摘要
用兔抗促黄体素生成素受体(LHR)或称绒毛膜促性腺激素受体(CGR)、雌激素受体(ER)和孕激素受体(PR)的抗体对LHR,ER和PR进行免疫组织化学定位。目的在于揭示外源性促性腺激素(鲤鱼脑垂体提取物,CPE和人绒毛膜促性腺激素,hCG)诱发日本鳗鲡卵子发生和卵母细胞成熟的内分泌机制。结果表明,注射激素前后卵巢发育和卵子发生出现了十分显著的变化。卵巢组织学切片观察显示激素处理前鳗鲡卵巢发育处于卵黄发生早期,卵母细胞平均直径(220±0.01)μm。第一次注射这两种激素后10d,实验组卵母细胞中卵黄核分散在核的周围,核仁数量显著增加,多达18～20个左右,而对照组8～10个。第3和4次激素处理后,卵母细胞发育进入卵黄发生早-中期至中期,卵黄颗粒数量增加。第6和7次激素处理后,卵母细胞进入卵黄发生中后期到成熟期,卵母细胞胞质中充满卵黄颗粒,胞径和核径增加,分别为(570±1.39)μm和(128±1.19)μm,而对照组没有变化。其次,免疫组织化学染色结果显示LHR、ER和PR均定位在鳗鲡卵巢中卵母细胞胞质、核膜、核质、卵被膜和体细胞上。这里值得指出的是,从第3次和第4次激素处理后,这三种受体的定位各有特点,LHR免疫活性主要定位在卵黄发生中期至中后期卵母细胞的体细胞(卵被膜内层为滤泡细胞和外层为膜细胞)的胞质,显示强的免疫阳性反应,而ER和PR则定位在卵母细胞的核内。这些结果首次揭示了外源激素(LH、FSH和hCG)诱导雌鳗卵母细胞发育和成熟的内分泌机制:首先,LH和hCG与体细胞膜上的LH受体结合,刺激其合成和分泌E2,接着E2与卵母细胞ER核受体结合,激发卵黄发生期卵母细胞生长发育和合成卵黄颗粒。另外,PR可能介导17α-羟基孕酮在卵母细胞的基因效应使之最后成熟。本研究提供确凿的形态学证据。
Immunohistochemical localization of LHR,ER and PR was performed using rabbit anti-serum against leuteinizing hormone receptor （LHR or choriogonadotropin receptor CGR）,estrogen receptor （ER） and progesterone receptor（PR）.The aim of study was to reveal the endocrine mechanism of exogenous gonadotropin （CPE and hCG） inducing oogenesis and oocytes maturation in ovary of Japanese eel.The results showed that the ovary development and oogenesis in Japanese eel displayed very marked changes before and after hormones injection.The histological section showed at the early phase of vitellogenesis stage before hormone treatment,oocytes average diameter is about （220±0.01）μm,ten days after the first injection of the combination of two gonadotropins.It differed from control group,yolk nucleus scattered in the surrounding of nucleus and number of nucleolus increased obviously,about 18-20 nucleoli,and about 8-10 nucleoli in the control group..After the third and fourth hormones treatment,the oocytes entered in the early-mid phase and middle phase of vitellogenesis stage and the number yolk granuleus was increased.After the sixth and seventh hormone treatment,the oocytes entered mature stage from mid-late phase of vitellogenesis stage,the cytoplasm of oocyte were full of yolk granules,cellular diameter and nuclear diameter were increased,about （570±1.39）μm and （128±1.19）μm,respectively,while no change in the control group.At the same time,the staining results of immunohistochemistry showed that LHR,ER and PR were located on the cytoplasm,nuclear membrane,nucleoplasm,egg envelope and somatic cell.It was worth to note that localization characteristic of the three receptors after the third and furth hormone treatment,LHR immunoreactivity was located mainly in the cytoplasm of somatic cells（follicle cell at inner layer and thecal cell outside）of oocyte and showed strong immuno-positive reaction,while ER and PR were located in the nucleus.Those results revealed that the endocrine mechanism of exogenous hormones （LH,FSH and hCG）inducing the development and maturation of oocyte in female eel were as follows：first, LH and hCG bound with LH receptor on the somatic cells,which stimulated somatic cells synthesis and secretion estradiol-17β.Then,E2 bound with its nuclear receptor in oocyte,stimulating growth and development as well as synthesis of yolk granules in the vitellogenesis stage oocyte.In addition,PR might mediate 17α-hydroxyprogesterone in the gene effects of oocyte and its final maturation.For the first time,the present study would provide morphological demonstration.
出处
《水产学报》
CAS
CSCD
北大核心
2010年第7期999-1010,共12页
Journal of Fisheries of China
基金
农业部公益性研究项目(nyhyzx07-043-16-02)
