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取出生后21~26天Sprague-Dawley未成熟雌性大鼠,分为实验与对照两组。实验组只/天注射己烯雌酚(DES)0.5mg(溶于芝麻油中),并按注射天数分为2、3、4、5天组;对照组注射等量的芝麻油。注射2天后,逐日取材固定,用光镜和电镜观察两组动物的卵巢和卵泡颗粒细胞。同时从各组卵巢中分离出卵泡颗粒细胞,用无血清McCoy 5a培养基培养,加入不同的激素,分为对照组、FSH组、DES+FSH组、FSH+hCG组。培养3天后,取材固定,用光镜、透射电镜、扫描电镜观察各组的细胞形态,结果为:1.实验组卵巢比对照组的大,卵泡也大。卵泡细胞增多,细胞体积大,分裂象多见。2.注射3天以上DES的卵巢中,闭锁卵泡和退化的卵泡细胞增多。3.电镜下,实验组卵泡细胞中与蛋白质合成有关的细胞器比对照组发达。4.体外培养的卵泡颗粒细胞,对照组细胞排列分散;FSH组细胞集聚成团,细胞表面微绒毛增多;细胞质内与合成蛋白质有关的细胞器比对照组的发达。DES+FSH组的细胞排列和结构与FSH组的相似。FSH+hCG组细胞体积增大,细胞表面微绒毛减少或消失。细胞质内出现滑面内质网,细胞开始黄体化。5.注射2~5天DES的各组细胞形态和结构均相似。从形态学来看,经DES处理的未成熟雌鼠的卵泡颗粒细胞的形态、排列和对不同激素作用下的变化,与文献报道的去垂体雌鼠的同类细胞相似。因此我们认为,未成熟雌鼠的卵泡颗粒细胞也能作为研究细胞分化的一种模型。
Sprague-Dawley immature female rats were removed 21 to 26 days after birth and divided into experimental and control groups. In the experimental group, diethylstilbestrol (DES) 0.5 mg (dissolved in sesame oil) was injected into the experimental group only on the day of injection. The rats in the experimental group were divided into groups of 2,3,4 and 5 days according to the number of injection days. The control group was injected with the same amount of sesame oil. Two days after injection, the materials were fixed daily, and the ovaries and follicular granulosa cells of the two groups were observed by light microscope and electron microscope. At the same time, follicular granulosa cells were isolated from ovaries of each group and cultured in serum-free McCoy 5a medium. Different hormones were added and divided into control group, FSH group, DES + FSH group and FSH + hCG group. After culturing for 3 days, the cells were fixed and the cell morphology of each group was observed by light microscopy, transmission electron microscopy and scanning electron microscopy. The results were as follows: 1. The ovary of the experimental group was larger than that of the control group with large follicles. Increased follicular cells, large cell size, split more common. 2. In the ovaries of DES over 3 days, the atresia follicles and degenerated follicles increased. Electron microscope, the experimental group of follicular cells and protein synthesis related organelles developed than the control group. In vitro culture of granulosa cells, the control group cells arranged in scattered; FSH group gathered into clusters, cell surface microvilli increased; cytoplasm and synthetic protein-related organelles developed than the control group. The cell arrangement and structure of DES + FSH group were similar to that of FSH group. FSH + hCG group of cells increased in size, cell surface microvilli reduced or disappeared. Sliding endoplasmic reticulum appears in the cytoplasm, the cells begin to luteinize. The morphology and structure of the cells in all groups injected with DES for 2 ~ 5 days were similar. Morphologically, the morphology, arrangement and changes of follicular granulosa cells of immature female mice treated with DES were similar to those of the pituitary female mice reported in the literature. Therefore, we believe that immature granulose cells of females can also be used as a model for studying cell differentiation.