p21、c-myc和S100蛋白在小鼠卵泡及黄体中的表达
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摘要
成年哺乳动物的卵泡发育、黄体形成及退化是一个周期性循环循环过程。黄体在生殖周期中起着“关卡”作用,它的正常退化是启动下一个生殖周期的前提。本研究的目的是探索凋亡相关的p21~(WAF1/CIP1)抑癌基因、c-myc癌基因和S100蛋白在卵泡和黄体中的表达及相互关联。本试验采用免疫组织化学超敏感SP法、相邻切片比较法结合图像分析法对成年小鼠卵泡和黄体中p21~(WAF1/CIP1)蛋白、c-myc癌基因蛋白和S100蛋白的分布和变化规律进行了研究,主要结果如下:
1.在成年小鼠卵泡发育和闭锁的各个时期卵母细胞和颗粒细胞均有p21~(WAF1/CIP1)蛋白的表达,黄体细胞中也有其表达,均定位于细胞核内,其相对表达量在该过程中大致呈先增高后降低的趋势,最大值出现在三级卵泡时期。提示,p21~(WAF1/CIP1)蛋白在卵母细胞中发挥一定作用,可能参与优势卵泡的选择,并对卵泡发育及黄体形成和退化有影响。
2.在卵泡发育、闭锁、黄体形成及退化过程中,除原始卵泡时期未见c-myc癌基因蛋白的表达外,其它时期都有它的微量表达,最大值出现在早期黄体中,并且表达部位由卵泡中的核周围转移到黄体中的胞浆内,在血管新生部位表达较强。这说明,c-myc癌基因蛋白作为卵巢组织内局部调控因子,对卵泡发育及闭锁有影响,促进黄体形成,还可能与血管新生有密切关联。
3. S100蛋白作为一种Ca2+结合蛋白,在卵泡发育、闭锁、黄体形成及退化过程中,除原始卵泡时期未见表达外,其它时期都有微量表达,定位于胞浆中,最大值出现早期黄体时期,并且该时期阳性细胞形态多样,比如圆形、椭圆形、梭形、部分还有明显突起,随后恢复到基准水平。这表明,S100蛋白可能对卵泡发育及闭锁有影响,为黄体形成完备的内分泌功能具有积极作用,同时还可能作为黄体结构性退化的标志之一,参与黄体退化。
4.相邻切片比较法及相关性分析表明,p21~(WAF1/CIP1)蛋白、c-myc癌基因蛋白、S100蛋白免疫阳性细胞在卵泡和黄体中存在部分相同区域,且有两种物质在同一细胞共存现象,p21~(WAF1/CIP1)蛋白的相对表达量与S100蛋白、c-myc癌基因蛋白分别呈高度、中度负相关, c-myc癌基因蛋白与S100蛋白的相对表达量之间呈中度正相关。
Follicular development, corpus luteum formation and regression are regarded as a cyclic process in adult mammalian. Corpus luteum plays a role of“checkpoint”function in reproductive cycle. And its degradation starts up next reproductive circle. The purpose of this study is to explore the expression and correlation of p21 WAF1/CIP1 apoptosis- related tumor suppressor gene, c-myc oncogene and S100 protein in follicle and corpus luteum. Our research studies the distributions and laws of these three proteins by immunohistochemical SP method, compare of adjacent slices and the image analysis. The main results are as follows.
1. p21~(WAF1/CIP1) protein expressd in all periods, including primordial, primary, secondary, tertiary, mature, atresic follicles, and forepart, metaphase and anaphase corpus luteums, and located in the nuclei. The tendency of its relative expression in this process generally increased first and then decreased. The peak value appeared in the period of the tertiary follicle. We conclude that p21~(WAF1/CIP1) protein plays a role in oocytes during follicular development, may participate in choosing dominant follicle, and has a potential influence on follicular development, corpus luteum formation and regression.
2. All periods existed the expression of c-myc, except the primordial follicle. The maximum appeared in the forepart corpus luteum. The distribution of the products changed from encircling nucleus of follicular cells to cytoplasm of corpus luteum cells. And the sites of angiogenesis aggregated more products. These results show that c-myc effects on the follicular development, promotes corpus luteum formation and may relate closely to angiogenesis.
3. S100, as one of Ca2+ binding proteins, located in the cytoplasm, expressed in all periods. Primordial follicle made an exception. The forepart corpus luteum expressed the max products. There were various morphological features of the positive cells during that period, such as round, oval, spindle, polygonal shapes, etc, and some of which had significant protrusions. Then the relative expression of S100 protein returned to the baseline lever. These results suggest that S100 protein influence follicular development and atresia, have a positive role in the formation of the complete endocrine function for corpus luteum, and as one of markers of luteal structural degradation, may involve in corpus luteum regression.
4. By compare of adjacent slices and the image analysis, we found that there existed overlapping regions between positive cells in follicle and corpus luteum, which play an important role in maintain ovarian function; the relative expression of p21~(WAF1/CIP1) protein had high and moderate negative correlations respectively with S100 protein and c-myc oncoprotein; there was a moderate positive relationship between S100 protein and c-myc oncoprotein.
1.在成年小鼠卵泡发育和闭锁的各个时期卵母细胞和颗粒细胞均有p21~(WAF1/CIP1)蛋白的表达,黄体细胞中也有其表达,均定位于细胞核内,其相对表达量在该过程中大致呈先增高后降低的趋势,最大值出现在三级卵泡时期。提示,p21~(WAF1/CIP1)蛋白在卵母细胞中发挥一定作用,可能参与优势卵泡的选择,并对卵泡发育及黄体形成和退化有影响。
2.在卵泡发育、闭锁、黄体形成及退化过程中,除原始卵泡时期未见c-myc癌基因蛋白的表达外,其它时期都有它的微量表达,最大值出现在早期黄体中,并且表达部位由卵泡中的核周围转移到黄体中的胞浆内,在血管新生部位表达较强。这说明,c-myc癌基因蛋白作为卵巢组织内局部调控因子,对卵泡发育及闭锁有影响,促进黄体形成,还可能与血管新生有密切关联。
3. S100蛋白作为一种Ca2+结合蛋白,在卵泡发育、闭锁、黄体形成及退化过程中,除原始卵泡时期未见表达外,其它时期都有微量表达,定位于胞浆中,最大值出现早期黄体时期,并且该时期阳性细胞形态多样,比如圆形、椭圆形、梭形、部分还有明显突起,随后恢复到基准水平。这表明,S100蛋白可能对卵泡发育及闭锁有影响,为黄体形成完备的内分泌功能具有积极作用,同时还可能作为黄体结构性退化的标志之一,参与黄体退化。
4.相邻切片比较法及相关性分析表明,p21~(WAF1/CIP1)蛋白、c-myc癌基因蛋白、S100蛋白免疫阳性细胞在卵泡和黄体中存在部分相同区域,且有两种物质在同一细胞共存现象,p21~(WAF1/CIP1)蛋白的相对表达量与S100蛋白、c-myc癌基因蛋白分别呈高度、中度负相关, c-myc癌基因蛋白与S100蛋白的相对表达量之间呈中度正相关。
Follicular development, corpus luteum formation and regression are regarded as a cyclic process in adult mammalian. Corpus luteum plays a role of“checkpoint”function in reproductive cycle. And its degradation starts up next reproductive circle. The purpose of this study is to explore the expression and correlation of p21 WAF1/CIP1 apoptosis- related tumor suppressor gene, c-myc oncogene and S100 protein in follicle and corpus luteum. Our research studies the distributions and laws of these three proteins by immunohistochemical SP method, compare of adjacent slices and the image analysis. The main results are as follows.
1. p21~(WAF1/CIP1) protein expressd in all periods, including primordial, primary, secondary, tertiary, mature, atresic follicles, and forepart, metaphase and anaphase corpus luteums, and located in the nuclei. The tendency of its relative expression in this process generally increased first and then decreased. The peak value appeared in the period of the tertiary follicle. We conclude that p21~(WAF1/CIP1) protein plays a role in oocytes during follicular development, may participate in choosing dominant follicle, and has a potential influence on follicular development, corpus luteum formation and regression.
2. All periods existed the expression of c-myc, except the primordial follicle. The maximum appeared in the forepart corpus luteum. The distribution of the products changed from encircling nucleus of follicular cells to cytoplasm of corpus luteum cells. And the sites of angiogenesis aggregated more products. These results show that c-myc effects on the follicular development, promotes corpus luteum formation and may relate closely to angiogenesis.
3. S100, as one of Ca2+ binding proteins, located in the cytoplasm, expressed in all periods. Primordial follicle made an exception. The forepart corpus luteum expressed the max products. There were various morphological features of the positive cells during that period, such as round, oval, spindle, polygonal shapes, etc, and some of which had significant protrusions. Then the relative expression of S100 protein returned to the baseline lever. These results suggest that S100 protein influence follicular development and atresia, have a positive role in the formation of the complete endocrine function for corpus luteum, and as one of markers of luteal structural degradation, may involve in corpus luteum regression.
4. By compare of adjacent slices and the image analysis, we found that there existed overlapping regions between positive cells in follicle and corpus luteum, which play an important role in maintain ovarian function; the relative expression of p21~(WAF1/CIP1) protein had high and moderate negative correlations respectively with S100 protein and c-myc oncoprotein; there was a moderate positive relationship between S100 protein and c-myc oncoprotein.
引文
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