Ghrelin protects against ischemia/reperfusion-induced hepatic injury via inhibiting Caspase-11-mediated noncanonical pyroptosis

Abstract

Background: Ischemia/reperfusion (I/R) injury is a complication of liver transplantation. I/R-induced inflammatory cell death, namely, pyroptosis, that is triggered by overactive inflammasomes results in the production of proinflammatory cytokines. Hepatic I/R injury correlates with the activation of the Caspase-11-mediated pyroptosis pathway. We investigated whether ghrelin, which is a pleiotropic gut hormone, may have anti-hepatic I/R injury effects, but the mechanism by which Ghrelin ameliorates hepatic I/R -induced injury remains a mystery.

Methods: Hepatic I/R injury was induced in a mouse model by clamping the left and right lobes of the liver for 90 min followed by reperfusion for 6 h, 12 h, or 24 h. As treatment, a saline with or without ghrelin was infused via the tail vain. Hepatocytes were isolated using a two-step collagenase liver perfusion method.

Results: In our study, treatment with ghrelin protected against hepatic I/R injury as shown by decreased alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) levels (p < 0.001) and reduced the histological injury in liver tissues compared with untreated controls. The LDH level of primary hepatocytes was increased by hypoxia/reoxygenation (H/R), and it was then restored to normal levels by ghrelin-treatment (p < 0.05). Western blotting analysis showed that ghrelin significantly inhibited the expression of pyroptosis-related proteins, including Caspase-11, GSDMD-N, NLRP3 and HMGB1, both in vivo and in vitro (all p < 0.05) compared with the untreated controls. Immunofluorescence showed that the expression of Gasdamin D (GSDMD) in hepatocytes was increased after I/R or H/R, whereas GSDMD expression was reduced by ghrelin treatment (p < 0.05).

Conclusions: Our findings suggest that ghrelin ameliorated I/R-induced hepatic injury by inhibiting Caspase-11-mediated pyroptosis. Ghrelin may be a potential therapeutic option to prevent hepatic I/R injury after liver transplantation.

Keywords: Caspase-11; Ghrelin; Hepatic ischemia-reperfusion injury; Inflammation; Pyroptosis.