activation [25–28]. The clinical outcomes in cases of non-infected-bullous (pemphigus) and hyperkeratotic (psoriasis) lesions, for example, have suggested that this SOS could exert a direct anti-inflammatory effect [27]. However, the use of neutral pH super-oxidized solution in any allergic and inflammatory condition will have to be evaluated in proper controlled studies. In summary, our data suggest that SOS is able to inhibit the cell machinery for granule secretion without altering the main signal transduction pathway induced by IgE-antigen receptor crosslinking. This is a relevant finding because the identification of the mechanism(s) of action of SOSs on mast cells could help to identify novel targets in the intricate pathways that control allergic and inflammatory reactions. This work was financed by CONACyT (Grant # 39726-Q) and the Third World Academy of Sciences (Grant # 02-073 RG/BIO/LA). CGE thanks Dr. Rafael Villalobos Molina from Cinvestav Sede Sur for his support allowing the authors to use of diverse equipment for cell culture and electrophoresis and Mr. Armando Resendiz for technical assistance. AAG thanks Dr Eileen Thatcher from Sonoma S University for reviewing the manuscript and Carlos Landa for technical support with flow cytometry analysis. J. Medina-Tamayo et al. / International Immunopharmacology 7 (2007) 1013–1024 References [1] Galli SJ, Nakae S, Tsai M. Mast cells in the development of adaptive immune responses. Nat Immunol 2005;6:135–42. [2] Galli SJ, Kalesnikoff J, Grimbaldeston MA, Piliponski AM, Williams CM, Tsai M. 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Antimicrobial activity of super-oxidized solution. J Hosp Infect 1996;34:43–9. [26] Dalla Paola L, Brocco E, Senesi A, Merico M, De Vido D, Assaloni R, et al. Super-oxidized solution (SOS) therapy for infected diabetic foot ulcers. Wounds 2006;18:262–70. [27] Bogiovanni C, Bomengen RW, Hughes MD. Superoxidized solution: spectrum of clinical applications. Diabetic Foot Global Conference Hollywood, Ca. USA 23rd–25th March; 2006. [28] Miranda-Altamirano A. Reducing bacterial infectious complications from burn wounds. A look at the use of Oculus Microcyn60 to treat wounds in Mexico. Wounds Jan. 2006:17–9 (Suppl). [29] Goretti C, Mazzurco S, Ambrosini Nobili L, Macchiarini S, Tedeschi A, Palumbo F, et al. Use of a novel antiseptic superoxidised solution (SOS) in the management of wide post-surgical lesion in the infected diabetic foot. Int J Low Extrem Wounds 2007;6:22–7. [30] Gutiérrez AA. The science behind stable, super-oxidized solution. Exploring the various applications of super-oxidized solutions. Wounds. Jan 2006 (Suppl), 7–10. [31] Huff TF, Lantz CS, Ryan JJ, Leftwich JA. Mast cell-committed progenitors. In: Kitamura Y, Yamamoto S, Galli SJ, Greaves MW, editors. Biological and molecular aspects of mast cell differentiation and function. New York: Raven Press; 1995. p. 105–17. [32] Saitoh S, Arudchandran R, Manetz TS, Sommers CL, Zhang W, Love PE, et al. LAT is essential for FcεRI-mediated mast cell activation. Immunity 2000;12:525–35. [33] Hidalgo E, Domínguez C. Growth-altering effects of sodium hypochlorite in cultured human dermal fibroblasts. Life Sci 2000:1331–4. [34] Gonzalez-Espinosa C, Odom S, Olivera A, Hobson PJ, CidMartinez ME, Oliveira-dos-Santos A, et al. Preferential signaling and induction of allergy-promoting lymphokines upon weak stimulation of the high affinity IgE receptor on mast cells. J Exp Med 2003;197:1453–65. [35] Hallet MB, Hodges R, Cadman M, Blanchfeld H, Dewitt S, Pettit EJ, Blanchfeld H, et al. Techniques for measuring and manipulating free Ca2+ in the cytosol and organelles of neutrophils. J Immunol Methods 1999;232:77–88. [36] Barker SA, Lujan D, Wilson BS. Multiple roles for PI3-Kinase in the regulation of PLC gamma activity and Ca2+ mobilization in antigen-stimulated mast cells. J Leukoc Biol 1999;65(3):321–9. [37] Common and distinct signaling pathways mediate the induction of TNF-α and IL-5 in IgE plus antigen-stimulated mast cells. J Immunol 1998;160(1):273–83. [38] Rivera J, Furumoto Y, Gonzalez-Espinosa C, Kovarova M, Odom S, Parravicini V. Functional coupling of FcεRI employs two proximal Src family kinases. A new paradigm in mast cell activation. In: Bienenstock J, Ring J, Togias AG, editors. Allergy 1024 J. Medina-Tamayo et al. / International Immunopharmacology 7 (2007) 1013–1024 frontiers and future. Proceedings of the 24th Symposium of the Collegium Internationale Allergologicum. Cambridge, MA: Hogrefe and Huber; 2004. p. 152–6. [39] Gordon JR. FcεRI-induced cytokine production and gene expression. In: Hamawy MM, editor. IgE receptor (FcεRI) function in mast cell and basophils. Austin, Tx: RG Landes Company; 1997. p. 209–42. [40] Carvalho H, Evelson P, Sigaud S, Gonzalez-Flecha B. Mitogenactivated protein kinases modulate H(2)O(2)-induced apoptosis in primary rat alveolar epithelial cells. J Cell Biochem 2004;92: 502–13. [41] Blank U, Rivera J. The ins and outs of mast cell exocytosis. Trends Immunol 2004;25:266–73. [42] Mancel E, Drouet M, Sabbah A. Membrane stabilizers (chromones and ketotifen). Allerg Immunol (Paris) 1999;31:103–5. [43] Suzuki T, Masuda M, Friesen MD, Fenet B, Ohshima H. Novel products generated from 2'-deoxyguanosine by hypochlorous acid or a myeloperoxidase -H2O2-Cl-system: identification of diimino-imidazole and amino-imidazolone nucleosides. Nucleic Acids Res 2002;30:2555–64. [44] Len SV, Hung YC, Erickson M, Kim C. Ultraviolet spectrophotometric characterization and bactericidal properties of electrolyzed oxidizing water as influenced by amperage and pH. J Food Prot 2000;63:1534–7. Microdacyn International Accreditations CE MicroSafe USA FDA (1) Bacterialcidal USA FDA (2) Virucidal - Human Corona Virus USA FDA (3) Sporicidal USA FDA (4) Clostridium difficile USA FDA (5) Fungicidal USA FDA (6) Tuberculicidal FDA Press release Journals Of Hospital Infection United States E.P.A.(1) 2006 MicroSafe Disinfectant (MSD) Bactericidal / Tuberculocidal / Virucidal / Sanitizer Disinfectant / Disinfectant Cleaner (Hospital –Grade / Households / Industrial Areas) United States E.P.A.(2) 2007 MicroSafe Sanitizer (MSS) Food Contact Sanitizer 2004 Debriding, irrigating and moistening acute and chronic wounds in comprehensive wound treatmentby reducing microbial load and creating moist environment European Union CE Mark Mexico ProductRegistration 2003 Antiseptic disinfection solution for high level disinfection of medical instruments, and/or equipmentand clean-rooms, areas of medical instruments, equipment and clean rooms. Mexico Product Registration 2004 Antiseptic treatment of wounds and infected areas. United Arab Emitates Registred – Ministry 2012 Management via debridement of wounds such as of Health UAE stage I-IV pressure ulcers, partial and full thickness wounds, diabetic foot ulcers, postsurgical wounds, first, second and thrid degree burns, grafted and donor sites Canada Class II Medical Device 2004 Moistening, irrigating, cleansing and debriding acute and chronic dermal lesions, diabetic ulcers and post-surgical wounds. ! United States Fulbright Scholar in Law United States Fulbright Senior Specialist Program Chairman American Board of Healthcare Law and Medicine " 1935 South Plum Grove Rd. #311 Newport Beach, California 92660 949-260-8487 760-683-3187 " 4630 Campus Drive, Suite #200 Palatine, Illinois 60067 949-260-8487 760-683-3187 ! # " & $ * / ( % + " & ,% ! & " " (% ' - ( " + & 4 * 2 " ) " ! !0 234 56( 67 2 ;/ " ! 5'/7 ! " " :! " 1 9 ! " 8 1 ( " " 2 < 1 1 " 91 ! & !0 8 <1 9+ 4 ( / 0# ! %& & + <+ (% " ! % ! ! > $ < ? . * & 0 4* $ ( A B @ ! 4 2 % "( ( & & 8 & ( " ( = ? 0 @ ! * + 4 + & ( . 1 Chairman, American Board of Healthcare Law and Medicine United States Fulbright Scholar in Law United States Fulbright PEER Review Committee – Global / Public Health UCLA Visiting Scholar in Public Health, UCLA School of Public Health Department of Health Services, Division of Healthcare Management and Policy President, Aristotle University Dean and Professor of Public Health, Aristotle University School of Public Health Fellow, International College of Surgeons * * & .@ 3D E E F 0 / .' ! ! ) ! 0 1 " " 4 " 7 ! ( ! ( ! = ? ! -# & 5 ( = ( 9+2 & & "- ) . & 0( ! ( . / D G C HH. ! I C. . Microdacyn60 Wound Care Tecnología Patentada El método más eficaz para el tratamiento de heridas crónicas y agudas NUEVAS INDICACIONES MUCOSAS: ORAL, NASAL, OCULAR Y VAGINAL NUEVAS PRESENTACIONES: HIDROGEL, SPRAY, NWT Beneficios: 1. Acción bactericida. También es efectivo contra los biofilms. 2. Efecto antiinflamatorio 3. Regeneración tisular activa 4. Se necesitan menos antibióticos 5. Seguro para las células sanas de la piel Microdacyn Wound Care Microdacyn ocupa una posición especial dentro de la categoría significa una curación más rápida de la herida. El producto no de productos para el tratamiento de heridas, debido a que su sólo es eficaz sino también muy fácil de usar. acción no se basa en componentes químicos sintéticos No es irritante de la piel, los ojos o la garganta y elimina el olor. específicos. Microdacyn es un tratamiento biológicamente activo para el desbridamiento, irrigación, humidificación y disminución de la carga microbiana de las heridas agudas y crónicas que son difíciles de curar, están infectadas y otras. Es totalmente seguro, tanto para el paciente como para el personal sanitario. Microdacyn está listo para su uso y es respetuoso con el medio ambiente. Tras su uso, los ingredientes activos se reducen a 0, lo que significa que no es necesario eliminar el producto como desecho hospitalario. Microdacyn se puede utilizar fácilmente en combinación con otras formas de tratamiento de heridas. Microdacyn disminuye la carga microbiana patógena eliminando Microdacyn60 ha demostrado su eficacia en el tratamiento de los microorganismos unicelulares patógenos, sin efectos tóxicos heridas en mucosas: boca, nariz, ojos y vagina. Con esta nueva o nocivos sobre los tejidos humanos. Además, Microdacyn indicación amplía su campo de acción, facilitando así la labor de contribuye a crear un entorno húmedo en la herida, lo que los profesionales. Descripción y uso del producto Microdacyn es la marca internacional de productos para el Microdacyn60 solución y Microdacyn gel son productos cuidado de heridas basados en la tecnología Microcyn, superoxidados que se producen mediante la electrólisis de agua desarrollada por la empresa estadounidense Oculus Innovative ultrapura y NaCl. Microdacyn desnaturaliza la pared celular del organismo unicelular patógeno, causándole fugas y provocando su desintegración. Los estudios demuestran que Microdacyn tiene efecto tanto antiinflamatorio como antialérgico, que acelera la curación de la herida. (Estudios disponibles ) Propiedades y ventajas para el paciente y durante el tratamiento: ‡ Antiséptico ‡ Utilizable en mucosas: boca, ojos, nariz y vagina. ‡ No citotóxico ‡ Combinable con otras formas de tratamiento de heridas. ‡ pH neutro ‡ Acción antiinflamatoria. ‡ No irritante ‡ Muy económico. ‡ Elimina el olor ‡ Reepitelizante. RECOMENDADO POR EL: AMERICAN BOARD OF HEALTHCARE POR SU EFICACIA Y SEGURIDAD En otro estudio comparativo, se observó también que el grupo de pacientes tratados con Microdacyn mostró una epitelización más rápida. Resultados de las investigaciones 1. Efecto antibacteriano En estudios in vitro se ha demostrado que Microdacyn mata bacterias, hongos, esporas y virus rápidamente, con una reducción logarítmica significativa. 2. También es eficaz contra las películas biológicas La investigación ha demostrado que Microdacyn tiene un efecto bactericida en cepas de pseudomonas y E. coli que producen biofilms. Después de 30 minutos de exposición a Microdacyn, la mayoría de microorganismos habían sido eliminados. 3. Efecto antiinflamatorio Ensayos in vitro han demostrado que 15 minutos de exposición al producto son suficientes para suprimir la liberación de histamina en un 90%. Además, Microdacyn inhibe la liberación de otras moléculas que prolongan la inflamación (entre otras interleucinas, interferón gamma y TNF-alfa), gracias a lo cual las citoquinas "buenas" tienen mayor capacidad para iniciar el proceso de cicatrización de la herida. 4. Regeneración tisular activa En un estudio clínico, se demostró que Microdacyn era casi dos veces más eficaz en el proceso de curación de úlceras de pie diabético que un grupo de ensayo tratado con una solución salina fisiológica (NaCl 0,9%). Los investigadores también observaron una reducción de la carga microbiana y un aumento en la formación de tejido de granulación. Indicaciones 5. Se necesitan menos antibióticos En dos estudios recientes se ha demostrado que los pacientes con úlceras de pie diabético que fueron tratados con Microdacyn necesitaron antibióticos durante un periodo considerablemente más corto que un grupo de control que se trató con povidona yodada. En uno de estos estudios, la diferencia fue de casi el 43% (74 frente a 129 días). Además, el cierre de la herida en el grupo de Microdacyn se produjo en promedio 2 meses antes que en el grupo de yodo (144 frente a 212 días, una reducción del 32%). 6. Seguro para las células sanas de la piel En comparación con otros productos (por ejemplo, a base de peróxido de hidrógeno) Microdacyn NO es citotóxico y por lo tanto es completamente seguro para el tejido humano. La investigación ha demostrado que incluso la exposición prolongada de fibroblastos humanos a Microdacyn no conduce a la muerte celular. A diferencia de los virus, esporas y bacterias, las células humanas tienen un mecanismo que regula la absorción de los ingredientes activos de Microdacyn. Esto evita daños en los tejidos. En comparación con otros tratamientos, como yodo, clorhexidina y sulfadiazina de plata, Microdacyn demuestra ser uno de los productos más seguros y suaves para piel. Más información www.sdomedical.com o [email protected] ‡ Heridas oncológicas ‡ Heridas agudas (profiláctico) ‡ Heridas traumáticas infectadas y necróticas ‡ Heridas quirúrgicas ‡ Úlceras crónicas ‡ Úlceras del pie diabético ‡ Ulcera crural (arterial y venosa) ‡ Heridas por decúbito infectadas ‡ Quemaduras (1 º y 2 º grado) Contraindicaciones No se conoce ninguna condición en la cual no se deba aplicar Microdacyn. Aplicación - Instrucciones de uso: Microdacyn Wound Care Solución directamente sobre la herida con la ayuda de unas gasas empapadas, en fomento, ‡ Aplicar durante 5-15 minutos. Microdacyn Wound Care Solución dos veces al día de la forma habitual, y cada vez que se cambie el apósito. No es nece‡ Utilizar sario limpiar la herida tras su uso. ‡ Aplicar Microdacyn Hidrogel directamente sobre la herida y cubrir con un apósito de cura húmeda. Repetir cada 24-48 hrs. ‡ Aplicar Microdacyn Spray y Microdacyn líquido directamente en la herida o inmersión siguiendo la pauta indicada. NUEVAS PRESENTACIONES Microdacyn60 Wound Care Microdacyn Hidrogel Descripción del producto Código Nacional Precio Venta Recomendado 120 ml / SPRAY, 6 botellas por caja (caja individual) 172386.4 15,75 € (10% IVA incluido) 250 ml / SPRAY, 12 botellas por caja (caja individual) 172388.8 21,60 € (10% IVA incluido) 500 ml / Solución, 24 botellas por caja (caja individual) 172389.5 29,70 € (10% IVA incluido) 990 ml/ Irrigación/NWT, 6 botellas por caja --- --- 5 L / Garrafa, 4 garrafas por caja --- --- 172398.7 28,35 € (10% IVA incluido) 120 g / 6 botellas por caja (caja individual) SDOMEDICAL Gran de Sant Andreu 159, 1º 2ª · 08030 Barcelona, Spain  The International Journal of Lower Extremity Wounds http://ijl.sagepub.com/ Super-Oxidized Solution (Dermacyn Wound Care) as Adjuvant Treatment in the Postoperative Management of Complicated Diabetic Foot Osteomyelitis : Preliminary Experience in a Specialized Department Javier Aragón-Sánchez, Jose L. Lázaro-Martínez, Yurena Quintana-Marrero, Irene Sanz-Corbalán, Maria J. Hernández-Herrero and Juan J. Cabrera-Galván International Journal of Lower Extremity Wounds published online 26 February 2013 DOI: 10.1177/1534734613476710 The online version of this article can be found at: http://ijl.sagepub.com/content/early/2013/02/01/1534734613476710 Published by: http://www.sagepublications.com Additional services and information for The International Journal of Lower Extremity Wounds can be found at: Email Alerts: http://ijl.sagepub.com/cgi/alerts Subscriptions: http://ijl.sagepub.com/subscriptions Reprints: http://www.sagepub.com/journalsReprints.nav Permissions: http://www.sagepub.com/journalsPermissions.nav >> OnlineFirst Version of Record - Feb 26, 2013 What is This? Downloaded from ijl.sagepub.com at INTERNATIONAL JOURNAL OF LOWER EXTREMITY WOUNDS on February 27, 2013 476710 IJLXXX10.1177/1534734613476710The International Journal of Lower Extremity WoundsAragón-Sánchez et al. © The Author(s) 2011 Reprints and permission: http://www. sagepub.com/journalsPermissions.nav Case Report The International Journal of Lower Extremity Wounds XX(X) 1–8 © The Author(s) 2013 Reprints and permission: sagepub.com/journalsPermissions.nav DOI: 10.1177/1534734613476710 ijl.sagepub.com Super-Oxidized Solution (Dermacyn Wound Care) as Adjuvant Treatment in the Postoperative Management of Complicated Diabetic Foot Osteomyelitis: Preliminary Experience in a Specialized Department Javier Aragón-Sánchez, MD, PhD1, Jose L. Lázaro-Martínez, DPM, PhD2, Yurena Quintana-Marrero, RN1, Irene Sanz-Corbalán, DPM2, Maria J. Hernández-Herrero, MD1, and Juan J. Cabrera-Galván, MD, PhD1,3 Abstract Surgery is usually used to treat diabetic foot osteomyelitis (DFO), whether primarily or in cases in which antibiotics are not able to control infection. In many cases, the bone is only partially removed, which means that residual infection remains in the bone margins, and the wound is left open to heal by secondary intent.The use of culture-guided postoperative antibiotic treatment and adequate management of the wound must be addressed. No trials exist dealing with local treatment in the postoperative management of these cases of complicated DFO. We decided to test a super-oxidized solution, Dermacyn Wound Care (DWC; Oculus Innovative Sciences Netherlands BV, Sittard, Netherlands) to obtain preliminary experience in patients in whom infected bone remained in the surgical wounds. Our hypothesis was that DWC could be useful to control infection in the residual infected bone and surrounding soft tissues and would thus facilitate healing. Fourteen consecutive patients who underwent conservative surgery for DFO, in whom clean bone margins could not be assured, were treated in the postoperative period with DWC. Eleven cases were located in the forefoot, 6 on the first ray and the rest in lesser toes, 1 in the Lisfranc joint, and 2 on the calcaneus. No side effects appeared during treatment. Neither allergies nor skin dermatitis were found. Limb salvage was successfully achieved in 100% of the cases. Healing was achieved in a median period of 6.8 weeks. Keywords diabetic foot, osteomyelitis, super-oxidized solution, diabetic foot infections Background The systematic surgical treatment of bone infection in the feet of patients with diabetes is currently under debate because some patients achieve remission exclusively with antibiotics.1 However, surgery is the usual approach for treating diabetic foot osteomyelitis (DFO), whether primarily or in cases in which antibiotics are not able to control the infection.2 The choice of surgical technique to treat DFO is not well standardized, and the goal is to remove the infected bone without reducing the functionality of the foot. Some teams perform amputations to removed the infected bone,3 whereas others use conservative surgery to conserve the architecture of the foot.4,5 When infection remains in 1 La Paloma Hospital, Las Palmas de Gran Canaria, Spain Complutense University, Madrid, Spain 3 Las Palmas de Gran Canaria University, Las Palmas de Gran Canaria, Spain 2 Corresponding Author: Javier Aragón-Sánchez, Hospital La Paloma, C/Maestro Valle, 20, 35005 Las Palmas de Gran Canaria, Canary Islands, Spain Email: [email protected] Downloaded from ijl.sagepub.com at INTERNATIONAL JOURNAL OF LOWER EXTREMITY WOUNDS on February 27, 2013 2 The International Journal of Lower Extremity Wounds XX(X) bone and/or soft tissues, the wound should not be closed.6-8 The open wound caused by surgery will undergo postoperative care to achieve healing by secondary intention or before undergoing reconstructive techniques. Our approach to treating DFO is to remove the infected bone, leave the wound open to heal by secondary intention, and use culture-guided postoperative antibiotic treatment.5,9 In many cases, we cannot be sure that all the infected bone has been removed and thus believe that wide drainage through the open wound and postoperative culture-guided antibiotics should be used to eradicate the infection. Theoretically, adjuvant treatment with topical antibiotics or antiseptics could help reduce the bacterial load, which is located in the residual infected bone and soft tissues. This could also help reduce the timing of postoperative antibiotic treatment but this has not been confirmed. In our experience, management of the postoperative wound following surgery for diabetic foot infections is always determined in an individualized way, and we frequently use silver-based dressings. However, this choice is empiric because no trials exist dealing with local treatment in the postoperative management of DFO. Based on the experience of other groups,10,11 we decided to test a superoxidized solution, Dermacyn Wound Care (DWC; Oculus Innovative Sciences Netherlands BV, Sittard, Netherlands), in order to obtain preliminary experience in patients in whom infected bone remained in the surgical wounds. Our hypothesis was that DWC could be useful for controlling infection in the residual infected bone and surrounding soft tissues and could thus facilitate healing. The aim of this work is thus to present our preliminary experience with a super-oxidized solution as adjuvant treatment in the postoperative wound management of complicated DFO in patients presenting a high risk of amputation in order to establish its safety and usefulness prior to designing a clinical trial. Patients and Methods Fourteen consecutive patients who underwent conservative surgery for DFO, in whom clean bone margins could not be assured, were treated in the postoperative period with DWC. All were assumed to have infected but viable bone remaining in the wounds after removing most of the infected bone. Conservative surgery in this case was defined as a procedure in which only the infected bone and nonviable soft tissue were removed, but no amputation of any part of the foot was undertaken. Conservative surgery preserves the soft-tissue envelope and more distal tissues, which means resection of the infected bone while preserving the soft-tissue envelope.5 Only cases with osteomyelitis without soft tissue infection were included in this preliminary study. Soft tissue infections were diagnosed either preoperatively or during surgical procedures according to previously published criteria.12,13 Diagnosis of osteomyelitis was based on our flow chart comprising a sequential combination of the probe-to-bone test and plain X-rays, as published elsewhere.14 A neurological examination was undertaken using SemmesWeinstein filaments (5.07 = 10 g). Neuropathy was diagnosed when the patient did not feel 3 or more sites. Peripheral arterial disease was diagnosed if the patient met the following criteria: absence of both distal pulses and/or ankle brachial index below 0.9. Ulcers in patients diagnosed for neuropathy were defined as neuropathic ulcers. If both neuropathy and peripheral arterial disease were present, the ulcer was defined as neuroischemic. During surgical intervention, bone samples were extracted for analysis by the microbiology and pathology laboratories. Only aerobic cultures were grown in this study. Surgical wounds were copiously irrigated with DWC in the operating room. DWC was irrigated daily through the wound with the purpose of washing the residual bone. According to our treatment protocol, cases with persistent postoperative infection either in bone or soft tissue that precluded wound healing and/or produced clinical infectious symptoms were reoperated. Reoperations included minor and major amputations if required. Based on our previous experience, infection control was considered successful once the surgical wound and the index ulcer that acted as the point of entry of the infection had healed.5,9 Patients were followed-up to detect recurrence of the infection, reulceration (whether complicated or not with new osteomyelitis), and the need for amputation. Limb salvage was achieved when the patient did not require a major amputation, which was defined as amputation through or above the ankle joint. The major endpoint to be evaluated was healing without recurrence of the infection and need for amputation. Any side effect associated with the treatment was assessed. Patients gave informed consent for surgery, photography, and inclusion in the study. The ethical research committee of the Materno-Insular Hospital of Las Palmas de Gran Canaria reviewed and approved the study protocol and gave consent for publication. Results Histopathology reports confirmed osteomyelitis in every case. Eleven cases were located in the forefoot, 6 on the first ray and the rest in lesser toes, 1 in the Lisfranc joint, and 2 on the calcaneus. Cases are presented in Table 1. Eight patients had previously undergone surgery for the same infection, which had not been resolved. Other teams had treated 6 patients, and amputations (2 major) had been indicated. Four patients had undergone surgery in our hospital but infection was not adequately resolved. DWC was not used in the postoperative period following the initial surgery. Cultures were negative in 3 cases, were not available in another one, and bacteria were isolated in the Downloaded from ijl.sagepub.com at INTERNATIONAL JOURNAL OF LOWER EXTREMITY WOUNDS on February 27, 2013 Table 1. Case Series Point of Entry to the Infection Case 1 Case 2 Downloaded from ijl.sagepub.com at INTERNATIONAL JOURNAL OF LOWER EXTREMITY WOUNDS on February 27, 2013 Neuroischemic interdigital ulcer Spreading of forefoot infection along plantar aponeurosis Case 3 Neuroischemic plantar ulcer beneath the first metatarsal– phalangeal joint Case 4 Neuropathic plantar ulcer beneath the first metatarsal– phalangeal joint Case 5 Location of Bone Infection Proximal phalanx Previous Surgery Due to the Same Infection No Yes. Forefoot Calcaneus. infection had been Attachment of operated. Calcaneus plantar aponeurosis osteomyelitis was to the calcaneus a consequence of spreading of the infection along plantar aponeurosis. DWC was not applied at the first surgery Medial sesamoid bone Yes. DWC was not applied at the first surgery Surgical Procedure Bacteria Involved Ostectomy of the phalanx NA Calcaneus curettage Escherichia coli, Streptococcus spp. DWC Use Lavage of the open wound and soaked gauzes with DWC Close lavage with DWC through the wound Need for Reoperation Postoperative and/or Amputation Antibiotic Treatment No 3 weeks No 7.1 weeks Time to Healing Sesamoidectomy. Keller’s arthroplasty. Internal fixation Escherichia coli, Methicillinresistant Staphylococcus aureus Infection of the surgical wound. Closed lavage with DWC through the surgical wound and gauzes soaked in DWC Yes 8.5 weeks Medial sesamoid bone Yes. DWC was not applied at the first surgery Sesamoidectomy. Keller’s arthroplasty. Internal fixation Staphylococcus aureus No 8.5 weeks Neuroischemic ulcer of the big toe as a consequence of critical ischemia Distal phalanx of the big toe. Big toe amputation had been indicated by another team Methicillinresistant Staphylococcus aureus No 4.3 weeks 3 weeks Case 6 Neuroischemic ulcer on the dorsum of the foot Case 7 Neuropathic ulcer on the fifth toe Yes. Another team Osteomyelitis of the performed Lisfranc joint. Major debridement amputation had been indicated by another team No Osteomyelitis of the proximal interphalangeal joint. Fifth toe amputation had been indicated by another team Removal of the distal phalanx. After the first procedure recurrence was found and treated with new curettage and DWC Curettage of the Lisfranc joint Infection of the surgical wound and reopening of the ulcer. Closed lavage with DWC through the index ulcer and gauzes soaked in DWC Lavage of the open wound and gauzes soaked in DWC Yes. DWC was not applied at the first surgery Curettage of the interphalangeal joint Follow-Up 8.5 weeks Patient died 1 month after healing following heart attack 7.1 weeks 13 months without recurrence 8.5 weeks Recurrence 2 months after apparent healing. Patient underwent new conservative surgical procedures. No amputation has been required in the year since the first surgery 6.8 weeks 12 months without recurrence 6 months without recurrence Yersinia enterocolitica Close intraosseus lavage with DWC through the wound No 4.1 weeks 9.2 weeks 8 months without recurrence No growth Lavage of the open wound and gauzes soaked in DWC No 0.3 weeks 2.8 weeks 9 months without recurrence (continued) 3 4 Table 1. (continued) Point of Entry to the Infection Downloaded from ijl.sagepub.com at INTERNATIONAL JOURNAL OF LOWER EXTREMITY WOUNDS on February 27, 2013 Case 8 Neuropathic ulcer on the big toe Case 9 Neuropathic ulcer over the first metatarsal– phalangeal joint Location of Bone Infection Osteomyelitis of the interphalangeal joint of the hallux. Big toe amputation had been indicated by another team Osteomyelitis of the first metatarsalphalangeal joint. Transmetatarsal amputation had been indicated by another team Osteomyelitis of the distal phalanx Case 10 Neuropathic ulcer on the tip of the third toe Osteomyelitis of the Case 11 Interdigital middle phalanx of neuroischemic the fourth toe ulcer Previous Surgery Due to the Same Infection Bacteria Involved DWC Use Need for Reoperation Postoperative and/or Amputation Antibiotic Treatment Time to Healing Follow-Up No Curettage of the Escherichia coli interphalangeal joint Lavage of the open wound and gauzes soaked in DWC No 5.4 weeks 6.1 weeks 8 months without recurrence Yes. Another team performed debridement during admission in another hospital Curettage of the first metatarsalphalangeal joint. External fixator. The index ulcer was left open to allow joint lavage Ostectomy of the distal phalanx Proteus morganii, Staphylococcus aureus Lavage through the ulcer and gauzes soaked in DWC No 4.4 weeks 7.1 weeks 8 months without recurrence No growth Lavage of the wound and gauzes soaked in DWC No 2.7 weeks 4.3 weeks 9 months without recurrence Yes. Progressive involvement of soft tissue with necrosis requiring amputation of the fourth toe No NA NA NA 7.1 weeks 11.4 weeks Three months without recurrence No No Open arthroplasty Acinetobacter baumanii Lavage of the open wound and gauzes soaked in DWC Yes. Another team performed debridement. Negative pressure with silver was applied during admission in another hospital No Curettage of the calcaneus and Chopart joint No growth Close intraosseus lavage with DWC through the wound Open arthroplasty Serratia marcescens Lavage of the open wound and gauzes soaked in DWC No 4.8 weeks 3.8 weeks Five months without recurrence Partial distal phalanx removal Serratia marcescens Debridement of the ulcer. Lavage of the open wound and bone exposed and gauzes soaked in DWC for 5 days. Bone surgery and closure of the wound No 3 weeks 2.1 weeks Three months without recurrence Case 12 Neuropathic ulcer on the heel Osteomyelitis of the calcaneus and Chopart joint. Major amputation had been indicated by another team Case 13 Interdigital neuropathic ulcer Osteomyelitis of the proximal interphalangeal joint of the fourth toe Yes. DWC was not Osteomyelitis of the applied at the first distal phalanx of the surgery big toe Case 14 Neuropathic plantar ulcer on the big toe Surgical Procedure Abbreviation: DWC, Dermacyn Wound Care (DWC; Oculus Innovative Sciences Netherlands BV, Sittard, Netherlands); NA, Not available. 5 Aragón-Sánchez et al. Figure 1. Case 2. Abscess on the heel after undergoing debridement of the plantar central compartment. Bone could be probed through the incision Figure 3. The wound was closed after new curettage of the calcaneus, leaving a Foley’s catheter inside for performing instilation with DWC Figure 2. X-ray of the case 2.Yellow arrow shows cortical defect where the abscess was drained Figure 4. Total healing of the surgical wound remainder, as illustrated in Table 1. Gram-negative bacteria were isolated in 8 cases and methicillin-resistant Staphylococcus aureus (MRSA) in 2 cases. Postoperative antibiotic treatment was given for a median of 4.4 weeks. DWC was used in different ways: closed lavage through a catheter inserted in the cavity or by inserting the tip of a syringe between the stitches, open lavage, or by placing gauzes soaked in DWC. Case 2 is shown in Figure 1. The patient had undergone debridement of the plantar central compartment and was readmitted for abscess on the heel. X-ray showed signs of osteomyelitis in the calcaneus (Figure 2, yellow arrow). He underwent debridement and bone curettage, which did not resolve the infection. He underwent a reoperation consisting of new curettage of the calcaneus and the wound was closed, leaving a Foley’s catheter inside (Figure 3). Postoperative culture-guided antibiotics were administered and lavage with DWC was carried out 3 times a day. The catheter was removed and healing was achieved without complications (Figure 4). Lavage through the stitches due to wound infection in case 3 is shown in Figure 5. The Kirschner wire was not removed due to infection. The Kirschner wire was removed 6 weeks after surgery, and the patient was discharged with only a minor defect in wound healing in order to undergo outpatient care (Figure 6A, white arrow). However, recurrence 3 months after apparent healing was found in case 3 (Figure 6B). The latter patient underwent new conservative surgical procedures using DWC in the postoperative period. No amputation has been required in the year since the first operation. One patient (case 11, Figure 7) required amputation of the toe due to uncontrolled infection by Acinetobacter baumanii and progressive destruction of the soft tissue envelope. Neither recurrence of Downloaded from ijl.sagepub.com at INTERNATIONAL JOURNAL OF LOWER EXTREMITY WOUNDS on February 27, 2013 6 The International Journal of Lower Extremity Wounds XX(X) Figure 5. Lavage through the stitches due to wound infection in case 3 the infection nor subsequent amputation was found in the rest of the patients. Limb salvage was successfully achieved in 100% of the cases. Healing was achieved in a median period of 6.8 weeks. No side effects appeared during treatment. Neither allergies nor skin dermatitis were found. Pain was not evaluable because of the neuropathic etiology of the ulcers. Discussion The role of antiseptics in wound care is controversial because in vitro studies have shown the cytotoxicity of such products.15,16 Despite these in vitro studies, the clinical relevance of this cytotoxic effect on wound healing is unknown and could be compensated for by their control of bacterial burden.17 Recently, the cytotoxic effect of 12 commonly used antiseptics on topical application in vitro to 2 human skin substitutes and a full-thickness autograft was assessed. Dermacyn was not cytotoxic in that study.18 Cytotoxicity was not assessed in the present study. However, we consider that time to healing could be considered as an indirect way to test the possible deleterious effect of DWC on wound healing. In our first series dealing with DFO, open wounds healed by secondary intention over a median period of 12.8 weeks. The median wound healing time in patients with successful conservative surgery was 11.4 weeks compared with 17.1 weeks for those who had minor amputations (P = .003).5 Wound healing was achieved by secondary intention after a median of 8 weeks in our more recent series.9 Healing time in cases of wide postsurgical wounds was 10.5 ± 5.9 weeks in the group treated with DWC versus 16.5 ± 7.1 weeks in the group treated with povidone iodine (P = .007).11 The median time to healing in the current investigation using DWC was 6.8 weeks, and this figure can be considered favorable. Even though we cannot state categorically that the improvement in healing time was due to the use of DWC, at least the healing time was not prolonged due to possible cytotoxicity. Our approach to dealing with DFO is to remove the infected bone if possible and leave the wound open to healing by secondary intention in most cases.2,5,19-21 We hypothesized that using a product that reduces the bacterial burden in bone and soft tissues could be beneficial. Two small single-center randomized controlled trials showed that superoxidized solution was useful for treating foot infections and wide postsurgical lesions.10,11 Another group reported that super-oxidized solution was at least as effective as oral levofloxacin for mild diabetic foot infections.22 In this small series, DWC was always in contact with the residual infected bone. It was applied by means of pulse irrigation or by filling the wound cavity with soaked gauzes. We also used closed lavage in cases in which the surgical wound had been closed; in 2 of these cases a wire had been inserted. Intraosseus lavage with DWC was used in midfoot and calcaneus osteomyelitis. This is a novel approach in our experience and the outcomes were favourable. Limb salvage was achieved in 100% of patients and only one required a minor amputation. Infection was eradicated in every patient included in this series and every wound healed. Only 1 recurrence was detected but the patient was treated with new conservative procedures, including the use of DWC through the wound. No amputation has been required to date in this complicated case. One of the possible advantages of using DWC as adjuvant treatment to surgery and antibiotics could be its potential to reduce the period of postoperative antibiotic treatment. IDSA guidelines recommend that in cases in which residual infected (but viable) bone remains, antibiotics should be administered for 4 to 6 weeks initially parenterally but possibly switching to oral administration depending on conditions.23 Definitive conclusions cannot be drawn from our preliminary experience but 4.4 weeks seems to be a short period compared with other investigations. Moreover, the microbiological isolates in this series were a little atypical. They included a high proportion of MRSA and gram-negative bacteria unlike most series in which Methicillin Sensitive Staphylococcus aureus (MSSA) was the most frequent isolate.1 Postoperative antibiotic treatment was given for a median of 5.1 weeks in our previously reported prospective series.9 The duration of antibiotic therapy was 10.1 ± 6.1 weeks in the group treated with DWC compared with 15.8 ± 7.8 weeks in the group treated with povidone iodine (P = .016) reported by Piaggesi’s group.11 The weaknesses of this study were the following: there was no control group, only short-term follow-up was conducted in 3 cases, and in our experience, at least 6 months should be considered to state that the bone infection has been eradicated. No bacteriological postoperative controls Downloaded from ijl.sagepub.com at INTERNATIONAL JOURNAL OF LOWER EXTREMITY WOUNDS on February 27, 2013 7 Aragón-Sánchez et al. Figure 6. (A) Minor defect in wound healing when the patient was discharged from hospital. (B) Recurrence of osteomyelitis after apparent wound healing was detected after 3 months of follow-up Figure 7. Progressive destruction of the soft tissue envelope and necrosis in case 11 requiring toe amputation were carried out. The strengths of the study are that it is the first time that DMC has been used in cases of residual bone infection after surgery for DFO, bone infections were always histopathologically confirmed, and patients underwent specialized treatment in cases with a high risk of amputation. Indeed, other teams had indicated several amputations, including 2 major. The good outcomes obtained in this preliminary study have encouraged us to design a trial to test the use of DWC in cases of bone infection. In conclusion, using DWC as adjuvant treatment in the postoperative period of surgery for DFO when the wound is open and bone margins may have residual infection is safe and may help eradicate the infection when combined with antibiotic treatment. Additional controlled studies are necessary to determine the precise role of DWC in the management of patients who have a high risk of amputation. Declaration of Conflicting Interests The author(s) declared the following potential conflicts of interest with respect to the research, authorship, and/or publication of this article: Free samples of Dermacyn Wound Care (Oculus Innovative Sciences Netherlands BV, Sittard, Netherlands) were received from the company to be tested in a series of patients. No payments or grants were received from the company. Funding The author(s) received no financial support for the research, authorship, and/or publication of this article. References 1. Game F. Management of osteomyelitis of the foot in diabetes mellitus. Nat Rev Endocrinol. 2010;6:43-47. 2. Aragon-Sanchez J. Treatment of diabetic foot osteomyelitis: a surgical critique. Int J Low Extrem Wounds. 2010;9:37-59. 3. Karchmer AW, & Gibbons GW. Foot infections in diabetes: evaluation and management. Curr Clin Top Infect Dis. 1994;14:1-22. 4. Ha Van G, Siney H, Danan JP, Sachon C, & Grimaldi A. Treatment of osteomyelitis in the diabetic foot. Contribution of conservative surgery. Diabetes Care. 1996;19:1257-1260. 5. Aragon-Sanchez FJ, Cabrera-Galvan JJ, & Quintana-Marrero Y, et al. Outcomes of surgical treatment of diabetic foot osteomyelitis: a series of 185 patients with histopathological confirmation of bone involvement. Diabetologia. 2008;51:1962-1970. Downloaded from ijl.sagepub.com at INTERNATIONAL JOURNAL OF LOWER EXTREMITY WOUNDS on February 27, 2013 8 The International Journal of Lower Extremity Wounds XX(X) 6. Frykberg RG, Wittmayer B, & Zgonis T. Surgical management of diabetic foot infections and osteomyelitis. Clin Podiatr Med Surg. 2007;24:469-482. 7. Zgonis T, Stapleton JJ, & Roukis TS. A stepwise approach to the surgical management of severe diabetic foot infections. Foot Ankle Spec. 2008;1:46-53. 8. Aragon-Sanchez J. Seminar review: a review of the basis of surgical treatment of diabetic foot infections. Int J Low Extrem Wounds. 2011;10:33-65. 9. Aragon-Sanchez J, Lazaro-Martinez JL, & HernandezHerrero C, et al. Does osteomyelitis in the feet of patients with diabetes really recur after surgical treatment? Natural history of a surgical series. Diabet Med. 2012;29:813818. 10. Martinez-De Jesus FR, Ramos-De la Medina A, & RemesTroche JM, et al. Efficacy and safety of neutral pH superoxidised solution in severe diabetic foot infections. Int Wound J. 2007;4:353-362. 11. Piaggesi A, Goretti C, & Mazzurco S, et al. A randomized controlled trial to examine the efficacy and safety of a new super-oxidized solution for the management of wide postsurgical lesions of the diabetic foot. Int J Low Extrem Wounds. 2010;9:10-15. 12. Aragon-Sanchez J, Quintana-Marrero Y, & Lazaro-Martinez JL, et al. Necrotizing soft-tissue infections in the feet of patients with diabetes: outcome of surgical treatment and factors associated with limb loss and mortality. Int J Low Extrem Wounds. 2009;8:141-146. 13. Eneroth M, Larsson J, & Apelqvist J. Deep foot infections in patients with diabetes and foot ulcer: an entity with different characteristics, treatments, and prognosis. J Diabetes Complications. 1999;13:254-263. 14. Aragon-Sanchez J, Lipsky BA, & Lazaro-Martinez JL. Diagnosing diabetic foot osteomyelitis: is the combination of probe-to-bone test and plain radiography sufficient for highrisk inpatients? Diabet Med. 2011;28:191-194. 15. Lineaweaver W, Howard R, & Soucy D, et al. Topical antimicrobial toxicity. Arch Surg. 1985;120:267-270. 16. Cooper ML, Laxer JA, & Hansbrough JF. The cytotoxic effects of commonly used topical antimicrobial agents on human fibroblasts and keratinocytes. J Trauma. 1991;31:775-782. 17. Hermanns JF, Paquet P, Arrese JE, Pierard-Franchimont C, & Pierard GE. The beneficial cytotoxicity of antiseptics. The controversial paradox of cutaneous ulcer treatment. Rev Med Liege. 1999;54:600-605. 18. Duc Q, Breetveld M, Middelkoop E, Scheper RJ, Ulrich MM, & Gibbs S. A cytotoxic analysis of antiseptic medication on skin substitutes and autograft. Br J Dermatol. 2007;157: 33-40. 19. Aragon-Sanchez J. Evidences and controversies about recurrence of diabetic foot osteomyelitis: a personal view and an illustrated guide for understanding. Int J Low Extrem Wounds. 2012;11:88-106. 20. Garcia-Morales E, Lazaro-Martinez JL, Aragon-Sanchez J, Cecilia-Matilla A, Garcia-Alvarez Y, & Beneit-Montesinos JV. Surgical complications associated with primary closure in patients with diabetic foot osteomyelitis. Diabet Foot Ankle. 2012;3. doi:10.3402/dfa.v3i0.19000. 21. Aragon-Sanchez J, Lazaro-Martinez JL, & HernandezHerrero C, et al. Surgical treatment of limb- and life-threatening infections in the feet of patients with diabetes and at least one palpable pedal pulse: successes and lessons learnt. Int J Low Extrem Wounds. 2011;10:207-213. 22. Landsman A, Blume PA, Jordan DA Jr, Vayser D, & Gutierrez A. An open-label, three-arm pilot study of the safety and efficacy of topical Microcyn Rx wound care versus oral levofloxacin versus combined therapy for mild diabetic foot infections. J Am Podiatr Med Assoc. 2011;101:484-496. 23. Lipsky BA, Berendt AR, & Cornia PB, et al. 2012 Infectious Diseases Society of America clinical practice guideline for the diagnosis and treatment of diabetic foot infections. Clin Infect Dis. 2012;54:e132-e173. 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Dermacyn® Effective in Treatment of Chronic Wounds with Extensive Bioburden while Reducing Local Pain Levels Debra J. Peterson, CCRN, MSN, ACNP-BC, Karre Hermann, MSN, APNP-BC, Jeffrey A. Niezgoda MD, FACEP, FACHM The Center for Comprehensive Wound Care and Hyperbaric Oxygen Therapy, Aurora St. Luke's Medical, Milwaukee, WI Hyperbaric & Wound Care Associates, Milwaukee, WI INTRODUCTION: Treatment of chronic and acute wounds requires comprehensive local wound care, which clearly must include bioburden control and pain management. Due to ease of use and relative cost efficiency, local antiseptic agents are widely used to assist with prevention of infection while maintaining a moist wound environment to promote healing. Super-Oxidized Water (SOW) has shown to be an effective antiseptic agent via a mechanism of denaturing cellular membrane proteins which results in the destruction of single-cell organisms while blocking the inflammatory process. Data suggests that SOW can reduce critical bioburden levels without affecting the host as human cell walls are not disrupted due to minimal cellular wall exposure.¹ This is a 57-year-old male with a 40 year history of painful bilateral lower extremity ulcerations that failed to heal despite topical enzymatic, cadexomer, and antimicrobial care. Also attempted were hyperbaric oxygen therapy, split thickness skin grafting, as well as vein stripping and compression therapy. Patient had multiple courses of systemic antibiotic therapy for MRSA, while pain management included scheduled doses of Oxycontin and Percocets for breakthrough pain. Dermacyn was initiated 6 months ago as a wound soak for 45 minutes and then the wounds were covered with a silver product. Pain levels were measured at each appointment. Figure 1 - Pre-Treatment R Medial Pain = 8 DISCUSSION: Historically SOW has had a short shelf life and was therefore under utilized. Recently, a stable, pH-neutral SOW was developed which has eliminated this storage problem. Dermacyn® is a SOW approved for the treatment of pressure ulcers, diabetic foot ulcers, stasis ulcers, post-surgical wounds, and burns. We present a series of 4 patients with recalcitrant wounds that failed to heal despite aggressive local wound care that included serial sharp debridement and local antimicrobial control. All wounds were subsequently treated with Dermacyn® soaks 30-45 minutes prior to resuming local wound care efforts. CASE #3 CASE #1 Figure 3 - Pre-Treatment L Medial Pain = 8 This is a 47-year-old male with a 7 year history of non-healing wounds secondary to venous insufficiency. Wounds failed to close despite compression therapy, Ultrasound Assisted Wound (UAW) therapy, cadexomer iodine and topical wound care as well as grafting. Pain was significantly reduced while wounds showed contracture while on Dermacyn®. Figure 7 - Pre-Dermacyn L Anterior Pain = 8 Figure 8 - 6 Weeks Post Dermacyn L Anterior Pain = 1 Figure 9 - Pre-Dermacyn L Lateral Pain = 8 Figure 10 - 6 Weeks Post Dermacyn L Lateral Pain = 1 Figure 2 - Current Appearance R Medial Pain = 0 (no meds) Figure 4 - Current Appearance L Medial Pain = 0 (no meds) CASE #2 This is a 79-year-old male seen on consult with a one year history of venous ulceration on the left medial ankle. Treatment prior to Dermacyn® included compression, vein stripping, ablation of deep vein, as well as enzymatic, cadexomer iodine and Negative Pressure Wound Therapy, all of which failed. Punch biopsy was performed to rule out malignancy. CONCLUSION: All patients showed contracture with progression to closure soon after the initiation of SOW. Additionally, these patients reported a significant reduction in pain levels with the addition of Dermacyn® therapy as measured using a numerical pain scale. We conclude that SOW is an effective antiseptic agent that assists with bioburden control and pain management while promoting wound healing. CASE #4 This is a 37-year-old female with a non-healing surgical abdominal wound. Wound was present for 18 months prior to consultation with healing compromised due to obesity and colonization with Pseudomonas. The wound failed to heal despite NPWT, local wound care, antibiotics and UAW therapy. Pain was also a problem prior to therapy, but patient was pain free with Dermacyn®. The wound progressed to closure in 2 months. REFERENCES: Bryant R. (2005). Super-oxidized water kills bacteria; demonstrates potential for healing. Dermatology Figure 5 - Pre-Dermacyn Pain = 4 Figure 6 - 12 Weeks Post Dermacyn Pain = 0 Figure 11 - Pre-Dermacyn Pain = 8 Figure 12 - 2 Months Post Initiation Pain = 0 Times. Efficacy and Compatibility of Combination Therapy with Super-Oxidized Solution* and a Skin Substitute§ for Lower Extremity Wounds Regulski M, Floros R, Petranto R, Migliori V, Alster H, Pfeiffer D Ocean County Foot & Ankle Surgical Associates. NJ. USA PURPOSE New technologically advanced wound care products, like skin substitutes and a super-oxidized solution, SOS*, have been shown to promote healing rates. Unfortunately, the evaluation of combined therapies is rarely undertaken. OBJECTIVE Evaluate the incidence of 100% closure (epithelialized) of non-infected, graft ready wounds treated with SOS and a skin substitute (SkS) § during the initial 12 weeks of the treatment period and additional 12 weeks of follow-up. METHODS Twenty-two patients with lower extremity wounds of at least 1 month were randomized to receive SOS or saline solution. SOS or saline solution (30 mL) was applied bid for three days before grafting the SkS and once everyday thereafter. A dressing saturated with either SOS or saline was placed in the wound bed and covered with a 2-layer compression system. The SkS was changed every week and assessments were conducted on Day 0, Day 3 and weekly thereafter for up to 12 weeks. Patient Demographics Control Group (N=9) Mean ± SD (Median) Study Group (N=9) Mean ± SD (Median) Age (in years) 70.22 ± 13.86 (72.00) 64.89 ± 18.81 (66.00) Gender (M/F) 7/2 1/8 Wound Type Control Group (N=9) Frequency Study Group (N=9) Frequency Post – traumatic 5 5 Post – surgical 1 3 Venous Ulcer 3 1 Baseline Wound Measurements Control Group (N=9) Mean ± SD (Median) Study Group (N=9) Mean ± SD (Median) Length (cm) 1.55 ± 0.91 (1.34) 2.22 ± 1.34 (1.83) Width (cm) 2.26 ± 1.04 (2.11) 1.94 ± 0.81 (1.69) 2 Area (cm ) 2.53 ± 1.37 (2.08) 3.39 ± 3.11 (1.62) Depth (cm) 2.89 ± 0.60 (3.00) 3.33 ± 0.71 (3.00) Volume (cm3) 0.71 ± 0.34 (0.62) 1.14 ± 1.21 (0.51) RESULTS Eighteen out of 22 patients were fully evaluable, 9 in each group (SOS or Saline). The median ulcer volumes were similar for the study group (0.51 cm3) and the control group (0.62 cm3). Despite of this the median number of weeks to complete epithelialization was shorter for the study group (6 weeks) than the control group (7 weeks). Before After 4 weeks Before After 4 weeks CONCLUSION SOS might induce a faster epithelialization than saline solution independently of the use of the OWM. Both technologies appear to be compatible. * Microcyn® Technology, Oculus IS, Petaluma, USA. § OASIS® Wound Matrix, HealthPoint, Texas, USA. The Heart Surgery Forum #2009-1162 13 (4), 2010 [Epub August 2010] doi: 10.1532/HSF98.20091162 Online address: http://cardenjennings.metapress.com Dermacyn® Irrigation in Reducing Infection of a Median Sternotomy Wound Abdul Ramzisham Rahman Mohd, MS, Mohd Khairulasri Ghani, MD, Raflis Ruzairee Awang, MBChB, Joanna Ooi Su Min, MMed, Mohd Zamrin Dimon, MS Heart and Lung Centre, Universiti Kebangsaan Malaysia Medical Centre, Kuala Lumpur, Malaysia Dr. Ramzisham ABSTRACT Background: Sternal wound infection is an infrequent yet potentially devastating complication following sternotomy. Among the standard practices used as preventive measures are the use of prophylactic antibiotics and povidone-iodine as an irrigation agent. A new antiseptic agent, Dermacyn® superoxidized water (Oculus Innovative Sciences), has recently been used as a wound-irrigation agent before the closure of sternotomy wounds. Methods: This prospective, randomized clinical trial was conducted to compare the effectiveness of Dermacyn and povidone-iodine in reducing sternotomy wound infection in patients undergoing coronary artery bypass graft surgery. Upon chest closure and after insertion of sternal wires, wounds were soaked for 15 minutes with either Dermacyn or povidone-iodine. Subcutaneous tissue and skin were then closed routinely. Patients were followed up, and any wound infection was analyzed. Results: Of the 178 patients, 88 patients were in the Dermacyn group, and 90 were in the povidone-iodine group. The mean (±SD) age of the patients was 61.1 ± 7.6 years. The incidence of sternotomy wound infection was 19 cases (10.7%). Five (5.7%) of these cases were from the Dermacyn group, and 14 (15.6%) were from the povidone-iodine group (P = .033). No Dermacyn-related complication was identified. Conclusion: We found Dermacyn to be safe and more effective as a wound-irrigation agent than povidone-iodine for preventing sternotomy wound infection. INTRODUCTION Patients undergoing coronary artery bypass grafting (CABG) surgery are exposed to possible sternal wound Received October 13, 2009; received in revised form November 8, 2009; accepted November 24, 2009. Correspondence: Associate Professor Dr. Mohd Ramzisham Abdul Rahman, Senior Lecturer/Consultant Cardiothoracic Surgeon, Heart and Lung Centre, Department of Surgery, Universiti Kebangsaan Malaysia Medical Centre, Jalan Yaacob Latif, 56000 Cheras, Kuala Lumpur, Malaysia; +603-91456201/2; fax: +603-91737831 (e-mail: [email protected]). E228 infection despite the procedure being categorized as clean surgery. Infections range from superficial infection to deeper infection of soft tissue, sternal osteomyelitis, and mediastinitis. The incidence of such infections ranges from 0.9% to 20% of cases [Loop 1990; Ulicny 1991], and the incidence of mediastinitis reported in most studies is 1% to 2% [El Oakley 1996; Bitkover 1998]. Although deep sternal wound infections, including mediastinitis, are infrequent complications, such infections are associated with significant morbidity and mortality (between 9.8% and 14% [Loop 1990; Ulicny 1991; El Oakley 1996]), prolonged hospitalization, and an increased financial burden on the health care system [Vegas 1993]. Therefore, it is vital to prevent infection of the sternotomy wound following surgery. Among the standard practices for prevention are the prophylactic use of antibiotics and placement of the pericardial drains to facilitate drainage of blood and serous fluid. These practices have been shown to lower the rate of deep sternal infection [Blanchard 1995]. Agents that irrigate the dermal wound have also been used to facilitate removal of debris before wound closure. One common practice is the use of saline and antiseptics such as povidoneiodine as wound-irrigation agents. Dermacyn® (Oculus Innovative Sciences, Petaluma, CA, USA), a superoxidized aqueous solution, is a new antiseptic agent cleared by the US Food and Drug Administration (FDA) that has recently been used in our center as an irrigation agent during closure of sternotomy wounds. The initial experience has been good with no adverse effects. Dermacyn is produced by means of a proprietary electrolysis process that uses a sophisticated, unique multichamber system. The solution is manufactured from purified water and sodium chloride. The electrolysis process separates and captures the ions to produce a sterile, pH-neutral solution consisting of a stable controlled formula of reactive oxygen and chlorine species [Oculus Innovative Sciences 2006]. The active compounds in Dermacyn are 99.9% water, hypochlorous acid, and sodium hypochlorite, and the inactive compounds are sodium carbonate, sodium hydroxide, hydrogen peroxide, chlorine dioxide, and ozone. Dermacyn wound-irrigation agent is applied to chemically debride, flush, and cleanse the wound surface. The reactive oxygen species assist in promoting the body’s own healing process by reducing the microbial load and via the agent’s Dermacyn Irrigation in Reducing Infection—Ramzisham et al moistening properties [Bongiovanni 2006]. Several studies have found Dermacyn to be safe and to exhibit a wide antimicrobial spectrum [Landa-Solis 2005]. Paola et al [2006] proved the effectiveness of Dermacyn over povidone-iodine and Dermacyn’s safety in treating the infected diabetic foot. A study of another series by Ohno et al [2000] showed that superoxidized water had no adverse effect on hemodynamics and was safe when used as a mediastinal irrigation solution during open heart surgery. Dermacyn has also been subjected to a series of biocompatibility tests. These tests were performed in accordance with the requirements set forth by International Standards Organization and FDA guidelines. The product has also been tested in animal models to satisfy FDA requirements and to ensure its safety and efficacy [Oculus Innovative Sciences 2006]. Several studies, however, have identified various factors that contribute to wound infection that always need to be considered [Zacharias 1996]. We therefore conducted this study to compare the effectiveness of Dermacyn irrigation and povidone-iodine with respect to reducing the incidence of sternotomy wound infection following CABG. We also evaluated our study population to identify the potential risk factors for sternotomy wound infection following CABG. MATERIALS AND METHODS This prospective randomized clinical trial took place between June 2007 and December 2008. All patients scheduled for elective CABG were included in this trial. Exclusion criteria included emergency cases, patients who underwent other surgical procedures in addition to CABG, patients allergic to Dermacyn, and those who had infective or other skin lesions over anterior chest wall area. Informed consent was obtained, and patients were consecutively randomized into 2 groups. Group A patients used Dermacyn as a woundirrigation agent, and group B patients used povidone-iodine. All patients received intravenous prophylaxis with 1.2 g Augmentin (amoxicillin and clavulanate) at induction. Before the surgery, the skin was cleaned with 10% povidone-iodine solution and covered with polyurethane drapes. CABG was performed through a standard median sternotomy by 3 cardiothoracic surgeons. Upon closure, bone wax and diathermy were used sparingly for hemostasis. Two drains were normally left in the mediastinal cavity. The sternum was closed primarily with no. 6 steel wires in a figure-of-eight manner. In both groups, the wounds were then soaked for 15 minutes after insertion of the sternal wires. The subcutaneous tissue was closed in 2 layers with polyglactin 1-0 suture, and the skin was closed subcuticularly with Vicryl 3-0 suture. Diagnosis and Follow-up The sternotomy wounds were inspected on postoperative day 2 and daily until discharge. Patients were then followed up at 2, 4, and 6 weeks postoperatively to assess for the presence of wound infection and Dermacyn side effects. The primary outcome was the presence of sternotomy wound infection, which was defined according to the Centers for Disease Control and Prevention in their National Nosocomial Infections © 2010 Forum Multimedia Publishing, LLC Table 1. Clinical Characteristics of the 178 Patients Involved in the Trial* Age, y 61.1 ± 7.6 Male-female ratio 3.24:1 COPD, n 66 (37.1%) Diabetes mellitus, n 79 (44.4%) ESRF, n 32 (18.0%) 2 Obesity (BMI >30 kg/m ), n 20 (11.2%) Smoking, n 66 (37.1%) On-pump/off-pump surgery, n 153 (86%)/25 (14%) Bypass time, min 99.0 ± 7.4 IMA harvested, n 171 (96.1%) *Data are presented as the mean ± SD where indicated. COPD indicates chronic obstructive pulmonary disease; ESRF, end-stage renal failure; BMI, body mass index; IMA, internal mammary artery. System [Horan 1992]. Wound infections were graded according to Horan et al [1992] as superficial (involving the skin and subcutaneous tissue of the incision), deep (involving fascia, muscle layers, and sternum), or deep organ space. The nature of discharge from the wounds was documented, and samples of the discharge were sent for culture and sensitivity analyses. The risk factors for wound infections were identified and analyzed. Ethical Considerations This trial involved no financial gain and was approved by the research and ethics committee of the Universiti Kebangsaan Malaysia Medical Centre (code no. FF-236-2007). Statistical Analysis Data were stored and analyzed with SPSS software (version 12.0; SPSS, Chicago, IL, USA). Continuous and categorical variables were analyzed with the independent Student t test and the chi-square test, respectively. With the use of 95% confidence intervals, a P value <.05 was considered statistically significant. RESULTS We recruited 190 patients for this trial, 95 patients in each group. Twelve patients, however, dropped out owing to postoperative mortality (4 cases, 2 deaths due to poor left ventricular function of <20% and 2 deaths due to cerebrovascular accident) and chest reopened for bleeding (8 cases), leaving 178 patients remaining in the study. Eighty-eight patients were in group A, and 90 were in group B. The mean (±SD) age of the patients was 61.1 ± 7.6 years. The male-female ratio was 3:1. Diabetes mellitus and chronic obstructive pulmonary disease (COPD) were the most common comorbidities (44.4% and 37.1%, respectively). End-stage renal failure (ESRF) and obesity were documented in 18% and 11.2% of cases, respectively. On-pump CABG surgery was performed in 86% of the patients with a mean cardiopulmonary bypass E229 The Heart Surgery Forum #2009-1162 Table 2. Comparison of Comorbidities and Intraoperative Details for the 2 Randomized Groups* Group A Group B P Age, y 62.61 ± 7.7 59.54 ± 7.2 .085 COPD, n 35 (39.8%) 31 (34.4%) .462 Diabetes mellitus, n ESRF, n 39 (44.3%) 40 (44.4%) .986 8 (9.1%) 24 (26.7%) .002 Obesity (BMI >30 kg/m2), n 7 (8.0 %) 13 (14.4%) .170 Smoking, n 35 (39.8%) 31 (34.4%) .462 73 (83%) 80 (88.9%) .255 Bypass time, min 98.14 ± 9.8 99.83 ± 9.9 .076 IMA harvested, n 84 (95.5%) 87 (96.7%) .677 On-pump surgery, n *Data are presented as the mean ± SD where indicated. COPD indicates chronic obstructive pulmonary disease; ESRF, end-stage renal failure; BMI, body mass index; IMA, internal mammary artery. time of 99 ± 7.4 minutes. A single internal mammary artery (IMA) was harvested in 171 patients (96.1%). Table 1 summarizes the patients’ clinical characteristics. The 2 groups were well matched with respect to demographic, comorbidity, and intraoperative variables. The only exception was ESRF, which was more prevalent in group B (26.7%) than in group A (9.1%). Table 2 summarizes the comparison of comorbidities and intraoperative details of the 2 groups. Sternal wound infections occurred in 19 patients (10.7%). Group B had a significantly higher incidence of infections (14 patients, 15.6%) than group A (5 patients, 5.7%) (P = .033, Table 3). All 5 patients who developed sternotomy wound infections in group A had superficial infections. Of the 14 patients in group B who developed infections, 10 had superficial infections, and the remaining 4 patients had deep sternal wound infections that led to sternal dehiscence requiring surgical debridement and repair (Table 4). Bacteriology Swabs for microbiology analyses were obtained from infected wounds. Of the 19 patients with infections, 11 patients had significant microbial growth. Six patients had Staphylococcus aureus infection, and Acinobacter sp was cultured in 2 patients. Pseudomonas aeroginosa, Staphylococcus epidermidis, and coagulase-negative Staphylococcus were isolated from 1 patient each. Risk Factors Risk factors were divided into preoperative, intraoperative, and postoperative factors. A univariate analysis of all preoperative variables revealed that COPD, diabetes mellitus, ESRF, obesity (body mass index >30 kg/m2), and smoking were associated with an increased risk of sternal wound infection at a P level of <.05. In this study, however, age was not significantly associated with sternal wound infection. Intraoperatively, we studied 3 variables: the CABG technique, the use of an IMA, and bypass time. Bypass time was significantly associated with E230 Table 3. Incidence of Sternal Wound Infection in the 2 Groups* Sternal Wound Infected Not Infected Total Group A, n 5 (5.7%) 83 (94.3%) 88 Group B, n 14 (15.6%) 76 (84.4%) 90 *P = .033. Table 4. Type of Infection Type of Infection Superficial Deep Total Group A, n 5 (100%) 0 (0%) 5 Group B, n 10 (71.4%) 4 (28.6%) 14 the risk of infection (P < .001). Patients who had an IMA harvested, however, were not associated with an increased risk of infection. The duration of mechanical ventilation and the length of intensive care unit (ICU) stay, which were studied as postoperative risk factors of sternotomy wound infection in this trial, were significantly associated with an increased risk of wound infection (Table 5). DISCUSSION It is imperative to reduce the risk of sternotomy wound infection, and many standard practices have been described, including meticulous scrubbing, skin preparation, use of disposable prefabricated drapes, use of prophylactic antibiotics, and placement of the pericardial drains to facilitate the drainage of blood and serous fluid. The standard practice of irrigating the wound before closure with either normal saline or povidone-iodine has long been used in most centers. The efficacy of Dermacyn for antiseptic wound irrigation during closure of the sternotomy wound following open heart surgery has yet to be proven, however. As an antiseptic solution, it exerts a wide range of antimicrobial activities [Landa-Solis 2005; Oculus Innovative Sciences 2006]. It can also be applied when chemically debriding and cleansing wound surfaces. The mechanism of action in eradicating various microorganisms is well documented [Landa-Solis 2005]. The active agents in Dermacyn consist of both chlorine and reactive oxygen ions. It is believed that the bactericidal action exhibited by Dermacyn is due to the combined effect of these species. The reactive oxygen species is also beneficial in the wound-healing process. Both ions are often referred to as free radicals. A number of different types of free radicals are also produced naturally by the body. Many free radicals provide significant health benefits, whereas free radicals produced by radiation can be destructive in nature. Conversely, Dermacyn ions and free radicals are biological and are not produced by radiation. Dermacyn technology has been tested in accordance with FDA requirements to ensure that such free radicals do not cause any harm. Dermacyn Irrigation in Reducing Infection—Ramzisham et al Table 5. Univariate Analysis of Preoperative Risk Factors for Sternal Infection* Infected (n = 19) Not Infected (n = 159) P Age, y 63.0 ± 7.5 60.8 ± 7.6 .257 COPD, n 11 (57.9%) 55 (34.6%) .047 Diabetes mellitus, n 13 (68.7%) 66 (41.5%) .026 ESRF, n 11 (57.9%) 21 (13.2%) .000 Obesity, n 16 (84.2%) 4 (5.2%) .000 Smoking, n 11 (57.9%) 55 (34.6%) .047 108.05 ± 11.5 96.4 ± 7.9 .000 IMA harvest, n 17 (89.5%) 154 (96.9%) .118 Ventilation duration, h 10.84 ± 6.1 4.21 ± 0.7 .000 Length of ICU stay, d 2.63 ± 0.76 2.06 ± 0.23 .000 Bypass time, min Infected ESRF *Data are presented as the mean ± SD where indicated. COPD indicates chronic obstructive pulmonary disease; ESRF, end-stage renal failure; IMA, internal mammary artery; ICU, intensive care unit. In this study, the 5 patients from the Dermacyn group with infected wounds had only superficial infections, and they required only a course of antibiotics and wound dressings to treat the infection and to promote wound healing. Four patients from the povidone-iodine group with wound infection, however, had associated deep wound infection and sternal dehiscence. These patients required surgical debridement and sternal rewiring. The remaining 10 patients had superficial infections and were successfully treated with antibiotics and wound dressings. Obviously, these findings indicate that the Dermacyn group not only had a lower infection rate but also had less severe infections than the povidone-iodine group. Many studies have demonstrated that the development of sternotomy wound infection following CABG is multifactorial. Most authors have classified the risk factors into 3 groups: preoperative factors pertaining to the patient, such as advanced age and comorbidities; intraoperative factors, such as a prolonged duration of operation and the need for reoperation; and postoperative factors, such as the use of an intra-aortic balloon pump, massive blood transfusion, and prolonged mechanical ventilation [Oschner 1972]. In the present study, we assessed 11 variables as potential risk factors for sternotomy wound infection. A univariate analysis was performed to determine the associations between these risk factors and wound infection (Table 5). In contrast to the results of other studies, we found that age was not significantly associated with sternotomy wound infection (P = .257). This result was probably because most of our patients were younger than 65 years, whereas several studies have shown that older patients are at high risk of developing wound infection. Our analysis revealed that patients who had diabetes mellitus and were obese were more likely to develop sternotomy wound infection, a result that is concordant with other studies [Moulton 1996]. Cigarette smoking and COPD were also identified as significant risk factors for © 2010 Forum Multimedia Publishing, LLC Table 6. Infected Cases among ESRF and non-ESRF Patients* Non-ESRF Total Group A, n 3 (60%) 2 (40%) 5 Group B, n 8 (57.1%) 6 (42.9%) 14 *Odds ratio, 1.1. sternal wound infection following CABG. More than 60% of our patients with infection were smokers and had COPD. The latter is frequently associated with an increased risk of postoperative mediastinitis, probably because of frequent coughing, which may contribute to wound dehiscence and thereby facilitate bacterial migration into the mediastinum. Patients with COPD also experience respiratory infections more frequently and prolonged weaning from mechanical ventilation [Grossi 1991]. The details of the pathophysiology causing the infection in these patients are beyond the scope of this report. In this trial, we were unable to match the 2 groups with respect to the number of ESRF patients. ESRF was more prevalent in group B. The overall incidence of sternal wound infection in ESRF patients was statistically significant (P < .001). Further analysis, however, revealed that 3 (60%) of the 5 patients in group A and 8 (57.1%) of the 14 patients in group B who developed infection had ESRF. The calculated odds ratio between these 2 groups was 1.1, implying that sternotomy wound infection is equally likely to occur in the 2 groups, regardless of the type of irrigation used (Table 6). We observed that 14 patients (82.4%) who developed wound infection had bypass times >110 minutes. This finding was supported by the statistical analysis, which showed bypass time to be significantly associated with the risk of sternal infection. This finding is similar to findings from most other studies, which showed a higher incidence of sepsis with longer bypass times [Loop 1990]. Many studies have linked the use of IMA grafts to a higher incidence of sternal infections [Hirotani 2002]. Of the 19 patients who developed sternotomy wound infection in the present study, 17 (89.5%) had a single IMA harvested. This incidence showed that this variable did not make a significant contribution to the risk of sternal infection. Decreased sternal perfusion has been hypothesized as a cause for an increased incidence of sternal infections following use of an IMA [Carrier 1992]. Sofer et al [1999] showed that after bilateral IMA dissection, sternal perfusion was reduced by a mean of 24% ± 6%. Four weeks postoperatively, sternal blood flow was still reduced by a mean of 2% ± 2%. This low sternal perfusion may lead to tissue necrosis, and hence to an impaired wound-healing process. We tried to avoid bilateral IMA use unless it was absolutely necessary. Our results also confirmed that prolonged mechanical ventilation and a longer ICU stay were significantly associated with wound infection. All of our patients were ventilated and observed postoperatively in the cardiac beds in the general ICU rather than in a dedicated cardiac ICU. The risk of infection is generally greater in the E231 The Heart Surgery Forum #2009-1162 general ICU. These patients not only are exposed to endogenous infection (ie, normal commensals) but also are prone to contract infections from exogenous sources, such as the staff, other patients, and visitors. This trial had certain limitations. The number of participants in this study was relatively small because our center is not the main center for cardiothoracic surgery in this region; thus, the logistics allowed for only a limited number of operations. In addition, the variables for risk factors for sternotomy wound infections were limited to 11. Although all patients underwent their operations with the same cardiothoracic surgical teams, we were not able to appoint a dedicated surgeon for closure of sternotomy wounds. Some nonobjective factors that might also have influenced this study include the number of staff and medical students entering and exiting the theater during surgery. This trial, however, suggests that the use of Dermacyn as a wound-irrigation agent at closure of sternotomy following CABG reduced the incidence of wound infection, compared with povidone-iodine. There were no adverse side effects associated with the use of Dermacyn. Therefore, it can be used without hesitancy in view of its effectiveness and safety as a wound-irrigation agent. REFERENCES Bitkover CY, Gårdlund B. 1998. Mediastinitis after cardiovascular operations: a case-control study of risk factors. Ann Thorac Surg 65:36-40. Blanchard A, Hurni M, Ruchat P, Stumpe F, Fischer A, Sadeghi H. 1995. Incidence of deep and superficial sternal infection after open heart surgery. A ten years retrospective study from 1981 to 1991. Eur J Cardiothorac Surg 9:153-7. Bongiovanni CM. 2006. Superoxidized water improves wound care outcomes in diabetic patients. Diabet Microvasc Complications Today May/ June:11-4. Carrier M, Grégoire J, Tronc F, Cartier R, Leclerc Y, Pelletier LC. 1992. Effect of internal mammary artery dissection on sternal vascularization. Ann Thorac Surg 53:115-9. El Oakley RM, Wright JE. 1996. Postoperative mediastinitis: classification and management. Ann Thorac Surg 61:1030-6. Grossi EA, Esposito R, Harris LJ, et al. 1991. Sternal wound infections and E232 use of internal mammary artery grafts. J Thorac Cardiovasc Surg 102:342-6. Hirotani T, Shirota S, Cho Y, Takeuchi S. 2002. Feasibility and suitability of the routine use of bilateral internal thoracic arteries. Ann Thorac Surg 73:511-5. Horan FC, Gaynes RP, Martone WJ, Jarvis WR, Emori TG. 1992. CDC definitions of nosocomial surgical site infections, 1992: a modification of CDC definitions of surgical wound infections. Infect Control Hosp Epidemiol 13:606-8. Landa-Solis C, González-Espinosa D, Guzmán-Soriano B, et al. 2005. Microcyn: a novel super-oxidized water with neutral pH and disinfectant activity. J Hosp Infect 61:291-9. Loop FD, Lytle BW, Cosgrove DM, et al. 1990. J. Maxwell Chamberlain memorial paper. Sternal wound complications after isolated coronary artery bypass grafting: early and late mortality, morbidity, and cost of care. Ann Thorac Surg 49:179-87. Moulton MJ, Creswell LL, Mackey ME, Cox JL, Rosenbloom M. 1996. Obesity is not a risk factor for significant adverse outcomes after cardiac surgery. Circulation 94(suppl):II87-92. Oculus Innovative Sciences. 2006. Technical description of DERMACYN™. Microcyn™ Technology, Inc. Petaluma, Calif.: Oculus Innovative Sciences. Ohno H, Higashidate M, Yokosuka T. 2000. Mediastinal irrigation with superoxidized water in open-heart surgery: the safety and pitfalls of cardiovascular surgical application. Surg Today 30:1055-6. Oschner JL, Mills NL, Woolverton WC. 1972. Disruption and infection of median sternotomy incision. J Cardiovasc Surg 13:394-9. Paola LD, Brocco E, Senesiet A, et al. 2006. Super-oxidized solution (SOS) therapy for infected diabetic foot ulcers. Wounds 18:262-70. Sofer D, Gurevitch J, Shapira I, et al. 1999. Sternal wound infections in patients after coronary artery bypass grafting using bilateral skeletonized internal mammary arteries. Ann Surg 229:585-90. Ulicny KS Jr, Hiratzka LF. 1991. The risk factors of median sternotomy infection: a current review. J Card Surg 6:338-51. Vegas AA, Jodra VM, García ML. 1993. Nosocomial infection in surgery wards: a controlled study of increased duration of hospital stays and directs cost of hospitalization. Eur J Epidemiol 9:504-10. Zacharias A, Habib RH. 1996. Factors predisposing to median sternotomy complications. Deep vs superficial infection. Chest 110:1173-8. Microcyn and HAIs Saturday, January 22, 2011 Expanded article - A New Approach to the Problem of Antibiotic Resistance A NEW APPROACH TO THE PROBLEM OF ANTIBIOTIC RESISTANCE AND HOSPITAL ACQUIRED INFECTIONS by Susanne Lewis Hospital-acquired infections, (HAIs), have become far too prevalent across the nation. The statistics are grim. Ten million patients will get an infection in a health care institution over the next five years and half a million of them will die. Many people fear going to the hospital because that is where the most virulent antibiotic-resistant microbes are encountered. A vast study conducted several years ago found that about half of the patients in ICU wards around the world are battling some kind of acquired infection. This study looked at data from 1,265 ICU units in 75 countries. The longer the patients stayed in the ICU, the greater the risk of becoming infected—from a 32% chance for patients staying one day or less to a 70% chance for patients staying more than seven days. These patients were more than twice as likely to die than non-infected patients. This problem accounts for about 40% of total ICU health care costs. There is a great need for new approaches to be developed to cope with resistant superbugs. Appropriately, our age is described as being on the cusp of the “post-antibiotic era.” Microbes in the community are also becoming resistant. Superbugs are on everyone’s mind. MRSA, multi-drug resistant staph aureus, has taken a terrible toll, and continues to proliferate, generating a wide range of afflictions. At this point four million people have MRSA on the skin or in the nostrils, and that is a very conservative estimate. New strains of resistant staph are continuing to show up, especially in livestock. It has been dubbed “the perfect pathogen.” But it has competition for that title. Pseudomonas Aeruginosa, C. Diff, actinobacter, e. coli and many others are developing super strains. One woman, who was infected in a wound care clinic where she had been referred for postoperative care, became colonized by three different superbugs—MRSA, pseudomonas and actinobacter. Five years later she still has an open wound in her abdomen where mesh to repair hernia had been inserted. She is on maintenance doses of very strong antibiotics and running out of options. After 60 years of reliance on antibiotics, we are slowly descending into an iatrogenic nightmare that has already reached epidemic proportions and seems to be heading in the direction of a pandemic of global proportions. How has this happened? We all know about the over-use of antibiotics as a causative factor. But that only accelerated the problem, it did not cause it. To understand the cause, we need to understand the nature of antibiotics themselves. Bacteria naturally produce their own antibiotics as a means of controlling competing bacteria for nutritional resources. In other words, they engage in a kind of “chemical warfare” against each other. Medical science simply learned to isolate the antibiotics directly from the microbes, and then to synthetically produce them. But microbes that produce antibiotics themselves have the ability to withstand them, at least potentially. This is one of their primary survival strategies, built into their DNA. And antibiotic resistance can be transmitted from one species to another via plasmids. So, in the long run, medical science can never win out against microbes with antibiotics because it is trying to fight them on their own turf with their own weapon. And superbugs are here to call our bluff. In The Coming Plague, Laurie Garrett writes: …the use of antibiotics in colonies of bacteria in which even 1% of the organisms were genetically resistant could have tragic results. The antibiotics would kill off the 99% of the bacteria that were susceptible, leaving a vast nutrient-filled Petri dish free of competitors for the surviving resistant bacteria. Like weeds that suddenly invaded an untended open field, the resistant bacteria rapidly multiplied and spread out.” At this point, we struggle with infections nearly impossible to treat as a direct result of the use of antibiotics—not just staph but pneumonia, strep, dysentery, even resistant strains of tuberculosis and gonorrhea. The list is getting long. The problem is further exacerbated by the widespread use of antiseptics, antibacterials and disinfectants, such as BSK (benzalkonium chloride), CHG (chlorhexidine gluconate), povidone iodine, dilute bleach, triclosan, and more. Unfortunately, microbes are able to develop resistance to these chemicals in the same way they develop resistance to antibiotics. A recent study showed microbes resistant to BSK could withstand concentrations of BSK up to 400 times greater than nonmutated strains. More importantly, these same microbes also developed resistance to ciprofloxacin, even though they had never been exposed to the drug. The conclusion is that these chemicals proliferate antibiotic resistance. In addition to this, some people are allergic to certain antibacterials, such as CHG, the severest reaction documented being anaphylactic shock. Some health care professionals are against the use of well known antiseptics in wound treatment because of the proven cytotoxicity in dermal and epidermal cells. Antibiotic toxicity is also becoming a serious issue. Antibiotics have been shown to be chondrotoxic, damaging the body’s cartilage; neurotoxic, carcinogenic, toxic to the retina, nephrotoxic, and more. In fact, antibiotics are the principle cause of drug-associated nephropathy. The FDA was compelled to put out a black-box warning on Cipro as well as Gentamicin, for tendon rupture, ototoxicity and renal damage. A black box warning is the strongest form of warning issued by the FDA about a drug, the step taken just short of removing the drug from the market. A 2008 study conducted by the CDC revealed more than 140,000 incidences of bad reactions to antibiotics which resulted in visits to the Emergency Room each year in the US. Far more reactions go unreported, so there’s really no way of telling what the actual figure is. It is not mere speculation that prolonged and repeated exposure to antibiotics induces cancer. A 2008 two-year study of three million people who were tracked for six years, concluded those who had taken two to five prescriptions during that two year period had a 27% increase in cancers compared to those who took none. Those who took six or more prescriptions had a 37% increase in cancers. This study was published in the International Journal of Cancer. And a National Cancer Institute study found that the number of breast cancers doubled among women who took more than 25 antibiotic prescriptions over 17 years—usually for recurrent urinary tract infections and acne. This study was published in the Journal of American Medical Association. Antibiotics tend to have a down-regulating effect on the immune system, some even interfering with lymphocyte blastogensis. Macrobid (nitrofurantoin), often used for urinary tract infections, is one of many implicated here. Destruction of intestinal microflora is also a contributing factor. A repetitive cycle of illness and dependency on repeated use of antibiotics results. People repeatedly treated with antibiotics have far more infections than those not treated this way, especially children with recurring ear infections. Studies have clearly demonstrated that long-term use of antibiotics does indeed suppress the immune system. This is in part due to the fact that antibiotics do not reinforce the immune system but rather replace one of its functions—the destruction of bacteria. This causes that arm of the immune system to weaken. Thus re-infection is easily succumbed to. Combine this syndrome with the problem of virulent resistant strains of microbes and you have a recipe for disaster. A paradigm shift is greatly needed in order to devise new approaches to the problems of infectious disease. One important idea is to shift emphasis from attacking the microbe at the expense of the immune system, to attacking the microbe in a way that enhances the immune system. There are many approaches. One is to use recombinant electrolyzed salt water which is laden with freefloating electrically charged oxygen ions with traces of hypochlorous acid--an antimicrobial solution that mimics the way neutrophils work to eradicate infection through what is known as oxidative burst. During oxidative burst, also known as respiratory burst, or phagocytosis, white blood cells surround the pathogen and release hypochlorous acid and reactive oxygen to burst through the cell wall of the pathogen, annihilating it, (lysis). This does not harm mammalian tissue, because its cells are tightly bound together in a matrix that cannot be affected by oxidative burst. Working in tandem with the immune system, this solution, dubbed nSOS, or pH neutral superoxidized solution, is highly effective against a very broad spectrum of pathogens, including abx resistant bacteria, viruses, fungi and spores. It is neither acidic nor alkaline, but has a neutral pH of between 6.2 and 7.8, depending on the formula. It is an entirely new class of antiseptic. In an article entitled, “The Science Behind Stable, Super-Oxidized Water,” Dr. Andres A. Gutierrez describes what he calls “the dawn of a new solution.” Tests have documented in-vitro antimicrobial activity of an astonishing thoroughness. Toxicology studies are cited that confirm nSOS is neither genotoxic nor cytotoxic. Worldwide approvals have been coming steadily forth for about eight years, in Europe, Canada, United States, Mexico, Russia, China, Middle Eastern countries, India, and more. It has been used in such a wide variety of applications as to be bewildering—everything from cleaning and treating cuts and scrapes to treating periodontal disease to mediastinal irrigation after open heart surgery. It has also been used to treat peritonitis and abscesses of the abdominal cavity, as well as bladder infections, urethritis, cystitis, ear infections, acne, and much much more. The list of potential uses is long indeed. Much of this is being done in an off-label capacity, pending clearance by regulatory agencies. But approvals are coming steadily through each year. The use of nSOS as a treatment for infection as well as a preventative to keep infection from occurring, is gradually being recognized by the medical community as a very promising alternative to over-reliance on antibiotics. It is being used in wound care clinics, limb salvage clinics, diabetic foot clinics, and burn clinics, with wonderful results. Much of the research to date has been with diabetic foot ulcers. These wounds are particularly difficult to treat and often result in amputation, which is known to significantly shorten the lifespan of many undergoing this procedure. Non healing diabetic wounds are highly vulnerable to opportunistic infections, which are more and more frequently turning out to be MDROs—multi drug resistant organisms, or superbugs. Treatment with neutral superoxidized water has been highly effective with these kinds of wounds, reducing bacterial load, enhancing local blood supply, promoting neovascularity and reducing inflammation. Neutral superoxidized water is marketed in Europe, Russia and China as Dermacyn, in India as Oxum, and in the United States and Mexico as Microcyn, Puracyn, Vetericyn, Periocyn and MyClynse. It is the first neutral superoxidized water to be stable enough to make it practical for widespread use, with a shelf life of approximately two years. One study demonstrated that Microcyn was effective in treating chronic wounds with extensive bioburden, and it discovered that local pain levels were substantially reduced when this treatment was used. And there have been a number of clinical studies comparing povidone iodine and Microcyn in wound care. They have been done in the United States, Italy, India and Mexico, to name just four. Wounds range from diabetic foot ulcers, venous stasis ulcers, pressure sores, surgical site infections and burns. Here is info on just one of those studies: STUTTGART, Germany -- 218-patient study indicates Microcyn(TM) Technology superior in microbial load reduction, healing time, surgical dehiscence and adverse side effects as compared to povidone iodine (10%). Oculus Innovative Sciences, Inc. announced that Dr. Luca Dalla Paola, a surgeon with the diabetic foot unit of the Abano Terme Hospital in Italy, recently presented the results of a 218-patient controlled clinical study that assessed the safety and efficacy of Microcyn(TM) Technology in treating diabetic foot ulcers as compared to povidone iodine (10%) antiseptic, which is often used as the "standard care" in treatment of open wounds. In the study, the Microcyn(TM) Technology proved superior to the iodine relative to the reduction of the number of bacterial strains, local adverse effects, surgical dehiscence (incidence of not healing after surgery due to infection or ischemia) and healing time. The key endpoint of the study was microbial load reduction at both entry and at surgery (or follow-up). The Microcyn(TM) Technology showed a significantly improved rate of reduction of microbial load and healing time in open wounds as compared to the povidone iodine group. 88.2% of ulcers in the Microcyn(TM) group had a negative microbiological specimen versus 68.5% of ulcers in the povidone iodine group. The Microcyn(TM) group showed no local adverse effects, while the povidone iodine group experienced 18 incidences of such effects. All studies indicated more complete wound healing, healthier appearance, absence of odor, reduction of inflammation, and greater presence of granulation tissue. Microcyn’s well-established “safe as saline” profile assures there will be no irritation or other adverse reactions when used. There is a lot of misinformation circulating about neutral superoxidized solution, but research clearly refutes these myths. First, nSOS is NOT dilute bleach in any way shape or form. Dakins solution is dilute bleach, and amongst enlightened wound care clinicians, it went out with the dinosaurs. Dakins solutions is damaging to mammalian tissue and is in well deserved disrepute because of this. It was used in the Vietnam war, but that was before research proved that it destroys fibroblasts, which are crucial to wound closure. When compared to dilute bleach, studies show that nSOS is actually a much more powerful antimicrobial solution of oxidants that are actually able to eliminate bleach resistant bacteria, without damaging fibroblasts in any way. Another myth circulating about nSOS is that it is only another manifestation of so-called ionized water. Medical grade nSOS is hardly the alkaline ionized water people drink thinking it will do everything from making them younger to curing cancer. Stable nSOS is made through an intricate proprietary method that requires considerable scientific sophistication to master. So far only one company has been able to do it successfully, Oculus Innovative Sciences, located in Petaluma, California. In-vitro studies have shown nSOS to eradicate viruses as well as bacteria--poliovirus-1, rhinovirus1A, herpes simplex viruses 1 & 2, influenza H1N1, West Nile virus, HIV, and many others. In addition to all the drawbacks I have mentioned in this article, one of the major limitations to antibiotic therapy is that antibiotics are helpless against viruses. Viruses have always been superbugs. In an article on bacterial complications from burn wounds by Dr. Ariel Miranda Altamirano, Microcyn60, the nSOS product used in Mexico, was evaluated. He indicated that infection by superbugs in burn wounds has been steadily increasing in recent years, retarding overall wound healing, elevating the rate of graft loss and increasing mortality from sepsis. He concluded that: As predicted from previous animal and clinical experience, the use of Oculus Microcyn60 was efficient and safe for the prevention of partial- and full-thickness burn infections in pediatric patients. Treatment with Oculus Microcyn60 reduced the microbial load in 90% of patients with partial- and full-thickness thermal injuries. Children also reported less pain during cleaning procedures. Application was easy and inexpensive. In addition, the length of hospital stay of patients treated with Oculus Microcyn60 was reduced by 50% relative to the control. Considering that the daily hospital cost at this facility is approximately $1,800 US per patient, treatment with Oculus Microcyn60 saved the institution an average of $24,660 US per patient. The results of this study also suggest that burns treated with Oculus Microcyn60 heal with better cosmetic results and less chelation relative to the previous standard burn treatment. Dr. David E. Allie said that infection plays a significant role in many chronic limb wounds, leading to amputation. In the United States the amputation rate has increased dramatically over the last two decades, and there is an excessively high mortality rate in the higher risk patients. “Successful rehabilitation after below-knee amputation is achieved in less than two-thirds of patients.” Statistics for above knee amputations are even worse. This is why there has been an increasing emphasis on limb salvage work. And nSOS is beginning to play a vital role in this area. In Dr. Allie’s study, limb salvage rate was 100% with Microcyn, there was a marked decrease in antibiotic use, both oral and IV, and far fewer hospitalizations were required. It’s hard to imagine a solution that not only eliminates bioburden but reduces inflammation while delivering a healing payload of oxygen to tissue, but this is actually the case. nSOS contains more than five times the amount of oxygen as tap water, and elevated tissue oxygen levels have been measured up to 72 hours after application. Just how this works in tandem with small amounts of hypochlorous acid to promote such rapid and thorough healing is not fully understood yet. But an increasing body of evidence indicates that it does possess a special kind of healing power. A group of scientists in Japan speculated that reactive oxygen species, shown to be electron spin resonance spectra present in nSOS, triggers would healing through fibroblast migration and proliferation. Whatever the reason, it works beautifully, and has saved a great many lives around the world. Dramatic stories abound. In one case, a seventeen year old developed necrotizing fasciitis, also known as flesh eating bacteria, after having a wisdom tooth extracted. The infection rapidly spread and he was in the ICU on a ventilator by the time his father, a surgeon who happened to be aware of the uses of nSOS, had several gallons of Microcyn overnighted to the hospital, with instructions to irrigate the wound around the clock. Within the next few days the young man made a remarkable recovery from a terrible infection that even the strongest IV antibiotics could not contain. Another case of necrotizing fasciitis happened recently in Florida. The patient had a leg wound that had become infected with this microbe, and the progress of the infection could not be contained. The doctor was on the verge of amputating when he was advised about nSOS. He had a shipment overnighted to his hospital and began irrigations as soon as he could. The leg was saved and the family was ecstatic. At a well known specialty wound care clinic that focuses on life-threatening wounds only, Lake Wound Care Clinic in Lakeview Oregon, Dr. Cheryl Bongiovanni uses nSOS extensively. Many patients are referred to her with surgical site infections that are wildly insensitive to just about every antibiotic in existence. She routinely saves lives and limbs by skillful use of nSOS. Her success rate is phenomenal. She says that she no longer uses hyperbaric oxygen, or saline solution, and the use of topical and systemic antibiotics has been greatly reduced in her clinic. Microcyn, the only stabile nSOS, with a shelf life of two years, has been evaluated by the USP <51> Antimicrobial Effectiveness Test and is now designated as a Category 1 product. The log reduction numbers are spectacular. As an antiseptic, it highly versatile and unparalleled in its effectiveness. Over just the past seven years, over two million people have been treated worldwide without a single report of a serious adverse effect. Health care professionals struggling with the enormous problem of HAIs are trying to establish higher standards of hygiene, especially hand washing technique, hoping to curb the epidemic. Though this is a good effort which will certainly help, it could never really solve the problem. It is impractical to expect extremely busy nurses and aides to be 100% thorough in hand washing as they struggle through another busy day. Somewhere along the line, they are all going to slip up. What needs to be done in addition to focusing on strict sanitation standards is to bring nSOS into our hospitals and clinics for a whole spectrum of treatment and techniques, thereby eliminating the danger of a missed hand wash. If appropriate protocol using nSOS were to be established in our health care institutions, the outcome would be far reaching indeed. It has the capability of preventing HAIs when used as a skin prep prior to surgery, an irrigant during surgery, and as an antiseptic for post-operative care. A highly qualified reconstructive surgeon in Cabo San Lucas related that he has been using nSOS in his practice for some time, finding it to be extremely effective. He uses it instead of saline solution during his surgeries. In the future he believes that Microcyn60, the nSOS product in Mexico, will become the only product used for treatment during surgeries. nSOS can also be used as a nasal spray, which could greatly reduce the danger of patients bringing staph into the hospital setting. And nSOS can eliminate catheter-associated urinary tract infections, (CAUTIs), by rinsing the bladder on a daily basis. Ventilator associated pneumonia could be prevented by routinely cleansing the both ventilator tubing, stoma and even the trachea. Neutral super-oxidized water could be to the epidemic of HAIs what quinine was to malaria and the Salk vaccine to polio. Unfortunately, history demonstrates that the obstacles of ignorance and oversight bias often create barriers to the timely emergence of such crucial medical breakthroughs. Back before Pasteur, circa 1785, when hundreds of women were dying from childbirth infection, Alexander Gordon noticed that the cleaner things were in the birthing room the less a women giving birth was susceptible to infection. Though he attempted to spread the word about the need for cleanliness, he was scorned and ignored. Seventy years later Ignaz Semmelweis, aware that scores of women were still dying from what was called childbirth fever, took extensive empirical procedures to test the cleanliness element in birthing rooms, especially having doctors wash their hands just before delivering a baby. Yet though Semmelweis did his best, even using empirical evidence to prove cleanliness was essential, he was rejected, ridiculed and ignored, thus thousands of women continued to die because of such arrogant self-willed ignorance. (Pushing Ultimates, 2006) Let’s hope this notorious syndrome does not prevent the timely recognition and acceptance of vital Email ThisBlogThis!Share to TwitterShare to Facebook Object 1 Friday, August 20, 2010 Microcyn and HAIs Hospital-acquired infections, (HAIs) have become far too prevalent across the nation. The statistics are grim. Ten million hospital patients will get an infection in a health care institution over the next five years and half a million of them will die. Many fear going to the hospital because that is where the most virulent microbes are encountered. Along with MRSA, many significant infection-causing bacteria in the world are also becoming resistant to the most commonly prescribed antimicrobial treatments. Antimicrobial resistance occurs when bacteria change or adapt in a way that allows them to survive in the presence of antibiotics designed to kill them. In some cases bacteria become so resistant that no available antibiotics are effective against them. People infected with antibiotic-resistant organisms like MRSA are more likely to have longer and more expensive hospital stays, and may be more likely to die as a result of the infection. When the drug of choice for treating their infection doesn’t work, they require treatment with second- or third-choice medicines that may be less effective, more toxic and more expensive. Antibiotic toxicity is just as serious a problem as resistance. Antibiotics have been shown to be chondrotoxic, damaging the body’s cartilage; neurotoxic, carcinogenic, nephrotoxic, and more. In fact, antibiotics are the principle cause of drug-associated nephropathy. But there is a new weapon in the fight against resistant microbes. Microcyn technology is a safe-assaline anti-infective that quickly eradicates a broad range of pathogens, including antibioticresistant bacteria (including MRSA and VRE), viruses, fungi and spores. Dual-action in nature, in addition to killing the infection, Microcyn also accelerates the wound-healing process by reducing inflammation in the wound and increasing nutrient-rich blood and oxygen flow to the wound bed. Twenty-five clinical studies have demonstrated Microcyn to be both safe and effective in killing pathogens. What makes Microcyn so remarkably effective is the way hypochlorous acid works in tandem with reactive oxygen species. This is also the way the immune system’s neutrophils work, through oxidative burst. This is something no microbe can stand up to, nor can they develop resistance to it. Microcyn has been evaluated by the USP <51> Antimicrobial Effectiveness Test, now designated as a Category 1 product. The log reduction rates are spectacular. As a hygiene product it is unparalleled. Microcyn has revolutionized wound care as we know it. A dramatic case of limb salvage when dealing with necrotizing fasciitis came to my attention just this week. A well known Infectious Disease specialist in Florida was fighting a losing battle against NF in a leg wound. Amputation was inevitable. However, the doctor became aware of Microcyn and called the company, requesting an emergency shipment. Microcyn irrigations were initiated and the leg was saved. The family was ecstatic. This is just one of many such cases. The medical applications for Microcyn are too numerous to list here. Over two million people have been treated worldwide without a single report of a serious adverse effect. FDA clearances are coming steadily through this year and next. We will most certainly be seeing more remarkable medical breakthroughs involving Microcyn. If appropriate protocol using Microcyn were to be established in our health care institutions, the outcome would be far reaching indeed. It has the capability of preventing HAIs when used as a skin prep prior to surgery, an irrigant during surgery, and as an antiseptic for post-operative care. It also can be used as a nasal spray, ear drops, eye drops, and can eliminate catheter-associated urinary tract infections, (CAUTIs), by rinsing the bladder on a daily basis. Microcyn is to the epidemic of HAIs what quinine was to malaria and the Salk vaccine to polio. There is a great deal more to be said about this, but a picture is worth a thousand words: http://www.youtube.com/watch?v=sAiWWNCfYH4 http://www.youtube.com/watch?v=O2saruj3qvQv http://www.youtube.com/watch?v=3b0gp2C8R5E Email ThisBlogThis!Share to TwitterShare to Facebook Object 2 Home Subscribe to: Posts (Atom) Irrigación con Solución Superoxidada (SSO) en el tratamiento del conducto radicular Miguel Angel Flores-Martínez1 1Universidad Michoacana de San Nicolás de Hidalgo, U.M.S.N.H. Morelia Michoacán, México. Palabras clave: agua superoxidada, radicales libres, estrés oxidativo, oxidación del ADN, agua electrolizada, tratamiento del conducto radicular. Introducción Las infecciones del conducto radicular son una enfermedad infecciosa de etiología bacteriana y constituyen una de las mayores causas de pérdida de piezas dentales en el mundo. Las modalidades terapéuticas actuales incluyen el raspado y alisado radicular de las superficies de los dientes para eliminar la placa bacteriana y los cálculos, así como el uso de soluciones antisépticas para combatir el proceso infeccioso causado por un amplio espectro de microorganismos orales. Sin embargo, estos antisépticos tienen una elevada toxicidad y por lo tanto no pueden ser utilizados durante periodos prolongados. Además, algunos de los antisépticos de uso común tienen efectos secundarios adversos tales como modificar el sabor o manchar los dientes. Entre los numerosos antisépticos alternativos propuestos hasta ahora se incluyen las soluciones superoxidadas (SSO), que podrían ser una buena opción. Horiba et al. (1998) mostraron que una SSO de pH neutro era eficaz contra cepas bacterianas obtenidas de conductos radiculares infectados. Una segunda publicación sobre este campo mostró que el agua electrolizada ácida podría ser un tratamiento aceptable para el enjuague de impresiones con hidrocoloides (Nakagawa et al, 2002). Lamentablemente no se publicó más información. La SSO ensayada aquí (Microcyn™, Oculus Innovative Sciences, EE.UU.) ha demostrado que ejerce una actividad microbicida amplia, que incluye bacterias, virus, hongos y esporas (Landa et al, 2005). El amplio espectro no puede explicarse únicamente en base a las especies de cloro, ya que su concentración es muy baja (contenido de cloro total <80 ppm). También ha superado ensayos de biocompatibilidad según los requisitos de la Food and Drug Administration para productos de cuidado de heridas en EE.UU.. Estos ensayos incluyeron pruebas de citotoxicidad, sensibilización, irritación y reactividad intracutánea, e incluso pruebas de genotoxicidad. Tras su aprobación en México, esta SSO ha sido utilizada de forma amplia y segura en todo tipo de heridas abiertas y lesiones mucosas en los últimos 24 meses. El único efecto secundario reportado hasta el momento ha sido una sensación de ardor en los pacientes hipersensibles, particularmente en aquellos con úlceras venosas. Nuestra experiencia en odontología comenzó con el éxito en el tratamiento de la enfermedad periodontal con la irrigación de esta SSO con un escarificador ultrasónico (Flores MA, 2005). Dado que esta novedosa solución superoxidada no era ni sensibilizante ni irritante en la enfermedad periodontal y su contenido de cloro estaba era como máximo de 80 ppm, ofrecía un mejor enfoque para tratar las infecciones del conducto radicular. Por lo tanto, este estudio se realizó para evaluar la eficacia de la SSO frente a irrigación con hipoclorito sódico al 2,5% en la prevención de reacciones agudas después del tratamiento del canal radicular. En conjunto estos resultados favorecen la terapia con SSO en comparación con la solución diluida de hipoclorito sódico como solución de irrigación en el tratamiento del conducto radicular. Un estudio prospectivo, controlado, multicéntrico, debe llevarse a cabo para confirmar plenamente estas observaciones iniciales. Pacientes y métodos Pacientes Todos los pacientes consecutivos atendidos en la Universidad Michoacana de San Nicolás de Hidalgo en Morelia, México, entre octubre de 2003 abril de 2005 y con diagnóstico de infección del conducto radicular se incluyeron en el grupo de SSO. La infección aguda del conducto radicular se define como una imagen de rayos X que muestra una lesión periapical, además de pruebas clínicas negativas de vitalidad pulpar y positivas de percusión. En el grupo de control se realizó un análisis retrospectivo de pares de casos que presentaron infecciones similares del conducto radicular entre 2002 y 2003 en la misma institución. Solución superoxidada (SSO) Se investigó la actividad antimicrobiana de una solución de pH neutro, con una cantidad controlada de especies reactivas de cloro y oxígeno (Microcyn™, Oculus Innovative Sciences, EE.UU.) en pacientes mexicanos con infecciones agudas del conducto radicular. Esta novedosa solución superoxidada (SSO) para el cuidado de heridas, ofrece un enfoque radicalmente mejorado en el tratamiento del conducto radicular. Esta SSO de pH neutro sin enjuague fue certificada inicialmente como antiséptico para tratar heridas en México en 2004. En primer lugar se realizaron ensayos en animales para mostrar que la SSO no induce irritación o sensibilización en la piel y mucosas (Gutiérrez, 2006). Los componentes activos de esta solución son 99,98% de agua superoxidada y <0,02% de diversas especies reactivas de cloro y oxígeno, incluyendo ácido hipocloroso, hipoclorito sódico, cloruro sódico, ozono y dióxido de cloro. Es de destacar que el contenido total de cloro libre disponible es bajo y oscila entre 50 y 80 ppm. Esta SSO tiene actividad bactericida, virucida, fungicida y esporicida y está lista para su uso sin dilución adicional o mezcla (Landa et al, 2006). Además, no requiere una manipulación o desecho especial y tiene una vida útil superior a 12 meses. Manipulación dental con SSO Durante el proceso, los conductos están aislados para evitar la contaminación. Se abre la cavidad se perfora la raíz. Es necesario localizar un conducto apropiado para garantizar una manipulación eficiente. La irrigación con SSO comienza con el proceso de perforación, en lugar de usar solución salina o agua. La instrumentación se lleva a cabo de acuerdo con la técnica de corona invertida utilizando instrumentos rotatorios de NiTi. La instrumentación se realiza de acuerdo con la extensión del conducto, según lo determinado por rayos X. La irrigación del conducto se realiza con SSO después de usar cada instrumento. Se realiza una irrigación final con SSO en la cavidad de la raíz durante un máximo de 15 minutos (tiempo de contacto); la inyección de SSO con una aguja hipodérmica para infiltrar el tejido blando que rodea la raíz es otra alternativa para descontaminar completamente el conducto. Este último se seca con papel dental y la oclusión se realiza por condensación lateral con hidróxido de calcio y resina epoxi. El tratamiento ambulatorio se compone de enjuague bucal con SSO durante 30 días, 3 veces al día durante 2 minutos (15 ml que se cambian cada 30 segundos). Después de 30 días, se recomienda enjuague bucal una vez al día por un tiempo indefinido. Ninguno de los pacientes tratados con la terapia de SSO recibió antibióticos o analgésicos. Manipulación dental con hipoclorito sódico al 2,5% (HS 2,5%) Se realizó el procedimiento típico de terapia con hipoclorito sódico para el tratamiento del conducto radicular (Walton y Torabinejad, 2002). Se prestó especial atención para evitar la infiltración de la solución a la región periapical durante la manipulación. Todos los pacientes de este grupo recibieron ampicilina (500 mg tres veces al día) o cefalexina (500 mg dos veces al día) durante 7 días, así como medicación antiinflamatoria (naproxeno 500 mg dos veces al día) según era necesario. Resultados Se completó un estudio en 238 pacientes con tratamiento de conducto radicular con HS 2,5% (129 pacientes) o SSO (109 pacientes). Las características demográficas fueron similares en ambos grupos (Figuras 1 y 2). La media de edad fue de 42,5 años (rango 1768) y 40 años (rango 13-67) para los grupos de control y de SSO, respectivamente. La relación varones:mujeres fue similar para los grupos de control y SSO (44:56 frente a 39:61, respectivamente). Sólo 2 pacientes tuvieron una reacción local aguda después del tratamiento del conducto radicular con SSO, por 16 pacientes en el grupo de HS 2,5%. Sin embargo, los resultados son más significativos teniendo en cuenta el número total de dientes afectados en el grupo de SSO (2) en comparación con los del grupo de HS 2,5% (23). Sólo se registraron pérdidas dentales en el grupo de hipoclorito, por ninguna en el grupo de SSO. Esto podría ser debido a que el re-tratamiento con SSO fue viable y exitoso. Por lo tanto, incluso en los dos casos con reacciones agudas, fue posible preservar los dientes. El dolor se resolvió en 24 horas en la mayoría de los casos. Los pacientes también pudieron masticar en un plazo de 24 horas, cuando generalmente se tarda 72 horas con el uso de otros antisépticos. Diez pacientes de los 109 experimentaron una ligera sensación de ardor en la lengua al realizar enjuagues con SSO. En estos casos, la sensación de ardor se produjo sólo en los dos primeros días de aplicación y sólo fue necesario indicar a los pacientes que escupieran la solución de enjuague sin más. El ardor se disipó sin ningún otro efecto secundario. No se observó ninguna contraindicación para el uso de SSO. Figura 1.- Irrigación con SSO en el tratamiento del conducto radicular Pacientes Varones Mujeres Número de tratamientos Ptes. con 1 tratamiento Ptes. con 2 tratamientos Ptes. con 3 tratamientos Ptes. con 4 tratamientos Dientes con reacción aguda Ptes. con reacción aguda Figura 2.- Irrigación con hipoclorito sódico 2,5% en el tratamiento del conducto radicular Pacientes Varones Mujeres Número de tratamientos Ptes. con 1 tratamiento Ptes. con 2 tratamientos Ptes. con 3 tratamientos Ptes. con 4 tratamientos Dientes con reacción aguda Ptes con reacción aguda Discusión La infección debida a un canal que no está totalmente limpio o lleno es una complicación que puede ocurrir en un 10 a un 30% de los casos sometidos a tratamiento del conducto radicular. Esto puede ser debido a la complejidad del sistema de conductos rediculares, por ejemplo algunos canales pueden ser muy estrechos o curvos. La infección también puede ser debida a un canal que no está en absoluto limpio o lleno, si los rayos X no mostraban todos los canales de un diente. Además, ciertas bacterias pueden no responder al tratamiento del conducto radicular, por lo que el tratamiento puede fallar (Walton & Torabinejad, 2002). Dado que la SSO ejerce una actividad antimicrobiana amplia sin sensibilizar o irritar la piel y las mucosas (Gutiérrez AA, 2006), se supuso que podría ser útil para la prevención de las reacciones agudas tras el tratamiento del conducto radicular. El control de infecciones en el conducto radicular se ha basado tradicionalmente en la irrigación con hipoclorito o clorhexidina. En ambos casos, la aplicación se ve limitada por los efectos secundarios limitantes. La solución de hipoclorito sódico al 2,5%, por ejemplo, es muy irritante para el conducto radicular y no puede entrar en contacto con el tejido blando circundante debido al potencial de necrosis. La irrigación con SSO, en cambio, tiene la ventaja de no ser irritante o sensibilizante para la piel y las mucosas. Por ejemplo, la SSO se ha utilizado de forma segura en el tratamiento de heridas diversas (por ejemplo, estasis venosa, quemaduras, úlceras de pie diabético, etc), así como en la cavidad peritoneal (Dalla Paola, 2005; Miranda, 2005; Barrera et.al., 2006). En comparación con la clorhexidina, la SSO tiene mejor sabor, no mancha los dientes y se puede utilizar por lo menos durante 30 días como enjuague bucal tras el procedimiento. Los pacientes tratados en este estudio con SSO no recibieron medicación con antibióticos, antiinflamatorios o analgésicos. Por el contrario todos los pacientes tratados con de hipoclorito sódico recibieron antibióticos y se añadió un agente antiinflamatorio en caso necesario. Pese a ello, el tratamiento con SSO fue superior para reducir la incidencia de las reacciones agudas tras el tratamiento del conducto radicular en comparación con la solución de hipoclorito sódico al 2,5%. Además, los pacientes con fallo del tratamiento en el grupo de SSO pudieron ser exitosamente tratados de nuevo con la solución. Por consiguiente, ninguno de los sujetos en el grupo de SSO perdió dientes. A destacar que los procedimientos agresivos de irrigación e infiltración realizados con la SSO no podrían haberse realizado con ningún otro antiséptico actualmente en uso debido a la posible necrosis de los tejidos circundantes. Lamentablemente, el grupo de control tratado con hipoclorito sódico era histórico. Será pues necesario realizar un correcto estudio controlado prospectivo aleatorizado para validar totalmente los resultados. Sin embargo, estos resultados apoyan la eficacia y falta de toxicidad de SSO para la prevención de las reacciones agudas después del tratamiento del conducto radicular. Bibliografía Barrera-Zavala A, Guillen-Rojas M, Escobedo Anzures J, Rendon J, Ayala O & Gutiérrez AA. A pilot study on source control of peritonitis with a neutral pH super oxidized solution. XV Congreso Nacional de Cirugía (AEC) Sevilla, España, 25-28 Oct, 2005. Dalla Paola, E. Brocco, A. Senesi, S.Ninkovic, M. Merico, D.DeVido. Use of dermacyn™, a new antiseptic agent, for the local treatment of diabetic foot ulcers. DGFW European Wound Healing Meeting. 2005. Stuttgart. 15-17 Sept, 2005. Flores-Martinez MA. Periodontic Applications of Microcyn. 2006. Presentado para publicación. Gutiérrez AA. The Science Behind Stable, Super-Oxidized Water. Exploring the various applications of super-oxidized solutions. Wounds. 2006, Jan. Suppl. 7-10. Horiba N, Hiratsuka K, Onoe T, Yoshida T, Suzuki K, Matsumoto T, Nakamura H. Bactericidal effect of electrolyzed neutral water on bacteria isolated from infected root canals. Oral Surg Oral Med Oral Pathol Oral Radiol Endod 1999, 87:83-87 Landa-Solis, González-Espinosa D, Guzman B, Snyder M, Reyes-Terán G, Torres K and Gutiérrez AA. MicrocynTM a novel super-oxidized water with neutral pH and disinfectant activity. J Hosp Infect 2005, 61: 291-299 Miranda A. Reducing Bacterial Infectious Complications from Burn Wounds. A look at the use of Oculus Microcyn60 to treat wounds in Mexico. Wounds. 2006, Jan. Suppl. 1719. Nakagawa H, Hiraguchi H, Uchida H, Tanabe N. Effect of rinsing hydrocolloid impressions using acidic electrolyzed water on surface roughness and surface hardness of stone models. J Oral Sci 2002, 44:141-146. Walton R, Torabinejad M (Eds). Principles and Practice of Endodontics, 3rd Edition. Elsevier. 2002. pp 331-344. ART 01 Dalla Paola IT Gutierrez USA Miranda Mx Super-Oxidized Solution (SOS) Therapy for Infected Wounds Diabetic Foot Ulcers (IWJ) The Science behind stable, Super-Oxidized water, Exploring the various applications of SOW Reducing Bacterial Infectious Complications from Burn Wounds Wounds (IWJ) Wounds (IWJ) 2006 x x DFU PI = Providone Iodine PVD = Peripheral Vacular Disease SH = Sodium Hypochlorite PU - Pressure Ulcers HP = Hydrogen Peroxide n.a. x x 2006 x x x Burns "Silver", Saline 2006 x DFU TWC = Traditional Wound Care x x P.I. x DFU, LU, PU, Burns 64 50% reduction 8 x DFU, burns, other n.a. x ART 03 Bongiovanni USA Nonsurgical Management of Chronic Wounds in Patients with Diabetes JVU 2006 x x x DFU, LU, PU, Burns n.a. x ART 04 Dalla Paola IT Super-Oxidized Solution (SOS) Therapy for Infected IWJ, AAWC Diabetic Foot Ulcers 2006 x x x DFU P.I. x ART 05 Dalla Paola IT Treatment of Diabetic Foot Ulcer: An Overview Strategies for Clinical Approach 2006 x x DFU IT Clinical Outcome of Wide Postsurgical lesions in the Infected Diabetic Foot Managed with 2 different local LEW treatment regimes compared using Quasi Experimental Study Design: A preliminary comm. 2007 NL A Cytotoxic analysis of antiseptic medication on skin BJOD substitutes and autograft 2007 JHI 2005 x x IWJ 2007 x x 2007 x x JCPSP 2007 x x IWJ 2007 x x ART 07 Le Duc ART 08 Landa-Solis Mx Microcyn: a novel super-oxidized water with neutral pH and disinfectant activity ART 09 Martinezde Jezus Mx Efficacy and Safety of Neutral pH superoxidized solution in severe diabetic foot infections ART 10 Medina, Gonzalez Mx Super-oxidized solution inhibits IgE-antigen-induced IntImp degranulation and cytokine release in mast cells ART 11 Hadi ART 12 Gonzalez Treating infected Diabetic Wounds with India Superoxidized water as anti-septic agent: a preliminary experience Effects of pH-neutral, super-oxidized solution on Mx human dermal fibrobvlasts in vitro Comparative Efficacy and Tolerability of Oxum India against Providone Iodine Application in the Postcaesarean Section Wound Management ART 13 Anand ART 14 Chittoria India ART 15 Dhusia Comparative Efficacy and Tolerability of Microcyn India Superoxidized Solution (Oxum) against Providone Iodine Application in Orodental Infections ART 16 Zhang China Indian Medical gazette The role of Super-Oxidized Solution in the Nepal Med. management of Diabetic Foot Ulcer: Our Experience Coll. J. The clinical efficacy of Dermacyn on deep partial thickness burn wounds 2007 x x 2007 Indian Medical gazette 2008 Report 2008 x x x x x x x x Reduction of Odor Skin Irritation Limb Salvage x decrease 108 MCN 43 P.I. 55 15 MCN 145 MCN 95% PI 212 PI 34% (P<0.003 (minor amp). ) n.a. x x MCN 0 PI 18 x MCN 0 PI 1 x n.a. P.I. x Allergy Sodium hypochl x Diabetic Wounds Saline x Fibroblast 16 x x x 25 x DFU n.a. x 20 x Dental P.I. x 20 x Burns Silver x 20 Improve Wound Bed Vascularity via increased capillary perfusion and accelerationof the development of neovascularity MCN62% Sal.20% (7days) Positive Wound dehiscence MCN 14 and PI 21 patients. x x P.I. 8 different cases where Super Oxidized Solution actually resolved the problems (also prevention of amputation) x x x x n.a. Medium HP Better cosmetic results and less chelation Article about general DFU wound care progressions where MCN has its standard place. n.a. Postx Caesarean Wounds x Method: Daily dressing changes. Healed at 6 months: MCN 87.5% and PI 51.4% (P<0.008). Reinfection : with MCN 4 and PI 9 patients. x 50 x n.a. x 50 Microbial kill of MCN against gram positive and gram negative bacteria, virusses and fungi. x PVD, DFU, MRSA x 21 x DFU, LU, PU, Burns x 18 x Ps, MRSA, DFU, burns MCN 100% decrease 110 x Additional Features Positive Wound dehiscence MCN 14 and PI 21 patients. Burns several x DFU x x x PVD, DFU, MRSA x Efficacy x x MCN 0 PI 18 CLI, CVI, trauma, surgery P.I. x Toxicity x x DFU Hospitalization 64 x x Control x x x x Product MCN 43 P.I. 55 2006 x decrease MCN 100% 108 DMCT x Positive Features 110 Superoxidized Water Improves Wound Care Outcomes in Diabetic Patients Goretti, Piagessi # subjects 40 x Bongiovanni USA ART 06 Industrial x ART 02 CDR Animal Care Usefull for: Human Care In Vivo Subject Focus Poster In Vitro Abstract Article x CVI = Chronic Venous Insufficiency LU = Leg Ulcers Decease of antibiotics 2006 BKA = Below Knee Amputations Simple Wounds (IWJ) Ps = Pseudomonas infection MCN = Microcyn Technology Platform Accepted by patients Super-Oxidized Dermacyn in Lower-Extremity Wounds Year MRSA = Methicillin resistant S.Aureas Safe (non-toxic) USA Source DFU = Diabetic Foot Ulcer CLI = Critical Limb Ischemia For variety of wounds Allie, D Title Case study Author Study report Doc# Definition: Control Product LITERATURE: Studies, Articles and Posters B-C Ulcers - Bactericidal (incl. B.Atrophaeus) & Fungicidal active - Inactivates HIV-1 completely (virucidal claim with EPA) - Reduces Viral Load completely after 10min. MCN after 2 days 100% odor reduction (PI: 25%). MCN 90% improved outcome (vs. 62% of control) MCN 81% Cellulitis reduction (vs. 44% of control) x n.a. MCN is a mast-cell stabilizer! MCN inhibits 90% of the histamine release induced by allergens or Calcium ionophores. And also the late secretion of pro inflammatory molecules. MCN is anti-allergic and anti-inflammatory! yes An economical alternative to the other available antiseptic agents n.a. x MCN does not induce cell death, accelerated senescence, DNA oxidation nor RNA degradation. n.a. x Safe and effective and faster healing time. n.a. x 21 n.a. x 20 n.a. x 25 no x x x x MCN debrides necrotic tissue, reduces microbial load, promotes granulation and decreases the healing time, without damaging the normal tissue or complications. Evident that MCN results in a significant greater reduction of pain after 3 and 7 days. Also all other symptoms show the same trend. 10% faster healing time due to control of infection, abating inflammation, improving topical circulation, promoting the movement of proliferation of fibroblastic cells. ART 17 Dharap India Efficacy and Safety of Oxum in Treatment of the Venous Ulcer ART 18 Piagessi IT A Randomized Controlled Trial to examine the efficacy and safety of a new Super-Oxidized Solution IJLEW for the management of wide Postsurgical lesions of the Diabetic Foot JIMA Super-Oxidized Solution (Dermacyn Wound Care) as Adjuvant Treatment in the Postoperative Management of Complicated Diabetic IJLEW Foot Osteomyelitis x x x 2010 x x x DFU ART 19 AragónSánchez ES PST 01 Allie, D USA Dermacyn: A novel superoxidized water facilitates wound healing. Early experience in lower extremity wounds and limb salvage SAWC 2006 x PST 02 Dalla Paola IT Use of Dermacyn, a new antiseptic agent, for the local treatment of Diabetic Foot Ulcers EWMA 2005 x PST 03 Gonzalez Mx PST 04 Goretti, Piagessi IT Effects of pH-neutral, super-oxidized solution on IWJ human dermal fibrobvlasts in vitro Clinical Outcome of Wide Postsurgical lesions in the Infected Diabetic Foot Managed with 2 different local LEW treatment regimes compared using Quasi Experimental Study Design: A preliminary comm. PST 06 Monaghan UK Dermacyn as the Local Treatment for infected Diabetic Foot Wounds a case series PST 07 Peterson USA PST 08 ISDF Venous Ulcers 2008 2013 2007 2007 x x x x x 52 52 x x DFU P.I. x 110 110 HP x x Fibroblast x DFU x DFU Dermacyn Effective in Treatment of chronic Wounds with extensive Bioburden while reducing Local Pain SAWC Levels 2007 x x x x Recalcitrant wounds Ramaswami USA The role of Super-Oxidized Solution in Wound Treatment SAWC 2007 x x x PST 09 Ricci IT Clinical Results about an antimicrobial solution in the EWMA treatment of infected chronic wounds 2007 x x PST 11 Suri India Role of Neutral pH Superoxidized Water in the healing of Diabetic Foot Ulcers 2007 PST 13 Uribe Mx Effect of a Neutral pH Super-Oxidized Solution in the ETRS & Healing of Diabetic Foot Ulcers ICAAC Methicilin-resistent Staphylococcus aureus - problem EWMA in health care Super-Oxidized water for mesh hernioplasty in AHS glossy contaminated fields. An experimental study. PST 14 Veverkova CZ MartinezMunive Mx PST 16 Thatcher USA The anti-viral Efficacy of a New Super-Oxidized Solution PST 17 Thatcher PST 18 20 TWC x PST 15 20 x DFU x x x x x 2007 P.I. no x 2007 DFU symp. 30 P.I. x x x x Diabetic Foot Osteomyelitis x x n.a. P.I. 14 MCN 31 MCN 98.0% MCN 0% TWC 64 TWC 92.3% TWC 13.4% x x 18 15 x n.a. x n.a. x CLI, DFU, CVI x MCN 0 PI 1 n.a. x n.a. x 15 several x 5 5 n.a. x Chronic Wounds several x 3 3 n.a. x x Chronic Wounds P.I. x 20 20 x x x DFU Saline x 50 50 x x x DFU Saline x 40 40 x 2005 x ICAAC 2007 x x USA The anti-bacterial Efficacy of a New Super-Oxidized ICAAC Solution 2007 x Thatcher USA Activity of a pH Neutral Super-Oxidized Solution against Bacteria Selected for Sodium Hypochlorite Resistance. ICAAC 2007 x PST 19 SanchezMiranda Mx Anti-inflammatory effects of an oxychlorine compound in a murine model of passive cutaneous anaphylaxis WUWHS 2008 PST 20 Regulski USA Efficacy and compatibility of combination Therapy with Super-Oxidized Solution and a Skin Substitute for Lower Extremity Wounds SAWC 2008 ABS 01 MirandaAltamirano Mx Treatment of 2nd and 3rd Degree Burns in 48 Pediatric Patients without Routine Antibiotics using New "Super-Oxidized water Technology" Microcyn TSS submission 2005 x x x x MRSA x other x x x Virusses Saline x x x Bacteria n.a. x x Resistant Bacteria 40% bleach x x MCN reduces in Ulcer Size, bacterial burden and the clinical signs of soft tissue infection Wounds failed to heal despite of aggressive local wound care, soon after initiating MCN the wounds closed. Also significant reduction of pain! x Wound treatments were unsuccesssful, and with MCN 1 wound healed after 1 month and 2 wounds after 3 months White blood cells decreased more significantly (45% vs. 20%). Very good results in reduction of clinical signs and pain relief. Odor with MCN gone after 2 days (control after 9 days) x Economic requirements for the treatment of MRSA are evident MCN log reduction far more better then Saline and much less abscess formation The load of enveloped and non-enveloped virusses exposed to MCN for 1 minute was reduced by a log10 factor > 5. Complete inactivation was seen after 5 min. n.a. n.a. x 48 48 n.a. x x 46 x x x 10 x Reduction in cellulites, decrease of Bioburden, and growth of good granulation tissue at the base of the ulcers. Early closure of the wounds (78% vs. 40%) and less other wound care procedures. Epithelization period 33% shorter with MCN (induced by 0.5-0.7mm per day). Total closure with MCN=35 days and with Saline 60 days. Wound healing effect not only antimicrobial activity! n.a. 12 Fourteen consecutive patients who underwent conservative surgery for DFO, were treated in the postoperative period with DWC. No side effects, no allergies nor skin dermatitis were found. Limb salvage was successfully achieved in 100% of the cases. Healing was achieved in a median period of 6.8 weeks. At surgical timw 75% of Ulcers in MCN group had negative Micro soecimen vs. 48% of Ulcers in PI group (P < 0.005) MCN does not induce cell death, accelerated senescence, DNA oxidation nor RNA degradation. Method: Daily dressing changes. Healed at 6 months: MCN 87.5% and PI 51.4% (P<0.008). Reinfection : with MCN 4 and PI 9 patients. Plantar Foot Ulcers MCN 35 10% BKA vs. Sal. 60 25% in contr. Method: Daily installation. Healed at 6 months: MCN 90% and PI 55% Reinfection : with MCN 1 and PI 9 patients. Does not oxidize fibroblast. Does not cause skin complications. Infected CW 12 n.a. A spectrum of 46 bacterial species exposed to MCN for at least 30 seconds were reduced by a log10 factor > 4.5. This was the limit of detection. 10 n.a. After 2 minutes all isolated bacteria had a reduction with MCN of a log10 factor > 5. 6 6 n.a. Can potentially be used to ameliorate the inflammatory process induced by IgE-antigen-induced mast cell degranulation or chemical irritation of the skin in atopic eczema. 9 n.a. MCN might induce faster epithelialization than Saline. x x Mast Cells Saline x x x Lower extr. Wounds Saline x 9 n.a. x 48 x Burns 40% reduction 27 Saline x Diabetic Foot Infections x 2005 Activity against micro-organisms, including MRSA and VREF. MCN has an anti-allergic roleby inhibiting Ig-E antigen induced granulation and cytokine release by acting as a mast cell stabiliser. MCN promotes tissue regenaration. x stage II/III B-C-D Ulcers MCN 0 PI 15 MCN 145 MCN 95% PI 212 PI 34% (P<0.003 (minor amp). ) n.a. MCN 6W Sal. 7W 50% no n.a. x x No antibiotics, either systematic or topical were used. No infections were encountered. Third degree burns of 10cm in diameter healed completely without skin grafts. Better cosmetic results and less chelation. Hospital days reduced from 20 to 10 days (cost saving = $19.000 per day) RPT 01 Martinezde Jesus Mx A Prospective, Randomized, Single-Blind, Clinical Investigation of a Neutral pH Superoxidized Water in Report Patients with Infected Diabetic Foot Ulcers 2004 RPT 02 Flores Mx Periodontic Applications of Microcyn Draft report 2005 RPT 03 Flores Mx Super-Oxidized Solution (SOS) irrigation in Root Canal Treatment Report 2005 RPT 04 Landsman USA An Open-Three Arm, Pilot Study Of The Safety And Clinical Efficacy Of Topical Dermacyn Wound Care DFCon Versus Oral Levofloxacin Versus Combined Therapy For Mild Diabetic Foot Inf. 2008 x DFU P.I & Saline x 24 x Dental Chlorhexidin e x 14 x x Dental SH x 109 129 n.a. x x DF Levofloxacin x 20 20 n.a. x x x 21 x no n.a. x n.a. x 100% MCN patients had reduction of Fetid Odor (c: 25%). MCN reduced cellulitis with 81% vs. 44% of c. 90% improvement of wound with MCN vs. 62% with c. 94% of MCN groupd had improved tissue and skin around wound instead of 31% with control. x x Resolution of the Gingivitus was only two to four weeks. Bleeding upon brushing resolved with 24 hours till 4 days. Within 2 weeks bone regenaration was observed. MCN works for Gingivitus and Periodontitus Acute local reaction after toot canal treatment occurred less with MCN (2 vs. 16 patients). Affected teeth with MCN only 2 while with control 23. Patients could chew within 24h (instead of 72h). x Clinical success; MCN: 93.3% ; Levo/Saline: 56.3% ; Levo/MCN: 83%. No serious adverse events reported