Aspirin reprograms platelet signaling and the intrahepatic microbiome to suppress RyR2-driven inflammation and fibrosis in preclinical chronic liver disease.

Abstract

Platelet deactivation by aspirin possibly helps in regression of liver fibrosis, though the mechanisms are unclear. We administered aspirin in a murine model of liver fibrosis and studied molecular signatures associated with fibrosis regression; both in vivo (murine model/ patients) and in vitro. Increase in intrahepatic PF4, p-selectin, PDGFR-&#x3b2; levels (platelet activation) correlated with increase in liver fibrosis (p&#x202f;<&#x202f;0.05, r>0.3). Aspirin reduced the number and activation of intrahepatic platelets, inflammation and fibrosis (p&#x202f;<&#x202f;0.05). Platelet deactivation using aspirin in murine model increased autophagy, glutathione, energy metabolism and decreased arachidonic acid and butanoate metabolism (p&#x202f;<&#x202f;0.05). Aspirin modulated liver microenvironment and showed decrease in intrahepatic immune cell activation (blood transcription module) which correlated with histidine and tryptophan metabolism (r2&#x202f;>&#x202f;0.5, p&#x202f;<&#x202f;0.05). The intrahepatic microbiome post-aspirin showed increased abundance of Firmicutes (Ruminococcaceae, Lachnospiraceae, and Clostridiaceae) and improved functionality (p&#x202f;<&#x202f;0.05). Aspirin caused decreased expression of Ryanodine-receptor-2 (RyR2), Arginase-1 and Kynurenine-3-monooxygenase, which correlate with reduction in &#x3b1;-SMA and degree of hepatic fibrosis (r2&#x202f;>&#x202f;0.75; p&#x202f;<&#x202f;0.05). In addition, pan specific blocking of RyR2 by carvedilol/flecainide markedly inhibited HSC activation and proliferation in-vitro by reducing Caoverload, ER/mitochondrial stress (p&#x202f;<&#x202f;0.05). Further, RyR2 blockade in HSCs reduced its activation by activated platelet secretome or TGF&#x3b2;1 (p&#x202f;<&#x202f;0.05). CONCLUSIONS: Platelet deactivation using aspirin regresses hepatic fibrosis by decreasing intrahepatic platelet accumulation/activation, inflammation and modulation of intrahepatic microbiome. Induction of RyR2 is critical for fibrosis development and pharmacological inhibition of RyR2 could ameliorate liver fibrosis.