Although potential association of hepatitis B virus (HBV) gene products and cellular lipid metabolism have been reported, the molecular interaction between hepatitis B surface antigen (HBsAg) and lipid metabolism remains unclear. This study revealed a regulatory mechanism of HBsAg on lipid metabolism through the analysis of cell transcriptome. The cell line HepG2-S-G2 stably expressing HBsAg and the control cell line HepG2-neo-F4 were selected for transcriptome analysis. The results showed that the genes involved in lipid metabolism were significantly changed, suggesting up-regulation of lipid synthesis pathways and down-regulation of lipid degradation pathways in host cells by HBsAg. The expression difference of two key differential genes, OXCT1 and CYP4F3 were confirmed by quantitative PCR for mRNA and western blot for proteins. To verify the phenotype on lipids, the two cell lines were stained with oil red O and the levels of fatty acid and total cholesterol were detected. The result demonstrated that the lipid droplet in HepG2-S-G2 cells was more obvious while the non-esterified fatty acid and total cholesterol were significantly increased. We also found that the extracellular HBsAg was dramatically reduced when stably transfected cells with HBV were treated with lipid-lowering method. These results revealed that HBsAg could up-regulate lipid metabolism and promote lipid synthesis, suggesting that the decrease of lipid may be an effective way to inhibit HBsAg.