4-Methylumbelliferone inhibits liver fibrogenesis via HA-independent, FXR-independent, gut microbiota-dependent mechanisms

4-Methylumbelliferone (4-MU) is the active component of hymecromone, a choleretic and antispasmodic treatment approved in European and Asian countries with a great safety profile. It is also widely used to inhibit hyaluronan (HA) synthesis. Given a positive correlation between circulating HA levels and the severity of liver fibrosis, 4-MU has been tested in several animal models of liver fibrosis. Regardless of the etiology of liver fibrosis, 4-MU showed great efficacy in reducing its severity, igniting interest in repurposing the compound for treating liver fibrosis. In our report, we primarily used a choline-deficient, amino acid-defined, high-fat diet (CDAHFD) treatment regimen to establish liver fibrosis in mice, which captures many characteristics of liver fibrosis in metabolic dysfunction-associated steatohepatitis (MASH). We showed 4-MU effectively inhibits liver fibrosis in mice treated with CDAHFD. To test whether 4-MU works through suppression of HA production, several transgenic mouse models with either enhanced HA production or over-digestion were exposed to CDAHFD, and all showed reduced liver fibrogenesis, highlighting a complicated role of HA in liver fibrogenesis. Even though 4-MU is a choleretic agent in humans, it did not modulate liver, gut, or stool bile acid (BA) content, nor affect intestinal FXR, to suppress liver fibrogenesis in mice. In contrast, 4-MU reshapes gut microbiota, and gut microbiota ablation significantly blunts 4-MU's anti-fibrotic properties. In summary, despite an increase in HA during the development of liver fibrosis, HA's role in fibrogenesis is complex, and 4-MU ameliorates liver fibrosis via FXR-independent, gut microbiota-dependent mechanisms.