Silencing of Fas, but not caspase-8, in lung epithelial cells ameliorates pulmonary apoptosis, inflammation, and neutrophil influx after hemorrhagic shock and sepsis.
Apoptosis and inflammation play an important role in the pathogenesis of direct/pulmonary acute lung injury (ALI). However, the role of the Fas receptor-driven apoptotic pathway in indirect/nonpulmonary ALI is virtually unstudied. We hypothesized that if Fas or caspase-8 plays a role in the induction of indirect ALI, their local silencing using small interfering RNA (siRNA) should be protective in hemorrhage-induced septic ALI. Initially, as a proof of principle, green fluorescent protein-siRNA was administered intratracheally into transgenic mice overexpressing green fluorescent protein. Twenty-four hours after siRNA delivery, lung sections revealed a significant decrease in green fluorescence. Intratracheally administered Cy-5-labeled Fas-siRNA localized primarily in pulmonary epithelial cells. Intratracheal instillation of siRNA did not induce lung inflammation via toll-like receptor or protein kinase PKR pathways as assessed by lung tissue interferon-alpha, tumor necrosis factor-alpha, and interleukin (IL)-6 levels. Mice subjected to hemorrhagic shock and sepsis received either Fas-, caspase-8-, or control-siRNA intratracheally 4 hours after hemorrhage. Fas- or caspase-8-siRNA significantly reduced lung tissue Fas or caspase-8 mRNA, respectively. Only Fas-siRNA markedly diminished lung tissue tumor necrosis factor-alpha, IL-6, IL-10, interferon-gamma, IL-12, and caspase-3 activity. Fas-siRNA also preserved alveolar architecture and reduced lung neutrophil infiltration and pulmonary epithelial apoptosis. These data indicate the pathophysiological significance of Fas activation in nonpulmonary/shock-induced ALI and the feasibility of intrapulmonary administration of anti-apoptotic siRNA in vivo.
Perl
Mario
M
Shock-Trauma Research Laboratories, Division of Surgical Research, Providence, RI 02903, USA.
Chung
Chun-Shiang
CS
Lomas-Neira
Joanne
J
Rachel
Tina-Marie
TM
Biffl
Walter L
WL
Cioffi
William G
WG
Ayala
Alfred
A
D000818:Animals; D017209:Apoptosis; D001483:Base Sequence; D053181:Caspase 8; D020169:Caspases / Q000235:genetics*; D017931:DNA Primers; D004195:Disease Models, Animal; D020868:Gene Silencing; D007249:Inflammation; D008171:Lung Diseases / Q000235:genetics* / Q000473:pathology; D055370:Lung Injury; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D009504:Neutrophils / Q000502:physiology*; D034741:RNA, Small Interfering / Q000235:genetics; D018805:Sepsis / Q000235:genetics* / Q000473:pathology / Q000628:therapy; D012771:Shock, Hemorrhagic / Q000235:genetics* / Q000473:pathology / Q000628:therapy; D019014:fas Receptor / Q000235:genetics*
D017931:DNA Primers; D034741:RNA, Small Interfering; D019014:fas Receptor; C505526:Casp8 protein, mouse; D053181:Caspase 8; D020169:Caspases