孕酮诱导非洲爪蟾卵母细胞生发泡破裂(GVBD)试验方法的优化及干扰物质筛查

Optimization of Xenopus laevis Oocyte Germinal Vesicle Breakdown(GVBD) Assay and Screening of Disrupting Chemicals

非洲爪蟾卵母细胞生发泡破裂(GVBD)是生物学上研究卵细胞成熟过程及其机制的良好模型。由于GVBD受激素的调控,曾有研究将孕酮诱导的非洲爪蟾GVBD试验用于内分泌干扰物的筛查。通过预注射人体绒毛膜促性腺激素(HCG)提高非洲爪蟾卵母细胞对孕酮的敏感性、缩小个体差异,缩短试验时间等优化GVBD试验方法;并选用甲氧氯作为阳性参照物对优化的方法进行验证,最后用此方法研究了米非司酮、来曲唑、咪鲜胺3种化学物质对孕酮诱导GVBD的影响。结果发现,甲氧氯对GVBD的发生具有明显抑制作用,随着剂量的增加抑制作用增强,显示优化方法的可靠性。米非司酮作为孕酮受体拮抗剂,没有表现出抑制GVBD的效应,证明孕酮诱导爪蟾GVBD并非完全由孕酮受体(PR)介导。同时,来曲唑和咪鲜胺对孕酮诱导爪蟾GVBD没有影响,显示此两种物质在孕酮诱导GVBD过程中不具有内分泌干扰作用。

Germinal vesicle breakdown （GVBD）ofXenopus laevis oocytes is a good model for studying the process and mechanism of oocyte maturation. Because the occurrence of GVBD is regulated by hormones, progesterone-induced Xenopus GVBD assay has been used for screening endocrine disruptors. In the present study, the GVBD assay was optimized by pre-injecting human chorionic gonadotrophin （HCG） to improve the sensitivity o＇fXenopus oocytes to progesterone and reduce the difference in the sensitivity among individu-als, and by shortening the experiment time. Methoxychlor, as a positive reference, inhibited progesterone-in- duced GVBD in a concentration-dependent manner, demonstrating that the optimized assay was reliable. Mife- pristone, as a progesterone receptor antagonist, had no inhibitory effect on progesterone-induced GVBD, sug- gesting that progesterone-induced GVBD could not be mediated completely by progesterone receptor. In addi- tion, prochloraz and letrozole had no influence on Xenopus GVBD, showing no endocrine-disrupting activity in the process ofXenopus GVBD.