The caspase inhibitor IDN-6556 prevents caspase activation and apoptosis in sinusoidal endothelial cells during liver preservation injury

Abstract

Cold ischemia (CI)-warm reperfusion (WR) liver injury remains a problem in liver transplantation. CI-WR initially causes sinusoidal endothelial cell (SEC) apoptosis through a caspase-dependent mechanism. We previously showed that the caspase inhibitor IDN-1965 prevents CI-WR-induced SEC apoptosis. However, this agent required to be administered to the donor, preservation solution, and recipient for efficacy. Here, we show that a second-generation caspase inhibitor, IDN-6556, effectively prevents CI-WR-induced SEC injury when added only to University of Wisconsin (UW) cold storage media. Rat livers were stored in UW solution for 24 hours at 4°C and reperfused for 1 hour at 37°C. Apoptosis was quantitated using terminal deoxynucleotide transferasemediated deoxyuridine triphosphate nick end labeling (TUNEL) assay and caspase 3 activation determined by biochemical measurement and immunohistochemical analysis. P rimary graft nonfunction caused by preservation injury remains a clinical problem in liver transplantation. 1,2 Moreover, this problem may potentially increase as non-heart-beating donors, marginal donors, and steatotic livers are used more widely in practice. Although preservation injury is multifactorial, ischemia-reperfusion injury of the liver is an important contributing factor. 3 Sinusoidal endothelial cells (SECs) are a cellular target of ischemia-reperfusion injury. 4-6 SEC injury causes microcirculatory disturbances, including leukocyte migration, platelet activation, and ultimately, secondary injury to hepatocytes leading to organ nonfunction and dysfunction. 1,6,7 We and others have shown that SEC injury in this process is caused by cell death by apoptosis. [8][9][10] SECs show the classic morphological feature of apoptosis after ischemia-reperfusion injury. 8,9 Biochemical hallmarks of apoptosis, including DNA fragmentation and caspase 3 activation, also are evident. Finally, protease inhibitors known to block apoptosis also protect against liver ischemia-reperfusion injury. 8,11 Thus, ischemia-reperfusion injurymediated apoptosis of SECs would appear to be a key feature of liver preservation injury.The best-characterized proteases contributing to cell injury by apoptosis are the caspases. 12,13 These proteases are synthesized as proenzymes and activated by proteolytic processing by other caspases. 12 Of caspases cloned and identified to date, caspases 2,3,6,7,8,9, and 10 have been implicated most strongly in apoptosis. 12 All these caspases have a cysteine residue in their active site and recognize aspartate in the p1 position of the substrate. 12 This unique substrate specificity has permitted the development of selective caspase inhibitors. Inhibition of caspases by these protease inhibitors often abrogates apoptosis. We recently showed that IDN-1965, a pan-caspase inhibitor, reduced liver injury in a rodent model of ischemia-reperfusion injury relevant to transplantation. 8 However, for optimal efficacy, the drug had to be included in the University of Wisconsin (UW) storage solution and admin...

Experimental study on the effects of massive bowel resection on liver function and hepatocyte apoptosis

“…In contrast to hepatocyte apoptosis, this response was significantly higher on POD7. This interesting finding resembled the effects of ischemia-reperfusion on the liver in previous reports (10)(11)(12). Thus, we postulate that a decrease in portal pressure after small intestinal resection creates a low-flow state that induces hypoxia in the liver, resulting in increased liver sinusoidal cell apoptosis.…”

Cold ischemic injury of transplanted kidneys: new insights from experimental studies

“…Currently, suppressing apoptosis in the acute injury setting is considered beneficial. Limited early reports from a liver reperfusion model suggest that antiapoptotic treatments might reduce dysfunction of coldstored organs (46). The events during cold storage that lead to the apoptotic form of cell death have not been studied in any detail.…”

Oral IDN‐6556, an antiapoptotic caspase inhibitor, may lower aminotransferase activity in patients with chronic hepatitis C†

“…3 In a rat model of ischemia/reperfusion injury during liver transplantation, IDN-6556 lowered aminotransferase activity and improved liver function. 4 In human hepatic disease the presence of increased levels of apoptotic cells in the liver has been demonstrated in hepatitis C, 5,6 hepatitis B, 7 nonalcoholic steatohepatitis (NASH), 8 acute alcoholic hepatitis, [9][10][11][12] primary biliary cirrhosis (PBC), autoimmune hepatitis, 13 and after liver transplantation. 14,15 Furthermore, in hepatitis C the number of apoptotic cells present in liver biopsies have been shown to correlate with the grade of inflammation.…”