Gonadotropin suppression of apoptosis in cultured preovulatory follicles: mediatory role of endogenous insulin-like growth factor I.
Gonadotropin suppression of apoptosis in cultured preovulatory follicles: mediatory role of endogenous insulin-like growth factor I.
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DOI：
10.1210/endo.135.5.7525255
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发表时间：
1994-11
期刊：
影响因子：
4.8
通讯作者：
S. Chun;H. Billig;J. Tilly;I. Furuta;A. Tsafriri;A. Hsueh
中科院分区：
文献类型：
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作者：
S. Chun;H. Billig;J. Tilly;I. Furuta;A. Tsafriri;A. Hsueh
虽然大多数卵泡通过涉及凋亡细胞死亡的机制发生闭锁，但由于卵巢中存在异质性卵泡群体，有关闭锁的激素调节的体内研究一直很困难。在本研究中，卵泡凋亡的调节促性腺激素，胰岛素样生长因子I（IGF-I），和IGF结合蛋白3（IGFBP-3）进行了检查使用无血清培养排卵前卵泡。26日龄的未成熟大鼠接受单剂量的PMSG。两天后，收集最大的排卵前卵泡进行体外培养（有或无激素）。培养24 h后，通过[32 P]双脱氧-ATP在3 '端标记的大小分级DNA的放射自显影术分析卵泡凋亡DNA片段。在不存在激素的情况下，培养24小时后发生凋亡DNA片段的自发增加，而用人CG（hCG）或FSH处理以剂量依赖性方式抑制卵泡凋亡，0.1 μ g/ml引起60- 62%的最大抑制。hCG和FSH的联合治疗没有额外的效果。与促性腺激素一样，IGF-I和胰岛素治疗也抑制了细胞凋亡的自发发生，IGF-I比胰岛素更有效。IGFBP-3和hCG剂量依赖性地逆转hCG对细胞凋亡的抑制作用达42%，表明内源性产生的IGF-I的介导作用。IGFBP-3的加入也阻断了49%的IGF-I的抑制作用，而它不影响IGF-I激动剂或胰岛素的抑制作用。单独用IGFBP-3处理对凋亡DNA片段化没有影响。用放射免疫分析法测定培养卵泡的雌激素和孕激素水平。促性腺激素处理导致两种类固醇产生的显著刺激。相反，IGF-I治疗引起雌激素的小幅增加，但孕酮的产生减少。虽然单独用IGFBP-3处理降低了雌激素和孕酮的产生，但用IGFBP-3和hCG共处理导致雌激素产生略有下降，但孕酮产生增加。此外，IGFBP-3不影响IGF-I对类固醇产生的作用。为了进一步证实IGFBP-3通过中和内源性产生的IGF-I来阻断hCG对细胞凋亡的抑制作用的假设，进行了溶液杂交分析，并且显示hCG处理使培养卵泡中的IGF-I信使RNA水平增加1.9倍。（400字处截断摘要）
Although the majority of ovarian follicles undergo atresia through a mechanism involving apoptotic cell death, in vivo studies concerning the hormonal regulation of atresia have been difficult due to the presence of heterogeneous population of follicles in the ovary. In the present study, the regulation of follicle apoptosis by gonadotropins, insulin-like growth factor I (IGF-I), and IGF-binding protein 3 (IGFBP-3) was examined using a serum-free culture of preovulatory follicles. Immature rats at 26 days of age received a single dose of PMSG. Two days later, the largest preovulatory follicles were collected for in vitro culture with or without hormones. After 24 h of culture, follicular apoptotic DNA fragmentation was analyzed by autoradiography of size-fractionated DNA labeled at 3'-ends by [32P]dideoxy-ATP. A spontaneous increase in apoptotic DNA fragmentation occurred after 24 h of culture in the absence of hormones, whereas treatment with human CG (hCG) or FSH suppressed follicular apoptosis in a dose-dependent manner, with 0.1 microgram/ml causing maximal suppression by 60-62%. Cotreatment with hCG and FSH had no additional effect. Like gonadotropins, treatment with IGF-I and insulin also suppressed the spontaneous onset of apoptosis, with IGF-I being more effective than insulin. Cotreatment with IGFBP-3 and hCG dose-dependently reversed the suppressive effect of hCG on apoptosis by 42%, suggesting a mediatory role of endogenously produced IGF-I. The addition of IGFBP-3 also blocked the suppressive action of IGF-I by 49%, whereas it did not affect the suppressive action of an IGF-I agonist or insulin. Treatment with IGFBP-3 alone had no effect on apoptotic DNA fragmentation. Estrogen and progesterone production by the cultured follicles were also analyzed by RIA. Gonadotropin treatment resulted in a marked stimulation of the production of both steroid productions. In contrast, treatment with IGF-I caused a small increase in estrogen but decreased progesterone production. Although treatment with IGFBP-3 alone decreased both estrogen and progesterone production, cotreatment with IGFBP-3 and hCG resulted in a slight decrease in estrogen production but an increase in progesterone production. Furthermore, IGFBP-3 did not affect IGF-I action on steroid production. To further substantiate the hypothesis that IGFBP-3 blocks the suppressive effect of hCG on apoptosis by neutralizing endogenously produced IGF-I, solution hybridization analysis was performed, and hCG treatment was shown to increase IGF-I messenger RNA levels in cultured follicles by 1.9-fold.(ABSTRACT TRUNCATED AT 400 WORDS)