Hyaluronan (HA) as a glycosaminoglycan canbind to cell-surface receptors, such as TLR4, to regulateinfl ammation, tissue injury, repair, and fi brosis. 4-methylumbelliferone(4-MU), an inhibitor of HA synthesis, is adrug used for the treatment of biliary spasms. Currently,therapeutic interventions are not available for non-alcoholicsteatohepatitis (NASH). In this study, we investigated theeff ects of 4-MU on NASH using a choline-defi cient aminoacid (CDAA) diet model. CDAA diet-fed mice showedNASH characteristics, including hepatocyte injury, hepaticsteatosis, infl ammation, and fi brogenesis. 4-MU treatmentsignifi cantly reduced hepatic lipid contents in CDAA dietfedmice. 4-MU reversed CDAA diet-mediated inhibitionof Ppara and induction of Srebf1 and Slc27a2 . Analysis ofserum ALT and AST levels revealed that 4-MU treatmentprotected against hepatocellular damage induced by CDAAdiet feeding. TLR4 regulates low molecular weight-HAinducedchemokine expression in hepatocytes. In CDAAdiet-fed, 4-MU-treated mice, the upregulated chemokine/cytokine expression, such as Cxcl1 , Cxcl2 , and Tnf wasattenuated with the decrease of macrophage infi ltration intothe liver. Moreover, HA inhibition repressed CDAA dietinducedmRNA expression of fi brogenic genes, Notch1 , and Hes1 in the liver. In conclusion, 4-MU treatment inhibitedliver steatosis and steatohepatitis in a mouse model ofNASH, implicating that 4-MU may have therapeutic potentialfor NASH.