MicroRNA 181b-3p and Its Target Importin α5 Regulate Toll-Like Receptor 4 Signaling in Kupffer Cells and Liver Injury in Mice in Response to Ethanol

Increased inflammatory signaling by Kupffer cells contributes to alcoholic liver disease (ALD). Here we investigated the impact of small, specific-sized hyaluronic acid of 35 kD (HA35) on ethanol-induced sensitization of Kupffer cells, as well as ethanol-induced liver injury in mice. Unbiased analysis of microRNA (miRNA) expression in Kupffer cells identified miRNAs regulated by both ethanol and HA35. Toll-like receptor 4 (TLR4)-mediated signaling was assessed in primary cultures of Kupffer cells from ethanol-and pair-fed rats after treatment with HA35. Female C57BL6/J mice were fed ethanol or pair-fed control diets and treated or not with HA35. TLR4 signaling was increased in Kupffer cells by ethanol; this sensitization was normalized by ex vivo treatment with HA35. Next generation sequencing of Kupffer cell miRNA identified miRNA 181b-3p (miR181b-3p) as sensitive to both ethanol and HA35. Importin alpha 5, a protein involved in p65 translocation to the nucleus, was identified as a target of miR181b-3p; importin alpha 5 protein was increased in Kupffer cells from ethanol-fed rats, but decreased by HA35 treatment. Overexpression of miR181b-3p decreased importin alpha 5 expression and normalized lipopolysaccharide-stimulated tumor necrosis factor a expression in Kupffer cells from ethanol-fed rats. In a mouse model of ALD, ethanol feeding decreased miR181b-3p in liver and increased expression of importin alpha 5 in non-parenchymal cells. Treatment with HA35 normalized these changes and also protected mice from ethanol-induced liver and intestinal injury. Conclusion: miR181b-3p is dynamically regulated in Kupffer cells and mouse liver in response to ethanol and treatment with HA35. miR181b-3p modulates expression of importin alpha 5 and sensitivity of TLR4-mediated signaling. This study identifies a miR181b-3p-importin alpha 5 axis in regulating inflammatory signaling pathways in hepatic macrophages.