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12 May 2011 Oocyte and Cumulus Cell Transcripts from Cultured Mouse Follicles Are Induced to Deviate from Normal In Vivo Conditions by Combinations of Insulin, Follicle-Stimulating Hormone, and Human Chorionic Gonadotropin
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Abstract

Gonadotropins and insulin are major regulators of cell proliferation, differentiation, and survival in cultured mouse ovarian follicles. Applications of variable doses of insulin in combination with follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were studied at the gene expression level in oocytes and cumulus cells. Early preantral follicles grown over 9 days were sequentially exposed to combinations of doses of insulin, FSH, and human chorionic gonadotropin (hCG). From culture Day 1 to 6 (preantral stage), two insulin concentrations (5 ng/ml and 5 μg/ml) were combined with 10 mIU/ml FSH. From Days 6 to 9 (antral stage), the three variable gonadotropin treatments set under each insulin condition were 10 mIU/ml FSH, 25 mIU/ml FSH, and 25 mIU/ml FSH plus 3 mIU/ml hCG. The Gdf9, Bmp15, Fgf8, Dazl, Pou5f1, and Pik3ca mRNA transcripts were quantified in oocytes, and the Amh, Lhcgr, Hsd3b1, Vegfa, and Insig1 mRNA transcripts were quantified in cumulus cells. In vivo controls were unprimed and eCG (equine chorionic gonadotropin)-primed prepubertal female mice. During the preantral stage, none except the Amh transcripts was regulated by insulin. Oocyte transcripts were not affected by the variable gonadotropin treatments on the last culture day but were upregulated in the combination of high insulin plus 25 mIU/ml FSH. Under low insulin conditions, high FSH levels increased levels of Lhcgr and Vegfa expression, and hCG abated this effect. However, under high insulin conditions, hCG upregulated levels of Lhcgr, Vegfa, and Insig1 mRNA. High insulin concentrations upregulated Hsd3b1 transcripts. These results demonstrate that in an in vitro follicle culture, a near physiological insulin background yields oocyte and cumulus cell transcript levels that are more similar to those in vivo.

Gene expression in the cumulus–oocyte complex during antral growth is differentially regulated by exposure to high levels of gonadotropins, depending on the background insulin concentration in culture medium.

Flor Sánchez, Sergio Romero, and Johan Smitz "Oocyte and Cumulus Cell Transcripts from Cultured Mouse Follicles Are Induced to Deviate from Normal In Vivo Conditions by Combinations of Insulin, Follicle-Stimulating Hormone, and Human Chorionic Gonadotropin," Biology of Reproduction 85(3), 565-574, (12 May 2011). https://doi.org/10.1095/biolreprod.111.091744
Received: 15 February 2011; Accepted: 1 April 2011; Published: 12 May 2011
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