Pediatric Endocrinology, Diabetology, and Metabolism, Department of Pediatrics, University Children's Hospital Bern, 3010, Bern, Switzerland; Department of Biomedical Research, University of Bern, 3010, Bern, Switzerland.
Pediatric Endocrinology, Diabetology, and Metabolism, Department of Pediatrics, University Children's Hospital Bern, 3010, Bern, Switzerland; Department of Biomedical Research, University of Bern, 3010, Bern, Switzerland.
J Steroid Biochem Mol Biol. 2020 Jun;200:105684. doi: 10.1016/j.jsbmb.2020.105684. Epub 2020 Apr 28.
Metformin is recommended as one of the first-line drugs for the treatment of type 2 diabetes and the metabolic syndrome. In addition to its insulin sensitizing effects, it has been shown to attenuate androgen excess in women with polycystic ovary syndrome (PCOS) or congenital adrenal hyperplasia (CAH), as well as to ameliorate obesity. The mechanisms of metformin action seem manifold. Preclinical studies suggest that it inhibits the cellular stress response at the level of the mitochondrial OXPHOS system and through AMPK dependent and independent mechanisms. Recent studies have shown that metformin decreases ACTH secretion from pituitary and reduces ACTH-stimulated adrenal secretion. In this study we investigated its specific effect through the melanocortin receptor 2 (MC2R) on signaling targeting adrenal steroidogenesis. To assess this effect, we used mouse adrenal OS3 cells, which do not express the MC2R. Cells were transfected with the MC2R and stimulated by ACTH. Downstream cyclic AMP production was then assessed by a co-transfected cAMP-responsive vector producing luciferase that was measured by a dual luciferase assay. The amount of luciferase produced in this assay corresponds to the amount of receptor activation with varying amount of ACTH. The effect of metformin was then tested in this system. We found a significant inhibition of ACTH induced MC2R activation and signaling with 10 mM metformin. The ACTH concentration response curve (CRC) was half-log shifted and a ∼30 % reduction in maximum receptor response (Rmax) to ACTH in presence of metformin was observed. This effect was dose dependent with an IC of 4.2 mM. qRT-PCR analyses showed that metformin decreased ACTH induced MC2R expression. Metformin did not affect cell viability and basal cAMP levels. We also tested the effect of metformin on homologous melanocortin receptors (MCRs). No significant effect was found on MC1R and MC4R activity. However, a log shift of EC of ACTH stimulation on MC3R was observed with metformin treatment. Metformin also inhibited melanocortin stimulating hormone (αMSH) induced MC3R activity. In conclusion, we show that metformin acts on MC2R and MC3R signaling directly. The role of MC2R for steroidogenesis is well established. MC3R is involved in energy balance and seems to act as a rheostat when the metabolism is challenged. Our study may explain how metformin helps in weight loss and attenuates the excess response to ACTH in androgen excess disorders such as PCOS and CAH.
二甲双胍被推荐作为治疗 2 型糖尿病和代谢综合征的一线药物之一。除了其胰岛素增敏作用外，它还被证明可以减轻多囊卵巢综合征（PCOS）或先天性肾上腺增生（CAH）妇女的雄激素过多，并改善肥胖。二甲双胍的作用机制似乎多种多样。临床前研究表明，它可以抑制线粒体 OXPHOS 系统水平的细胞应激反应，并通过 AMPK 依赖和非依赖机制。最近的研究表明，二甲双胍可减少垂体分泌的 ACTH，并减少 ACTH 刺激的肾上腺分泌。在这项研究中，我们通过黑素皮质素受体 2（MC2R）研究了其对靶向肾上腺类固醇生成的信号的具体作用。为了评估这种作用，我们使用了不表达 MC2R 的小鼠肾上腺 OS3 细胞。将 MC2R 转染到细胞中，并通过 ACTH 进行刺激。然后通过共转染产生荧光素酶的 cAMP 反应性载体评估下游 cAMP 产生，通过双荧光素酶测定法进行测量。该测定法中产生的荧光素酶的量与受体激活的量相对应，激活量与不同量的 ACTH 相对应。然后在该系统中测试了二甲双胍的作用。我们发现，10mM 二甲双胍可显著抑制 ACTH 诱导的 MC2R 激活和信号传导。ACTH 浓度反应曲线（CRC）发生半对数偏移，并且在存在二甲双胍时观察到最大受体反应（Rmax）对 ACTH 的约 30％降低。该作用呈剂量依赖性，IC 为 4.2mM。qRT-PCR 分析显示，二甲双胍降低了 ACTH 诱导的 MC2R 表达。二甲双胍不影响细胞活力和基础 cAMP 水平。我们还测试了二甲双胍对同源黑素皮质素受体（MCR）的作用。未发现 MC1R 和 MC4R 活性的显着影响。然而，在用二甲双胍处理时，观察到 ACTH 刺激 MC3R 的 EC 对数偏移。二甲双胍还抑制了促黑激素（αMSH）诱导的 MC3R 活性。总之，我们表明二甲双胍直接作用于 MC2R 和 MC3R 信号传导。MC2R 对类固醇生成的作用已得到充分证实。MC3R 参与能量平衡，并且在代谢受到挑战时似乎充当变阻器。我们的研究可以解释二甲双胍如何帮助减轻体重，并减轻雄激素过多症（如 PCOS 和 CAH）中对 ACTH 的过度反应。
