Aldosterone Insufficiency Contributes to Calcineurin Inhibitor‐induced Hyperkalemia

Abstract

The mineralocorticoid aldosterone promotes renal K+ excretion and Na+ reabsorption; thereby it is critical for the regulation of ion homeostasis and blood pressure. Interestingly, immunosuppression therapy with protein phosphatase 3 (calcineurin) inhibitors often results in rather low plasma aldosterone levels despite a concomitant hyperkalemia and hyperreninemia. Calcineurin (Cn) is a calcium and calmodulin-dependent protein phosphatase expressed in the adrenal cortex. We tested the hypothesis that Cn participates in the signal transduction pathway mediating the K+-dependent stimulation of aldosterone production. To address this question, we used the adrenocortical cell model NCI-H295R, mouse and human ex vivo adrenal preparations and a ZG-specific and inducible Cn-beta subunit 1 (CnB1) knockout mouse model (ZG-CnB1-KO). Inhibition of Cn with tacrolimus abolished the K+-stimulated expression of CYP11B2 in NCI-H295R cell line as well as in mouse and human adrenal pieces, ex vivo. Moreover, high K+ diet-dependent increase in aldosterone production was blunted in wild type mice treated with tacrolimus. Furthermore, male ZG-CnB1-KO mice fed a high K+ diet exhibited a decreased aldosterone excretion compared to control animals while ZG-CnB1-KO females became hyperkalemic. Using a phosphoproteomics analysis, we identified the nuclear factor of activated T-cells, cytoplasmic 4 (NFATc4) as a downstream factor regulated by Cn. The genetic deletion of NFATc4 in NCI-H295R cells reduced the basal expression of CYP11B2 and blunted the K+-stimulated expression of this gene. Conversely, the expression of a constitutively active form of NFATc4 in NCI-H295R cells drastically increased the expression of CYP11B2 which remained unaltered upon treatment with K+ or tacrolimus. Altogether, our data indicate that the calcineurin-NFATc4 pathway is activated by K+ to promote adrenal aldosterone production. Moreover, our data indicate that insufficient aldosterone production contributes to Cn inhibitor-induced hyperkalemia.