金黄地鼠腹侧前列腺来源蛋白结合精子表面的实验研究

Identification of Sperm-binding Proteins in the Ventral Prostate of the Golden Hamster

目的:探讨前列腺分泌蛋白是否可结合于精子表面。方法:以金黄地鼠作为研究对象,应用间接免疫荧光和亲和素标记的蛋白转印方法检测前列腺分泌蛋白是否可结合于精子。制备的抗前列腺粗提取物多克隆抗体,间接免疫荧光技术检测体外与前列腺分泌物孵育的附睾精子,以及体内分别与含前列腺及去除前列腺雄鼠交配后收集的子宫腔内和输卵管腔内精子的前列腺成分抗原。前列腺提取物经电泳分离转膜后和生物素标记附睾精子膜蛋白作用,测定在体外前列腺分泌蛋白能否与附睾精子膜结合。实验分为对照组、附属性腺全去组、腹侧前列腺组、去除腹侧前列腺组。结果:前列腺抗原成分的免疫反应局限在精子体中部表面,而精子头、颈部未见阳性反应。在体外(80±5)%附睾精子与前列腺分泌蛋白结合,在体内与对照组雄鼠交配后收集的子宫腔内精子与前列腺分泌蛋白结合精子阳性率为(30.0±4.6)%,与去除腹侧前列腺组(3.6±1.4)%比较差异有显著性(P<0.01),在体内与对照组雄鼠交配后收集的输卵管腔内精子与前列腺分泌蛋白结合精子阳性率为(16.0±3.6)%,与去除腹侧前列腺组精子(3.2±1.4)%比较差异有显著性(P<0.01)。前列腺分泌蛋白的电泳印记膜与生物素标记附睾精子膜蛋白孵育后显色分析结果可见5条阳性反应带。结论:金黄地鼠的前列腺分泌蛋白可结合于精子体中部表面,前列腺分泌蛋白中至少有5个组分可与精子膜蛋白结合。

Objective： To investigate the binding of secretory proteins in the ventral prostate to the surface of sperm. Methods：We used different techniques to demonstrate the possibility of ventral prostate secretory proteins binding to sperm in golden hamsters. Polyelonal antibodies against crude secretion of the ventral prostate cultured in rabbits were used to detect the antigens in hamster epididyreal, uterine and oviduetal spermatozoa by indirect immunofluoreseenee technique. The uterine and oviduetal spermatozoa were collected after mating with the males with or without ventral prostate glands. The ventral prostate secretory proteins were isolated and trans- blotted to the membrane, which was incubated with the biotinylated epididymal sperm membrane proteins, and then the biotinylated binding proteins were stained. Results ： An immunoreaetion restricted to the middle piece was observed in the sperm incubated with the ventral prostate secretion and ejaculated sperm recovered from the uteri and oviducts. The rate of the epididymal sperm bound with the ventral prostate secretory proteins was （ 80 ± 5 ） % , and the rats of the sperm binding to the ventral protstae secretory proteins were （ 30.0 ± 4.6） % from the uterus and （ 16.0 ± 3.6） % from the oviduct after mating with the males with ventral prostate glands, significantly higher than after mating with those without prostate glands （P 〈0.01 ）. Five bands were identified by Western blot analysis in vitro of the ventral prostate secretory proteins incubated with biotinylated epididymal sperm membrane proteins. Conclusion： The present data indicate that ventral prostate secretory proteins bind to the middle piece of sperm in golden hamsters.