DNA methylation status of the SPHK1 and LTB genes underlies the clinicopathological diversity of non-alcoholic steatohepatitis-related hepatocellular carcinomas

Abstract

AbstractPurposeThis study was performed to identify the DNA methylation profiles underlying the clinicopathological diversity of non-alcoholic steatohepatitis (NASH)-related hepatocellular carcinomas (HCCs). MethodsGenome-wide DNA methylation analysis of 88 liver tissue samples was performed using the Infinium assay.ResultsPrincipal component analysis revealed that distinct DNA methylation profiles differing from such profiles in normal control liver tissue had already been established in non-cancerous liver tissue showing NASH, which is considered to be a precancerous condition. Hierarchical clustering separated 26 NASH-related HCCs into Cluster I (n = 8) and Cluster II (n = 18). Such epigenetic clustering was significantly correlated with histopathological diversity, i.e. poorer tumor differentiation, tumor steatosis and development of a scirrhous HCC component. Significant differences in DNA methylation levels between the two clusters were accumulated in molecular pathways participating in cell adhesion and cytoskeletal remodeling, as well as cell proliferation and apoptosis. Among tumor-related genes characterizing Clusters I and II, differences in the levels of DNA methylation and mRNA expression for theSPHK1,INHBA,LTBandPDE3Bgenes were correlated with poorer tumor differentiation. 5-Aza-2′-deoxycytidine treatment of HCC cells revealed epigenetic regulation of theSPHK1andLTBgenes. Knockdown experiments showed that SPHK1 promotes cell proliferation, represses apoptosis and enhances migration, whereas LTB enhances migration of HCC cells. DNA hypomethylation resulting in increased expression ofSPHK1andLTBin poorly differentiated HCCs may underlie the aggressive phenotype of such HCCs.ConclusionThese data indicate that DNA methylation profiles may determine the clinicopathological heterogeneity of NASH-related HCCs via alterations of tumor-related gene expression.