Neferine inhibits angiotensin Ⅱ-stimulated proliferation in vascular smooth muscle cells through heme oxygenase-1

摘要

Aim: To explore the effect of neferine on angiotensin Ⅱ (Ang Ⅱ)-induced vascular smooth muscle cell (VSMC) proliferation.Methods: Human umbilical vein smooth muscle cells (HUVSMCs) were used. Cell proliferation was determined by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry analysis. Heme oxygenase (HO)-1 protein expression was tested by Western blot analysis. Extracellular signal-regulated protein kinase 1/2 (ERK1/2) activation was determined by using immunoblotting.Results: Pre-incubation of HUVSMCs with neferine (0.1, 0.5, 1.0, and 5.0 μmol/L) significantly inhibited Ang Ⅱ-induced cell proliferation in a concentration-dependent manner and neferine 5.0 μmol/L increased HO-1 expression by 259% compared with control. The antiproliferative effect of neferine was significantly attenuated by coapplication of zinc protoporphyrin X (ZnPP Ⅸ, an HO-1 inhibitor) with neferine. Ang Ⅱ-enhanced ERK1/2 phosphorylation was markedly reversed by neferine. By inhibiting HO-1 activity with ZnPP Ⅸ, the inhibitive effect of neferine on ERK1/2 phosphorylation was significantly attenuated. Cobalt-protoporphyrin (CoPP), an HO-1 inducer,significantly decreased Ang Ⅱ-induced ERK1/2 phosphorylation and inhibited Ang Ⅱ-induced cell proliferation. The ERK1/2 pathway inhibitor PD98059 significantly blocked Ang Ⅱ-enhanced ERK1/2 phosphorylation and inhibited cell proliferation.Conclusion: These findings suggest that neferine can inhibit Ang Ⅱ-induced HUVSMC proliferation by upregulating HO-1, leadingto the at least partial downregulation of ERK1/2 phosphorylation.