cultured alveolar epithelial cells from septic rats mimic in vivo septic lung培养从脓毒性大鼠肺泡上皮细胞模拟体内感染性肺.pdf

Cultured Alveolar Epithelial Cells From Septic Rats Mimic In Vivo Septic Lung Taylor S. Cohen, Gladys Gray Lawrence, Susan S. Margulies* Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America Abstract Sepsis results in the formation of pulmonary edema by increasing in epithelial permeability. Therefore we hypothesized that alveolar epithelial cells isolated from septic animals develop tight junctions with different protein composition and reduced barrier function relative to alveolar epithelial cells from healthy animals. Male rats (200–300g) were sacrificed 24 hours after cecal ligation and double puncture (2CLP) or sham surgery. Alveolar epithelial cells were isolated and plated on fibronectin- coated flexible membranes or permeable, non-flexible transwell substrates. After a 5 day culture period, cells were either lysed for western analysis of tight junction protein expressin (claudin 3, 4, 5, 7, 8, and 18, occludin, ZO-1, and JAM-A) and MAPk (JNK, ERK, an p38) signaling activation, or barrier function was examined by measuring transepithelial resistance (TER) ˚ or the flux of two molecular tracers (5 and 20 A). Inhibitors of JNK (SP600125, 20 mM) and ERK (U0126, 10 mM) were used to determine the role of these pathways in sepsis induced epithelial barrier dysfunction. Expression of claudin 4, claudin 18, and occludin was significantly lower, and activation of JNK and ERK signaling pathways was significantly increased in 2CLP monolayers, relative to sham monolayers. Transepithelial resistance of the 2CLP monolayers was reduced significantly compared to sham (769 and 1234 ohm-cm2, respectively), however no significant difference in the flux of either tracer was