OC 29.5 - Dysregulated fibrinogen γ-chain cross-linking in FibγΔ5 mice drives acute liver injury after acetaminophen overdose

Background: Hepatic fibrin(ogen) deposition and platelet accumulation are hallmarks of liver injury driven by acetaminophen (APAP) overdose. The mechanisms mediating initial platelet accumulation in the injured liver are not understood.

Aims: We tested the hypothesis that hepatic platelet accumulation in the APAP-injured liver is mediated by fibrin(ogen) engagement of the platelet integrin αIIbβ3.

Methods: Male wild-type mice and mice expressing normal levels of a mutant fibrinogen incapable of engaging integrin αIIbβ3 through the c-terminal domain of the fibrinogen γ chain (FibγΔ5 mice) were challenged with APAP (300 mg/kg, i.p.), and liver tissues were collected 24 hours after challenge.

Results: Contrary to our hypothesis, hepatic platelet accumulation was modestly increased in FibγΔ5 mice compared to wild-type mice after APAP challenge. Earlier reports indicate similar fibrin(ogen) cross-linking occurs in wild-type and FibγΔ5 mice. Surprisingly, fibrinogen γ-chain cross-linking, assessed by capillary western blotting, was unique in livers of APAP-challenged FibγΔ5 mice compared to wild-type, favoring accumulation of high molecular weight cross-linked γ-chain complexes (~220 and 350 kDa) and absence of γ-γ dimer. Moreover, we discovered using chain-selective antibodies that γ-γ dimer does not form in vitro in FibγΔ5 plasma or purified fibrinogen with addition of thrombin. Additionally, scanning electron microscopy revealed that in vitro clots formed from purified FibγΔ5 fibrinogen appeared to be composed of a denser network of thinner fibers compared to wild-type. Finally, APAP-induced liver necrosis was increased in FibγΔ5 mice compared to wild-type mice. This affect appeared to be independent of platelet integrin αIIbβ3-fibrinogen interactions, as treatment of wildtype mice with an αIIbβ3 inhibitor had no effect on APAP-induced hepatic necrosis.

Conclusion(s): These results indicate that FibγΔ5 mice have defective fibrinogen γ-chain cross-linking with a complete absence of γ-γ dimer formation and increased high molecular weight γ-chain cross-linking. Furthermore, this aberrant γ-chain cross-linking may exacerbate liver injury following APAP overdose.