Chen, Yang

Aldosterone is a critical driver for cardiovascular disease (CVD). We recently discovered that MANP, a novel atrial natriuretic peptide (ANP) analog, possessed more potent aldosterone inhibitory action than ANP. MANP is currently entering clinical trials for hypertension and thus understanding its aldosterone suppressing mechanism is important. The mechanism of aldosterone inhibition by natriuretic peptides (NPs) remains to be clearly defined. Conflicting results were reported on the roles of particulate guanylyl cyclase A receptor (pGC-A) and NP clearance receptor (NPRC) in aldosterone inhibition. Furthermore, the functions of protein kinase G (PKG) and phosphodiesterases (PDE) on aldosterone regulation are not clear. Herein, we investigated the molecular mechanism of aldosterone regulation in the human adrenocortical cell line H295R and in mice. We firstly showed that pGC-A mediates aldosterone inhibition. In contrast, with NPRC agonist and antagonist, we showed that NPRC did not inhibit aldosterone. Next, we confirmed that MANP inhibits aldosterone via PDE2, not PKG, with specific agonists, antagonists, siRNA silencing, and fluorescence resonance energy transfer (FRET) experiments. Specifically, MANP suppressed ANGII mediated activation of aldosterone (fold change) MANP+ANGII 3.2±0.1* vs. ANGII 3.8±0.1 (*p<0.05) with IBMX, a PDEs inhibitor and the PDE2 antagonist Bay 60-7550 reversed MANP-mediated aldosterone suppression (IBMX+MANP+ANGII 3.9±0.2 and Bay+MANP+ANGII 4.1±0.1). With PKG agonists and inhibitors, aldosterone levels were not changed. In PDE2 activity FRET studies, aldosterone control was 3.7±0.4 and with MANP 0.9±0.2* supporting PDE2 activation by MANP. Further, the inhibitory effect of PDE2 is mediated by a reduction of intracellular Ca2+ concentration (~22%). We then showed that MANP directly reduced aldosterone synthase CYP11B2 expression in vitro. Lastly, in PDE2 knockout mice (embryonic lethal), embryonic adrenal CYP11B2 expression is markedly increased (wild type: 1±0.2, KO: 2.8±0.5*). Our findings innovatively elucidate the pGC-A/cGMP/PDE2 pathway in aldosterone inhibition by MANP in vitro and in vivo. Additionally, our data also support the development of MANP as a novel ANP analog drug for CVD.

The well-balanced secretion between insulin and growth hormone (GH) is essential in regulating substrate metabolism, energy metabolism, and body composition. High insulin and low GH levels are often observed in obesity, contributing to reduced energy expenditure and further fat accumulation. Although suppression of hyperinsulinemia is proposed as a treatment for obesity, changes in GH secretion and energy metabolism following this treatment are not thoroughly studied. This leaves unexplained observations, such as unchanged lean mass following insulin reduction. In this study, high-fat diet-induced obese (DIO) and normal chow-fed lean mice on a C57BL/6J background were treated for 7 weeks with diazoxide (1250 mg/kg in food), a K ATP channel opener that suppressed insulin secretion. Diazoxide treatment for 10 days was sufficient to increase pulsatile GH secretion in DIO mice before any significant body weight change. The restored insulin-GH balance in DIO mice was followed by improvement in substrate and energy metabolism in a prolonged treatment period (4–6 weeks), including reduced fat mass, increased lipid oxidation and energy expenditure, as well as improved insulin sensitivity and metabolic flexibility. These metabolic benefits occurred along with the changes in the expression level of genes regulated by the insulin-GH balance. When applying diazoxide to normal chow-fed normoinsulinemic lean mice, none of the above metabolic effects was observed, suggesting that the metabolic changes following diazoxide treatment were mediated through the suppression of hyperinsulinemia. These results suggest that suppression of hyperinsulinemia by diazoxide restores GH secretion and improves substrate and energy metabolism in DIO mice.

Abstract Insulin and growth hormone (GH) are crucial counter-regulatory hormones in regulating glucose and lipid metabolisms. Insulin promotes fat storage, while GH promotes lipolysis and fat oxidation. In obese individuals, reduced GH secretion (hyposomatotropism) and increased insulin secretion (hyperinsulinemia) co-exist. The imbalance of these two hormones exacerbates fat accumulation. Therapeutic approaches to correct such hormonal imbalance in obesity are limited. The sodium/glucose cotransporter 2 inhibitor (SGLT2i), which promotes urinary glucose excretion, is a novel drug for overt type 2 diabetes (T2D). However, little is known about its efficacy in obese individuals without T2D in the clinic, in particular with hormonal imbalance. By applying SGLT2i (dapagliflozin, 1 mg/kg/d for 10 weeks) to a hyperphagic obese melanocortin 4 receptor knockout (MC4RKO) mouse model, we observed a significant reduction of hyperinsulinemia (fasting: 1.36±0.19 vs. 4.93±1.04 ng/ml, p&lt;0.01; fed: 9.50±3.37 vs. 31.11±5.85 ng/ml, p&lt;0.05, n=8) and restored pulsatile GH secretion without changing secretion pattern (pulsatile GH: 185.3±18.37 vs. 56.28±13.22 ng/ml per 6h, p&lt;0.001; GH mass per secretion pulse: 50.31±8.20 vs. 15.55±3.18 ng/ml, p&lt;0.01; number of secretory pulse per 6h: 3.71±0.29 vs. 3.57±0.43, p=0.78, n=8) as early as 4 weeks after the initiation of the treatment. Lipolysis and lipid oxidation-related gene expression levels were increased by SGLT2i treatment, whereas lipogenesis and inflammation gene expression levels were reduced, leading to decreased whole-body fat mass. Following the treatment, glucose tolerance and insulin sensitivity were both improved. Although a null effect was observed in food intake and daily activity, the treatment significantly promoted lipid usage and shifted energy metabolism towards negative energy balance. In conclusion, 10-week SGLT2i treatment improved glucose and lipid metabolisms in the hyperphagic obese MC4RKO mice. Such improvement occurs alongside reduced hyperinsulinemia and restored pulsatile GH secretion. This work provides insights for the potential use of SGLT2i in obese individuals prior to overt T2D. The final version of this work is published (1). Acknowledgements: grant (NHMRC, University of Queensland) and scholarship (CSC and UQ International scholarship) Reference: (1) Huang, Zhengxiang, et al. “Dapagliflozin restores insulin and growth hormone secretion in obese mice.” Journal of Endocrinology 245.1 (2020): 1-12. Unless otherwise noted, all abstracts presented at ENDO are embargoed until the date and time of presentation. For oral presentations, the abstracts are embargoed until the session begins. Abstracts presented at a news conference are embargoed until the date and time of the news conference. The Endocrine Society reserves the right to lift the embargo on specific abstracts that are selected for promotion prior to or during ENDO.