Metformin Inhibits StAR Expression in Human Endometriotic Stromal Cells via AMPK-Mediated Disruption of CREB-CRTC2 Complex Formation

Context: Endometriosis is an estrogen-dependent disease affecting reproductive women. Metformin could have a therapeutic effect on endometriosis through regulation of local estrogen production. Objects: The aim of this study was to investigate the molecular and cellular mechanism by which metformin...

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Published in:The journal of clinical endocrinology and metabolism 2014-08, Vol.99 (8), p.2795-2803
Main Authors: Xu, Jia-Ning, Zeng, Cheng, Zhou, Yan, Peng, Chao, Zhou, Ying-Fang, Xue, Qing
Format: Article
Language:English
Subjects:
Adult
AMP-Activated Protein Kinases - metabolism
Biological and medical sciences
Cells, Cultured
Cyclic AMP Response Element-Binding Protein - metabolism
Down-Regulation - drug effects
Down-Regulation - genetics
Endocrinopathies
Endometrium - drug effects
Endometrium - metabolism
Feeding. Feeding behavior
Female
Fundamental and applied biological sciences. Psychology
Humans
Medical sciences
Metformin - pharmacology
Phosphoproteins - genetics
Phosphoproteins - metabolism
Phosphorylation - drug effects
Protein Binding - drug effects
Signal Transduction - drug effects
Signal Transduction - genetics
Stromal Cells - drug effects
Stromal Cells - metabolism
Transcription Factors - metabolism
Vertebrates: anatomy and physiology, studies on body, several organs or systems
Vertebrates: endocrinology
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Summary:Context: Endometriosis is an estrogen-dependent disease affecting reproductive women. Metformin could have a therapeutic effect on endometriosis through regulation of local estrogen production. Objects: The aim of this study was to investigate the molecular and cellular mechanism by which metformin regulates StAR expression in human endometriotic stromal cells (ESCs). Methods: ESCs derived from ovarian endometriomas were cultured with metformin and prostaglandin E2 (PGE2). StAR mRNA was measured by quantitative PCR; pregnenolone, progesterone, and estrogen production were measured by ELISA kits; steroidogenic acute regulatory protein (StAR), AMP-activated protein kinase, cAMP response element binding protein (CREB), and CREB-regulated transcription coactivator 2 (CRTC2) protein expression were measured by Western blot assay; and CRTC2 translocation and its association with CREB were assessed by coimmunoprecipitation assay and CRTC2-CREB complex binding by a chromatin immunoprecipitation assay. Results: 1) StAR mRNA levels in ESCs are 264 times higher than those in endometrial cells. 2) Metformin downregulates the StAR mRNA expression (maximum 31.7%) stimulated by PGE2 (2.4-fold) in ESCs. 3) PGE2 induces CRTC2 translocation and enhances its association with CREB to form a transcription complex that binds to the StAR promoter region. 4) Metformin prevents the nuclear translocation of CRTC2 by increasing AMP-activated protein kinase phosphorylation. This inhibits transcription of StAR by disrupting formation of the CREB-CRTC2 complex, involved in activation of the StAR promoter cAMP response element. Conclusions: We have demonstrated a detailed mechanistic analysis of StAR expression regulated by metformin in ESCs. Our data highlight a role for CRTC2 in the mechanism by which metformin inhibits StAR expression.
ISSN:0021-972X
1945-7197
DOI:10.1210/jc.2014-1593