[Effects and mechanisms of resveratrol on vascular smooth muscle cell proliferation and neointimal hyperplasia in a carotid artery injury model].

Objective: To investigate the effects of resveratrol on vascular smooth muscle cell (VSMC) proliferation and neointimal hyperplasia in the rat carotid artery balloon injury model after endovascular intervention, and to explore the underlying mechanisms. Methods: Drug safety assessment: VSMC were assigned to a control group, a low-dose resveratrol group (10 μmol/L), and a high-dose resveratrol group (20 μmol/L). Cell viability was evaluated using the cell counting Kit-8 (CCK-8) assay. Pharmacodynamic assessment: VSMC were divided into a control group, a model group [20 ng/ml platelet-derived growth factor-BB (PDGF-BB)], a PDGF-BB+low-dose resveratrol group (20 ng/ml PDGF-BB+10 μmol/L resveratrol), a PDGF-BB+high-dose resveratrol group (20 ng/ml PDGF-BB+20 μmol/L resveratrol), and a positive control group (20 ng/ml PDGF-BB+20 nmol/L rapamycin). Relative cell proliferation was measured by CCK-8. Additionally, cell migration and Ki-67 expression were evaluated in the control, model, and resveratrol-treated groups (all at the previously described dose) using wound healing assays and immunofluorescence respectively. Mechanistic investigation: Western blotting was performed to quantify mammalian target of rapamycin (mTOR) pathway-related proteins, including mTOR, phosphorylated mTOR (p-mTOR), ribosomal S6 kinase (p70S6K), Cyclin D1, and proliferating cell nuclear antigen (PCNA). Rescue experiment: VSMC were divided into a control group, a model group, a resveratrol treatment group, and a mTOR agonist rescue group (10 μmol/L MHY1485 was added to the resveratrol treatment group). Cell proliferation and mTOR pathway protein levels (mTOR、p-mTOR and p70S6K) were assessed. In vivo study: the rat carotid artery balloon injury model was established. Rats were randomly allocated by a random number table to a sham-operated group, an injury group, and a treatment group (50 mg·kg-1·d-1 resveratrol by intragastric administration for 21 days post-injury). On the day 21, hematoxylin-eosin (HE) staining was performed to calculate the intima/media (I/M) ratio. Ki-67 expression and mTOR pathway proteins (mTOR、p-mTOR and p70S6K) in arterial tissue were quantified by immunofluorescence and Western blotting, respectively. Results: In the drug safety assessment, no significant differences in cell proliferation were observed among the control and resveratrol-only groups (P>0.05). In the pharmacodynamic assessment, compared with the model group, relative cell proliferation, migration rate, and Ki-67 fluorescence intensity were reduced in the PDGF-BB+low-dose and PDGF-BB+high-dose resveratrol groups (all P<0.01). There was no statistically significant difference in cell proliferation between the high-dose resveratrol group and the positive control group (P>0.05). In the mechanistic investigation, the expression of mTOR, p-mTOR, p70S6K, Cyclin D1, and PCNA was lower in the resveratrol-treated groups than the model group (all P<0.01). In the rescue experiment, the level of cell proliferation the expression of p-mTOR and p70S6K were both higher in the mTOR agonist rescue group than the resveratrol treatment group (all P<0.01). In vivo, the treatment group exhibited a lower I/M ratio, reduced Ki-67 fluorescence intensity, and decreased expression of mTOR pathway proteins compared with the injury group (all P<0.01). Conclusion: Resveratrol may inhibit VSMC proliferation and migration by suppressing mTOR signaling activity, thereby reducing neointimal hyperplasia after carotid balloon injury and alleviating post-intervention restenosis.