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Stimulation of Lactate Production in Human Granulosa Cells by Metformin and Potential Involvement of Adenosine 5' Monophosphate-Activated Protein Kinase

Developmental Origins of Health and Disease Research Division (M.C.R., I.T.C., R.S., A.V., L.S., S.C., C.D.B.), Institute of Developmental Sciences, University of Southampton, Southampton General Hospital, Southampton SO16 6YD, United Kingdom; and Wessex Fertility Unit (S.I., C.D.S.), Southampton SO15 5QS, United Kingdom

Address all correspondence and requests for reprints to: Dr. Malcolm C. Richardson, Institute of Developmental Sciences, Mailpoint 887, Southampton General Hospital, Tremona Road, Southampton SO16 6YD, United Kingdom. E-mail: mcr2{at}soton.ac.uk.

Context: Production of 3-carbon units (as lactate) by granulosa cells (GCs) is important in follicular and oocyte development and may be modulated by metformin.

Objective: 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).

Design: GCs were prepared from follicular aspirates. After exposure to metformin and other potential modulators of AMPK in culture, aspects of cellular function were examined.

Setting: The study was conducted in a private fertility clinic/university academic center.

Patients: Women undergoing routine in vitro fertilization participated in the study.

Interventions: All agents were added in culture.

Main Outcome Measures: 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.

Results: Metformin led to a rapid increase in lactate production by GCs [minimum effective dose, 250 µM; 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 µM; maximum effect, 500 µM (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 µM; maximum effect, 250 µM; P < 0.001). A level of metformin (125 µM) insufficient for the stimulation of lactate output when used alone potentiated the effects of suboptimal doses of insulin on lactate production. Adiponectin (2.5 µg/ml) had a small but significant effect on lactate output.

Conclusions: 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.