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BrdU labelling of oocyte in the rat fetus ovary

大鼠胎儿卵巢卵母细胞的 BrdU 标记

基本信息

批准号：
01570914
负责人：
FUJIMORI Katsuhiko
金额：
$ 1.28万
依托单位：
Yamagata University School of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1989
资助国家：
日本
起止时间：
1989 至 1991
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-01570914/
关键词：
Folliculogenesis First estrus Rat Immunohistochemistry

项目摘要

INTRODUCTIONIn order to analyze the follicular development dynamically, it is effective to set up a cohort by labelling a part of follicles and to follow up their growth. It is widely known that rat's oocyte doubles its DNA in the embryonal stage and its mitosis has been arrested until the second meiosis just before ovulation. We noticed this phenomenon is available to determine the possibility of labelling a certain group of oocyte's nuclei in the embryonal stage.MATERIALS AND METHODSThe pregnant rats were divided into four groups (A, B, C and D group, each consisted of 4 animals) and transabdominally injected with bromodeoxyuridine (BrdU) 5 g/body weight (g) at one time in the 17th day of pregnancy. They were reared to delivery. Thereafter, the female newborn animals were reared and sacrificed at 1week (group A), 2week (B), 3week (C) and 4week (D) to excise bilateral ovaries and liver. These organs were fixed in formalin, embedded in paraffin, and then serially sliced at 5 m in thick … More ness. Every seventh section was mounted on slide glass. Immunohistochemical staining was employed with anti-BrdU monoclonal antibody (Boehringer-Mannheim Biochemica) by using Biotin-streptAvidin's method. We investigated the localization of BrdU which was incorporated into the cell.RESULTS AND DISCUSSION1) A part of fetal oocytes were labelled by BrdU in the 17th day of pregnancy and their labelling in the nuclei were maintained up to 4th week after delivery (group A, B, C, D). On the contrary, liver cells showed positive staining of BrdU in group A, however, BrdU labelling was not detectable in group B, C, and D. Therefore, it was clarified that BrdU-uptake in the liver cell was so diluted at its every mitosis to the degree which could not be demonstrated.2) BrdU labelling was evident mainly in the primordial follicle which did not have granulosa cells. At 1 week of delivery (group A), the follicles labelled by BrdU were 100% of the primordial follicles. The primordial follicles occupied 96.3% of the tagged follicles at 2 week of delivery (group B) and 46.7% at 3 week (group C). This decreased incidence of BrdU-labelled primordial follicles presented that some follicles in given BrdU-labelled cohort were developed into the next category of primary follicles accompanied with a few layers of granulosa cells. However, there were only 1 or 2 vesicular follicles showing BrdU incorporation through 1 to 4 week of delivery. Less

为了动态分析卵泡的发育，通过标记部分卵泡建立队列并跟踪其生长是有效的。众所周知，大鼠的卵母细胞在胚胎期使其DNA加倍，并且其有丝分裂一直被阻止，直到排卵前的第二次减数分裂。材料与方法将孕鼠分为A、B、C、D四组，每组4只，于孕17 d一次性经腹注射溴脱氧尿苷（BrdU）5g/g体重。他们是被养大的。然后饲养雌性新生动物，分别于1周（A组）、2周（B组）、3周（C组）和4周（D组）处死，切除双侧卵巢和肝脏。将这些器官固定在福尔马林中，包埋在石蜡中，然后连续切成5 m厚的切片 ...更多信息奈斯将每第七个切片固定在载玻片上。免疫组化染色采用抗BrdU单克隆抗体（Boehringer-Mannheim Biochemica），生物素-链霉亲和素法。结果与结论1）孕17天部分胚胎卵母细胞被BrdU标记，并持续标记至产后4周（A、B、C、D组）。相反，A组肝细胞BrdU染色呈阳性，而B、C、D组肝细胞BrdU标记均未检测到。因此，这是明确的，BrdU-摄取在肝细胞中是如此稀释，在其每一个有丝分裂的程度，不能证明。2）BrdU标记是明显的，主要是在原始卵泡，没有颗粒细胞。分娩1周时（A组），BrdU标记的卵泡占原始卵泡的100%。分娩2周时（B组）原始卵泡占标记卵泡的96.3%，分娩3周时（C组）原始卵泡占标记卵泡的46.7%。BrdU标记的原始卵泡的发生率降低，表明给定BrdU标记队列中的一些卵泡发育成下一类初级卵泡，伴有几层颗粒细胞。然而，只有1或2个泡状卵泡显示BrdU掺入通过1至4周的交付。少