Abstract: Objective To study the association between the expression of peritumoral circWDR25 (hsa-circRNA-0004310) secreted by hepatic stellate cells (HSCs) and the prognosis of the patients with hepatocellular carcinoma (HCC) after curative resection. Methods HSCs cell line LX-2 was co-cultured separately with 3 liver cancer cell lines (Hep3B, SMMC-7721, and HCCLM3) in Transwell chambers to obtain tumor cell-activated HSCs. The supernatants of HSC cultures were collected to isolate the exosomes, from which total RNA was extracted to detect circRNA expression profile. We also collected specimens of paracancerous liver tissues from 288 HCC patients undergoing radical resection in our department from January, 2014 to October, 2015, and the expression levels of circWDR25 and α-SMA were detected with in situ hybridization. Log-rank test and Cox regression analysis were used for univariate and multivariate analysis of the factors affecting the patients' prognosis, respectively. Results Gene expression profiling revealed that the expression of circWDR25 was the most obviously up-regulated in the exosomes isolated from tumor-activated LX-2 cells. The expression of peritumoral circWDR25 was positively correlated with HSCs adjacent to the cancer loci (r=0.156, P=0.008). Multivariate analysis showed that a preoperative AST level >36 g/L, multiple tumors, a tumor diameter >5 cm, HSC>70, and circWDR25>190 were independent risk factors affecting the overall survival of HCC patients after radical resection; a preoperative AST level >36 g/L, multiple tumors, a tumor diameter >5 cm, presence of tumor thrombus, HSC>70, and circWDR25>190 were all independent risk factors for tumor-free survival in patients with liver cancer. Conclusion Peritumoral circWDR25 and HSCs are factors affecting the prognosis of HCC patients after radical hepatectomy, and their high expression in the adjacent tissues is closely related to a poor prognosis of the patients.

Key words: hepatocellular carcinoma; hepatic stellate cells; exosomes; circRNA; circWDR25