1,25-dihydroxyvitamin D3 causes ADAM10-dependent ectodomain shedding of tumor necrosis factor receptor 1 in vascular smooth muscle cells.

Abstract

1,25-Dihydroxyvitamin D3 (1,25D3) has a potential antiatherosclerotic effect through anti-inflammatory actions. We investigated how 1,25D3 regulates tumor necrosis factor-α (TNF-α)-induced lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) expression in cultured human aortic smooth muscle cells. TNF-α activated Rac1/reactive oxygen species/spleen tyrosine kinase and transcriptional factors, activator protein-1, and nuclear factor κB, which led to LOX-1 expression. 1,25D3 inhibited TNF-α-induced LOX-1 expression by inhibiting Rac1 activation and thereby its downstream signals. 1,25D3 rapidly induced extracellular Ca(2+) influx. Verapamil, an inhibitor of L-type calcium channels, inhibited 1,25D3-induced Ca(2+) influx and counteracted the inhibitory effects of 1,25D3 on Rac1 activation, whereas Bay K8644 [1,4-dihydro-2,6-dimethyl-5-nitro-4-[2-(trifluoromethyl)phenyl]-3-pyridinecarboxylic acid, methyl ester], an L-type calcium channel agonist, attenuated TNF-α-induced Rac1 activation, as 1,25D3 did. 1,25D3 induced the ectodomain shedding of TNF receptor 1 (TNFR1), which was abolished by verapamil and in Ca(2+)-free media. Like 1,25D3, Bay K8644 induced the ectodomain shedding of TNFR1. Both 1,25D3 and Bay K8644 caused the translocation of a disintegrin and metalloprotease (ADAM) 10 from the cytoplasm to the plasma membrane, which was dependent on extracellular Ca(2+) influx. In contrast, depletion of ADAM10 by transfection of ADAM10-small interfering RNA prevented 1,25D3- or Bay K8644-induced ectodomain shedding of TNFR1 and abolished the suppressive effect of 1,25D3 on TNF-α-induced Rac1 activation. Taken together, these findings suggest that 1,25D3 induces extracellular Ca(2+) influx via L-type calcium channel, triggering ADAM10-mediated ectodomain shedding of TNFR1, and it thereby decreases responsiveness to TNF-α. By shedding TNFR1 from the cell surface, 1,25D3 may regulate inflammation and atherogenesis, whereas this effect could be attenuated by calcium channel blockers.