Farrerol Inhibits Vascular Smooth Muscle Cell Proliferation and Protects Them From Oxidative Injury via Bidirectional Modulation of the PI3K/Akt/mTOR Signaling Pathway

The inhibition of intimal hyperplasia (IH) is an effective strategy to improve the long-term outcome of endovascular therapy and prevent restenosis. Farrerol, a naturally occurring dihydroflavone with a variety of bioactivities, exerts inhibitory effects against balloon injury-induced IH in rats. In the present study, bioinformatics analysis, in combination with in vitro experimental validation, was performed to elucidate the underlying inhibitory mechanisms. The protein-protein interaction (PPI) network was assessed to identify farrerol-related protein targets in the context of IH, based on which biological functions and pathway enrichment were analyzed. The proliferation and cell cycle distribution of vascular smooth muscle cells (VSMCs) were investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide and 5-ethynyl-2-deoxyuridine incorporation assays and flow cytometric analysis, respectively. The level of pro-inflammatory cytokines in the cell culture medium was estimated using an enzyme-linked immunosorbent assay (ELISA). Protein expression in A7r5 cells was determined by western blotting. Forty-six IH-related targets of farrerol were identified, and the PI3K/Akt/mTOR pathway was highly enriched among the 43 predicted pathways (P < .05). In serum (10% fetal bovine serum)-induced A7r5 cells, farrerol inhibited proliferation through non-cytotoxic effects, induced cell cycle arrest in the G(0)/G(1) phase, and suppressed the activation of the PI3K/Akt/mTOR pathway. In H2O2 (300 mu M)-induced A7r5 cells, farrerol reduced the release of IL-1 beta and TNF-alpha and reversed the suppressive effect on the PI3K/Akt/mTOR pathway in response to H2O2 stimulation. In conclusion, farrerol inhibits the proliferation of VSMCs and protects VSMCs from oxidative injury via the bidirectional modulation of the PI3K/Akt/mTOR signaling pathway, which might contribute to the suppression of neointima formation.