Altered DNA Methylation Sites in Peripheral Blood Leukocytes from Patients with Simple Steatosis and Nonalcoholic Steatohepatitis (NASH)

BACKGROUND The aim of this study was to identify DNA methylation sites in peripheral blood leukocytes from patients with histologically confirmed nonalcoholic fatty liver disease (NAFLD) that included simple hepatic steatosis and nonalcoholic steatohepatitis (NASH). MATERIAL AND METHODS DNA was isolated from peripheral blood leukocytes from patients with histologically diagnosed NAFLD (n=35), including simple hepatic steatosis (n=18) and NASH (n=17). Healthy controls included individuals without liver disease (n=30). DNA was hybridized, and DNA methylation was interrogated in an epigenome-wide association study (EWAS). DNA methylation levels (β-values) were correlated with serum lipid profiles, liver enzymes, and liver histology. RESULTS Circulating blood leukocytes from 35 patients with NAFLD (simple steatosis and NASH) contained 65 CpG sites, which represented 60 genes that were differentially methylated when compared with healthy controls. In the simple hepatic steatosis group (n=18), 42 methylated CpG sites were found to be associated with increased levels of serum alanine aminotransferase (ALT), and 32 methylated CpG sites were associated with increased serum lipid profiles. In the NASH group (n=17), when compared with the simple hepatic steatosis group, methylated CpG sites showed significant correlations with the presence of lobular inflammation compared with hepatic steatosis and fibrosis. Six differentially methylated CpG sites were identified in the ACSL4, CRLS1, CTP1A, SIGIRR, SSBP1 and ZNF622 genes, which were associated with histologically confirmed simple hepatic steatosis and NASH. CONCLUSIONS The study identified some key methylated CpG sites from peripheral blood leukocytes, which might be used as serum biomarkers to stratify NAFLD patients into simple hepatic steatosis and NASH.