Accumulating evidence has implicated that the activation of signal transducer and activator of transcription 3 (STAT3) contributes to the progression of liver cancer by affecting the expression of proliferation‑associated genes. A previous study reported that elevated levels of 17β‑hydroxysteroid dehydrogenase 4 (HSD17B4) are observed in patients with liver cancer. The current study investigated how upregulated HSD17B4 expression promoted the expression of proliferation‑associated genes in rats with liver cancer. HSD17B4 expression in rats with liver cancer was significantly increased compared with the control group as determined by reverse transcription‑quantitative polymerase chain reaction and western blot assays. Immunohistochemical results revealed that STAT3 activation was positively correlated with increased HSD17B4 expression in tumor tissues from patients with liver cancer. Western blot results further suggested that HSD17B4 overexpression increased STAT3 activation via the protein kinase B and the mitogen‑activated protein kinase/extracellular‑signal‑regulated kinase signaling pathways in HepG2 cells. The present study suggested that overexpression may promote HepG2 proliferation by enhancing expression of various downstream targets of STAT3. Targeted inhibition of HSD17B4 may describe a novel approach in the prevention and treatment of liver cancer.