Abstract
Vascular disease remains a leading cause of morbidity and mortality and is driven by maladaptive vascular remodeling following injury. Stent-induced vascular injury induces vascular smooth muscle cell (VSMC) phenotypic switching from a contractile to a proliferative state, resulting in intimal hyperplasia (IH), restenosis and compromised vessel function. Nitrite, an endogenous oxidation product of nitric oxide and a dietary constituent, attenuates IH after vascular injury; however, its underlying mechanisms remain incompletely understood. Nitrite is known to modulate mitochondrial structure and function, and dysregulated mitochondrial dynamics have independently been implicated in VSMC proliferation. We therefore hypothesized that nitrite attenuates IH by modulating mitochondrial dynamics to suppress VSMC proliferation. Using rat aortic smooth muscle cells (RASMCs), we demonstrate that nitrite treatment inhibits cell cycle progression and cell proliferation through upregulation of mitofusin-1 (Mfn1), a GTPase that catalyzes mitochondrial fusion. Mechanistically, nitrite increased Mfn1 protein levels by inhibiting Mfn1 proteasomal degradation. Mfn1 deletion resulted in enhanced proliferation, loss of contractile gene expression, and decreased expression of antioxidant enzymes including catalase and glutathione peroxidase. Restoration of cellular antioxidant capacity significantly attenuated proliferation and preserved contractile gene expression in Mfn1-deficient cells. Smooth muscle cell-specific Mfn1 knockout mice subjected to carotid artery ligation injury exhibited exacerbated IH compared to wildtype mice. Nitrite administration significantly decreased IH in wildtype mice but not Mfn1-deficient mice. These findings identify endogenous Mfn1 as a critical regulator of VSMC cell cycle progression and as an essential mediator of the vasoprotective effects of nitrite.