β-Sitosterol activates autophagy to inhibit the development of hepatocellular carcinoma by regulating the complement C5a receptor 1/alpha fetoprotein axis

Hepatocellular carcinoma (HCC) is highly refractory. beta-Sitosterol has been reported to suppress proliferation and migration as well as interfere with cell metabolism in tumors. However, there is limited information on the effects of beta-sitosterol on HCC. Herein, we used a xenograft mouse model to investigate the effects of beta-sitosterol on HCC tumor growth. The molecular mechanism was elucidated using quantitative real-time PCR, western blotting, lentiviral transfection, CCK8, scratch, Transwell, and Ad-mCherry-GFP-LC3B assays. The results showed that HepG2 cells highly expressed complement C5a receptor 1. beta-Sitosterol antagonized complement component 5a and exerted inhibitory effects on the proliferation and migration of HepG2 cells. The inhibitory effect of beta-sitosterol was reversed by the knockdown of complement C5a receptor 1. Bioinformatics analysis suggested alpha fetoprotein (AFP) as a downstream factor of complement C5a receptor 1. beta-Sitosterol inhibited AFP expression, which was reversed by complement C5a receptor 1 knockdown. The inhibitory effects of beta-sitosterol on cell proliferation and migration were weakened by AFP overexpression. Furthermore, beta-sitosterol induced autophagy in HepG2 cells, which was reversed by complement C5a receptor 1 knockdown and AFP over-expression. Blockade of autophagy by 3-MA attenuated beta-sitosterol inhibition of proliferation and migration in HepG2 cells. Moreover, beta-sitosterol inhibited HCC progression in vivo. Our findings demonstrate that beta-sitosterol inhibits HCC advancement by activating autophagy through the complement C5a receptor 1/AFP axis. These findings recommend beta-sitosterol as a promising therapy for HCC.