Changes in localization of cytochrome P450 cholesterol side-chain cleavage (P450scc) in Japanese eel testis and ovary during gonadal development
Changes in localization of cytochrome P450 cholesterol side-chain cleavage (P450scc) in Japanese eel testis and ovary during gonadal development
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DOI：
10.1016/j.ygcen.2005.07.005
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发表时间：
2006-01-01
影响因子：
2.7
通讯作者：
Yamauchi, K
中科院分区：
文献类型：
--
作者：
Ijiri, S;Takei, N;Yamauchi, K
本研究以重组蛋白为抗原，制备并鉴定了鳗鱼胆固醇侧链裂解酶P450（P450SCC）的多克隆抗血清。我们研究的定位和丰富的P450SCC在日本鳗鲡睾丸和卵巢在人工诱导性腺发育的免疫组织化学。原位杂交检测P450SCC mRNA的定位。在雄性鳗鱼中，睾丸发育是通过单次注射人绒毛膜促性腺激素（HCG）诱导的。在雌性动物中，通过每周注射鲑鱼垂体匀浆（SPH）诱导卵巢发育。在注射HCG之前，睾丸含有主要为A型精原细胞的生殖细胞。此外，间质间质细胞中有数簇P450 SCC免疫反应阳性细胞，但未检测到P450 SCC mRNA信号。这表明P450SCC是一种相对稳定的蛋白质，或者它是由mRNA产生的，其水平太低而无法检测。单次注射HCG后不久，P450 SCC mRNA的表达被刺激，免疫反应簇的数量和染色强度均增加。P450SCC mRNA在激素刺激后3天下降到检测不到的水平。虽然P450SCC蛋白也与mRNA同时下降，但在此期间仍保持在可检测的水平。在精子细胞和精子中也检测到P450SCC mRNA，但未检测到P450SCC蛋白。P450SCC mRNA在此阶段表达的生物学意义尚不清楚。在实验之前，卵巢含有发育到油滴阶段的卵母细胞，其中几簇免疫反应细胞位于卵泡膜层和产卵层上皮中。卵巢中SPH注射也刺激P450SCC mRNA的表达。与睾丸相反，在整个人工诱导成熟过程中，在卵泡膜细胞层中连续检测到P450SCC mRNA，这可能是由于每周注射SPH的重复刺激。随着卵巢发育的进行，卵泡膜细胞层中的免疫反应细胞簇数量增加。P450SCC mRNA和蛋白的这种增加可以解释，至少部分地，雌性鳗鱼血清类固醇激素的增加。P450SCC抗血清清楚地免疫染色肾间类固醇生成细胞的头肾，不仅鳗鱼，但也金鱼，表明这种抗体也可用于其他硬骨鱼类。(c)2005年爱思唯尔公司All rights reserved.
In this study, we generated and characterized a polyclonal antiserum against eel P450 cholesterol side-chain cleavage (P450scc) using a recombinant protein as the antigen. We examined the localization and abundance of P450scc by immunohistochemistry in Japanese eel testes and ovaries during artificially induced gonadal development. P450scc mRNA localization was also examined by in situ hybridization. In male eels, testicular development was induced by a single injection of human chorionic gonadotropin (HCG). In females, ovarian development was induced by weekly injections of salmon pituitary homogenate (SPH). Before HCG injection, the testis contained germ cells that were primarily type A spermatogonia. Additionally, several clusters of immunoreactive cells for P450scc were localized in the interstitial Leydig cells, but no P450scc mRNA signals were detected. This suggests that P450scc is either a relatively stable protein or it is produced by a mRNA that is present at too low a level to detect. Shortly after a single injection of HCG, expression of P450scc mRNA was stimulated and the number of immunoreactive clusters and their staining intensity were both increased. P450scc mRNA fell to an undetectable level 3 days after hormonal stimulation. Although the P450scc protein also decreased at the same time as the mRNA, it remained at a detectable level throughout this period. P450scc mRNA, but not the P450scc protein, was also detected in the spermatids and spermatozoa. The biological significance of P450scc mRNA expression at this stage is unknown. Prior to experimentation, the ovary contained oocytes that were developed to the oil-droplet stage, with several clusters of immuno reactive cells localized in the thecal layer and ovigerous lamella epithelium. Expression of P450scc, mRNA was also stimulated by SPH injections in the ovary. In contrast to the testis, P450scc mRNA was continuously detected in the thecal cell layer throughout artificially induced maturation, possibly due to a repeated stimulus by the SPH injection every week. Clusters of immunoreactive cells in the thecal cell layer increased in number as ovarian development progressed. This increase ill P450scc mRNA and protein may explain, at least in part, the increase in serum steroid hormones in female eels. The P450scc antiserum clearly immunostained interrenal steroidogenic cells in the head kidney of not only eel but also goldfish, indicating that this antibody could also be used in other teleost species. (c) 2005 Elsevier Inc. All rights reserved.