Division of Human Reproduction, IVF Unit, First Department of Obstetrics and Gynaecology, Alexandra Hospital, Athens University Medical School, 11528 Athens, Greece.
Endocrinology. 2013 Jan;154(1):222-31. doi: 10.1210/en.2012-1838. Epub 2012 Dec 4.
In vitro growth systems of preantral follicles allow studying the effect of various endocrine, paracrine, and autocrine factors on follicular growth and oocyte maturation. CRH is a 41-amino-acid neuropeptide responsible for endocrine, autonomic, immunological, and behavioral responses of mammals to stress and has two receptors, CRH receptor type 1 (CRH-R1) and CRH-R2. Antalarmin, a CRH-R1 antagonist, has been used to elucidate the role of CRH in stress, inflammation, and reproduction. The present study describes in vitro growth of mouse preantral follicles, early embryo development, and steroidogenesis in the presence of CRH and its antagonist antalarmin. We cultured 732 follicles in control media, 1306 in CRH 10(-7) mol/liter, and 1202 in CRH 10(-7) plus antalarmin 10(-6) mol/liter. The culture medium was assayed on alternate days for 17β-estradiol, progesterone, and β-human chorionic gonadotropin. Total RNA was extracted from preantral follicles as well as early preimplantation embryos and was assessed by real-time RT-PCR for the expression of CRH-R1 and CRH-R2 mRNAs. Hormone analysis showed that the CRH group had lower levels of 17β-estradiol, progesterone, and β-human chorionic gonadotropin as the culture progressed, in comparison with the other two groups. RT-PCR demonstrated the presence of CRH-R1 and CRH-R2 in all stages of preantral follicle culture. Morula/blastocyst-stage embryos expressed only CRH-R1. In conclusion, CRH has an inhibitory effect on in vitro fertilized oocytes, resulting from cultured preantral follicles at all stages of preimplantation embryo development. Furthermore, the presence of CRH in the culture medium inhibits steroidogenesis by preantral mouse follicles cultured in vitro.
在体外培养的原始卵泡生长系统可以研究各种内分泌、旁分泌和自分泌因素对卵泡生长和卵母细胞成熟的影响。CRH 是一种由 41 个氨基酸组成的神经肽，负责哺乳动物对压力的内分泌、自主、免疫和行为反应，有两种受体，CRH 受体 1（CRH-R1）和 CRH-R2。抗促肾上腺皮质激素释放因子，一种 CRH-R1 拮抗剂，已被用于阐明 CRH 在应激、炎症和生殖中的作用。本研究描述了在 CRH 及其拮抗剂安塔拉明存在的情况下，小鼠原始卵泡的体外生长、早期胚胎发育和类固醇生成。我们在对照培养基中培养了 732 个卵泡，在 CRH10(-7)mol/l 中培养了 1306 个卵泡，在 CRH10(-7)加安塔拉明 10(-6)mol/l 中培养了 1202 个卵泡。每隔一天检测培养基中的 17β-雌二醇、孕酮和β-人绒毛膜促性腺激素。从原始卵泡和早期植入前胚胎中提取总 RNA，并用实时 RT-PCR 评估 CRH-R1 和 CRH-R2 mRNA 的表达。激素分析表明，与其他两组相比，CRH 组随着培养的进行，17β-雌二醇、孕酮和β-人绒毛膜促性腺激素的水平较低。RT-PCR 显示 CRH-R1 和 CRH-R2 在原始卵泡培养的所有阶段均存在。桑椹胚/囊胚阶段的胚胎仅表达 CRH-R1。总之，CRH 对体外受精的卵母细胞有抑制作用，这是由于培养的原始卵泡在植入前胚胎发育的所有阶段。此外，培养基中存在 CRH 会抑制体外培养的小鼠原始卵泡的类固醇生成。
