Digoxin improves steatohepatitis with differential involvement of liver cell subsets in mice through inhibition of PKM2 transactivation

Abstract

The cardiac glycoside digoxin was identified as a potent suppressor of pyruvate kinase isoform 2-hypoxia-inducible factor-α (PKM2-HIF-1α) pathway activation in liver injury mouse models via intraperitoneal injection. We have assessed the therapeutic effects of digoxin to reduce nonalcoholic steatohepatitis (NASH) by the clinically relevant oral route in mice and analyzed the cellular basis for this effect with differential involvement of liver cell subsets. C57BL/6J male mice were placed on a high-fat diet (HFD) for 10 wk and started concurrently with the gavage of digoxin (2.5, 0.5, 0.125 mg/kg twice a week) for 5 wk. Digoxin significantly reduced HFD-induced hepatic damage, steatosis, and liver inflammation across a wide dosage range. The lowest dose of digoxin (0.125 mg/kg) showed significant protective effects against liver injury and sterile inflammation. Consistently, digoxin attenuated HIF-1α sustained NLRP3 inflammasome activation in macrophages. We have reported for the first time that PKM2 is upregulated in hepatocytes with hepatic steatosis, and digoxin directly improved hepatocyte mitochondrial dysfunction and steatosis. Mechanistically, digoxin directly bound to PKM2 and inhibited PKM2 targeting HIF-1α transactivation without affecting PKM2 enzyme activation. Thus, oral digoxin showed potential to therapeutically inhibit liver injury in NASH through the regulation of PKM2-HIF-1α pathway activation with involvement of multiple cell types. Because of the large clinical experience with oral digoxin, this may have significant clinical applicability in human NASH. tocytes via the regulation of PKM2-HIF-1α axis pathway activation. Oral digoxin may have significant clinical applicability in human NASH.