Question:

Klotho, a transmembrane protein, protease and hormone, is expressed in several tissues with highest expression in the kidney. Klotho is associated with various protective effects on kidney, vasculature and cancer metastasis, whereas Klotho deficiency results in accelerated aging and early death, effects at least partially due to hyperphosphatemia. Renal klotho expression is inhibited by aldosterone. The present study explored the interaction of the mineralocorticoids aldosterone and DOCA as well as the moderately selective mineralocorticoid receptor blocker spironolactone on Klotho gene expression.

Methods:

Transcript levels of Klotho and Cyp27b1 were determined utilizing quantitative RT-PCR in human embryonic kidney cells (HEK293) without and with aldosterone and/or spironolactone treatment or in renal tissue from mice with or without prior DOCA and/or spironolactone treatment. The experiments in HEK293 cells were performed without or with silencing of 25-hydroxyvitamin D₃ 1-alpha-hydroxylase, of vitamin D₃ receptor or of mineralocorticoid receptor (NR3C2).

Results:

In HEK293 cells aldosterone and and in mice DOCA significantly decreased Klotho gene expression, effects opposed by spironolactone treatment. Spironolactone treatment alone significantly increased Klotho and Cyp27b1 transcript levels in HEK293 cells (24 hours) and mice (8 hours or 5 days). Silencing of CYP27B1 and VDR, but not of NR3C2, significantly blunted the stimulating effect of spironolactone on klotho transcript levels in HEK cells.

Conclusions:

Besides blocking the effects of aldosterone, spironolactone upregulates Klotho gene expression by a mechanism not depending on the mineralocorticoid receptor, but involving upregulation of 25-hydroxyvitamin D₃ 1-alpha-hydroxylase with subsequent activation of the vitamin D₃ receptor by 1,25(OH)₂D₃.