Bedeutung der Kaspasenaktivität für das hepatische Mikrozirkulationsversagen nach Ischämie und Reperfusion

Abstract

Background: Sinusoidal perfusion failure (no-reflow), leukocyte accumulation in the hepatic microvasculature (reflow-paradox), and Kupffer cell dysfunction are critical determinants of liver ischemia/reperfusion (I/R) injury. Most recent studies indicate that caspase-mediated apoptosis of hepatic parenchymal and non-parenchymal cells contributes to post-ischemic liver dysfunction. Methods: To study the role of caspase activity for hepatic microvascular injury after I/R, C3H/HeN mice were laparotomized under rompun/ketanest anesthesia (90/25 mg/kg b.w.) and subjected to 60 min left lobar liver ischemia followed by 90 min of reperfusion in the presence (I/R + z-VAD, n = 6) or absence (I/R, n = 6) of the caspase inhibitor z-VAD-fmk (z-VAD). Sham-operated animals (n = 6) served as a control. Hepatic microcirculation was repetitively analyzed at 10, 45, and 90 min of reperfusion using intravital fluorescence microscopy, including quantitative analysis of sinusoidal perfusion (sodium fluorescence), leukocyte adherence in post-sinusoidal venules (rhodamine-6G), and phagocytic activity of Kupffer cells (fluorescence-labeled latex beads). Sinusoidal endothelial cell injury was analyzed using electron microscopy. Data are mean ± SEM; ANOVA and Student-Newman-Keuls-test. Results: Hepatic microcirculation after I/R was characterized by severe sinusoidal perfusion failure during early reperfusion (37.2 ± 3.1% and 12.6 ± 2.8% of non-perfused sinusoids at 10 and 90 min of reperfusion vs. sham: < 5%; p < 0.01). Adherence of leukocytes to the venular wall was significantly (p <0.05) increased (e.g. 10,798 ± 3,415 cells/mm2 (10 min) endothelial surface vs. sham: < 20 cells/mm2) during reperfusion while phagocytic activity was significantly (p <0.05) suppressed. Repetitive administration of z-VAD prevented (p < 0.01) sinusoidal perfusion failure (< 5% of non-perfused sinusoids) and leukocyte accumulation in post-sinusoidal venules < 20 cells/mm2), while Kupffer cell activity was not altered by z-VAD. Transmission electron micrographs of post-ischemic livers revealed early signs of apoptosis in sinusoidal endothelial cells which were prevented by z-VAD. Conclusion: These data suggest that protection of hepatic microcirculation by caspase inhibition may represent a therapeutic approach to counteract I/R-mediated liver injury.