The novel mineralocorticoid receptor modulator balcinrenone protects against diet-induced cardiac microvascular dysfunction and plasma potassium elevation in mouse models

Figures

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  Fig 1. MPR analysis in mice with diet-induced HFpEF: (A) nominal and achieved doses of balcinrenone and eplerenone, (B) rest and stress perfusion values and derived MPR, and (C) MPR of treatment groups. (A) The amount of food consumed during the main study was monitored weekly and used to calculate the average doses of balcinrenone and eplerenone that each mouse received. (B) Myocardial perfusion before (rest) or after (stress) administration of 0.1 μg/g regadenoson. (C) MPR was calculated as: stress perfusion in mL/g/min divided by rest perfusion in mL/g/min, in mice after 20 weeks on the respective diets. Data are presented as means ± standard error. N = 14 or 15 for vehicle groups, N = 9 or 10 for treatment groups. bal, balcinrenone; eple, eplerenone; HFD, high-fat diet; MPR, myocardial perfusion reserve; RD, regular diet.

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  Fig 2. Balcinrenone and eplerenone attenuate cardiac perivascular fibrosis in mice with diet-induced HFpEF. Cardiac tissue samples from mice fed an RD, HFD, or HFD with 100 mg/kg/day balcinrenone or eplerenone were examined for structural changes by histology. (A) Representative images of cardiac slides stained with Masson’s Trichrome (three to five hearts per group and two to eight slides per heart). (B) Quantification of histology using ImageJ color deconvolution and the blue channel to set thresholds. Data are presented as means ± standard error. *P < 0.05. HFD, high-fat diet; ns, not significant; RD, regular diet.

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  Fig 3. Balcinrenone and eplerenone inhibit aldosterone-induced expression of MR target genes and markers of cardiac remodeling in H9C2/MR+ cells. H9C2/MR+ cells were stimulated with 1 nM aldosterone or vehicle for 24 hours, followed by the addition of balcinrenone or eplerenone to final concentrations of 0.1, 0.5, 2.5, or 12.5 µM. Quantitative reverse transcription polymerase chain reaction was performed to assess gene expression. Data are presented as means ± standard deviations. *P < 0.05 versus control; #P < 0.05 versus 1 nM aldosterone. Aldo, aldosterone; Ctrl, control; MR, mineralocorticoid receptor; mRNA, messenger RNA.

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  Fig 4. Balcinrenone and eplerenone inhibit (A) fibrotic (collagen 1) and (B) inflammatory (IL-6) processes in primary human cardiac fibroblasts. Primary human cardiac fibroblasts were stimulated for 24 hours with or without 10 nM aldosterone, and balcinrenone or eplerenone added to final concentrations of 0.5, 2.5, or 12.5 µM. (A) Collagen 1 and (B) IL-6 concentrations in the supernatant were measured using an enzyme-linked immunosorbent assay. Data are presented as means ± standard deviations. *P < 0.05 versus control; #P < 0.05 versus 10 nM aldosterone. Aldo, aldosterone; Ctrl, control; IL-6, interleukin-6.

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  Fig 5. K+ homeostasis in the mouse model of nephrectomy-induced CKD: Plasma K+ levels in (A) sham-operated mice and (B) mice with nephrectomy-induced CKD before and after K+ challenge, (C) creatinine-normalized urinary K+ excretion, and (D) fecal K+ excretion. Twenty-four mice had CKD induced through 5/6 nephrectomy or had sham surgery. Three groups of eight mice (with CKD and sham operated) received parallel treatment with either vehicle, eplerenone, or balcinrenone; eplerenone and balcinrenone were administered at doses of 100 mg/kg, twice daily for 4 days, then once on day 5. Overnight K+ challenge was performed between 6 PM on day 4 and 9 AM on day 5. Plasma K+ levels were measured on day 5 (4 hours after last administration of vehicle, eplerenone, or balcinrenone). Urinary and fecal K+ levels were measured using flame photometry. K+ homeostasis data are presented as means ± standard error. *P < 0.05; #P < 0.05 versus vehicle. CKD, chronic kidney disease; K+, potassium; KCI, potassium chloride; ns, not significant.