Spironolactone ameliorates PIT1-dependent vascular osteoinduction in klotho-hypomorphic mice

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Abstract

Klotho is a potent regulator of 1,25-hydroxyvitamin D3 [1,25(OH)₂D₃] formation and calcium-phosphate metabolism. Klotho-hypomorphic mice (kl/kl mice) suffer from severe growth deficits, rapid aging, hyperphosphatemia, hyperaldosteronism, and extensive vascular and soft tissue calcification. Sequelae of klotho deficiency are similar to those of end-stage renal disease. We show here that the mineralocorticoid receptor antagonist spironolactone reduced vascular and soft tissue calcification and increased the life span of kl/kl mice, without significant effects on 1,25(OH)₂D₃, FGF23, calcium, and phosphate plasma concentrations. Spironolactone also reduced the expression of osteoinductive Pit1 and Tnfa mRNA, osteogenic transcription factors, and alkaline phosphatase (Alpl) in calcified tissues of kl/kl mice. In human aortic smooth muscle cells (HAoSMCs), aldosterone dose-dependently increased PIT1 mRNA expression, an effect paralleled by increased expression of osteogenic transcription factors and enhanced ALP activity. The effects of aldosterone were reversed by both spironolactone treatment and PIT1 silencing and were mitigated by FGF23 cotreatment in HAoSMCs. In conclusion, aldosterone contributes to vascular and soft tissue calcification, an effect due, at least in part, to stimulation of spironolactone-sensitive, PIT1-dependent osteoinductive signaling.

Authors

Jakob Voelkl, Ioana Alesutan, Christina B. Leibrock, Leticia Quintanilla-Martinez, Volker Kuhn, Martina Feger, Sobuj Mia, Mohamed S.E. Ahmed, Kevin P. Rosenblatt, Makoto Kuro-o, Florian Lang