Centre of Reproductive Medicine and Andrology, University Hospital of Münster, Domagkstrasse 11, D-48149 Münster, Germany.
Reprod Biomed Online. 2011 Aug;23(2):196-203. doi: 10.1016/j.rbmo.2011.04.009. Epub 2011 May 12.
The study was designed to evaluate in vitro the cellular mechanisms of the single nucleotide polymorphism (SNP) p.N680S of the FSH receptor gene (FSHR) in human granulosa cells (GC) and included patients homozygous for the FSHR SNP (NN/SS) undergoing ovarian stimulation. GC were isolated during oocyte retrieval and cultured for 1–7 days. Basal oestradiol and progesterone concentrations were measured after short-term culture. The kinetics of cAMP, oestradiol and progesterone concentrations in response to various amounts of FSH were analysed in a 6–7 day culture. Basal oestradiol, but not progesterone, concentrations on day 1 of GC culture, were significantly higher in NN compared with SS (P = 0.045), but non-responsive to FSH stimulation. Immunofluorescence microscopy demonstrated the re-appearance of FSHR expression with increasing days in culture. Upon stimulation with FSH, GC cultured for 6–7 days displayed a dose-dependent increase of cAMP, oestradiol and progesterone but no difference in the EC50 values between both variants. Primary long-term GC cultures are a suitable system to study the effects of FSH in vitro. However, the experiments suggest that factors down-stream of progesterone production or external to GC might be involved in the clinically observed differences in an FSHR variant-mediated response to FSH.
本研究旨在评估人颗粒细胞（GC）中促卵泡激素受体基因（FSHR）单核苷酸多态性（SNP）p.N680S 的细胞机制，该 SNP 为纯合子（NN/SS）的患者正在接受卵巢刺激。GC 在取卵过程中分离出来，并培养 1-7 天。短期培养后测量基础雌二醇和孕酮浓度。在 6-7 天的培养中分析了不同剂量 FSH 对 cAMP、雌二醇和孕酮浓度的动力学反应。与 SS 相比，NN 组 GC 培养第 1 天的基础雌二醇浓度显著升高（P = 0.045），但对 FSH 刺激无反应。免疫荧光显微镜显示，随着培养天数的增加，FSHR 表达重新出现。经 FSH 刺激后，培养 6-7 天的 GC 显示出 cAMP、雌二醇和孕酮的剂量依赖性增加，但两种变体的 EC50 值无差异。原发性长期 GC 培养是体外研究 FSH 作用的合适系统。然而，实验表明，孕激素产生下游或 GC 以外的因素可能参与了临床观察到的 FSHR 变异体介导的 FSH 反应差异。
