Nifedipine inhibits vascular smooth muscle cell dedifferentiation via downregulation of Akt signaling.

Calcium is an essential signaling molecule that controls vascular smooth muscle cell (VSMC) contraction, proliferation, and differentiation. Here, we show that the calcium antagonist nifedipine inhibits VSMC dedifferentiation in vitro and in vivo. Differentiated VSMCs cultured on laminin-coated dishes were transferred to laminin-free dishes to induce dedifferentiation. Induction of dedifferentiation resulted in the upregulation of nonmuscle myosin heavy chain expression, a marker of dedifferentiation, and the downregulation of smooth muscle myosin heavy chain expression, a marker of differentiation. Nifedipine significantly inhibited both the induction of these phenotypic changes and upregulation of Akt signaling in these cells. Administration of nifedipine at a low concentration that did not affect blood pressure could inhibit the increase in nonmuscle myosin heavy chain expression and decrease in smooth muscle myosin heavy chain expression in a rat balloon-injury model. Furthermore, nifedipine suppressed neointimal hyperplasia and upregulation of Akt signaling. However, phospho-Akt expression was not suppressed in the regenerating arterial endothelium of the nifedipine-treated rats. The inhibitory effect of the downregulation of Akt signaling by dominant-negative Akt on the induction of VSMC dedifferentiation in the intima was identical to that of nifedipine. In contrast, upregulation of Akt signaling by transfection of the cells with a constitutively active Akt reversed the nifedipine-induced inhibition of VSMC dedifferentiation. In conclusion, nifedipine inhibits VSMC dedifferentiation by suppressing Akt signaling, thereby preventing neointimal thickening.

Homocysteine enhances endothelial apoptosis via upregulation of Fas-mediated pathways.

Hyperhomocysteinemia is an independent risk factor for the development of atherosclerosis. However, the underlying mechanism of endothelial cell injury in hyperhomocysteinemia has not been elucidated. In this study, we examined the effect of homocysteine (Hcy) on Fas-mediated apoptosis in endothelial cells. Hcy-induced upregulation of Fas in endothelial cells (ECs) in a dose-dependent manner. At the same time, Hcy increased intracellular peroxide in ECs. Hcy-induced Fas expression was inhibited by the treatment with catalase. Hcy increased NF-kappaB DNA binding activity, and adenovirus-mediated transfection of a Ikappa-B mutant (Ikappa-B mt) gene inhibited Hcy-induced Fas expression. ECs were sensitive to Fas-mediated apoptosis when exposed to Hcy. Under these condition, Ikappa-B mt protected ECs from Fas-mediated apoptosis. In addition, Hcy inhibited expression of the caspase-8 inhibitor FLICE-inhibitory protein (FLIP). Adenovirus-mediated transfection of constitutively active Akt gene abolished the Hcy-mediated downregulation of FLIP. These data suggest that upregulation of Fas expression and downregulation of FLIP is a mechanism through which Hcy induces EC apoptosis.

Nifedipine upregulates manganese superoxide dismutase expression in vascular smooth muscle cells via endothelial cell-dependent pathways.

Calcium antagonists normalize endothelial dysfunction and improve the clinical outcome in patients with hypertension. However, the mechanism underlying these beneficial effects remains to be elucidated. Here, we show that the calcium antagonist nifedipine upregulates the expression of manganese superoxide dismutase (Mn SOD), an endogenous antioxidant enzyme, in vascular smooth muscle cells (VSMC) via cellular interactions between VSMC and endothelial cells (EC). Nifedipine induced upregulation of Mn SOD activity and expression in VSMC when cocultured with EC but not when cultured individually. NG-Monomethyl-L-arginine (L-NMMA), an inhibitor of nitric oxide (NO) synthesis, inhibited the upregulation of Mn SOD expression induced by nifedipine. Additionally, N-ethyl-2-(1-ethyl-2-hydroxy-2-nitrosohydrazino) ethanamine, a NO donor, reversed this inhibition by L-NMMA, indicating that NO may be involved in the mechanism underlying the nifedipine-induced upregulation of Mn SOD in VSMC. Preincubation of VSMC with Mn SOD antisense oligodeoxyribonucleotides (ODN) blocked the suppressive effects of nifedipine on DNA synthesis in VSMC cocultured with EC, whereas sense ODN had no effect. We conclude that the calcium antagonist nifedipine induces upregulation of Mn SOD expression in VSMC via NO derived from EC. This finding may provide some insight into the mechanism underlying the beneficial effects of calcium antagonists in patients with hypertension.