Alterations in amounts of different forms of inhibin during follicular atresia.
Alterations in amounts of different forms of inhibin during follicular atresia.
复制标题
卵泡闭锁期间不同形式的抑制素含量的变化。
DOI：
10.1095/biolreprod50.6.1265
复制
发表时间：
1994
影响因子：
3.6
通讯作者：
Ireland,JJ
中科院分区：
文献类型：
--
作者：
Ireland,JL;Good,TE;Knight,PG;Ireland,JJ
用抑制素免疫印迹法测定牛卵泡液（bFF）中抑制素（αβ）和抑制素α亚基。在奶牛发情早期，从非排卵卵泡（直径≥6mm）中去除牛FF。孕酮(P)与雌二醇(E)浓度的比值将卵泡分为三个不同的分化和闭锁阶段：雌激素活性阶段（EA, PE < 1），闭锁阶段（PE = 1 - 100），高度闭锁阶段（PE = 100 - 100）。牛抑制素（bINH）采用免疫印迹法、与抑制素αβ二聚体和α亚基交叉反应的RIA法和αβ二聚体特异性免疫放射测定法（IRMA）进行分析。免疫印迹在每个卵泡bFF中一致检测到9种不同的bINH形式。bINH形式的平均分子质量在29 ~ 160 kDa之间，与纯化的bINH和bINH α亚基相似。bINH的主要形式≥122 kDa。在EA卵泡闭锁期间，大多数bINH形式的数量减少了几倍。然而，34 kda的形式增加了1.9倍。为了检验每种bINH形式的数量在闭锁期间是否独立变化，统计分析每种bINH形式所代表的bINH免疫印迹活性占总bINH活性的比例。结果表明，在闭锁期间，34-、68-和> 160-kDa bINH型的总免疫印迹活性比例增加，而48-、49-和122-kDa型的比例下降。大多数bINH形式的量的变化与滤泡内E浓度呈正相关，与p呈负相关。相反，34-kDa bINH形式与E呈负相关，但与p呈正相关。然而，除了29 kda形式（r= 0.72）外，大多数形式的bINH的数量与RIA结果呈相对弱相关（r< 0.41）或不相关。与RIA结果相比，IRMA测量的bINH二聚体浓度在闭锁期间增加了2.5倍，但只有34 kda形式的bINH与IRMA结果相关（r= 0.77）。我们得出结论，在雌激素活性卵泡的分化和闭锁过程中，1)34-kDa形式的bINH的产生被选择性地增强，2)大多数形式的blNH的量被差异调节，3)34-kDa bINH和E的产生没有同时增强，4)RIA和IRMA都不能准确测量bFF中所有形式抑制素的变化。
An inhibin immunoblot procedure was validated for measurement of inhibins (αβ) and inhibin α subunits in bovine follicular fluid (bFF) from individual follicles. Bovine FF was removed from nonovulatory follicles (≥ 6 mm in diameter) of cows during early diestrus. Ratio of progesterone (P) to estradiol (E) concentrations was used to classify follicles into three different stages of differentiation and atresia: estrogen-active (EA; PE ratio < 1), atretic (PE = 1–100), and highly atretic (PE = > 100). Bovine inhibin (bINH) was analyzed by an immunoblot procedure, an RIA that cross-reacts with inhibin αβ dimers and α subunits, and a two-site αβ dimer-specific immunoradiometric assay (IRMA). Immunoblots consistently detected 9 different bINH forms in bFF of each follicle. The average molecular mass for the bINH forms ranged from 29 to > 160 kDa, which was similar to those for purified bINH and bINH α subunits. The predominant bINH forms were ≥ 122 kDa. Amounts of most bINH forms decreased several fold during atresia of EA follicles. However, the 34-kDa form increased 1.9-fold. To examine whether amounts of each bINH form varied independently during atresia, proportion of total bINH immunoblot activity represented by each bINH form was statistically analyzed. The results indicated that the proportion of total immunoblot activity for the 34-, 68-, and > 160-kDa bINH forms increased during atresia, whereas proportions for the 48-, 49-, and 122-kDa forms decreased. Alterations in amounts of most bINH forms were positively correlated with intrafollicular concentrations of E and negatively correlated with P. In contrast, the 34-kDa bINH form was negatively correlated with E, but positively correlated with P. Total bINH concentrations determined by RIA decreased 3-fold during atresia. However, amounts of most forms of bINH, except the 29-kDa form (r= 0.72), were relatively weakly correlated (r< 0.41) or not correlated with the RIA results. In contrast to RIA results, concentration of dimeric forms of bINH measured by IRMA increased 2.5-fold during atresia, but only the 34-kDa form of bINH was correlated (r= 0.77) with IRMA results. We concluded that during differentiation and atresia of estrogen-active follicles, 1 ) production of a 34-kDa form of bINH was selectively enhanced, 2) amounts of most forms of blNH were differentially regulated, 3) production of 34-kDa bINH and E was not enhanced simultaneously, and 4) neither RIA nor IRMA accurately measured alterations in all forms of inhibin in bFF.