Journal of Burns and Wounds, 2005; 4: (más artículos en esta revista)

La cicatrización de heridas cutáneas de mostaza de azufre lesiones

Abrir las ciencias Company, LLC
John S. Graham [a], p. Robert Chilcott [b], Paul Rice [b], Stephen M. Milner [c], Charles G. Hurst [a], Beverly I. Maliner [a]
[a] comparativa rama patología, la medicina comparativa y la división química de atención de urgencias División de Ejército de los EE.UU. Instituto de Investigaciones Médicas de defensa química, de pruebas Aberdeen, MD
[b] Departamento de Ciencias Biomédicas, de Defensa de Ciencia y Tecnología de Laboratorio, Porton Down, Salisbury, Wiltshire, UK
[c] El Instituto de Cirugía Plástica, en el sur de Illinois University School of Medicine, Springfield

Se trata de un acceso abierto mediante el cual el artículo los autores conservan los derechos de autor de la obra. El artículo se distribuye bajo la licencia Creative Commons Attribution License, que permite el uso ilimitado, distribución y reproducción en cualquier medio, siempre que la obra original esté debidamente citados.

Resumen

La mostaza de azufre es un alquilantes agentes de guerra química que afecta principalmente a los ojos, la piel y vías respiratorias. La mostaza de azufre lesiones pueden tomar varios meses para sanar, requieren largas hospitalizaciones, y se traducen en importantes cosméticos y / o déficit funcional. Históricamente, blister aspiración y / o deroofing (epidérmica eliminación), físico desbridamiento, la irrigación, antibióticos tópicos, los apósitos estériles y han sido los principales cursos de acción en el manejo médico de mostaza de azufre cutáneo lesiones. El tratamiento actual consiste la estrategia de gestión sintomático y está diseñado para aliviar los síntomas, prevenir las infecciones, y promover la cicatrización. Actualmente no hay normalizados o métodos de optimización de gestión de siniestros que prevenir o minimizar los déficit y para proporcionar rápida cicatrización de la herida. Varios laboratorios están buscando activamente para mejorar los tratamientos para las lesiones cutáneas vesicant, con el objetivo de devolver a la piel dañada óptima apariencia y función normal en el menor tiempo. La mejora de trato se traducirá en una mejor cosmético y funcional para el paciente, y permitirá a la víctima a regresar a las actividades normales antes. Esta editorial ofrece breves reseñas de mostaza de azufre uso, su toxicidad, los conceptos de medidas de lucha, los tratamientos actuales, y estrategias para el desarrollo de mejores terapias.

Perspectiva histórica de mostaza de azufre uso

La mostaza de azufre [bis (2-chloroethyl) sulfuro; o HD] es un potente vesicating (ampollas) de agentes de guerra química que se utilizó por primera vez en la Primera Guerra Mundial por Alemania contra las tropas francesas en Ypres, Bélgica (1917). Desde entonces, ha habido pruebas o alegaciones de uso HD en 11 conflictos, incluida la de Italia contra Etiopía en 1936, por el Japón contra China en 1937, contra Polonia por Alemania en 1939, de Yemen contra Egipto de 1963 a 1967, y de Iraq contra Irán en la década de 1980 1. Además, su uso se ve amenazada a comienzos de 1990 durante la guerra del Golfo Pérsico. No hay paz para los usos industriales HD. Envejecimiento HD existencias siguen siendo peligrosas para los civiles. Estas municiones y arsenales fueron descartados por la tierra y en el océano durante y después de la Segunda Guerra Mundial. Periódicamente, la superficie agrícola o durante las actividades de pesca, causando lesiones graves. Amplia producción y el almacenamiento durante la Segunda Guerra Mundial, junto con los efectos conocidos de HD en los tejidos epiteliales, se han impulsado acuerdos para prohibir su producción y uso. Estos acuerdos incluyen la 1993 Convención sobre las Armas Químicas (CAQ) 2 y agente de programas de destrucción. Sin embargo, este agente de guerra química sigue siendo una amenaza debido a su popularidad en algunos países que no son signatarios de la Convención y las exposiciones accidentales. Dispersos como un vapor, aerosoles, líquidos o en gotas, HD sigue siendo una amenaza para los combatientes de guerra y civiles en todo el mundo.

Como una clase, vesicants incluyen la mostaza de azufre, las mostazas de nitrógeno (HN 1, el agente quimioterapéutico HN 2 o "Mustargen", y HN 3), arsenicals como lewisita, halogenados y oximas como el fosgeno oxima. Una plétora de información sobre vesicants está disponible en la literatura publicada. Este informe es una revisión de algunas de las publicaciones disponibles en HD.

La mostaza de azufre toxicidad
General toxicología

El conocido los mecanismos moleculares de acción, la química, toxicodynamics, y genotoxicidad de HD, así como la patogenia y la histopatología de HD lesiones han sido ampliamente descritos. 1, 3 - 16 Debido a sus acciones en parte se asemejan a las de las radiaciones ionizantes, HD es a veces considerado un radiomimetic compuestos. Papirmeister et al 1 escribió un extenso examen de HD investigación, que ofrece varias teorías de su citotoxicidad, y resumió lo que se conoce de su absorción, distribución, biotransformación y excreción. Somani et al 3 también han proporcionado una excelente visión general de la toxicodynamics de HD.

La investigación actual no es sólo la investigación de diversos aspectos de las teorías de Papirmeister, sino también para impulsar el desarrollo y la utilización de protectores de piel de actualidad, la descontaminación de agentes, profilácticos y terapéuticos contramedidas, y la mejora de las terapias para las lesiones establecidas. La mayor parte de la investigación hasta la fecha se ha centrado en la mitigación de las lesiones cutáneas. Más grandes lagunas de conocimiento existentes por lesiones a los ojos y el sistema respiratorio, y para efectos sistémicos.

General manifestaciones clínicas

La mostaza de azufre es un agente alquilante bifuncional que provoca graves lesiones incapacitantes para la piel, vías respiratorias y los ojos. Se reacciona con una gran variedad de moléculas biológicas de interés, que van desde los compuestos de bajo peso molecular para macromoléculas como ADN, ARN y proteínas. 1 Al igual que otros agentes alquilantes, se pueden cruzar-link complementarias de ADN y, por tanto, inhibe la célula División. Aunque el perjuicio se produce muy rápidamente después de los tejidos en contacto con HD, los signos clínicos de la lesión se demoran de 2 a 24 horas después de la exposición, con la duración del retraso están inversamente proporcional al nivel de exposición y otros factores. 16 A causa de este retraso y la falta de dolor en contacto, la exposición puede ir no reconocidos, y las personas expuestas a menudo puede dejar de tomar medidas inmediatas de descontaminación y las medidas de protección, han dado lugar a la exposición y lesiones graves.

Características lesiones son una función de la dosis y el tiempo después de la exposición. 1 A bajas dosis tóxicas, lesiones cutáneas se caracterizan por un primer período asintomático de duración muy variable seguida de eritema localizado pruriginosa. 1, 15 dosis más altas inicialmente producir pequeñas vesículas dentro o sobre la periferia de las áreas eritematosas que pueden fusionarse después para formar grandes, pendulares ampollas 15. Vesication puede tardar varios días. Dosis muy altas producen necrosis de coagulación, que a menudo es caracterizada como "rosquilla quemaduras" debido a la aparición de una región central necrótico de la piel rodeada por una periferia de circular menos tejido dañado. 15 infecciones bacterianas secundarias de este tipo de herida plantean una grave amenaza . En los ojos, bajo vapor de las exposiciones se caracterizan por enrojecimiento. Dado que la concentración aumenta, la conjuntivitis se hace progresivamente más graves, con fotofobia, blefaroespasmo, dolor y daño de la córnea visto a dosis más altas. 1, 15 líquida o vapor gran exposición a los ojos resultados en grave daño de la córnea, con potencial de ceguera 15. La inhalación de vapor HD induce cambios en la laringe y tracheobronchial mucosa, con leve a severa inflamación. 1, 15 Dependiendo del grado de exposición, los síntomas pueden ir desde una leve irritación de las vías respiratorias superiores a bronquiolares grave daño epitelial conduce a necrosis, hemorragia , Exudado inflamatorio, y pseudomembrane formación en el tracheobronchial árbol. 1, 15 Si bien raros, la exposición pulmonar masivo puede resultar en edema pulmonar hemorrágico. En la Primera Guerra Mundial, las lesiones respiratorias de vapor exposiciones resultado de muerte debido a la neumonía secundaria a neumonitis química. 15 Más profundo lesiones respiratorias de la exposición de aerosoles, como en la guerra Irán-Irak de finales del decenio de 1980, han dado lugar a olas de 2 de la muerte 17. La primera oleada se produjo dentro de 3 dias del ataque de insuficiencia respiratoria debido a la extrema perjuicio a epitelio respiratorio y los alvéolos. La segunda oleada de muertes ocurrieron entre el 1 y 3 semanas y post-secundaria como resultado de la bronconeumonía y la sepsis secundaria a la médula fracaso. Las dosis altas puede producir una toxicidad sistémica que incluye la destrucción de la médula ósea células precursoras. Leucopenia comienza típicamente de 3 a 5 días después de la exposición 15 y puede progresar a la pancitopenia. Muy altas dosis puede provocar la destrucción de los órganos linfoides y la mucosa del intestino delgado y producen efectos en el sistema nervioso central, incluyendo apatía, depresión, hyperexcitability, movimientos musculares anormales, y convulsiones. 1, 15

La mostaza de azufre ha demostrado ser carcinógeno y mutagénico en estudios con animales, y estudios epidemiológicos de trabajadores de fábricas que participan en la producción HD que fueron expuestos crónicamente a bajas dosis de HD se han implicado en el cáncer humano (principalmente respiratorias carcinomas). 1, 15, 16 Cánceres de piel se han generado experimentalmente en ratas y ratones expuestos a HD y pueden surgir en HD-inducida por tejido cicatrizal en los seres humanos. 15, 16 No hay pruebas de que HD es un motivo de los eventos reproductivos adversos, aunque estudio epidemiológico de afectados civil puede alterar las poblaciones actuales.

Problemas persistentes de los soldados y la fábrica los trabajadores expuestos durante la Primera Guerra Mundial incluyen la bronquitis crónica, asma, laringitis, neumonía recurrente, ya largo plazo queratitis. 16 clínicas recientes informes de seguimiento de los exámenes de los supervivientes del ataque de 1987 iraquí en Sardasht, incluyendo quienes eran niños en aquel momento, indican dermatológicas crónicas, oftálmica, y problemas respiratorios. 17, 18 disfunción pulmonar es el problema más común. Se observó que los efectos crónicos tienden a ser significativamente más pronunciado en los adultos, posiblemente porque los mecanismos de reparación celular son más dinámicos en los niños 18.

Las hipótesis de citotoxicidad

Hay varias hipótesis de citotoxicidad HD: (1), el poli (ADP-ribosa) polimerasa hipótesis, (2) el tiol-Ca 2 + hipótesis, y (3) la hipótesis de la peroxidación lipídica. 1 Ninguna de estas hipótesis ha sido plenamente aceptado , Y los mecanismos propuestos por estas teorías pueden ser activos al mismo tiempo. En la actualidad, el mecanismo exacto de acción de HD no se conoce.

La primera hipótesis, propuesta originalmente por Papirmeister, 1 se ha acuñado la poli (ADP-ribosa) polimerasa (PADPRP) hipótesis. La mostaza de azufre lesión se inicia por una rápida alquilación de ADN. Alkylated ADN purinas someterse espontánea depurination enzimática, por lo que en numerosos sitios apurinic que posteriormente sean exfoliados a través de apurinic endonucleasas de ADN para formar los descansos. La acumulación de estas rupturas cromosómicas se activa la enzima PADPRP, que utiliza NAD + como sustrato para ADP-ribosylate varias proteínas nucleares, lo que supone una reducción de NAD + celular. NAD + agotamiento parece ser el resultado de un ritmo acelerado de uso, por oposición a la interferencia con su síntesis. Este agotamiento se traduce en la inhibición de glycolysis y la posterior estimulación de la NADP + dependiente de shunt hexosa monofosfato (HMS), debido a la acumulación de glucosa-6-fosfato. La activación de la HMS resultados en la inducción y secreción de proteasas, lo que a su vez dar lugar a la patología típica de los cambios se ha señalado anteriormente. Los resultados de los estudios in vivo, utilizando un injerto de piel humana y el modelo in vitro de trabajo mediante los linfocitos humanos, los queratinocitos humanos, y la piel humana órgano culturas sugieren que los niveles de NAD + que disminuir por debajo de un valor crítico llegar a ser irreversible y el perjuicio que producen. Nuclear patología parece preceder frente al citoplasma de los daños. Este daño nuclear se caracteriza por la pérdida de eucromatina y la condensación y marginación de la heterocromatina. 1 graves daños a la envoltura nuclear, como blebbing y rotura, se ha demostrado por microscopía electrónica, junto con la formación de vacuolas perinuclear. El grado de daño nuclear ha demostrado ser la dosis y el tiempo relacionados con el 1.

La prueba de que puede contradecir la hipótesis PADPRP incluye los hechos que (1) NAD + agotamiento no se produce en ratas queratinocitos hasta que la concentración de HD es suficiente para inhibir la reparación del ADN 19; (2) la reparación del ADN en los queratinocitos humanos puede lograrse dentro de los 90 minutos de la exposición, 20 Considerando los importantes NAD + agotamiento no se observa hasta el 1 a 3 horas postexposición, y (3) elevación o el mantenimiento de NAD + en queratinocitos humanos no confiere protección 21.

La segunda hipótesis de HD citotoxicidad, el tiol-Ca 2 + teoría, se basa en un mecanismo propuesto originalmente por Orrenius y Nicotera en sus estudios de hepatocitos de rata. 22 El caso de iniciar en este caso es una reducción de la proteína celular tiol niveles, dando lugar a tóxicos aumentos en los tratados de libre citosólico de Ca 2 +. El tiol niveles de agotamiento como resultado directo de las reacciones de glutation (GSH) y de proteínas sulfhydryls con oxidantes u otras electrophilic xenobióticos. Glutatión intracelular es un scavenger de HD, lo que puede ser la causa de agotamiento de GSH. El agotamiento de GSH se produce después de reacción con electrophiles, exponiendo así a las proteínas sulfhydryls a los daños causados por los xenobióticos o de manera endógena producido especies tóxicas de oxígeno que surgen como subproductos de metabolismo oxidativo. Por otra parte, HD también puede reaccionar directamente con la proteína thiols a causar la inactivación de enzimas. Un grupo de enzimas afectadas por la modificación de su sulfhydryl grupos es Ca 2 + translocases, por lo que la alteración en la acumulación intracelular de Ca 2 + homeostasis. Esto conduce a un aumento citosólico libre en Ca 2 +, lo que a su vez conduce a la activación de Ca 2 + dependientes de los procesos catabólicos, incluyendo la estimulación de las proteasas, endonucleasas, y phospholipases. Proteasa actividad conduce a la degradación de las proteínas y, en última instancia perturbación del citoesqueleto. Los cambios en el citoesqueleto también pueden surgir de alquilación directa de HD a microfilamentous proteínas, aunque esto no parece desempeñar un papel decisivo. Endonucleasa actividad da lugar a rupturas de ADN, dando lugar a condensación de la cromatina y la pérdida de energía en la célula. La activación de phospholipases conduce a la hidrólisis de fosfolípidos, con las consiguientes alteraciones en la fluidez de membrana y la pérdida de la función de las proteínas de membrana y la integridad. El citoesqueleto, las armas nucleares, y todos los cambios de membrana conducir a la muerte celular. 1 No se sabe si vesicating dosis de HD agotan GSH lo suficiente como para producir estos efectos citotóxicos. Baja GSH agotamiento puede ser menos tóxicas, ya que permite tiempo para la adaptación de células. El prolongado período de latencia antes de la manifestación clínica de las lesiones HD es incompatible con la lesión más rápido esperar de una alquilación GSH inducida por agotamiento, lo que sugiere que el daño celular a un objetivo distinto de GSH es responsable de iniciar la citotoxicidad y que cualquier participación de la tiol - Ca 2 + citotóxicos vía resultados de este daño inicial.

La tercera hipótesis supone la peroxidación lipídica de las distintas tiol-Ca 2 + hipótesis, donde la principal consecuencia de tóxicos GSH agotamiento es la formación de peróxidos lípidos tóxicos. La muerte celular es, pues, que se propone debe a la acumulación de oxidantes endógenos (por ejemplo, H 2 O 2 en la acumulación resultante hidroxilo y perferryl formación de iones), dando lugar a la peroxidación lipídica y el daño irreversible de la membrana. 1

Más recientes teorías para tener en cuenta el mecanismo de toxicidad HD incluirá un melanocyte-radicales libres hipótesis 23 y uno en el que HD requiere activación metabólica 24. Debido a las múltiples dianas moleculares de alta definición, múltiples vías para acceder a la muerte celular puede ser iniciado. Estas múltiples vías puede tener algunos pasos comunes, por lo que todo lo anterior hipótesis puede desempeñar un papel en la citotoxicidad de mostaza de azufre. Papirmeister 25 también explicó cómo HD podría inducir la apoptosis (muerte celular programada) y postula la forma en que podría contribuir a modificar o el HD patogénesis de la lesión. Kan et al 26 postulan que HD-inducida por la muerte de las células implica la apoptosis temprana (6-12 horas postexposición) y la necrosis tarde (24 horas), que temporalmente se superponen para producir una única célula de muerte a lo largo de una vía apoptótica necrótico-continuo. Varias líneas de investigación se están realizando para esclarecer aún más el papel de la apoptosis en HD toxicidad.

Patogénesis de ampollas

Los efectos citotóxicos de HD en la piel han sido ampliamente descritos por un número de especies. 1, 4, 6, 10, 27 - 35 El principal cutáneo de células población objetivo de HD es el de células basales de la epidermis. Ampollas (seres humanos) y microblisters (modelos animales) se producen en la separación de la epidermis de la dermis a la epidermis cutánea de cruce. Esta separación depende de la pérdida de la integridad de células basales y filamentos de anclaje. 4 En estudios con animales modelo, el desarrollo de una aparente inicial nuclear condición patológica de células basales del estrato germinativum fue seguida por la progresiva citoplásmica, la cual conduciría a la eventual muerte afectados de células basales. Petrali et al 4 describen los efectos degenerativos subcelular de HD en la piel de pelo conejillos de 24 horas después de la exposición. Degeneración También se observó en linfocitos humanos y queratinocitos in vitro de Petrali et al 4, en una gran variedad de modelos animales de Papirmeister et al, 1, 34 y en conejos y cerdos de guinea Vogt et al. 6 Si bien los cambios degenerativos se observan en la capas superiores de la epidermis, así como en células epidérmicas de los folículos pilosos, los cambios son más prevalentes entre las células basales, con la primera y la más grave degeneración visto en las células situadas por encima de las papilas dérmicas. 1 Microblisters se observó que se deriven de las áreas focales de la epidermis - cutánea en las zonas de separación generalizada de células basales pyknosis, de 24 a 48 horas después de la exposición HD, como se ha visto por microscopio de luz. Cambios progresivos en las células epiteliales basales incluyen la formación de perinuclear o paranuclear vacuolas, una disminución en la intensidad de la tinción nuclear, citoplásmica hinchazón, la reubicación de la cromatina a la periferia del núcleo, la pérdida de la cromatina, y pyknosis 33. Estos cambios son seguido o acompañado de necrosis, vacuolización, o degeneración hidrópica del citoplasma. 33 Si bien el alcance de los daños nucleares es la dosis y el tiempo relacionados con el, no se llevará a cabo simultáneamente en todas las células basales examinados, probablemente refleje diferencias en eficiencia de la reparación de células en distintas fases de la del ciclo celular. 1 Nuclear pyknosis También se ha descrito tras la exposición HD en la piel humana o equivalente. 36, 37 También se describe en lugares aislados y perfundidos porcina colgajos de piel expuesta al HD simulante 2-chloroethyl sulfuro de metilo, un monofunctional analógica de HD. 38 La tiempo de aparición de las armas nucleares o citoplasmáticas cambios es cada especie. En general, pyknotic núcleos comienzan a aparecer en la capa de células basales 3 a 6 horas después de la exposición HD. En 12 horas, áreas focales de pyknosis se ven y se generalizan por 24 horas, con los núcleos de muchas suprabasal queratinocitos participar.

La patogenia de microblisters no se entienden plenamente. Petrali et al 5, 39 encontraron indicios de que las proteínas extracelulares de matrices de la zona de la membrana basal (BMZ) se verán afectadas durante el desarrollo del HD-patología inducida por la piel sin pelo en cobayas y el postulado de que puedan contribuir a la formación de microblisters. Tinción inmunohistoquímica para antígeno de penfigoide ampolloso, una proteína noncollagenous compartida entre hemidesmosomes de células basales y la lámina lucida, puso de manifiesto una disminución del antígeno de penfigoide ampolloso reactividad en los primeros tiempos y la posterior pérdida de antigenicidad en períodos de tiempo más tarde después de 8 minutos de exposición al vapor HD (tejidos fueron cosechadas en determinados períodos de tiempo post-hasta 24 horas). La laminina, el principal glicoproteína de la lámina lucida, mostró escaso immunolocalization a los períodos de tiempo más tarde, conforme al alterado estructuralmente lámina lucida en microblister lesión sitios. La reactividad del colágeno tipo IV, una proteína ubicua asignado a la lámina densa de la membrana basal, es inalterable a lo largo antisueros específicos prevesication y vesication períodos de tiempo. La influencia de estas macromoléculas alterado en los mecanismos de reparación después de HD toxicidad no es conocido. Otras proteínas de la BMZ aún debe ser investigado o descubierto probablemente juegan un papel en la patogénesis de HD-inducida microblisters.

Propiedades químicas

Un resumen de los agentes químicos, físicos, ambientales y propiedades biológicas de HD y otros agentes vesicating (lewisita, fosgeno oxima) se puede encontrar en el libro de texto de Medicina Militar 15. Mostaza de azufre es un amarillo pálido a marrón oscuro líquido aceitoso, con un el olor de ajo o mostaza. Tiene baja solubilidad en agua y es altamente lipofílicas, por lo que fácilmente particiones en la piel. Tarifas de 150 μ g cm -2 min -1 a través de la piel humana in vivo 40 y 157 μ g cm -2 min -1 a través de calor separados por la piel humana in vitro 41 han sido reportadas. La penetración se ve reforzada por la humedad y el calor, y en piel fina. La DL 50 de líquido HD es de unos 100 mg kg -1. Esto es bastante fluido (5-6 ml) para cubrir aproximadamente el 25% del total de superficie corporal (TBSA) en un adulto. 15 A 10 - μ g gota es suficiente para causar vesication. El umbral para la mezcla vapor / aerosol para inducir a los daños es de 200 a 2000 mg m -3 min (tiempo de concentración, Ct Ct), dependiente de la ubicación anatómica, las condiciones ambientales (temperatura y humedad), sudoración, y otros factores. 15 Vapor lesiones en general induce superficial o parcial de espesor lesión cutánea. Liquid HD pueden producir espesor total del perjuicio. Debido a su alto punto de congelación (14 ° C), HD es muy persistente y fría en los climas templados. Su persistencia es menor en climas más cálidos, donde el agente se vaporiza con mayor facilidad.

Clínica cutánea efectos

Clínicamente, eritema es el primer síntoma observado, por lo general con un retraso en la aparición de 4 a 8 horas. Prurito, ardor, picazón o puede acompañar al eritema 15. Leve edema puede también estar presente. Si la exposición es pequeña, el eritema no avanzará a vesication. Si la lesión progresa, muy pequeñas vesículas se desarrollará dentro o en la periferia de las áreas eritematosas que comienza en alrededor de 2 a 18 horas post-15. Estas vesículas pueden más tarde se unen para formar grandes vesículas y bullas. Vesication puede tardar varios días. Adicional (nueva) ampollas pueden surgir una semana más tarde. Extremadamente altas dosis puede inducir a una zona central de necrosis de coagulación, con formación de ampollas a lo largo de la periferia. La mostaza de azufre son ampollas subepidérmicas. Los más pequeños son muy duraderas grandes bullas, pero son vulnerables a la fricción. El blister fluido es inicialmente delgada y clara, o ligeramente de color paja, y más tarde se vuelve amarillento y tiende a coagular. No contiene y que no hayan agente no es un vesicant sí, a diferencia del líquido en lewisita inducida por ampollas. 15 Alta temperatura ambiente, hidrata la piel, delgada o delicada piel, la piel o la ropa ocluida por lo general presentan lesiones más graves y más corto a veces inicio de los síntomas que más frescas y menos hidratada, más grueso, o bajo la piel unoccluded niveles similares de exposición. La cara, cuello, fosa antecubital, axila, periné y genitales externos suelen ser zonas muy sensibles. Las diferencias de sensibilidad individual a HD se han observado. El color de la piel no se ha demostrado que afectan a los índices de penetración HD 42. Hiperpigmentación transitoria se informó de bajas en el decenio de 1980 Irán-Iraq War, principalmente debido a la acumulación de melanina muertos derivados de melanocitos en la base de una epidermis a punto de desquamate, y de opacificación y el oscurecimiento de nonviable células epidérmicas. 15, 43, 44 A raíz de desquamation, la piel puede aparecer hipopigmentado. Es nuestra opinión que a largo plazo hiperpigmentación resultados de la estimulación de los melanocitos y la hipopigmentación de melanocyte destrucción. Estos cambios pigmentarios suele invertir en 6 a 12 meses, pero puede ser permanente.

Curación tiempo de estas lesiones por vía cutánea depende de la gravedad, con eritema solo teniendo varios días para disminuir las lesiones graves y teniendo en semanas o meses, dependiendo de la ubicación anatómica, la profundidad de la lesión, y el tamaño. Las lesiones pueden ser dolorosas y la infección bacteriana secundaria es un problema común (sólo en los seres humanos). Si bien el pleno reepithelialization veces se ha considerado como criterio de valoración para la curación, la función de barrera debe regresar para la lesión que se considera completamente curado. Los datos preliminares de nuestro laboratorio indican que la función de barrera, como se indica por la pérdida de agua transepidermal (TEWL) las mediciones, se ve gravemente perturbado ventral abdominal en la piel de cerdo de 3 a 7 días después del 30 a 120 minutos de exposición a líquidos HD. Si bien el pleno reepithelialization de estas profundas cutánea / espesor total lesiones aparece groseramente a ocurrir de 35 a 56 días después de la exposición, TEWL valores no vuelven a su nivel inicial hasta el 63 a 70 días postexposición (JS Graham et al, datos no publicados, 1998). El retraso en la barrera tras la interrupción agente de la exposición puede explicarse por la existencia in situ de un estrato córneo intacto durante los primeros días tras la exposición. El estrato córneo se mantiene intacto durante 2 a 3 días, tras lo cual se convierte en función de barrera comprometida (es decir, las tasas de TEWL aumentar en gran medida), debido a la pérdida de sloughing epidermis o deroofing de una ampolla. Tras el cierre de heridas (por ejemplo, cobertura completa de la zona dañada por la migración de queratinocitos), más tiempo se necesita para formar una epidermis plenamente estratificado con un bien formado estrato córneo. La mala función barrera se atribuye generalmente a un desequilibrio en el contenido de agua, la falta de una organización adecuada, y / o adhesión de la corneocytes que componen el estrato córneo.

A largo plazo efectos cutáneos

Residual de los efectos sobre la salud de la exposición significativa HD suelen ser respiratorias, oculares, cutáneas o. La permanente consecuencias de las lesiones cutáneas pueden incluir hipopigmentación, hiperpigmentación, la fragilidad de la piel que es fácilmente dañado por un traumatismo, y cicatrices hipertróficas. Cuidado de la Piel de hipersensibilidad y problemas de ulceración crónica también se ha informado de 16. Cicatriz hipertrófica es el resultado de la actividad fibroblástica incontrolada y sobrecrecimiento del tejido conectivo durante la reparación de heridas 16. Excesiva contracción de la herida se ha observado en un modelo porcino weanling siguientes profunda cutánea / espesor total HD lesiones 45 y observados en víctimas humanas. 44 cicatrización excesiva y / o contracción de la herida es más articulaciones e impide desfigurar la destreza y la locomoción. Si el tratamiento inicial no es lo suficientemente agresivo y la excesiva contracción de la herida y la formación de tejido cicatrizal se producen, liberación quirúrgico puede estar indicado. Como se acepta generalmente que los cánceres de piel pueden surgir en tejido cicatrizal, HD-inducida por las cicatrices se puede considerar que tienen un potencial carcinogénico. Ha sido previamente observado que los cánceres cutáneos derivados de la exposición aguda HD suele localizar en cicatrices. 16 neoplasias cutáneas Aunque se ha informado, parecen ser un poco consecuencia de la exposición HD si es que se debieron a la exposición por sí sola HD 46. Teorías sobre la mecanismo de expresión maligna abundan.

Conceptos para medidas de lucha contra las mostaza de azufre

Conceptos de medidas de lucha contra las vesicant agentes se han desarrollado y activa los programas de investigación han puesto en marcha una serie de Norte Organización del Tratado del Atlántico (OTAN). Institutos participan activamente en esta área de investigación incluyen el Ejército de los EE.UU. Instituto de Investigaciones Médicas de Defensa de la Química, con sede en Aberdeen de pruebas, Md, el Departamento de Ciencias Biomédicas de la Defensa Ciencia y Tecnología de Laboratorio, Porton Down, Salisbury, Wiltshire, Reino Unido, y la Defensa Investigación y Desarrollo de Canadá-Suffield Medicine Hat, Alberta. Desarrollado conceptos de investigación incluyen la eliminación de contacto corporal, la mejora de descontaminación, la intervención farmacológica, química y gestión de siniestros.

Eliminación de contacto

El primer paso en la protección de una persona de los efectos nocivos de la HD es eliminar el contacto con el agente. Los respiradores y máscaras protectoras pueden ser donned para eliminar respiratoria y ocular exposiciones, 47 especializados ropa de protección (incluyendo traje, guantes y protección a los pies) puede ser donned para prevenir el agente de llegar a la piel, 47 y tópica piel protectores 48 - 62 puede ser utiliza para proteger las zonas de la piel vulnerable a la agente (por ejemplo, muñeca, tobillo, cintura, cuello y cruces en la ropa de protección).

Descontaminación

Lesiones de la piel HD pueden evitarse por completo a través de la rápida eliminación física, lo ideal sería que en cuestión de minutos. La eliminación en menos de 2 minutos previene totalmente el daño, sino incluso más tarde disminuye la eliminación de perjuicio. 15 eliminación física sin agente de neutralización (por ejemplo, trapos de limpieza con un paño o gasa, raspado con depresor de lengua) puede eliminar los agentes a granel de la piel y ayudan a reducir la gravedad de cualquier lesión resultante. Copiosas cantidades de agua con jabón parecen ser bastante eficaz en la eliminación de líquidos físicamente HD de la superficie de la piel. 63 descontaminación también se puede realizar por adsorción física con o sin productos químicos de inactivación. Hay una serie de descontaminación de agentes disponibles para eliminar el agente que se basan en polvo reactivo (por ejemplo, Ambergard XE-555 de resina [Rohm and Haas Company, Philadelphia, Pa] 63], neutralizando las soluciones (por ejemplo, el 0,5% de hipoclorito de 63], lociones para la piel reactiva (por ejemplo, reactiva la piel Descontaminación Loción [RSDL, O'Dell Ingeniería Ltd / EZ-EM, Inc, Lake Success, NY] 64], o polvos absorbentes (por ejemplo, la tierra más completa). Igualmente eficaz, sin embargo, se dispone de productos para el hogar, tales como la harina de panificación seguido por las toallitas húmedas. 65 disolventes como el queroseno y el espíritu de cirugía también puede ser eficaz para eliminar la piel de HD 66.

La cicatrización de la herida utilizando estudios weanling la peste porcina han demostrado que tras la exposición a puro líquido HD de 120 minutos, hay un período significativo de fuera de gaseamiento de agente no encuadernado, medido por un monitor MINICAMS aire (OI Analítica, College Station, Tex; JS Graham et al, datos no publicados, 2001). La cuantificación y localización de los HD depósito responsable de este largo fuera de gaseamiento en este modelo animal no se ha realizado. La existencia de un depósito por vía cutánea de HD en los seres humanos se propuso en la Primera Guerra Mundial 1 de Smith et al, 66 que demostraron que HD lesiones podrían prevenirse mediante el lavado de la piel contaminada con un disolvente apropiado hasta 45 minutos postexposición.

Por otra parte, Smith et al demostraron que la piel embalse de HD puede ser transferido a una segunda persona, incluso después de la superficie expuesta ha sido descontaminado. Sin embargo, los estudios realizados durante la Segunda Guerra Mundial, informó el efecto contrario en HD que fue rápidamente "fijo" de los mandantes piel tales como las proteínas. 40 Contemporáneas los estudios in vitro han confirmado la conclusión original de Smith et al que una importante reserva de HD está formado en la piel humana que puede representar hasta el 35% de la dosis aplicada después de 24 horas. 41 Este embalse ha sido localizado en el estrato córneo y la epidermis superior. Esto corrobora el trabajo realizado por Cullumbine 67 lo que demuestra que el proceso de vesication podría ser bloqueada por la oportuna aplicación del "pelado" (keratinlytic) agentes de hasta 14 horas postexposición. The existence of an HD depot in human skin for a period of time following exposure has implications for the safety of medical emergency personnel treating HD casualties; for example, use of protective apparel may be warranted. Prior to medical treatment or casualty transport in enclosed vehicles, thoroughness of cleansing should be ensured through multiple washings and/or use of a detector. The existence of an agent depot could also influence the design of decontaminating agents. Any decontaminating agent that is capable of pulling the targeted agent out of the skin as well as neutralizing it on the surface of the skin has the potential to decrease HD-induced pathology, even beyond the 2-min efficacy window 15 of conventional decontamination procedures.

Understanding the kinetics of HD skin absorption, and the amount and persistence of unbound HD in putative agent reservoirs, will aid in choosing the most appropriate decontaminating agent and in determining its application doctrine and its window of effective use. The best agent will be the one that not only decontaminates HD sitting on the surface of the skin, but also is capable of fully penetrating the stratum corneum and neutralizing any unbound agent reservoir located there, unlike previous reactive therapies. 68 Research is underway to develop an in vitro model for efficacy testing of advanced decontaminating agents capable of pulling HD out of the skin reservoir and neutralizing HD on the surface of the skin, to identify the most appropriate animal model for extrapolation of animal data to humans, and to conduct in vivo efficacy tests of candidate decontamination systems.

Pharmacological intervention

Because HD is not painful on contact, the exposed person may not be aware of the exposure until symptoms begin to appear after the latent period. Pharmacological approaches are being studied for their efficacy in minimizing or preventing that damage. 6971 The most successful strategies to date have included the use of anti-inflammatories, protease inhibitors, intracellular scavengers, cell cycle inhibitors, PADPRP inhibitors, and calcium modulators, (Dr William J. Smith, PhD, US Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, Md, oral communication, 2004). It is interesting to note that pretreatments (eg, antioxidants) that reside in the extracellular matrix are generally more effective than those designed for intracellular protection against HD (Defence Science and Technology Laboratory, Porton Down, unpublished data, 2004). This may imply that the cytotoxic effects of HD are mediated via interaction with cell surface components rather than intracellular targets. Thus, further investigations of the interaction of cell-surface molecules with HD may provide a new insight into the mechanisms of HD toxicity.

Chemical casualty management

When HD comes in contact with the skin, decontamination is not performed in a timely fashion, and pharmacological intervention is absent or not adequately effective, a chemical casualty will be produced that requires medical attention. Casualty management now comes into play, discussed in depth in the next section. Educational material and training courses are available through the Chemical Casualty Care Division of the US Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, Md (on the Web at https://ccc.apgea.army.mil/default.htm , by phone at 410-436-2230/3393, or by e-mail at ccc@apg.amedd.army.mil ).

CURRENT TREATMENTS FOR CUTANEOUS SULFUR MUSTARD INJURIES

There are currently no standardized or optimized methods of casualty management and no drugs available to prevent the effects of HD on skin and mucous membranes. Historically, blister aspiration and/or deroofing (epidermal removal), physical debridement, irrigation, topical antibiotics, and sterile dressings have been the main courses of action in the medical management of cutaneous HD injuries. 12 , 14 , 15 , 72 , 73 Current treatment strategy consists of symptomatic management and is designed to relieve symptoms, prevent infections, and promote healing. The general recommendations listed below are fully described in the Textbook of Military Medicine , 15 the Field Management of Chemical Casualties Handbook , 74 the Medical Management of Chemical Casualties Handbook , 75 and the NATO Handbook on the Medical Aspects of NBC Defensive Operations . 76

The decision to evacuate and hospitalize an HD casualty is based on the extent and severity of the skin lesions, in consideration with other injuries that may be present (eg, respiratory, ocular). For patients experiencing only cutaneous HD injuries, erythema covering more than 5% TBSA in noncritical areas requires hospitalization. Erythema covering less than 5% TBSA may require hospitalization, depending on the site of the injury (eg, face, inguinal area) and level of impairment (eg, limitation of limb movement due to pain, edema). Total body surface area can be determined using Wallace's Rule of Nines and the Lund and Browder chart for estimating burn severity. 77 , 78 Multiple or large areas of vesication will also require hospitalization. Since blister formation may initially be slight, the patient should be watched for a progression in the size and number of blisters. Topical antibacterial creams such as silver sulfadiazine or 10% mafenide acetate can be prescribed to patients not requiring close medical monitoring, with instructions to apply a thin layer to the affected area 4 times a day. Following application of the cream, the area should be covered with a petrolatum gauze bandage.

Not all burn injuries require specialized care in a burn center. The American Burn Association has well-defined criteria for patient transfer to such a center, 78 which should serve as additional guidance in deciding where to hospitalize an HD casualty.

Before commencement of any treatment, clothing should be carefully removed and treated as potentially contaminated, and the patient thoroughly decontaminated. For a general overview of decontamination procedures, the reader is directed to Hurst's chapter on decontamination in the Textbook of Military Medicine . 63

Direct comparisons in the literature between HD and thermal burns are scarce. Papirmeister noted that disintegration of the basal cell layer caused by thermal burns has been shown to produce an intraepidermal blister that contains fragments of the basal cell layer attached to the basal lamina, unlike the almost totally denuded basement membrane in HD lesions. 1 An argument against the adage “a burn is a burn” is that HD initially targets a specific cell type (epithelial basal cells), unlike a thermal burn that begins damage at the stratum corneum and then works its way downward. Since stratum corneum is the structure largely responsible for barrier function, water loss rates are very high immediately after a thermal burn (140–180 gm −2 h −1 in humans). 79 The stratum corneum remains intact for 2 to 3 days after a cutaneous HD injury, after which barrier function becomes compromised by loss of sloughing epidermis or deroofing of the blister. The systemic fluid derangement seen in cutaneous HD injury is less than that seen with thermal burns. Fluids and electrolytes should be closely monitored, since fluids may be lost to edematous areas, with resultant dehydration. Medical personnel are cautioned not to overhydrate the patient; hypervolemia and pulmonary edema can be iatrogenically induced in HD casualties. 15 , 44 Fluid requirements in Iranian casualties during the Iran-Iraq war appeared to have been relatively independent of TBSA. 44 The recommended infusion rates and formulas used to calculate total volume requirements for thermal burn patients, based on body weight and TBSA, 80 should not be routinely applied in HD casualty management. The exact fluid replacement requirements for cutaneous HD injuries should be based on patient status and considered on a case-by-case basis. The fluids used in replacement fluid therapy for non-HD burns, which would likely be appropriate for use in HD injuries if fluid replacement is required, are described by Settle, 80 Brisebois, 81 and Thomas et al. 82

Sulfur mustard casualties should be kept comfortable and their lesions regularly cleansed to prevent infections. Limbs may need to be immobilized, as movement of joints can aggravate existing lesions. Blisters arising on the trunk require protective dressings to avoid or minimize damage as a result of friction with clothing or bedding. As burning and itching sensations are typically present after the appearance of erythema is noted, topical antipruritics are applied (eg, calamine lotion, 0.25% camphor, menthol, corticosteroidal preparations, and silver sulfadiazine cream). Systemic analgesics and antipruritics may be indicated, depending on the discomfort level of the patient.

Infection is a significant factor in causing delayed healing of cutaneous HD injuries. These injuries are covered by necrotic debris, which is a nidus for infection. There is no consensus, however, on whether intact blisters should be deroofed. Blister fluid from intact blisters provides a sterile wound covering, but the blisters are fragile and easily ruptured. Once blisters have broken, ragged roofs should be removed and sterile dressings put in place as soon as possible. Wounds should be inspected periodically for signs of infection.

It is generally recommended in military medical manuals to deroof blisters that are greater than 1 cm in diameter, irrigating the underlying area 2 to 4 times per day with saline, sterile water, clean soapy water, or Dakin's solution. Following cleansing, the area should be liberally covered with a topical antibiotic cream (eg, silver sulfadiazine; mafenide acetate; bacitracin; and triple combination preparations of neomycin sulfate, polymyxin B sulfate, and bacitracin zinc [Neosporin, Pfizer Inc, New York, NY ]). A sterile dressing should then be put in place. Blisters that have already broken should have their ragged edges removed and the area irrigated, treated with antibiotic, and dressed with a sterile dressing. Blisters less than 1 cm in diameter should be left intact, with the area surrounding the blister irrigated at least once per day followed by application of a topical antibiotic. A petrolatum gauze bandage can be put in place over these unbroken blisters, if desired. Any such dressings should be changed every 3 to 4 days.

While these handbooks recommend the use of bacitracin and triple combination preparations following cleansing of deroofed blisters, they do not provide specific application guidelines. We feel that the use of these ointments should be limited to small wounds (less than 1% TBSA) and employed for very brief periods (3–5 days) because of their high capacity to provoke allergic cutaneous reactions. Likewise, the use of 10% mafenide acetate cream should be avoided because of the severe pain that it causes when applied to partial-thickness wounds and the possibility of metabolic derangements. Such problems are not encountered with the use of 5% mafenide acetate solution, which should be used instead of the 10% cream.

STRATEGIES FOR THE DEVELOPMENT OF IMPROVED THERAPIES

Previous animal studies have shown that surgically aggressive approaches are needed to prevent or minimize significant cosmetic and functional deficits that result from deep HD injury. For the best outcome, deep dermal/full-thickness cutaneous HD injuries require full-thickness debridement followed by autologous split-thickness skin grafting. 45 , 83 These surgically aggressive approaches to deep HD injuries resulted in the return of barrier function, skin color, and mechanical properties (hardness and elasticity) to near-normal levels within 15 days of treatment in a weanling pig model. 83 To be successful, the skin grafts must be placed on a hemostatically secure wound bed, devoid of blood clots, debris, or necrotic tissue. The recipient bed must have an adequate blood supply to nourish the skin grafts, and the grafts must be protected from shearing forces, motion, and mechanical disruption. A variety of modalities is available and may be employed in achieving initial graft adherence and subsequent acceptance (“take”). These include sutures, surgical staples, fibrin glue, tie-over bolsters, compression dressings, and a variety of antishear dressing techniques. The choice of fixation and dressing technique is determined by the size and location of the wounds, and the experience and preferences of the surgeon.

Split-thickness skin grafting was used late in a few cases during the Iran-Iraq conflict where healing was particularly slow 12 and for the late treatment of some poorly healing, deep injuries sustained in 1992 by a civilian who came across an unexploded artillery shell from the First World War. 72 The NATO Handbook on the Medical Aspects of NBC Defensive Operations states that grafting has rarely been required in the past and when it was attempted, graft acceptance has been poor. 76 Surgical details of the grafting procedures used are not readily available, and the procedures may not have been optimal. In contrast to this handbook, Graham et al 45 noted equally high graft acceptance rates following either full-thickness sharp surgical tangential excision or laser debridement using a deep dermal/full-thickness HD injury model in weanling pigs. In thermal burns management, deep burns are grafted to promote timely wound closure and improve outcome with minimal cosmetic and functional deficits. The decision to graft is based on depth of injury. As with thermal burns, depth of HD injury should be accurately assessed before treatment begins. Reported long-term effects such as fragile skin and scarring likely indicate that injury depth was not accurately diagnosed and treatment was not sufficiently aggressive. As with deep thermal burns, deep HD injuries will require surgically aggressive approaches.

While past HD wound-healing research in swine has concentrated on deep dermal/full-thickness injuries, superficial (epidermis only) and superficial dermal injuries may have greater clinical relevance on the battlefield. Partial-thickness injuries will likely not require such surgically aggressive approaches (eg, split-thickness skin grafting). Treatment strategies for improved healing of partial-thickness cutaneous HD injury have recently been formulated by a working group of researchers and physicians at government laboratories in the United States and United Kingdom. The strategies are described below. Research is underway to experimentally support these strategies and determine which medical devices, supplies, and pharmaceuticals are most efficacious.

It is important to recognize that for any therapeutic regimen to be successful, a healthy immunological, 84 nutritional/metabolic, 85 and psychological 86 status needs to be maintained. Infections and perturbations in organ function also need to be closely monitored and addressed as needed.

The ultimate goal is to determine the most efficacious treatment regimen to be applied in the clinical management of HD casualties. The ideal regimen should return damaged skin to optimal appearance and normal function in the shortest time. Improved treatment will result in a better cosmetic and functional outcome for the patient and will enable the casualty to return to normal activities sooner.

The pig as an animal model for efficacy testing of candidate treatment regimens

Since human testing with vesicating agents such as HD is unethical, candidate treatment regimens need to be tested in an appropriate animal model. The animal model of choice is the pig, due to the similarities between human and porcine skin. 8797 The comparable histological characteristics of pig and human skin are similarities in epidermal thickness and composition, epidermal enzyme patterns, epidermal tissue turnover time, lipid content , character of keratinous proteins, pelage density and pattern of hair growth, dermal structure, deposition of subdermal fat, and general morphology. 9092 In addition, pig skin is antigenically closer to human skin than is rodent skin. A number of human antibodies have been shown to cross-react in pig skin. 98 US Environmental Protection Agency guidelines for dermal exposure assessment state that the percutaneous absorption of many compounds in the pig is similar to that found in humans. 93 Dick and Scott 90 found that pig skin permeability to selected lipophilic penetrants was closer to that of human skin than was rat skin. Klain et al 94 concluded that pig skin was a good model for human skin metabolic studies. Meyer et al 95 concluded that among the domestic species, the pig provides the most suitable experimental model for dermatological research on humans. Results from studies of experimental treatments in porcine models of partial-thickness wound healing have correlated well with results of clinical studies. 91 These findings suggest that the pig is a suitable research animal to use for predicting cutaneous effects of xenobiotics in humans. Some differences between human and porcine skin have been noted, however. Pigs lack eccrine sweat glands, 92 although hair and sebaceous gland number and distribution are similar. 91 Tubular apocrine glands are present and lie adjacent to hair follicles, and are more numerous than in humans. 92 The subepidermal vascular network is less dense than it is in humans; however, the pattern of vascularization in the lower region corresponds to that found in humans. 92 In addition, the permeability of pig skin to HD may be significantly higher than that of human skin under certain conditions. 96

Pigs have been widely used in vesicant research. 10 , 11 , 38 , 45 , 83 , 96109 Weanling pigs have often been used for their ease of handling during lengthy wound healing studies. They are small enough to be easily placed under chemical fume hoods during agent exposures and have a large enough body surface to allow multiple, large (3-cm-diameter) lesions to be placed on either the dorsum or the ventral abdominal surface. While they do grow during wound healing studies, functional data (eg, microcutaneous blood flow, skin color) can be normalized to surrounding unaffected skin 83 and morphometric data can be normalized to total body surface area 45 to take this growth into account. The healing process appears to be optimal in young pigs. 92 The healing rate is faster in Yorkshire piglets than in mature Yucatan miniature pigs, 91 and young pigs are highly resistant to contamination and infection. 92 Furthermore, pigs are amenable to habituation and can be trained to allow noninvasive biophysical skin measurements to be obtained from HD-exposed sites on the dorsum without the need for restraint or anesthesia. 110 Finally, the use of pigs in cutaneous ulcer and burn wound research is supported by the US Food and Drug Administration. 111 Thus, weanling pigs appear to be a suitable model for examining the efficacy of treatment regimens without significant interference caused by high infection rates or slow healing rates. However, they would not likely be the ideal model for studying wound colonization or infection.

While there is no common laboratory animal species, including the pig, that generates frank blisters, as do humans, HD has been noted to induce microblisters in pigs. 10 This lack of frank blistering is thought to be the result of a diminished superficial dermal vascular plexus, a densely arranged dermis, and lack of loose areolar tissue that precludes intercellular fluid accumulation. 92

Immediate treatment of cutaneous sulfur mustard casualties

This section describes the recommendations of the US-UK working group for the immediate treatment of HD casualties. The uses of anti-inflammatory agents, antioxidants, and occlusive/semiocclusive dressings are discussed. The potential need for replacement fluid therapy and management of intact blisters are also addressed.

For those patients who are beginning to present with erythema or those who are in the latent period and suspect an exposure may have occurred, systemic administration of an anti-inflammatory agent will likely help to decrease the amount of damage ultimately induced. The pro-inflammatory mediators IL-1β, IL-6, IL-8, and TNF-α are released by normal human epidermal keratinocytes in culture on exposure to HD. 112 Sulfur mustard has also been shown to provoke an edema response and release of IL-6 in 2 different mouse models. 113 Sulfur mustard–induced inflammatory responses themselves likely contribute to the severity of the pathology, and numerous animal studies have shown the benefits of prophylactic or therapeutic use of anti-inflammatory agents. 43 , 6971 There are active programs researching a variety of nonsteroidal anti-inflammatory drugs (NSAIDs) administered topically or systemically, alone and in various combinations. Top candidates that have shown efficacy in a mouse ear model include indomethacin, fluphenazine dihydrochloride, olvanil, retro olvanil, octyl homovanillamide, and other analogs of capsaicin. 6971 It remains to be determined which NSAID (or combination), route of administration, length of administration, and dosing regimen is the most efficacious in preventing or ameliorating the effects of HD on skin. It is likely that administration for 2 to 5 days will be required for an NSAID. Topically delivered intracellular scavengers such as 4-methyl-2-mercaptopyridine-1-oxide 69 and dimercaprol 69 , 71 have proved effective in animal experiments in reducing the severity of HD-induced cutaneous injuries, and concurrent use of one of these agents with an NSAID may yield the best results (Dr William J. Smith, PhD, US Army Medical Research Institute of Chemical Defense, Aberdeen Proving Ground, Md, oral communication, 2004). (It should be noted that mouse skin is very thin and permeable. Any topical agent showing efficacy in a mouse model should also be tested in another animal model such as the weanling pig.) Corticosteroid anti-inflammatory agents such as hydrocortisone (given systemically or topically for cutaneous HD injuries) 70 , 71 and dexamethasone (tested in vitro on primary alveolar macrophages and given topically for ocular HD injuries) 114 , 115 also appear to be promising therapeutic agents. There are other topical steroidal anti-inflammatory agents of much greater potency that would likely be very efficacious if used early in the lesion development stage, such as betamethasone dipropionate, clobetasol propionate, and diflorasone diacetate. Superpotent (Class 1), potent (Class 2), and upper mid-strength (Class 3) topical corticosteroids should be tested for their efficacy in ameliorating HD-induced cutaneous injury.

As previously discussed, depletion of GSH and accumulation of endogenous oxidants and ultimate formation of potent oxidizing species (eg, toxic lipid peroxides) may be contributory factors in HD-induced cytotoxicity. 1 Topically applied HD has been shown to negatively affect antioxidant enzymes in blood cells and body tissues of rats. 116 Several antioxidants have been shown to protect liver and lung from oxidative damage following inhalation or percutaneous exposure to HD in a mouse model. 117 It has been suggested that administration of antioxidants may be protective and useful. 118 Thus , initial antioxidant treatment aimed at affecting the progression of lesions that is instituted during the erythema phase may prove to be of benefit. The effectiveness and role of the interruption of the inflammatory cascade by the inclusion of topical and systemic antioxidant agents as well as a determination of the optimal timing for such therapy are important and intriguing avenues for investigation.

Placement of an occlusive or semiocclusive dressing will likely prove to be helpful in promoting autolytic debridement and preventing desiccation. Debridement will play a central role in improving the healing of cutaneous HD lesions, and beginning the process early may be beneficial. How soon following exposure these dressings can be applied remains to be determined. While maintaining a moist environment has long been known to facilitate wound healing, 119122 caution needs to be observed since very early occlusion that builds up moisture levels in the skin will exacerbate the lesion. In addition, there is a period following exposure to sulfur mustard at which off-gassing of unbound HD occurs following a vapor 104 or liquid exposure in weanling pigs (JS Graham et al, unpublished data, 2001). The existence of a reservoir of unbound HD in human skin was previously discussed. These studies have suggested that off-gassing can continue for 24 to 36 hours, given a large exposure. Limiting the escape of this unbound HD by using an occlusive dressing may exacerbate the lesion. Placement of any occlusive dressing should probably be postponed for at least 24 hours following exposure. Keeping clothing off the exposed area, thereby not allowing vapors to build up, may also be of benefit.

The potential need for replacement fluid therapy (RFT) and caution in avoiding overhydration was previously discussed. Requirements for RFT can be studied experimentally. A scientific study using an appropriate animal model needs to be conducted to determine whether body weight and percent TBSA affected should be taken into consideration when trying to determine the total volume of fluid required. It is also important to determine which fluids are most appropriate, what infusion rates are needed, and when to commence RFT. Using instrumented pigs, large areas of injury can be induced, followed by close monitoring over time of important physiological parameters. The effects of different RFT protocols can then be determined. To generate large TBSA injuries, exposures with partial-body vapor chambers can be used. Alternatively, liquid HD can be diluted to volumes greater than that allowed under current surety regulations and fast-wicking material used to spread the liquid out over a large area. Previous research has enumerated a number of stable vehicles that are suitable for cutaneous exposure with liquid HD. 108

Management of casualties presenting with intact blisters varies according to the situation and level of care. Avoidance of infection is paramount so that the depth of injury does not increase. Small blisters should not be disrupted until deroofing can be done under controlled conditions. Larger blisters that are already flaccid may require deroofing or collapsing under sterile conditions. Current data does not permit recommendation of a single best approach. Once the necessary conditions and skills are reached, intervention should be more aggressive, with the dual goals of avoiding wound infection and optimizing recovery. As detailed below, aggressive management of the cutaneous HD injury, as opposed to that conducted on thermal or toxic epidermal necrolysis blisters, will require removal of the wound edges into normal-appearing skin along the periphery of the lesion and debridement of the base of the blister through the damaged BMZ into healthy dermis. Accurately determining depth of injury will influence this step. These patients will also require pain management and close observation for the systemic effects of HD exposure. Absence of thorough removal of damaged tissues will greatly slow healing and will enhance scarring and contracture in all but the most minor injuries.

For patients presenting with intact frank blisters, it may be beneficial to aspirate the blister fluid with a sterile needle and syringe and allow the roof of the blister to act as a sterile dressing until a physician can remove it. Reattachment of blister roofs has been noted to occur in the treatment of vitiligo via epidermal grafting using the tops of suction blisters 123 , 124 and in experimental suction blisters in humans following aspiration of blister fluid. 125 The roofs of HD blisters, however, are not expected to reattach to the blister floor owing to HD-induced damage to basal cells and BMZ components. Sloughing is expected to eventually occur. For patients presenting with ruptured HD-induced blisters, careful removal of the blister roof with scissors, application of an antibiotic ointment, and placement of a sterile dressing is warranted. For both of these scenarios, more complete debridement will be necessary for large lesions, as described later.

Treatment of established partial-thickness lesions: overview of the approach

The remaining sections of this review describe the recommendations of the US-UK working group for the treatment of established partial-thickness cutaneous HD lesions. The general approach that is being taken is to perform adequate debridement of partial-thickness injuries, then treat the lesions like chronic cutaneous ulcers or partial-thickness thermal burns using contemporary medical approaches.

Before treatment can begin, the extent and severity of cutaneous HD injuries must be determined. Following assessment of injury and deroofing of frank blisters, adequate wound debridement needs to be performed, followed by 1 or more treatment adjuncts. Examples of adjuncts under consideration are dressings, growth factors, skin substitutes, topical nutritional support, and Vacuum Assisted Closure™ (VAC®), Kinetic Concepts, Inc (KCI), San Antonio, Tex.

Injury assessment

Before HD injuries can be appropriately treated, assessment of the injuries must be made. Total body surface area of the injuries should be established and depth of injury determined. Total body surface area can be determined using Wallace's Rule of Nines and the Lund and Browder chart for estimating burn severity, as previously discussed. 77 , 78 Determination of injury depth is a much more challenging task. Accurate depth assessment is important because it dictates how aggressive treatment needs to be to minimize or prevent cosmetic and functional deficits.

In thermal burns, depth of injury is typically assessed by physical examination. Surface appearance, the pinprick test to assess pain, the “blanch-capillary return test” to evaluate microcirculation, and surface temperature difference between burned and unburned skin are often used in diagnosis of depth. 126 Using these methods, diagnosing very superficial and very deep burns is relatively easy for the experienced burn surgeon. Burns of intermediate depth are often problematic in determining the need for grafting. Determining depth of HD injuries is more challenging. First, the full extent of cutaneous injury can take several days to manifest. Second, superficial appearances do not accurately predict depth of injury or need for grafting. While the presence of blisters in thermal burns is generally associated with superficial dermal injuries, blistering in HD injuries can occur in deep dermal/full-thickness injuries because of the unique nature of the agent and the unique progression of the injury.

Noninvasively examining cutaneous blood flow using available bioinstrumentation can greatly assist the physician in determining depth of injury. Laser Doppler perfusion imaging and indocyanine green fluorescence imaging may prove to be very valuable tools in prognosticating optimal wound healing of both thermal burns and cutaneous HD injuries.

Laser Doppler flowmetry and laser Doppler perfusion imaging (LDPI) have been used for prolonged, noninvasive monitoring of tissue viability and wound healing and for the assessment of peripheral vascular disease, inflammation, ischemia, reperfusion, skin graft acceptance (take), and burn depth . 127142 Laser Doppler perfusion imaging may prove useful in delineating the areas of HD damage that need to be debrided, thereby avoiding areas with sufficient blood flow. Brown et al 101 found that laser Doppler perfusion images of vesicant vapor burns on the backs of swine correlated well with histopathological findings (thrombosis and necrosis of subepidermal capillaries) between 1 hour and 7 days postexposure and suggested that clinical management decision making for treatment of early vesicant burns could be aided by LDPI. Chilcott et al 102 used several noninvasive bioengineering methods to monitor wound healing in a large white pig model for 7 days following exposure to HD and Lewisite vapors. They concluded that while reflectance colorimetry and TEWL measurements could provide quantitative, noninvasive methods for determining efficacy of candidate treatment regimens, neither is comparable to the prognostic capabilities of LDPI. Graham et al 83 found LDPI to be useful in examining blood flow in grafted and ungrafted sites following treatment of deep dermal/full-thickness liquid HD injuries in a weanling swine model (Fig 1 ). Laser Doppler perfusion imaging is currently rather time consuming if there are multiple sites to be evaluated and/or large images to be collected at high resolution. The length of scanning procedures could be decreased by increasing scanning speed (thus decreasing flux resolution), decreasing the size of the scan area, and/or decreasing the number of lines scanned within the scanning area (scan resolution). Improvements in the technology that will speed up LDPI without compromising image resolution are being developed.

Indocyanine green fluorescence imaging has also shown promise in determining burn depth based on microcutaneous blood flow. It is a minimally invasive procedure that requires the placement of an intravenous line. Indocyanine green (ICG) is US Food and Drug Administration–approved for use in humans to determine cardiac output, hepatic function and blood flow, and for ophthalmic angiography. The fluorescence of intravenous ICG has been shown to estimate burn depth in small animals. 143 In contrast to fluorescein fluorescence, 144 ICG fluorescence is capable of distinguishing superficial and deep partial-thickness burns from full-thickness burns. The fluorescence intensity of ICG decreases exponentially with burn depth for burns of similar age. 145 Indocyanine green fluorescence was used to estimate burn depth in a porcine model. 146 An imaging system with a diagnostic algorithm was developed at the Wellman Laboratories of Photomedicine, Boston, Mass, that accurately diagnosed burns that healed within 21 days with minimal scarring from those that took longer to heal by secondary means. Measurements were made on burns created 2, 24, 48, and 72 hours prior to imaging. The algorithm was shown to be dependent on the age of the burn but not on location. This technology showed promise in plastic surgical applications 147 , 148 and for accurate determination of thermal burn depth in humans. 147 , 149 Indocyanine green fluorescence imaging also shows promise in diagnosing depth of HD injury (Fig 2 ; JS Graham et al, unpublished data, 1999). The advantage that this technology has over LDPI is the speed of image capture. Multiple images over large areas can be captured in a relatively short period of time. Images are typically collected 5 to 10 minutes after ICG injection to allow uptake and distribution. The dye is then excited (eg, 780 nm) and the resultant fluorescence emission (eg, 825 nm) immediately captured and saved by a computer and analyzed for burn-to-normal skin fluorescence ratio. Indocyanine green binds strongly to plasma globulins, limiting both extravasation within burn-injured vascular epithelia and extravascular transport to areas nearby. 145 Large signals are thought to be the result of vasodilation and hyperemia, and smaller signals are thought to be attributable to vascular occlusion and edema. 143 , 145 With this technique, live streaming video can also be captured immediately after injection, allowing the physician to watch the dye flowing through viable tissue and around nonviable tissue in real time.

It is of fundamental importance that any noninvasive or minimally invasive technique used to assess lesion severity be fully validated in order that results may be correctly interpreted. For example, a recent study has demonstrated that TEWL may not necessarily correlate with skin “barrier function.” 150

Debridement

Experimental approaches to vesicant wound debridement have included powered dermabrasion, 99 , 106 , 107 sharp surgical excision, 45 , 83 , 151 laser debridement, 45 , 83 , 100 , 107 , 109 , 152 and enzymatic debridement. 152

Powered dermabrasion has been shown to speed up the reepithelialization process of cutaneous HD injuries. 99 , 106 , 107 Kjellstrom et al 151 found sharp surgical excision with primary suturing of the skin defect to be effective in decreasing healing time of HD vapor lesions in guinea pigs . Powered dermabrasion, pulsed CO 2 laser ablation, and erbium:yttrium-aluminum-garnet (Er:YAG) laser ablation have been shown to accelerate the rate of healing of full-thickness cutaneous Lewisite vapor burns in swine without the need for split-thickness skin grafting. 107 , 109 Eldad et al 152 found that excimer laser ablation and debrase (Debridase, MediWound Ltd, Yavne, Israel) enzymatic debridement were efficacious in improving the healing of partial-thickness nitrogen mustard burns in a guinea pig model.

Because of positive results achieved by laser and enzymatic debridement of vesicant injuries, our future research efforts will concentrate on the use of usthese methods to debride partial-thickness cutaneous HD injuries prior to the application of treatment adjuncts. The treatment regimen that is found to be most efficacious in the pig will ultimately be recommended for use in treating human casualties. Because humans form frank blisters, unlike the pig, these regimens would follow deroofing of any frank blisters present.

LASER DEBRIDEMENT OF CUTANEOUS VESICANT WOUNDS

Laser debridement of cutaneous vesicant wounds has proven to be an effective method of improving the rate of wound healing in pig models. Graham et al 100 showed that viability, thickness, and organization of the epidermis were all significantly improved by partial-thickness pulsed CO 2 laser debridement of small, mild to moderately severe cutaneous HD vapor injuries. Laser debridement followed by skin grafting was as efficacious in improving the wound healing of deep HD burns as sharp surgical tangential excision followed by grafting (the gold standard in human deep dermal/full-thickness thermal burns medicine). 45 , 83 Middermal debridement by sharp excision or laser ablation without grafting produced less desirable results but was better than no treatment. 45 , 83 A 4-fold improvement in reepithelialization of Lewisite injuries was achieved at 1 week following laser dermabrasion, with almost 100% reepithelialization by 3 weeks. 109 It is not apparent why these full-thickness Lewisite injuries (10 cm 2 ) did not require grafting, as did HD injuries (12.6 cm 2 ) 45 , 83 or as would a full-thickness thermal burn. There are differences in biochemical action and rates of spontaneous reepithelialization between Lewisite and HD injuries. 109 Further studies need to be conducted to fully examine the comparative healing of deep Lewisite, HD, and thermal injuries.

Laser debridement offers additional benefits including hemostatic control during surgery, minimal risk of exposure to aerosolized pathogens, and time efficiency. Another major advantage to the use of lasers is the ability to control the amount of normal perilesional skin that is removed. Eldad et al 152 noted that it is technically difficult to control the amount of tissue to be removed by surgical tangential excision and that laser ablation of nitrogen mustard burns in a guinea pig model enabled both controlling the amount of tissue to be removed and minimizing blood loss . Minimizing the amount of tissue removed will be of a cosmetic benefit to the patient.

Types of lasers available

Pulsed CO 2 lasers have been used for a variety of dermatological applications, including skin resurfacing, 153155 excision of burn eschar, 156159 preparation of adequate graft beds, 160 and conservative ablation of skin lesions. 161 They are designed to promote rapid healing by minimizing laser-induced residual thermal damage, 153 , 154 , 160 , 162166 and offer precise, micrometer-depth removal of tissue. 166 They vaporize tissue rapidly and efficiently, with minimal blood loss. 159 , 166 , 167 However, because of the low average power available from a pulsed laser system, these lasers may be inefficient while performing full-thickness debridement of deep burns or while debriding burns covering large body surface areas. Continuous wave (cw) lasers can be quite efficient in removing tissue but tend to create significant amounts of thermal damage, sometimes creating more damage than the initial burn being treated. Domankevitz and Nishioka 167 demonstrated that under appropriate conditions, a scanned cw CO 2 laser could ablate tissue with a zone of residual thermal injury less than 200 μm, making it useful for cutaneous surgery and debridement of burn wounds prior to skin grafting. Residual thermal damage of cw lasers can be minimized if the laser is scanned over the surface rapidly enough that the amount of time the laser spends on any 1 point mimics a short laser pulse. 167 , 168 Use of such lasers has proven efficacious in pigs 45 , 169 and humans. 170 Glatter et al 169 usused a prototype cw CO 2 laser in a thermal burn model in pigs and found that long-term scarring, based on Vancouver scar assessments, was equivalent at 6 months postsurgery in both laser-ablated + grafted and sharply excised + grafted burns. In addition, they noted no significant difference in engraftment rates between the 2 methods of debridement. Graham et al 45 had similar success in engraftment rates using this same laser to treat HD injuries in pigs. In an initial clinical trial, Sheridan et al 170 used a similar, commercially available system to perform full-thickness ablations of thermal burns in children. They found that no bleeding occurred in laser-ablated sites, that engraftment rates for both laser-ablated sites and sharply excised sites were equally high, and that there were no significant difference in Vancouver scar scores at an average follow-up of 32.0 ± 5.2 weeks.

There are a number of lasers manufactured in the United States, Canada, and Europe that could be considered for routine debridement of vesicant injuries. Acland and Barlow 171 have provided a review on the current uses of lasers in dermatological practice and a list of the types of lasers used for specific procedures. They list CO 2 and Er:YAG lasers as being the most appropriate for cutaneous resurfacing. While rapid-scanning, high-powered cw CO 2 lasers would provide time-efficient ablation of damaged tissue, 45 they are no longer commercially available. Pulsed CO 2 lasers such as the UltraPulse (Lumenis Inc, Santa Clara, Calif) are in common use in dermatology and plastic surgery and have proved effective in improving wound healing of cutaneous vesicant injuries. 100 , 109 Er:YAG lasers are also commercially available and have been used for a wide variety of procedures, ranging from facial resurfacing to burn debridement. 171175 They have been shown to be particularly useful in the debridement of partial-thickness burns 175 and in the management of deep Lewisite injuries. 109 Unlike the Gaussian beam profiles created by CO 2 lasers, Er:YAG laser beams tend to be uniform and produce uniform depths of ablation. 175 One commercially available unit, the Sciton PROFILE (Sciton Inc, Palo Alto, Calif), can be configured as a high-powered, dual-mode long-pulse Er:YAG laser that allows independent control of both depth of coagulation and depth of ablation. This versatility would be very advantageous to any clinic or hospital that treats burns and a variety of dermatological disorders. Er:YAG lasers will play a central role in future HD wound healing studies at our facilities.

ALTERNATIVE METHODS OF DEBRIDEMENT UNDER CONSIDERATION

There are alternatives to using a laser to debride vesicant injuries. Sharp surgical tangential excisions and powered dermabrasion have proved effective. 45 , 99 , 106 , 107 , 152 Curettage, cryotherapy, larval therapy, and enzymatic debridement may be effective, lower-cost alternatives.

Sharp surgical tangential excisions have been effective in the treatment of sulfur mustard 45 and nitrogen mustard 152 injuries. Powered dermabrasion has been shown to speed up the reepithelialization process of cutaneous sulfur mustard 99 , 106 and Lewisite 107 injuries. There are drawbacks with this method, however, including lack of uniform depth control and risk of aerosolizing pathogens.

Scraping, using dermal curettes, may be a viable option for removing desiccated material from the surface of the skin. Cryotherapy using ice, liquid nitrogen, or Peltier coolers may also be efficacious if applied early. The therapeutic effects of cooling pig skin soon after exposure to HD vapor has recently been reported. 176 It may be possible to superficially freeze HD lesions before they proceed to vesication, thereby retarding the activity of the HD and progression of the lesion. The frozen tissue could then be removed using appropriate means, and the wound bed dressed with an antibiotic and sterile dressing until healed. Caution would have to be observed in the length of time the cooling agent is in direct contact with the skin and its potential to induce hypopigmentation taken into consideration. Finally, the use of larval therapy (maggots), while unconventional, has undergone a renaissance in the past few years and has proven to be very effective in debriding and improving the healing rate of hard-to-heal wounds (eg, chronic leg and foot ulcers). 177184 The success of this approach may warrant study as a possible treatment of small TBSA sulfur mustard injuries.

A final alternative under consideration for debridement of HD injuries is enzymatic debridement. These enzymes are categorized as proteolytics, fibrinolytics, and collagenases, and are designed to dissolve necrotic tissue from wounds. 185 They are often used to debride chronic wounds (eg, decubitus ulcers, venous stasis ulcers, arterial insufficiency ulcers, diabetic foot ulcers). Many have been found to be safe and effective in removing devitalized tissue and accelerating healing in burns. 186193 Any burn eschar present is typically cross-hatched to allow the agent to penetrate into the wound. Other agents, such as the bacterial proteolytic enzymes streptokinase and streptodornase, have given disappointing results in deep burns because they do not break down the collagen that separates vital from nonvital tissue. 194 Use of fibrinolysins may impair wound healing of HD lesions, as fibrin is an early matrix protein that is essential for wound healing. In addition, fibrinolysins are typically combined with deoxyribonuclease (DNase) and as such will also digest DNA in the dividing fibroblasts, which play a role in healing. 185 Some effective enzymes have produced better results than others, with enzyme concentration, skin moisture level, and the presence of certain antibacterial agents affecting results. Secondary dressings are needed to keep the wound moist and to allow these agents to work. 185 Klasen 194 offers an excellent review of the use of enzymatic debridement agents in burns. The most popular and effective agents on the market today are collagenases (eg, Collagenase Santyl ointment, Ross Products Division, Abbott Laboratories Inc, Columbus, Ohio) and papain/urea combinations (eg, Accuzyme and Panafil, Healthpoint Ltd, Fort Worth, Tex ; and Gladase Papain-Urea Debriding Ointment, Smith & Nephew Inc, Largo, Fla). In addition, a promising proteolytic enzyme extracted from the stem of the pineapple plant is in Phase II clinical trials in the United States and Europe for the treatment of deep partial- and full-thickness burns (Debrase Gel Dressing, MediWound Ltd, Yavne, Israel ). Enzymatic debridement of HD injuries is a promising and cheaper alternative to laser debridement, albeit more time consuming. Research is planned for determining which available enzymatic debridement product is most efficacious in debriding partial-thickness HD injuries. The specific application regimen, the time required to reach adequate debridement, and potential adverse effects (eg, conversion to a deeper injury, infection) need to be determined in an appropriate animal model.

Burn wound sepsis and bacteremias have been noted in burn patients undergoing enzymatic debridement. 185 , 194 Concomitant use of a topical antibiotic that does not interfere with the action of the enzyme under study may be warranted as a preventative measure.

EXTENT OF DEBRIDEMENT REQUIRED

In addition to vesication and death of epidermal keratinocytes, HD exposure results in sublethal damage to keratinocytes along the periphery of the gross lesion. Damage to the basement membrane zone and underlying collagen in the papillary dermis has also been noted. Deroofing frank blisters followed by timely removal of this adjacent and subjacent damage will likely improve the rate of reepithelialization.

Nonlethal damage is clearly noted at the periphery of cutaneous HD lesions and has been reported previously. 11 , 105 , 195 Nikolsky's sign, 196 characterized by separation and loss of the epidermis from the dermis when the skin is pressed with a sliding or twisting motion, has been demonstrated in weanling pig skin following HD vapor exposure. 11 , 195 These weakened areas of the dermal-epidermal junction occurred along the periphery of the gross lesions and are indicative of sublethally damaged basal cells and/or altered proteins of extracellular matrices of the BMZ. Sublethally injured cells at the periphery of an HD lesion and in hair follicles and other adnexal structures may be partly responsible for the slow rate of reepithelialization seen in these injuries. Rice et al 106 suggested that the level of damage to cellular DNA at the margins of HD lesions may be sufficient to delay or prevent effective replication of those keratinocytes. Removal of these sublethally damaged keratinocytes at the margins of the lesions by debridement beyond the visible borders of the lesion will likely speed up the reepithelialization process.

As previously discussed, HD induces damage to the BMZ at the level of the lamina lucida. 5 , 39 The floor of the blister retains portions of the damaged BMZ and needs to be removed to provide an adequate scaffold over which keratinocytes feeding the reepithelialization process can migrate. Thus, at minimum, debridement needs to proceed down into the papillary dermis after removal of the blister roof. Beyond the BMZ, dermal collagen itself is affected by HD exposure and can itself impede the wound healing process. 97 , 106 , 197 Brown and Rice 197 reported coagulation and hypereosinophilia of the papillary dermis in Yucatan minipig skin 12 to 24 hours following saturated HD vapor exposure, with the deeper reticular dermis unaffected. Rice et al 106 and Lindsay and Rice 97 suggested that following exposure to HD, papillary dermal collagen is altered and may no longer function normally as a healthy scaffold over which epidermal cells can migrate.

The question of how deep to debride needs to be addressed. Ablative lasers that create less than 160 ± 60 μm of residual thermal damage permit optimal skin graft take and healing. 160 Domankevitz and Nisioka 167 concluded that lasers that induce residual thermal damage zones of less than 200 μm are useful for cutaneous surgery and burn wound debridement prior to skin grafting. Lam et al 109 were able to improve wound healing of full-thickness cutaneous Lewisite injuries in pigs by partial-thickness laser debridement. Graham et al 45 were also able to improve wound healing of deep cutaneous HD injuries in pigs by partial-thickness debridement without grafting, albeit not to the extent attained by full-thickness debridement followed by grafting. These studies thus indicate that retaining some amount of damaged dermal tissue does not significantly impede wound healing. Complete debridement of partial-thickness injury, therefore, will likely not be required. Debridement of partial-thickness HD injury into the papillary dermis or upper reticular dermis will likely be adequate.

Dressings

Following wound debridement of HD injuries, an appropriate dressing will be needed to promote moist wound healing. Beneficial effects of such dressings include prevention of tissue dehydration and cell death, accelerated angiogenesis, increased breakdown of dead tissue and fibrin (eg, pericapillary fibrin cuffs), significant reduction in pain, and potentiation of the interaction of growth factors with their target cells. 122 Helfman et al 119 and Singhal et al 185 have provided overviews of the various types of occlusive and semiocclusive dressings. Hydrocolloids, hydrogels, foam dressings, alginates, and transparent film dressings are commercially available from a large number of manufacturers. As foam dressings and alginates are designed to control moderate to heavy exudates, they will likely not be needed for covering debrided cutaneous HD injuries. Silver impregnated dressing materials may be of great potential benefit in treating these wounds owing to their antimicrobial efficacy 198200 and demonstrated ability to enhance rates of reepithelialization. 201 , 202 A number of these dressing materials are currently employed in burn and chronic wound care, while other more advanced silver dressings are in various stages of development. Thus, research on finding the most appropriate dressing during wound healing of these lesions should concentrate on hydrocolloids, hydrogels, thin films, and silver-impregnated dressings. Testing these products in a pig model should be adequate, since similar responses to occlusive and semiocclusive dressings on wound healing have been noted in pigs and humans. 203

Weak attachment of the neoepidermis to the underlying dermis has been noted in human HD casualties 44 and experimentally exposed weanling pigs (JS Graham et al, unpublished data, 2004). Once the lesions have fully reepithelialized, protective dressings may be needed to avoid or minimize damage as a result of friction with clothing or bedding.

Growth factors

During cutaneous wound healing, growth factors play dominant roles in regulating cell proliferation, differentiation, and synthesis of extracellular matrix. 91 Epidermal growth factor (EGF), transforming growth factor-β (TGF-β), platelet-derived growth factor (PDGF) , insulin-like growth factor (IGF), keratinocyte growth factor (KGF), hepatocyte growth factor (HGF), granulocyte-macrophage colony-stimulating factor (GM-CSF), and fibroblast growth factors (FGFs) play important and critical roles in the healing of cutaneous wounds. 203241 Reviews of the effects of these growth factors on wound healing have been previously published. 203208 Improved wound healing has been reported for topical applications of EGF, 203220 , 222 , 223 PDGF, 203208 , 214 , 219 , 224229 KGF, 206 , 207 , 231233 and IGF-I. 235 , 236 There have been some negative reports on the effectiveness of EGF 221 and KGF 230 in improving wound healing of experimental split- thickness skin wounds in humans and esophagogastric anastomotic wounds in rats, respectively. Epidermal growth factor has been shown to improve the healing of graft donor sites, 210 corneal burns, 217 , 220 and cutaneous burns, 218 , 219 whereas PDGF and KGF have been shown to improve the healing of burns 219 and skin-grafted lesions. 224 , 233

Human leptin is a 146–amino acid residue, nonglycosylated polypeptide involved in body weight regulation. It is released from white adipose tissue and exerts its effect via receptors in the hypothalamus. While not a growth factor per se but rather characterized as a satiety-regulating cytokine, leptin has been shown to be a potent mitogenic stimulus to keratinocytes during skin repair. 237 , 238

Results from these reports suggest that topical application of healing-enhancing factors alone or in combination may be beneficial in improving the wound healing of cutaneous HD injuries following wound debridement. When to commence such treatment following agent exposure needs careful consideration and experimental testing. During the early phases of wound healing, chemokines and cytokines regulate the chemotaxis and activation of inflammatory cells, along with synthesis of proteases and protease inhibitors. 91 Early application of growth factors would be ineffective in a milieu of proteases. Application of such growth factors or combinations thereof will likely require a delay of 3 to 7 days following HD exposure until the inflammatory response has subsided. Concomitant use of protease inhibitors or a dressing that binds matrix metalloproteases and protects growth factors (eg, PROMOGRAN* Matrix Wound Dressing, Johnson & Johnson Wound Management Worldwide, Somerville, NJ) may be necessary. A number of these growth factors are commercially available for efficacy testing in animal models (EGF, PDGF, leptin). Of these 3, only PDGF has been approved for human use by the US Food and Drug Administration. Recombinant human EGF (β-urogastrone) is available from Roche Diagnostics Corp, Indianapolis, Ind. Regranex Gel, a recombinant human PDGF-BB, is available from Johnson & Johnson Wound Management Worldwide, Somerville, NJ. Recombinant human leptin is available from R&D Systems Inc, Minneapolis, Minn. Amgen Inc (Thousand Oaks, Calif) has recently completed Phase III clinical trials of a recombinant human KGF (Palifermin) that significantly reduces the duration and incidence of radiation- and chemotherapy- induced oral mucositis.

In addition to these growth factors, topical application of extracellular matrix components such as fibronectin, which supports fibroblast, keratinocyte, and endothelial cell adhesion and movement, 242 , 243 may be of benefit following wound debridement. Fibronectin can serve as a template for collagen deposition 242 and is a key component of the provisional matrix during wound repair. 243 Exogenous application of intact fibronectin has been shown to be beneficial in helping to close human skin and corneal wounds. 244 The ability of keratinocytes to spread on fibronectin may require the presence of serum or epibolin/vitronectin. 244 Addition of another adhesive glycoprotein, laminin, may actually be detrimental to the reepithelialization process. When human keratinocytes are placed in apposition with collagen, they attach and begin migrating. 244 This migration is inhibited by the addition of laminin, which acts as a major cell adhesion factor for keratinocytes. Laminin 5 has been shown to inhibit human keratinocyte migration and strongly promotes keratinocyte attachment. 244 It is believed to anchor the keratinocytes to the substratum, via α 3 β 1 -integrin receptors. 244

Skin substitutes

Skin substitutes may provide an excellent temporary wound dressing for debrided HD injuries. Permanent wound closure can only be achieved by spontaneous reepithelialization or by the provision of autologous skin by means of skin grafting. The use of skin substitutes to temporarily restore the multiple functions of normal skin may be of substantial benefit in the management of cutaneous HD injuries.

For a skin substitute application to be successful, the same conditions required for successful autograft “take” must be created and maintained. The selection of the most suitable and effective temporary skin substitute will require a critical assessment of its comparative attributes when applied to HD wounds as well as the issues of cost, ease of use, availability, and consistency of results. Skin substitutes are widely used in human thermal burns management and can be (1) temporary or permanent; (2) epidermal, dermal, or composite; and (3) biologic or synthetic. 245253 They have also been shown to be effective in speeding up time to closure of chronic leg and foot ulcers, 254269 surgical excision sites, 269 and partial-thickness donor sites. 269 They may be a source of growth factors and are generally semiocclusive in nature. They can provide barrier function; add tensile strength to the wound; are generally flexible and pliable; markedly reduce pain, inflammation, and drainage; and provide a moist wound healing environment. They do not control deep bacterial infections; can seal bacteria in; and, being a biologic, they can transmit infection. Hence, the wound surface must not be infected for application of a skin substitute. A number of skin substitutes are available on the market and should be tested for their efficacy in improving wound healing of cutaneous HD injuries. Marketed products currently under consideration include (1) living bilayered skin substitutes (APLIGRAF [Organogenesis Inc, Canton, Mass], designed for the treatment of venous leg ulcers and diabetic foot ulcers; and OrCel [Ortec International Inc, New York], designed for the management of split-thickness donor site wounds and for the treatment of epidermolysis bullosa), (2) bilayered composites consisting of a synthetic epidermal analog and a biologic (collagen-based) dermal analog (TransCyte [Smith & Nephew Inc, Hull, United Kingdom] and Biobrane [Bertek Pharmaceuticals Inc, Morgantown, Wva], both designed for partial-thickness wounds), (3) complex weaves of biopolymers that produce a thin protective membrane (Silon-TSR [Bio Med Sciences Inc, Allentown, Pa] , designed for use on partial-thickness burns, donor sites, and laser-resurfaced skin), and (4) acellular dermal matrices designed to aid in the natural healing of partial-thickness injuries of limited depth (SkinTemp, BioCore Medical Technologies Inc, Silver Spring, Md). Permanent skin substitutes that are designed for treating deep injuries and require application of a thin epithelial autograft will likely be inappropriate for use in treating partial-thickness HD injuries (eg, AlloDerm [LifeCell Corporation, Blanchburg, NJ] and INTEGRA dermal regeneration template [Integra LifeSciences Holdings Corporation, Plainsboro, NJ]).

Cryopreserved and glycerol-preserved cadaver skin has been used as a temporary dressing in the treatment of burns for a number of years. 270277 Similarly, xenografts from a variety of animal species, especially pig, have been used as temporary cover to treat burns . 278280 While pig skin is antigenic and would ultimately get sloughed, portions of the dermis may become incorporated in a healed wound and elicit an unwanted granular response. 278

Cultured epithelial allografts and autografts have been used for about 2 decades as a treatment for chronic ulcers and thermal burns. 248 , 281312 Keratinocytes can be harvested from skin biopsies and grown to confluence by the method originally described by Rheinwald and Green. 313 Large amounts of stratifying epidermis can thus be grown in the laboratory in short periods of time and used to restore defects in the epidermis. 314 Such grafts can be used immediately or cryopreserved and used at a later date. In addition to their usefulness in improving the healing of deep ulcers and burns, they have shown efficacy in improving the rate of reepithelialization of partial-thickness burns 286 , 290 , 300 and split-thickness skin graft donor sites. 289 , 291 , 307 There is no evidence that cultured allografts survive permanently on the wound bed. 289 Kaawach et al 286 showed that allografted cells were not present between 8 and 100 days postgrafting and suggested that the newly formed epithelium was of host origin. Cultured keratinocyte allografts speed healing by providing cover and producing growth factors and extracellular matrix proteins. 291 Because these coverings can be produced in large quantities and would thus be more readily available than cadaver skin, their application in the treatment of debrided partial-thickness HD injuries should be considered. Cultured epidermal autografts (CEAs) would be safer to use, from the perspective of disease transmission, and would not require donor-screening procedures. They do, however, require small punch biopsies to be collected from the patient and a lag time of about 2 weeks to grow the graft material. Several laboratories in the United States perform this service for their local burn centers (eg, Living Skin Bank, University Hospital, SUNY, Stony Brook, NY). Genzyme Corporation (Cambridge, Mass) has shown that CEA (Epicel) can be commercially produced. Despite their theoretical usefulness, CEAs are rather limited in their clinical effectiveness because they are unable to withstand even very low levels of bacterial wound contamination and do not provide a durable epithelial surface. Wounds covered by this modality are unstable and are subject to frequent epithelial disruption as a result of minor mechanical trauma. Durability has been increased by placing the CEA on a scaffolding of widely meshed autograft. 315 Alternatively, CEA placed over deepithelialized allograft (ie, engrafted allodermis) has also proved successful. 284

Finally, application of keratinocytes in suspension has shown to improve epidermal wound healing in pig 316 , 317 and mouse 318 , 319 models. Reconstitution of the dermal-epidermal junction was significantly enhanced in an athymic mouse model by suspending the cells in a fibrin-glue matrix. 319 Using a pig model, Currie et al 320 recently compared the effects of keratinocyte cell sprays with and without fibrin glue. No differences in mean epithelial area or quality of epithelium were noted at 3 weeks. Keratinocyte suspension technology shows promise in that it does not require the length of time necessary to produce cultured epidermal sheets. Use of this technology has proven efficacious in the treatment of thermal burns in humans. 321 A small biopsy is collected and the cells cultured and expanded in a clinical laboratory, then placed into a syringe-like spraying mechanism and sprayed onto the wound 2 to 5 days following biopsy. This technology is currently available (CellSpray and CellSpray XP, Clinical Cell Culture, Bentley, Western Australia). These products are designed for use in partial, deep partial, and full-thickness burns, donor sites, scar treatment, chronic ulcers, pigment loss, and cosmetic skin rejuvenation following laser resurfacing, dermabrasion, or chemical peels. A similar spray-on product in development that delivers allogeneic keratinocytes, fibroblasts, and fibrin to wounds has recently shown positive results in Phase II trials in Europe and the United States (Allox, IsoTis OrthoBiologics, Irvine, Calif). An innovative medical device (ReCell, Clinical Cell Culture, Bentley, Western Australia) has been developed that will allow rapid harvesting of cells from a thin split-thickness biopsy followed by spray application onto wounds within 30 minutes of collecting the biopsy, without the need of culturing the keratinocytes in a clinical laboratory. This single-use device is designed for injuries up to 2% TBSA. It may prove beneficial in the treatment of small TBSA HD injuries and is worthy of laboratory investigation.

Topical nutritional support

Boyce et al 322 noted that application of topical nutrients supports keratinocyte viability during graft vascularization of cultured skin substitutes and inhibits wound contraction. There are a large number of “cosmeceutical” products on the market designed to enhance the appearance, feel, flexibility, and function of skin by supplying moisturizing and nutritive substances. Amino-Plex Spray (biO 2 Cosmeceuticals International Inc, Beverly Hills, Calif) is such a product that is designed to increase oxygen in cells, stimulate ATP synthesis, improve glucose transportation, stimulate collagen formation, and promote angiogenesis. It is a mixture of over 100 low-molecular-weight ingredients, including amino acids, trace minerals, nucleotides, nucleosides, oligopeptides, electrolytes, glycosaminoglycans, and glycolipids. According to the company, this product has been shown to reduce irritation and improve results in laser resurfacing, chemical peels, microdermabrasion, hair transplantation, and hair removal. A new product (Oxy-Mist, biO 2 Cosmeceuticals International Inc, Beverly Hills, Calif) combines the ingredients of Amino-Plex Spray with micellized vitamin E and sterile mineralized water and uses medical-grade oxygen as the delivery source. The manufacturer reports that used after facial resurfacing with a pulsed CO 2 or erbium laser, Oxy-Mist has been clinically shown to accelerate reepithelialization, minimize pain, and decrease the period of postlaser erythema. The oxygen itself likely contributed to the improved healing noted, as both hyperbaric 323 and topical 324 oxygen therapies have been shown to facilitate wound healing. Because of its reported benefits in dermatology following laser resurfacing procedures, it would be advantageous to determine whether these topical nutritional products will improve the healing of debrided HD injuries.

Vacuum assisted closure

Application of topical negative pressure in the management of chronic wounds and burns has gained popularity in the last 5 years. 325332 Also known as Vacuum Assisted Closure (VAC), the procedure involves placing an open-cell foam into the wound bed (cut to conform to the shape of the wound), sealing it with an adhesive drape and applying subatmospheric pressure (125 mm Hg below ambient) that is transmitted via an evacuation tube by a computerized vacuum pump. 326 , 327 The procedure is becoming widely used for the closure of chronic wounds such as stage III and IV pressure ulcers; venous, arterial, and neuropathic ulcers; and subacute and acute wounds such as dehisced incisions, split-thickness meshed skin grafts, and muscle flaps. 326 , 327 VAC is also gaining popularity in the management of complex orthopedic wounds. 329 , 330 This methodology increases local blood perfusion and nutrient delivery to the wound, accelerates the rate of granulation tissue formation, and decreases wound tissue bacterial levels. 326 , 327 Per the manufacturer's recommendations, wounds must be debrided of all necrotic tissue prior to application of VAC, and it is contraindicated with the presence of nonenteric and unexplored fistulas, osteomyelitis (untreated), exposed organs or blood vessels, or malignancy in or around the wound. The dressings are typically changed every 1 to 4 days until wound closure. VAC has been shown to be effective in preventing progression of partial-thickness burns to a deeper injury in a swine model, 333 likely the result of helping to deliver oxygen and nutrients to the zone of stasis. The method has also been shown to increase the rate of skin graft donor site reepithelialization in pigs and humans 331 and is a safe and effective method for securing split-thickness skin grafts, providing improved graft survival. 332 Following debridement of partial-thickness HD injuries , VAC may prove efficacious in significantly speeding the reepithelialization process in these lesions. Recently, the US Food and Drug Administration approved the use of VAC in treating partial-thickness burns. The expedited closure of HD wounds by means of a mechanical force is an area that merits further consideration and investigation. Several VAC therapy systems are available from Kinetic Concepts Inc, San Antonio, Tex. One lightweight portable system is available for ambulatory care.

Use of noninvasive bioengineering methods to assess treatment efficacy

During efficacy testing of candidate treatment regimens, it is important to examine a number of parameters besides reepithelialization. While coverage of the wound by a new epithelium is important, there are a number of other skin characteristics that are important from a functional and cosmetic point of view. Surface contour and general appearance, epidermal hydration, epidermal barrier function, pH, mechanical properties, cutaneous blood flow, transcutaneous oxygen tension, neural supply/sensory function, and hair growth are all important characteristics that bear examination. While routine histopathology, immunohistochemistry, and electron microscopy are all valued tools in determining morphology and understanding the pathophysiology of HD wound development and healing, they do not directly measure physiological parameters or function. For these, a variety of noninvasive bioengineering methods are available. In support of HD wound healing research, laboratories in the United States and United Kingdom have used reflectance colorimetry to evaluate erythema, skin hue, chroma, and lightness 11 , 27 , 83 , 100 , 102 ; LDPI to examine cutaneous blood flow, depth of injury, neovascularization, and skin graft viability 83 , 101 ; torsional ballistometry to evaluate the mechanical properties of skin firmness and elasticity 83 ; evaporimetry to examine transepidermal water loss as a way to evaluate skin hydration/barrier function 83 , 102 ; 2-dimensional and 3-dimensional high-frequency (20-MHz) ultrasound to examine edema formation 11 and scar tissue thickness (JS Graham et al, unpublished data, 2002); and image analysis to evaluate wound size, shape morphometry, and wound contraction. 45 , 100 While not yet used in HD wound healing research efforts, instrumentation is also available for evaluating surface contour, pH, and sensory function.

SUMMARY

The toxicity of sulfur mustard has been widely described. 1 , 316 Cutaneous HD injuries can take several months to heal, may necessitate lengthy hospitalizations, and can result in significant cosmetic and/or functional deficits. There are currently no standardized or optimized methods of casualty management that prevent or minimize deficits and provide for speedy wound healing.

Research laboratories in the United States, United Kingdom, and Canada have developed concepts for medical countermeasures to vesicant agents. The initial step in protecting a person from the deleterious effects of HD is to eliminate contact with the agent. Protective gear and topical skin protectants have been designed for this purpose. Should the agent come in contact with the skin, it needs to be removed within 2 minutes to fully prevent damage. 15 Decontamination is generally performed by physical removal. Sulfur mustard is not painful on contact and the exposed person may not be aware of the exposure until symptoms begin to appear after a latent period. Pharmacological approaches are being studied for their efficacy in minimizing or preventing damage. At this time, it seems clear that the earliest possible application of anti-inflammatory agents, including cold packs, topical and systemic steroids, and nonsteroidal anti-inflammatory drugs, is beneficial. Should HD come in contact with the skin, were decontamination not performed timely, and should pharmacological intervention be absent or ineffective, a chemical casualty will be produced that requires medical attention. Casualty management now comes into play. There are no antidotes to HD. Therapy therefore rests on management of symptoms and consequences of exposure with the intent to reduce long-term morbidity. Historically, blister aspiration and/or deroofing (epidermal removal), physical debridement, irrigation, topical antibiotics, and sterile dressings have been the main courses of action in the medical management of cutaneous HD injuries. New strategies to relieve symptoms, prevent infections, and promote healing have been formulated. Deep cutaneous HD injuries will require aggressive surgical intervention, including skin grafting, if cosmetic and functional deficits are to be avoided. Our future research efforts will concentrate on partial-thickness injury that will not require such aggressive approaches.

Assessment of the injuries must occur early in the process. Total body surface area of the injuries should be established and depth of injury determined. Laser Doppler perfusion imaging and ICG fluorescence imaging show promise in prognosticating optimal wound healing of HD injury on the basis of examination of microcutaneous blood flow. Following assessment of HD injury, adequate wound debridement needs to be performed. At the minimum, debridement needs to proceed into normal-appearing skin along the periphery of the lesion and down through the base of the blister (eg, damaged BMZ) into the papillary dermis. Debridement is then followed by 1 or more treatment adjuncts. Such adjuncts under consideration are dressings, growth factors, skin substitutes, topical nutritional support, and Vacuum Assisted Closure.

The ultimate goal is to determine the most efficacious treatment regimen to be applied in the clinical management of HD casualties. The ideal regimen should return damaged skin to optimal appearance and normal function in the shortest time. Improved treatment will result in a better cosmetic and functional outcome for the patient, and will enable the casualty to return to normal activities sooner.