The Role of Sphingosine Kinase 2 in Chronic Alcohol‐Induced Liver Injury and Disease

Abstract

IntroductionAlcoholic liver disease (ALD) is one of the most common liver diseases worldwide. Currently, there are no effective therapeutics against ALD and liver transplantation remains the only curative option for end stage liver disease. Hepatic steatosis is a characteristic hallmark of alcohol‐induced hepatocyte injury and we have previously reported that sphingosine kinase 2 (SphK2) deficient mice on an alcohol diet develop overt fatty liver compared to wild type in the chronic and binge drinking NIAAA mouse model. In addition, there was significant downregulation of hepatic lipid genes and upregulation of pro‐inflammatory mediators in SphK2 deficient mouse livers. However, there are currently no effective mouse models that can recapitulate ALD progression and our aim is to determine whether SphK2 deficient mice on a chronic alcohol diet can produce advanced ALD pathology.MethodsMouse primary hepatocytes derived from wild type or SphK2 knockout (KO) mice were treated with different concentrations of ethanol (0–200 mM) for various time points (0–24h). For in vivo studies, C57BL/6J wild type and SphK2 KO mice (10–14‐week‐old, male and female) were fed a Lieber‐DeCarli 5% ethanol diet for 60 days. Cultured hepatocytes and liver tissue were harvested for RNA and protein using standard laboratory protocols. Quantitative RT‐PCR and western blot analysis were used to determine relative mRNA levels and protein levels, respectively. Hematoxylin and eosin (H&E) staining was performed on liver tissue for pathological analysis using standard laboratory protocols. Immunofluorescence was performed by staining frozen liver sections with anti‐CD11b antibody conjugated with FITC.ResultsEthanol increased the mRNA and protein expression levels of SphK2 in mouse primary hepatocytes (MPH). H&E staining revealed that SphK2 KO mice on an 8‐week alcohol diet produced extensive steatosis and steatohepatitis compared to wild type mice and the NIAAA binge model. In addition, liver enzyme assay levels of ALT, AST and ALP are significantly increased in SphK2 KO mice suggesting increased liver injury and inflammation. The mRNA expression levels of hepatic lipid genes and bile acid synthetic gene mediators (SIRT1, CYP7A1, CYP7B1 and FXR) are downregulated and pro‐inflammatory mediators (IL‐1β, F4/80 and TNFα) are upregulated in SphK2 KO mice compared to wild type. Immunofluorescent staining of CD11b reveals increased macrophage infiltration in SphK2 KO mouse livers.ConclusionSphK2 is involved in hepatic lipid metabolism and plays an important role in ameliorating alcohol‐induced hepatic injury. Our results suggest that SphK2 KO mice on a chronic alcohol diet exacerbate alcohol‐induced liver injury and promotes ALD progression.Support or Funding InformationNIDDK R01NIAAA F30This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.