Identification of proteins in pooled human follicular fluid which suppress follicular response to gonadotropins.
Identification of proteins in pooled human follicular fluid which suppress follicular response to gonadotropins.
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鉴定汇集的人卵泡液中抑制卵泡对促性腺激素反应的蛋白质。
DOI：
10.1210/jcem-56-1-35
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发表时间：
1983
期刊：
影响因子：
--
通讯作者：
O. Kling
中科院分区：
文献类型：
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作者：
G. diZerega;R. Marrs;P. Roche;J. Campeau;O. Kling
为了评估非甾体卵泡液蛋白在卵泡发生中的作用，使用 10,000 分子量排阻膜将汇集的人卵泡液中的 10-55% 饱和硫酸铵部分对 0.025 M Tris/HCl (pH 7.5) 进行透析，然后通过琼脂糖固定化纺织染料。通过垂体切除、己烯雌酚处理的 25 日龄雌性大鼠中人绝经期促性腺激素（2 U 人 LH/FSH·天）的测试分数抑制、诱导卵巢重量和血清雌二醇增加来确定活性。在用 KCl（1.5 M，pH 6.8）从 Orange A 柱洗脱的材料（2 ml）中发现对卵巢重量增加的特异性抑制（89 +/- 6.8% SEM）。通过标准化 Sephadex G-50 柱洗脱的 Orange A 结合材料的抑制活性对应于 13,000-25,000 的分子量。 Orange A 结合材料的 Sephadex G-15 支持床或两性电解质置换色谱等电聚焦显示在 pH 3.5-4.5 和 6.5-7.0 下具有抑制活性。在添加 α-甲基-D-甘露糖苷（2 M，pH 7）之前或之后，通过 Concanavalin A-Sepharose 4B 柱洗脱的类似级分中没有发现明显的活性。当活性级分被加热（56°C，1小时）或暴露于胰蛋白酶（10mg/100ml）时，活性丧失。当使用凝胶排阻柱通过高效液相色谱分离等份的饱和硫酸铵提取、透析的橙色A结合洗脱液时，生物测定中的活性在13,000-35,000分子量范围内恢复。尽管验证数据有待进一步研究，但人们很容易推测这种蛋白质可能是卵泡对促性腺激素反应的重要的卵巢间和/或卵巢内调节剂。
To evaluate the role of nonsteroidal, follicular fluid proteins in folliculogenesis, the 10-55% saturated ammonium sulfate fraction of pooled human follicular fluid was dialyzed against 0.025 M Tris/HCl (pH 7.5) using 10,000 molecular weight exclusion membranes, then passed through agarose immobilized textile dye. Activity was determined by test fraction inhibition of human menopausal gonadotropin (2 U human LH/FSH . day), induced ovarian weight, and serum estradiol increase in hypophysectomized, diethylstilbesterol-treated, 25-day-old female rats. Specific inhibition (89 +/- 6.8% SEM) of ovarian weight increase was found in the material (2 ml) eluted from an Orange A column with KCl (1.5 M, pH 6.8). Inhibitory activity of the Orange A-bound material, which eluted through a standardized Sephadex G-50 column, corresponded to a molecular weight of 13,000-25,000. Isoelectric focusing on a Sephadex G-15 support bed or ampholyte displacement chromatography of Orange A bound material demonstrated inhibitory activity at pH 3.5-4.5 and 6.5-7.0. No demonstrable activity was found in similar fractions eluted through a Concanavalin A-Sepharose 4B column before or after addition of alpha-methyl-D-mannoside (2 M, pH 7). When active fractions were heated (56 C, 1 h) or exposed to trypsin (10 mg/100 ml), activity was lost. When aliquots of the saturated ammonium sulfate-extracted, dialyzed, Orange A-bound eluent were separated by high performance liquid chromatography using gel exclusion columns, activity in the bioassay was recovered in the 13,000-35,000 molecular weight range. Although confirmatory data await further studies, it is tempting to speculate that this protein(s) may be an important inter- and/or intraovarian regulator of follicular response to gonadotropins.