Sexual Dimorphic Regulation of MicroRNAs Alters Sodium Transport in the Kidney Distal Nephron

Abstract

BackgroundHypertension affects more than one billion people worldwide. A key regulator of blood pressure homeostasis is aldosterone, the final signaling hormone of the renin‐angiotensin‐aldosterone‐signaling (RAAS) system. Aldosterone increases sodium (Na+) transport in the kidney distal nephron to regulate blood volume. Premenopausal women are less likely to develop hypertension than age‐matched men, due to both lower aldosterone levels and estrogen signaling. We previously demonstrated that aldosterone alters the expression of microRNAs (miRs) in collecting duct epithelial cells to modulate the Na+ transport response to aldosterone. However, the sex‐specific regulation of miRs and role of estrogen to alter miR expression in the kidney distal nephron has not been explored. This study investigated the hypothesis that estrogen alters aldosterone signaling in the distal nephron by regulating the expression of miRs in collecting duct epithelia.MethodsPrimary cortical collecting duct (CCD) cells derived from male and female mice as well as the mCCD‐cl1 cell line were cultured on permeable filter supports to measure Na+ transport by short‐circuit current recordings in Ussing chambers. Cells were incubated with estrogen for time and dose responses to determine the impact on aldosterone stimulation and Na+ transport. RT‐qPCR quantified miR and mRNA expression and western blot analysis quantified protein expression after aldosterone and estrogen stimulation. In vivo regulation of miRs by aldosterone was confirmed using freshly isolated CCD cells from male and female mice placed on low Na+ diets.ResultsA sex‐specific upregulation of the miR‐17~92 cluster was observed in female mice placed on a low‐Na+ diet to stimulate aldosterone release. MiRs‐19a&b were also upregulated in mCCD cells stimulated with estrogen. Estrogen pretreatment blunted aldosterone stimulation, by targeting the serum and glucocorticoid induced kinase (SGK1) mRNA. Luciferase assays demonstrated that miRs‐19a&b bind to the 3’‐UTR of SGK1. Overexpression of miR‐19 in mCCD cells using miR mimics significantly inhibited aldosterone stimulation of Na+ transport. Conversely, aldosterone stimulation was greater in mCCD cells transfected with a miR‐19 inhibitor (antagomir).ConclusionThe miR‐17~92 cluster is regulated by aldosterone and estrogen. Mir‐19 targets SGK1 and may account for estrogen’s inhibition of aldosterone signaling. This miR cluster may be responsible, in part, for the observed sex‐specific differences in aldosterone signaling. Ongoing studies aim to determine if altering the expression of miR‐17~92 disrupts RAAS signaling in the kidney in vivo.