Su1717 Host Response to Translocated Microbial Products, sCD163 and sCD14, Predicts Excessive Alcohol Consumption

Background: Alcohol abuse leads alcoholic liver disease (ALD). ALD is a step-progressing disease that starts with hepatic steatosis (HS), followed by steatohepatitis (SH). MaxiK's have diverse signaling effects and regulate inflammation. The Aim of our study was to evaluate the role of MaxiK channels in ALD. Methods: We fed alcohol (Lieber-deCarli) or control diet to C57Bl6 and MaxiK/beta subunit (MaxiK/β)-knockout (KO) mice. Liver macrophages Kupffer cells (KC) and hepatocytes (Hpt) were isolated by enzymatic digestion and gradient centrifugation. RAW264.7 andHepa 1.6 cell lines, as counterparts of KC andHpt correspondingly, were cultured with Iberiotoxin (Ibtx; MaxiK blocker), NS1619 (MaxiK opener) and l palmitic acid (PA), for induction of steatosis, or lipopolysaccharide (LPS) as surrogate of inflammation. Livers were analyzed by histology, RNA by PCR, protein by western blot, cytokines by ELISA and Multiplex. Results: Alcohol, but not control, diet led to significant increase in serum ALT, suggestive of liver injury, liver Tg and OilRedO liver tissue staining, suggestive of steatosis, and serum cytokines TNFa, IL1, IL6, suggestive of inflammation, in C57Bl6 mice. Liver Tg and OilRedO liver tissue staining were comparable between C57Bl6 and MaxiK/β-KO mice, indicating that MaxiK/β do not participate in development of hepatic steatosis phase (HS) of ALD. In contrast, serum ALT and cytokines were significantly lower in MaxiK/β-KO compared to C57Bl6 mice, suggestive protection from inflammation and liver injury. Both KCs and Hpt express MaxiK channel; there was a significant upregulation of individual components of MaxiK in alcoholcompared to control diet-fed mice however there were no differences between KC and Hpt in either mouse strain, indicating that level of receptor expression and cell-specific distribution are not responsible for the protective role of MaxiK/β in ALD. RAW264.7 cells produced significant amounts of cytokines in response to LPS; chronic alcohol pre-exposure lead to significantly higher levels of LPSinduced cytokines. NS1619 potentiated LPS-induced cytokine production similar to chronic alcohol exposure. Ibtx inhibited LPS-induced cytokine production in both alcohol-naive and in chronic alcohol-exposed RAW264.7 cells. Hepa 1.6 cells exposed to PA showed increased OilRedO staining, suggestive of steatosis; exposure to either Ibtx or NS1619 did not affect the degree of steatosis indicating that modulation of MaxiK is unlikely to affect fat deposition. In conclusion, we report novel finding that MaxiK channel is key to development of alcohol-induced liver tissue injury. We further detail that MaxiK/ β subunit is important in regulation of inflammation but not steatosis in ASH in mice. These results suggest potential therapeutic targets in transition from hepatic steatosis (HS) to steatohepatitis (SH) phase of ALD.