Prenatal metformin treatment improves ovarian function in offspring of obese rats

in Journal of Endocrinology
Correspondence should be addressed to G Cruz:

*(D Álvarez and K Ceballo contributed equally to this work)

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Maternal obesity causes a wide range of impairment in offspring, such as metabolic and reproductive dysfunctions. We previously demonstrated that female offspring of obese rats have increased serum estradiol levels during early postnatal life, probably because of decreased hepatic cytochrome P450 3A2 levels, which could lead to early onset of puberty and polycystic ovary condition in adulthood. Using metformin during pregnancy and nursing to improve the metabolic status of obese mothers could prevent the sequence of events that lead to an increase in postnatal serum estradiol levels in female offspring and, hence, reproductive dysfunction. We found that metformin prevented an increase in serum estradiol levels at postnatal day 14 in female offspring of obese mothers, which was associated with a restoration of hepatic cytochrome P450 3A2 levels to control values. Treatment using metformin could not prevent advanced puberty, but we observed that the number of antral follicles, follicular cysts and multi-oocyte follicles returned to control values in the female offspring of obese mothers treated with metformin. We also observed an increase in the levels of norepinephrine and the norepinephrine metabolite 3-methoxy-4-hydroxyphenylglycol in the ovaries, indicating increased sympathetic activity in female offspring induced by an obesogenic uterine environment. We found that this effect was prevented by metformin administration. From the results of this study, we concluded that metformin administration to obese mothers during pregnancy and nursing partially prevents ovarian dysfunction in female offspring during adulthood.


      Society for Endocrinology

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    Experimental design. Adult virgin rats were divided into three groups: one group received a CD and the other two groups received an HFD. The HFD was administered for 1 month before pregnancy, during pregnancy and until weaning. One of the two groups fed the HFD received metformin in tap water, starting from 1 week before mating, during pregnancy and until weaning. The offspring of all groups were fed a CD from weaning until the end of the experiments (PND 60). Superior boxes represent mothers, and inferior boxes represent offspring. The dashed fill represents the time of HFD consumption alone, while the gray fill represents the simultaneous consumption of HFD and metformin.

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    Offspring body weight. (A) Body weight curve from birth until PND 60 and (B, C, D and E) average body weight at PNDs 1, 7, 14 and 60. Data are shown as mean ± s.e.m. **P < 0.01 and ****P < 0.0001 for Control vs HFD; #P < 0.05, and ###P < 0.001 for HFD vs HFD + MET; ++P < 0.01 and ++++P < 0.0001 for Control vs HFD + MET. Control, N = 7; HFD, N= 5; HFD + MET, N = 4.

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    Puberty onset in offspring of mothers fed a CD, an HFD or HFD + MET. (A) Mean day of age of vaginal opening, (B) percentage of rats reaching vaginal opening through time and (C) mean day of age of first estrus. Data are shown as mean ± s.e.m. *P < 0.05 for Control vs HFD; +P < 0.05 and ++P < 0.01 for Control vs HFD + MET. Control, N = 7; HFD, N = 5; HFD + MET, N = 4.

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    Hepatic serum estradiol and CYP3A2 levels. (A) Serum estradiol and (C) serum estriol at PND 14; N = 4 for group. (B) Serum estradiol and (D) serum estriol at PND 60; N = 5 for control and HFD and N = 4 for HFD + MET. (E) Hepatic CYP3A2 level at PND 14; N = 4 for each experimental group. (F) Hepatic CYP3A2 level at PND 60; N = 5 for control and HFD and N = 4 for HFD + MET. (G) Representative image of CYP3A2 western blot at PND 14. (H) Representative image of CYP3A2 western blot at PND 60. Data are shown as mean ± s.e.m. *P < 0.05 for control vs HFD; #P < 0.05 for HFD vs HFD + MET.

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    Sympathetic activity in the ovary. (A) Total ovarian NE levels per ovary at PND 60 and (B) total ovarian MHPG levels per ovary at PND 60. Control, N = 7; HFD, N = 5; HFD + MET, N = 4. Data are shown as mean ± s.e.m. **P < 0.01 for Control vs HFD; #P < 0.05, and ##P < 0.01 for HFD vs HFD + MET.

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    Ovarian follicular development. The counting of ovarian follicles at different stages is shown. Black bars: offspring of Control rats; white bars: offspring of HFD rats; gray bars: offspring of HFD + MET rats. (A) Healthy antral follicles, (B) size distribution of healthy antral follicles, (C) atretic antral follicles, (D) corpora lutea, (E) MOFs, and (F) follicular cysts. N = 4 for Control, HFD and HFD + MET. Data are shown as mean ± s.e.m. *P < 0.05, **P < 0.01 and ***P < 0.001 for Control vs HFD; #P < 0.05 for HFD vs HFD + MET in each graph.

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    MOF and follicular cyst histology. (A and B) Oocyte nest–like structure, (C) secondary MOF, (D) healthy antral MOF and (E) follicular cyst. All the images are of ovaries from the HFD offspring.


AhimaRSDushayJFlierSNPrabakaranDFlierJS 1997 Leptin accelerates the onset of puberty in normal female mice. Journal of Clinical Investigation 99 391395. (

AmbrosettiVGuerraMRamirezLAReyesAAlvarezDOlguinSGonzalez-MananDFernandoisDSotomayor-ZarateRCruzG 2016 Increase in endogenous estradiol in the progeny of obese rats is associated with precocious puberty and altered follicular development in adulthood. Endocrine 53 258270. (

CatalanoPde MouzonSH 2015 Maternal obesity and metabolic risk to the offspring: why lifestyle interventions may have not achieved the desired outcomes. International Journal of Obesity 39 642649. (

ConnorKLVickersMHBeltrandJMeaneyMJSlobodaDM 2012 Nature, nurture or nutrition? Impact of maternal nutrition on maternal care, offspring development and reproductive function. Journal of Physiology 590 21672180. (

CruzGBarraRGonzalezDSotomayor-ZarateRLaraHE 2012 Temporal window in which exposure to estradiol permanently modifies ovarian function causing polycystic ovary morphology in rats. Fertility and Sterility 98 12831290. (

CheongYSadekKHBruceKDMacklonNCagampangFR 2014 Diet-induced maternal obesity alters ovarian morphology and gene expression in the adult mouse offspring. Fertility and Sterility 102 899907. (

ChunSYEisenhauerKMMinamiSBilligHPerlasEHsuehAJ 1996 Hormonal regulation of apoptosis in early antral follicles: follicle-stimulating hormone as a major survival factor. Endocrinology 137 14471456. (

DissenGAGarcia-RudazCParedesAMayerCMayerhoferAOjedaSR 2009 Excessive ovarian production of nerve growth factor facilitates development of cystic ovarian morphology in mice and is a feature of polycystic ovarian syndrome in humans. Endocrinology 150 29062914. (

Engen HanemLGStridsklevSJuliussonPBSalvesenORoelantsMCarlsenSMOdegardRVankyE 2018 Metformin use in PCOS pregnancies increases the risk of offspring overweight at 4 years of age; follow-up of two RCTs. Journal of Clinical Endocrinology and Metabolism 103 16121621. (

FernandoisDLaraHEParedesAH 2012 Blocking of beta-adrenergic receptors during the subfertile period inhibits spontaneous ovarian cyst formation in rats. Hormone and Metabolic Research 44 682687. (

GarrisDR 2004 Variable onset determinants and consequences of diabetes (db/db) obesity mutation expression: adrenergic promotion of utero-ovarian dysfunction. Hormone and Metabolic Research 36 312318. (

GlintborgDPetersenMHRavnPHermannAPAndersenM 2016 Comparison of regional fat mass measurement by whole body DXA scans and anthropometric measures to predict insulin resistance in women with polycystic ovary syndrome and controls. Acta Obstetricia et Gynecologica Scandinavica 95 12351243. (

HarrisKDesaiNGuptaMXueXChatterjeePKRochelsonBMetzCN 2016 The effects of prenatal metformin on obesogenic diet-induced alterations in maternal and fetal fatty acid metabolism. Nutrition and Metabolism 13 55. (

HealDJProwMRBuckettWR 1989 Measurement of 3-methoxy-4-hydroxyphenylglycol (MHPG) in mouse brain by h.p.l.c. with electrochemical detection, as an index of noradrenaline utilisation and presynaptic alpha 2-adrenoceptor function. British Journal of Pharmacology 96 547556. (

HeiderUPedalISpanel-BorowskiK 2001 Increase in nerve fibers and loss of mast cells in polycystic and postmenopausal ovaries. Fertility and Sterility 75 11411147. (

HirshfieldANMidgleyARJr 1978 The role of FSH in the selection of large ovarian follicles in the rat. Biology of Reproduction 19 606611. (

JanssonTPowellTL 2007 Role of the placenta in fetal programming: underlying mechanisms and potential interventional approaches. Clinical Science 113 113. (

JeffersonWNCouseJFPadilla-BanksEKorachKSNewboldRR 2002 Neonatal exposure to genistein induces estrogen receptor (ER)alpha expression and multioocyte follicles in the maturing mouse ovary: evidence for ERbeta-mediated and nonestrogenic actions. Biology of Reproduction 67 12851296. (

KezelePSkinnerMK 2003 Regulation of ovarian primordial follicle assembly and development by estrogen and progesterone: endocrine model of follicle assembly. Endocrinology 144 33293337. (

KippJLKilenSMBristol-GouldSWoodruffTKMayoKE 2007 Neonatal exposure to estrogens suppresses activin expression and signaling in the mouse ovary. Endocrinology 148 19681976. (

KnehansAWRomsosDR 1983 Norepinephrine turnover in obese (ob/ob) mice: effects of age, fasting, and acute cold. American Journal of Physiology 244 E567E574. (

KnehansAWRomsosDR 1984 Effects of diet on norepinephrine turnover in obese (ob/ob) mice. Journal of Nutrition 114 20802088. (

KovoMKogmanNOvadiaONakashIGolanAHoffmanA 2008 Carrier-mediated transport of metformin across the human placenta determined by using the ex vivo perfusion of the placental cotyledon model. Prenatal Diagnosis 28 544548. (

LansdownAReesDA 2012 The sympathetic nervous system in polycystic ovary syndrome: a novel therapeutic target? Clinical Endocrinology 77 791801. (

LaraHEDissenGALeytonVParedesAFuenzalidaHFiedlerJLOjedaSR 2000 An increased intraovarian synthesis of nerve growth factor and its low affinity receptor is a principal component of steroid-induced polycystic ovary in the rat. Endocrinology 141 10591072. (

LiLLiuDWYanHYWangZYZhaoSHWangB 2016 Obesity is an independent risk factor for non-alcoholic fatty liver disease: evidence from a meta-analysis of 21 cohort studies. Obesity Reviews 17 510519. (

LinYJTsaiCCHuangLTSheenJMTiaoMMYuHRChenCCTainYL 2017 Detrimental effect of maternal and post-weaning high-fat diet on the reproductive function in the adult female offspring rat: roles of insulin-like growth factor 2 and the ovarian circadian clock. Journal of Assisted Reproduction and Genetics 34 817826. (

LookinglandKJIrelandLMGunnetJWManzanaresJTianYMooreKE 1991 3-Methoxy-4-hydroxyphenylethyleneglycol concentrations in discrete hypothalamic nuclei reflect the activity of noradrenergic neurons. Brain Research 559 8288. (

MaharjanSSerovaLSabbanEL 2005 Transcriptional regulation of tyrosine hydroxylase by estrogen: opposite effects with estrogen receptors alpha and beta and interactions with cyclic AMP. Journal of Neurochemistry 93 15021514. (

Martinez-PintoJPiquerBTiszavariMLaraHE 2018 Neonatal exposure to estradiol valerate reprograms the rat ovary androgen receptor and anti-Mullerian hormone to a polycystic ovary phenotype. Reproductive Toxicology 75 127135. (

MouralidaraneASoedaJVisconti-PugmireCSamuelssonAMPomboJMaragkoudakiXButtASaraswatiRNovelliMFusaiGet al. 2013 Maternal obesity programs offspring nonalcoholic fatty liver disease by innate immune dysfunction in mice. Hepatology 58 128138. (

ObenJAMouralidaraneASamuelssonAMMatthewsPJMorganMLMcKeeCSoedaJFernandez-TwinnDSMartin-GronertMSOzanneSEet al. 2010 Maternal obesity during pregnancy and lactation programs the development of offspring non-alcoholic fatty liver disease in mice. Journal of Hepatology 52 913920. (

OrioFMuscogiuriGNeseCPalombaSSavastanoSTafuriDColarietiGLa SalaGColaoAYildizBO 2016 Obesity, type 2 diabetes mellitus and cardiovascular disease risk: an uptodate in the management of polycystic ovary syndrome. European Journal of Obstetrics and Gynecology and Reproductive Biology 207 214219. (

PadmanabhanVVeiga-LopezA 2011 Developmental origin of reproductive and metabolic dysfunctions: androgenic versus estrogenic reprogramming. Seminars in Reproductive Medicine 29 173186. (

PaliyOPiyathilakeCJKozyrskyjACelepGMarottaFRastmaneshR 2014 Excess body weight during pregnancy and offspring obesity: potential mechanisms. Nutrition 30 245251. (

PellattLRiceSDilaverNHeshriAGaleaRBrincatMBrownKSimpsonERMasonHD 2011 Anti-Mullerian hormone reduces follicle sensitivity to follicle-stimulating hormone in human granulosa cells. Fertility and Sterility 96 12461251.e1241. (

PeplingME 2012 Follicular assembly: mechanisms of action. Reproduction 143 139149. (

PeplingMESundmanEAPattersonNLGephardtGWMedicoLJr.WilsonKI 2010 Differences in oocyte development and estradiol sensitivity among mouse strains. Reproduction 139 349357. (

PicutCADixonDSimonsMLStumpDGParkerGARemickAK 2015 Postnatal ovary development in the rat: morphologic study and correlation of morphology to neuroendocrine parameters. Toxicologic Pathology 43 343353. (

RahmouniK 2010 Leptin-induced sympathetic nerve activation: signaling mechanisms and cardiovascular consequences in obesity. Current Hypertension Reviews 6 104209. (

RobertsVHFriasAEGroveKL 2015 Impact of maternal obesity on fetal programming of cardiovascular disease. Physiology 30 224231. (

RosaESAGuimaraesMAPadmanabhanVLaraHE 2003 Prepubertal administration of estradiol valerate disrupts cyclicity and leads to cystic ovarian morphology during adult life in the rat: role of sympathetic innervation. Endocrinology 144 42894297. (

RosenthalMRothJStorrBZeisbergerE 1996 Fever response in lean (Fa/-) and obese (fa/fa) Zucker rats and its lack to repeated injections of LPS. Physiology and Behavior 59 787793. (

SalomakiHVahataloLHLaurilaKJappinenNTPenttinenAMAilanenLIlyasizadehJPesonenUKouluM 2013 Prenatal metformin exposure in mice programs the metabolic phenotype of the offspring during a high fat diet at adulthood. PLoS ONE 8 e56594. (

SalomakiHHeinaniemiMVahataloLHAilanenLEerolaKRuohonenSTPesonenUKouluM 2014 Prenatal metformin exposure in a maternal high fat diet mouse model alters the transcriptome and modifies the metabolic responses of the offspring. PLoS ONE 9 e115778. (

SamuelssonAMMorrisAIgoshevaNKirkSLPomboJMCoenCWPostonLTaylorPD 2010 Evidence for sympathetic origins of hypertension in juvenile offspring of obese rats. Hypertension 55 7682. (

ShalitinSPhillipM 2003 Role of obesity and leptin in the pubertal process and pubertal growth--a review. International Journal of Obesity and Related Metabolic Disorders 27 869874. (

SlobodaDMHowieGJPleasantsAGluckmanPDVickersMH 2009 Pre- and postnatal nutritional histories influence reproductive maturation and ovarian function in the rat. PLoS ONE 4 e6744. (

Sotomayor-ZarateRDorfmanMParedesALaraHE 2008 Neonatal exposure to estradiol valerate programs ovarian sympathetic innervation and follicular development in the adult rat. Biology of Reproduction 78 673680. (

TaylorPDSamuelssonAMPostonL 2014 Maternal obesity and the developmental programming of hypertension: a role for leptin. Acta Physiologica 210 508523. (

TellecheaMLMensegueMFPirolaCJ 2017 The association between high fat diet around gestation and metabolic syndrome-related phenotypes in rats: a systematic review and meta-analysis. Scientific Reports 7 5086. (

TsoulisMWChangPEMooreCJChanKAGohirWPetrikJJVickersMHConnorKLSlobodaDM 2016 Maternal high-fat diet-induced loss of fetal oocytes is associated with compromised follicle growth in adult rat offspring. Biology of Reproduction 94 94. (

ValladaresMPlaza-ParrochiaFLepezMLopezDGablerFGayanPSelmanAVegaMRomeroC 2017 Effect of estradiol on the expression of angiogenic factors in epithelial ovarian cancer. Histology and Histopathology 32 11871196. (

VisserJADurlingerALPetersIJvan den HeuvelERRoseUMKramerPde JongFHThemmenAP 2007 Increased oocyte degeneration and follicular atresia during the estrous cycle in anti-Mullerian hormone null mice. Endocrinology 148 23012308. (

WojtkiewiczJJanaBKozlowskaACraytonRMajewskiMZaleckiMBaranowskiWRadziszewskiP 2014 Innervation pattern of polycystic ovaries in the women. Journal of Chemical Neuroanatomy 61–62 147152. (

YorkDAMarchingtonDHoltSJAllarsJ 1985 Regulation of sympathetic activity in lean and obese Zucker (fa/fa) rats. American Journal of Physiology 249 E299E305. (

ZambranoEIbanezCMartinez-SamayoaPMLomas-SoriaCDurand-CarbajalMRodriguez-GonzalezGL 2016 Maternal obesity: lifelong metabolic outcomes for offspring from poor developmental trajectories during the perinatal period. Archives of Medical Research 47 112. (


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