Richardson Malcolm C, Ingamells Susan, Simonis Chantal D, Cameron Iain T, Sreekumar Rajiv, Vijendren Ananth, Sellahewa Luckni, Coakley Stephanie, Byrne Christopher D
Developmental Origins of Health and Disease Research Division, Institute of Developmental Sciences, University of Southampton, Southampton General Hospital, Tremona Road, Southampton SO16 6YD, United Kingdom.
J Clin Endocrinol Metab. 2009 Feb;94(2):670-7. doi: 10.1210/jc.2008-2025. Epub 2008 Nov 11.
Production of 3-carbon units (as lactate) by granulosa cells (GCs) is important in follicular and oocyte development and may be modulated by metformin.
The aim of the study was to examine the action of metformin on GC lactate production and potential mediation via AMP-activated protein kinase (AMPK).
GCs were prepared from follicular aspirates. After exposure to metformin and other potential modulators of AMPK in culture, aspects of cellular function were examined.
The study was conducted in a private fertility clinic/university academic center.
Women undergoing routine in vitro fertilization participated in the study.
All agents were added in culture.
Lactate output of GCs was measured. Cell extracts were prepared after culture, and phosphorylated forms of AMPK and acetyl CoA carboxylase (ACC) were assayed using Western analysis.
Metformin led to a rapid increase in lactate production by GCs [minimum effective dose, 250 microm; maximum dose studied, 1 mm (1.22-fold; P < 0.01)]. This dose range of metformin was similar to that required for stimulation of phospho-AMPK in GCs [minimum effective dose, 250 microm; maximum effect, 500 microm (2.01-fold; P < 0.001)]. Increasing phospho-ACC, as a representative downstream target regulated by AMPK, was apparent over a lower range (minimum effective dose, 31 microm; maximum effect, 250 microm; P < 0.001). A level of metformin (125 microm) insufficient for the stimulation of lactate output when used alone potentiated the effects of suboptimal doses of insulin on lactate production. Adiponectin (2.5 microg/ml) had a small but significant effect on lactate output.
Metformin activates AMPK in GCs, stimulating lactate production and increasing phospho-ACC. Metformin also enhances the action of suboptimal insulin concentrations to stimulate lactate production.
颗粒细胞（GCs）产生三碳单位（如乳酸）在卵泡和卵母细胞发育中很重要，且可能受二甲双胍调节。
本研究旨在探讨二甲双胍对GCs乳酸生成的作用以及通过AMP激活蛋白激酶（AMPK）的潜在介导作用。
从卵泡抽吸物中制备GCs。在培养中使其暴露于二甲双胍和其他潜在的AMPK调节剂后，检测细胞功能方面。
该研究在一家私立生育诊所/大学学术中心进行。
接受常规体外受精的女性参与了研究。
所有药物均在培养中添加。
检测GCs的乳酸产量。培养后制备细胞提取物，使用蛋白质免疫印迹分析检测AMPK和乙酰辅酶A羧化酶（ACC）的磷酸化形式。
二甲双胍导致GCs乳酸生成迅速增加[最小有效剂量，250微摩尔；研究的最大剂量，1毫摩尔（1.22倍；P<0.01）]。该剂量范围的二甲双胍与刺激GCs中磷酸化AMPK所需的剂量相似[最小有效剂量，250微摩尔；最大效应，500微摩尔（2.01倍；P<0.001）]。作为受AMPK调节的代表性下游靶点，磷酸化ACC的增加在较低剂量范围内明显（最小有效剂量，31微摩尔；最大效应，250微摩尔；P<0.001）。单独使用时不足以刺激乳酸产量的二甲双胍水平（125微摩尔）增强了次优剂量胰岛素对乳酸生成的作用。脂联素（2.5微克/毫升）对乳酸产量有小但显著的影响。
二甲双胍激活GCs中的AMPK，刺激乳酸生成并增加磷酸化ACC。二甲双胍还增强次优胰岛素浓度刺激乳酸生成 的作用。
