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)

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.


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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.


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