Abstract: Objective To investigate whether circular RNA circRSF1 regulates radiation-induced inflammatory phenotype of hepatic stellate cells (HSCs) by binding to HuR protein and repressing its function. Methods Human HSC cell line LX2 with HuR overexpression or knockdown was exposed to 8 Gy X-ray irradiation, and the changes in the expression of inflammatory factors (IL-1β, IL-6 and TNF-α) were detected by qRT-PCR. The expressions of IκBα and phosphorylation of NF-κB were detected with Western blotting. The binding of circRSF1 to HuR was verified by RNA pull-down assay and RNA-binding protein immunoprecipitation (RIP). The expressions of inflammatory factors, IκBα and the phosphorylation of NF-κB were detected after modifying the interaction between circRSF1 and HuR. Results Knockdown of HuR significantly up- regulated the expressions of IL-1β, IL-6 and TNF-α, decreased IκBα expression and promoted NF-κB phosphorylation in irradiated LX2 cells, whereas overexpression of HuR produced the opposite changes (P<0.05). Overexpression or knockdown of circRSF1 did not significantly affect the expression of HuR. RNA pull-down and RIP experiments confirmed the binding between circRSF1 and HuR. Overexpression of circRSF1 significantly reduced the binding of HuR to IκBα and down-regulated the expression of IκBα (P<0.05). Overexpression of circRSF1 combined with HuR overexpression partially reversed the up-regulation of the inflammatory factors, down-regulated IκBα expression and increased phosphorylation of NFκB in LX2 cells, while the opposite effects were observed in cells with knockdown of both circRSF1 and HuR (P<0.05). Conclusion circRSF1 reduces IκBα expression by binding to HuR to promote the activation of NF-κB pathway, thereby enhancing radiation-induced inflammatory phenotype of HSCs.

Key words: radiation; hepatic stellate cells; circular RNA; HuR; inflammatory phenotype