Aybar M J, Genta S B, Villecco E I, Riera A N, Sánchez S S
Departamento de Biología del Desarrollo, Instituto Superior de Investigaciones Biológicas (INSIBIO), Tucumán, Argentina.
Int J Dev Biol. 1996 Oct;40(5):997-1008.
In the present study, we analyzed the localization of vitronectin-like protein in oocytes during oogenesis as well as in the serum and liver tissue of the amphibian Bufo arenarum. Vitronectin-like protein was purified from serum by heparin-affinity chromatography and showed to have the two biological properties in common with most animal vitronectins (VN): heparin binding activity and an RGD-dependent cell-spreading activity. SDS-PAGE of vitronectin-like protein revealed that it consists of two bands of 64 kDa and 72 kDa, while immunoblotting analyses showed that this protein strongly cross-reacts with two monoclonal antibodies against human VN. No immunofluorescent staining of vitronectin-like protein was observed in previtellogenic oocytes (stages I and II). In vitellogenic oocytes (stages III, IV and V) fluorescence was observed in the cortical cytoplasm localized in yolk platelets, extending concomitantly with the vitellogenic process. When we examined the yolk platelet formation pathway by immunoelectron microscopy, gold particles indicated that vitronectin-like protein was located on the yolk platelet precursors: multivesicular bodies and primordial yolk platelets. Gold particles also were seen sparsely distributed in all oocyte investing layers. The mean serum vitronectin-like protein concentration in amphibian animals was 127.8 +/- 11.6 micrograms/ml in adult males and 181.5 +/- 14.3 micrograms/ml in adult females. Serum vitronectin-like protein of males and females was susceptible to hormonal stimulation (17-beta estradiol). These results suggest that vitronectin-like protein is stored in the yolk platelets and may be involved in the later events of amphibian development.
在本研究中，我们分析了类玻连蛋白在非洲蟾蜍卵子发生过程中的卵母细胞以及血清和肝脏组织中的定位。通过肝素亲和层析从血清中纯化出类玻连蛋白，并显示其具有与大多数动物玻连蛋白（VN）相同的两种生物学特性：肝素结合活性和RGD依赖性细胞铺展活性。类玻连蛋白的SDS-PAGE显示它由64 kDa和72 kDa的两条带组成，而免疫印迹分析表明该蛋白与两种抗人VN的单克隆抗体强烈交叉反应。在卵黄发生前的卵母细胞（I期和II期）中未观察到类玻连蛋白的免疫荧光染色。在卵黄发生的卵母细胞（III期、IV期和V期）中，在位于卵黄小板的皮质细胞质中观察到荧光，其随着卵黄发生过程而延伸。当我们通过免疫电子显微镜检查卵黄小板形成途径时，金颗粒表明类玻连蛋白位于卵黄小板前体上：多囊泡体和原始卵黄小板。金颗粒也稀疏地分布在所有卵母细胞包被层中。两栖动物成年雄性血清中类玻连蛋白的平均浓度为127.8±11.6微克/毫升，成年雌性为181.5±14.3微克/毫升。雄性和雌性的血清类玻连蛋白对激素刺激（17-β雌二醇）敏感。这些结果表明类玻连蛋白储存在卵黄小板中，可能参与两栖动物发育的后期事件。
