MicroRNA‐21 Ablation Exacerbates Aldosterone‐Mediated Renal Injury
Authors: Njemanze Stephanie L , Syed Maryam , Nair Lakshmi Ramachandran , Ball Jana P , Mathis Keisa W , Ryan Michael J , Rothenberg Marc E , Cardozo Licy L Yanes , Romero Damian G 
Published: November 2023

Primary aldosteronism (PA) is characterized by excess autonomous secretion of aldosterone (ALDO) independent of the renin‐angiotensin system and accounts for ~10% of hypertensive patients. Excess ALDO causes hypertension and cardio‐renal hypertrophy, inflammation and fibrosis that lead to progressive cardio‐renal dysfunction. The molecular mechanisms involved in target organ injury induced by excess ALDO are poorly understood. MicroRNAs (miRNAs) are endogenous, small, non‐coding RNAs that downregulate the expression levels of specific proteins by translational repression or mRNA degradation. Multiple miRNAs have been implicated in renal physiological and pathological processes. In particular, microRNA‐21 (miR‐21) has been associated with renal ischemia/reperfusion injury, fibrosis and inflammation. Despite extensive studies, the role of miR‐21 in renal patho‐physiology remains highly controversial. The aim of the present study is to elucidate the role of miR‐21 in excess ALDO‐mediated renal injury. First, we use a transgenic reporter mouse that expresses the β‐galactosidase (LacZ) reporter gene under the endogenous miR‐21 promoter to analyze renal miR‐21 expression at the cellular level. Second, we use transgenic mice with genetic ablation of miR‐21 (miR21KO) to analyze renal injury and function in an experimental model of PA.To analyze renal miR‐21 expression, kidney samples from 10‐week old male miR‐21 reporter mice (lacZ‐KO‐Mir21tm1Mtm/Mmjax) were embedded in OCT, cryosectioned and LacZ detected by histochemistry. Histological analysis showed that miR‐21 is ubiquitously expressed in the kidney parenchyma, but its expression is highly enriched in the renal collecting ducts.To analyze the role of miR‐21 in ALDO‐mediated renal damage, 8‐week old UNX male miR21KO or their wild‐type (WT) controls were treated with ALDO (0.15 μg/h) and SALT (1.0 % NaCl + 0.3 % KCl) or vehicle for 8 weeks (n=12/group). ALDO/SALT caused renal hypertrophy (13.2 ± 0.1 vs. 8.4 ± 0.2 g/g BW*1,000, p<0.05) in WT mice and was exacerbated in miR21KO mice (14.1 ± 0.3 vs. 13.2 ± 0.1 g/g BW*1,000, p<0.05). ALDO/SALT increased blood pressure to the same level independently of miR‐21 genotype (WT: 139 ± 10 vs. 106 ± 2 mm Hg, miR21KO: 143 ± 8 vs. 114 ± 4 mm Hg) suggesting a blood pressure independent effect of miR‐21 in ALDO/SALT‐induced renal hypertrophy. Analysis of fibrosis markers' mRNA expression showed that ALDO/SALT increased renal collagen III (Col3a1) mRNA expression levels in miR21KO mice (1.85 ± 0.46 vs. 0.56 ± 0.06 AU, p<0.05) with no significant effect in WT mice (1.12 ± 0.10 vs. 0.55 ± 0.04 AU, p=NS). Analysis of inflammation markers' mRNA expression showed that ALDO/SALT increased renal osteopontin (Spp1) mRNA expression levels in miR21KO mice (4.70 ± 1.30 vs. 0.90 ± 0.06 AU, p<0.05) with no significant effect in WT mice (1.45 ± 0.13 vs. 0.98 ± 0.11 AU, p=NS). No significant genotype effects were observed in other fibrosis (Ctgf, Cola1a1, Fn1, Lox) or inflammation (TGF‐β, TNF‐α, IL1β, IL6, IL10, IL12) markers. No genotype effects were observed in urinary daily excretion of total protein, KIM‐1 or NGAL levels. Furthermore, no genotype effects were observed in glomerular filtration rate (GFR).Our results suggest that miR‐21 ablation exacerbates ALDO‐mediated renal hypertrophy, inflammation and fibrosis. These findings suggest that miR‐21 supplementation may be a novel therapy to mitigate excess ALDO‐mediated renal injury in PA patients.Support or Funding InformationSupported by American Heart Association Grant 12SDG8980032 (DGR).