title: The effect of metformin treatment in vivo on acute and long-term energy metabolism and progesterone production in vitro by granulosa cells from women with polycystic ovary syndrome.
abstract: OBJECTIVE
  What are the consequences of polycystic ovary syndrome (PCOS) pathology and metformin-pretreatment in vivo in women with PCOS on the metabolism and steroid production of follicular phenotype- and long-term cultured-granulosa cells (GC)?
  
  
  CONCLUSIONS
  PCOS pathology significantly compromised glucose metabolism and the progesterone synthetic capacity of follicular- and long-term cultured-GCs and the metabolic impact of PCOS on GC function was alleviated by metformin-pretreatment in vivo.
  
  
  BACKGROUND
  Granulosa cells from women with PCOS have been shown to have an impaired insulin-stimulated glucose uptake and lactate production in vitro. However, these results were obtained by placing GCs in unphysiological conditions in culture medium containing high glucose and insulin concentrations. Moreover, existing data on insulin-responsive steroid production in vitro by PCOS GCs vary.
  
  
  METHODS
  Case-control experimental research comparing glucose uptake, pyruvate and lactate production and progesterone production in vitro by GCs from three aetiological groups, all undergoing IVF; healthy control women (Control, n = 12), women with PCOS treated with metformin in vivo (Metformin, n = 8) and women with PCOS not exposed to metformin (PCOS, n = 8). The study was conducted over a period of 3 years between 2007 and 2010.
  
  
  METHODS
  Rotterdam criteria were used for the diagnosis of PCOS; all subjects were matched for age, BMI and baseline FSH. Individual patient cultures were undertaken with cells incubated in a validated, physiological, serum-free culture medium containing doses of 0-6 mM glucose and 0-100 ng/ml insulin for 6 h and 144 h to quantify the impact of treatments on acute and long-term metabolism, respectively, and progesterone production. The metabolite content of spent media was measured using spectrophotometric plate reader assay. The progesterone content of spent media was measured by enzyme-linked immunosorbent assay. Viable GC number was quantified after 144 h of culture by the vital dye Neutral Red uptake assay.
  
  
  RESULTS
  Granulosa cells from women with PCOS pathology revealed reduced pyruvate production and preferential lactate production in addition to their reduced glucose uptake during cultures (P < 0.05). Metformin pretreatment alleviated this metabolic lesion (P < 0.05) and enhanced cell proliferation in vitro (P < 0.05), but cells retained a significantly reduced capacity for progesterone synthesis compared with controls (P < 0.05).
  
  
  CONCLUSIONS
  Although significant treatment effects were detected in this small cohort, further studies are required to underpin the molecular mechanisms of the effect of metformin on GCs.
  
  
  CONCLUSIONS
  The individual patient culture strategy combined with multifactorial experimental design strengthens the biological interpretation of the data. Collectively, these results support the notion that there is an inherent impairment in progesterone biosynthetic capacity of the GCs from women with PCOS. The positive, acute metabolic effect and the negative long-term steroidogenic effect on GCs following metformin exposure in vivo may have important implications for follicular development and luteinized GC function when the drug is used in clinical practice.
  
  
  BACKGROUND
  No competing interests. This work was supported by the UK Medical Research Council Grant Reference number G0800250.
authors.affiliation: The Leeds Centre for Reproductive Medicine, Seacroft Hospital, York Road, Leeds LS14 6UH, UK maruthini@yahoo.co.uk.
authors.forename: D
authors.initials: D
authors.lastname: Maruthini
authors.affiliation: Division of Reproduction and Early Development, Leeds Institute for Genetics, Health and Therapeutics, University of Leeds, Clarendon Way, Leeds LS2 9JT, UK.
authors.forename: S E
authors.initials: SE
authors.lastname: Harris
authors.affiliation: Department of Clinical Biochemistry, Leeds Teaching Hospitals NHS Trust, Leeds General Infirmary, Great George Street, Leeds LS1 3EX, UK.
authors.forename: J H
authors.initials: JH
authors.lastname: Barth
authors.affiliation: The Leeds Centre for Reproductive Medicine, Seacroft Hospital, York Road, Leeds LS14 6UH, UK.
authors.forename: A H
authors.initials: AH
authors.lastname: Balen
authors.affiliation: Division of Human Development at Nottingham University Hospitals NHS Trust, Department of Obstetrics and Gynaecology, Queen's Medical Centre Campus, Nottingham NG7 2UH, UK.
authors.forename: B K
authors.initials: BK
authors.lastname: Campbell
authors.affiliation: Division of Reproduction and Early Development, Leeds Institute for Genetics, Health and Therapeutics, University of Leeds, Clarendon Way, Leeds LS2 9JT, UK.
authors.forename: H M
authors.initials: HM
authors.lastname: Picton
mesh_terms: D000328:Adult; D016022:Case-Control Studies; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D005260:Female; D015571:Follicular Fluid / Q000378:metabolism; D005947:Glucose / Q000378:metabolism*; D006107:Granulosa Cells / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D007328:Insulin / Q000378:metabolism; D019344:Lactic Acid / Q000096:biosynthesis; D008687:Metformin / Q000494:pharmacology / Q000627:therapeutic use*; D006080:Ovarian Follicle / Q000033:anatomy & histology / Q000187:drug effects / Q000378:metabolism; D011085:Polycystic Ovary Syndrome / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D011374:Progesterone / Q000096:biosynthesis*; D019289:Pyruvic Acid / Q000378:metabolism
chemical_list: D007328:Insulin; D019344:Lactic Acid; D011374:Progesterone; D019289:Pyruvic Acid; D008687:Metformin; D005947:Glucose
keywords: energy metabolism; human granulosa cells; metformin; polycystic ovary syndrome; progesterone