Does necroptosis have a crucial role in hepatic ischemia-reperfusion injury?

Background: Previous studies have demonstrated protective effects of anti-receptor interacting protein kinase 1 (RIP1), a key necroptosis molecule. However, it is uncertain whether necroptosis has a crucial role in hepatic IR injury. Therefore, we evaluated the role of necroptosis in hepatic IR injury.

Method: The IR mice underwent 70% segmental IR injury induced by the clamping of the hepatic artery and portal vein for 1 hr followed by reperfusion for 4 hr. The key necroptosis molecules (RIP1, RIP3, and MLKL) and other key molecules of regulated necrosis (PGAM5 and caspase-1) were evaluated in the warm IR injury model. A RIP1 inhibitor (necrostain-1s) and/or an anti-mitochondrial permeability transition (MPT)-mediated necrosis mediator (cyclosporine A, CyA) were administered before clamping. Necrotic injury was quantified using Suzuki's scoring system. qRT-PCR and western blot were performed to evaluate RIP1, RIP3, MLKL and PGAM5 expressions.

Results: RIP1, RIP3, MLKL and PGAM5 expression did not change in the hepatic IR injury model. Moreover, Nec1s pretreatment did not improve histology or biochemical markers. The overall Suzuki score (cytoplasmic vacuolization, sinusoidal congestion and hepatocytes necrosis) was increased in the RIP3(-/-) mice compared to the IR group (3.5 vs. 5, p = 0.026). CyA pretreatment and/or RIP3(-/-) mice decreased Bax/Bcl2 expression; however, it did lead to an overall change in the levels of AST, ALT and LDH or necrotic injury. The Bax/Bcl2 ratio and the expression of caspase-1 and caspase-3 did not increase in our hepatic IR injury model.

Conclusion: Key necroptosis molecules did not increase in the necrosis-dominant hepatic IR injury model. Anti-necroptosis and/or cyclosporine-A treatment did not have an overall protective effect on necrosis-dominant hepatic IR injury.