Tsubota T, Taki S, Nakayama K, Mason J I, Kominami S, Harada N, Kita I
Laboratory of Theriogenology, Faculty of Agriculture, Gifu University, Gifu 501-11, Japan.
Reproduction. 2001 Apr;121(4):587-94. doi: 10.1530/rep.0.1210587.
The Japanese black bear, Ursus thibetanus japonicus, is a seasonal breeder and shows delayed implantation for several months during pregnancy. The objective of this study was to clarify the steroidogenic capability of the corpus luteum and placenta during pregnancy, including both delayed implantation and fetal development, by immunolocalization of steroidogenic enzymes in these organs of the Japanese black bear. Ovaries and placentae from 15 wild Japanese black bears, which had been killed legally by hunters and were thought to be pregnant, were used in an immunocytochemical study to localize the cholesterol side chain cleavage cytochrome P450 (P450scc), 3beta-hydroxysteroid dehydrogenase (3betaHSD), 17alpha-hydroxylase cytochrome P450 (P450c17) and aromatase cytochrome P450 (P450arom) by the avidin-biotin-peroxidase complex method using polyclonal antisera raised in mammals against P450scc, 3betaHSD, P450c17 and P450arom. P450scc and 3betaHSD were localized in all luteal cells throughout pregnancy. P450c17 was present in a few luteal cells, especially in the outer area of the corpus luteum throughout pregnancy, but the number of positively immunostained cells decreased during the post-implantation period. Cells positively immunostained for P450c17 were significantly smaller than negatively immunostained cells (P < 0.01). P450arom was present sporadically in a few luteal cells throughout pregnancy, but the number of positively immunostained cells decreased during the post-implantation period. The size of cells positively immunostained for P450arom was not significantly different from that of negatively immunostained cells. The whole placenta was negatively immunostained for P450scc, 3betaHSD and P450c17, but P450arom was present in the syncytiotrophoblasts and endothelial cells of maternal blood vessels. These results indicate that, in the Japanese black bear, corpora lutea are a source of progesterone which may play an important role in the maintenance of delayed implantation and fetal development during pregnancy. Corpora lutea have a minimum capability to synthesize androgen in small luteal cells and oestrogen in normal-sized luteal cells during pregnancy, and placentae have the ability to synthesize oestrogen during late pregnancy.
日本黑熊（Ursus thibetanus japonicus）是季节性繁殖动物，孕期会有几个月的胚胎延迟着床。本研究的目的是通过对日本黑熊这些器官中类固醇生成酶进行免疫定位，来阐明孕期包括胚胎延迟着床和胎儿发育阶段黄体和胎盘的类固醇生成能力。本免疫细胞化学研究使用了15只野生日本黑熊的卵巢和胎盘，这些黑熊由猎人合法捕杀，据认为处于孕期。采用抗P450scc、3βHSD、P450c17和P450arom的哺乳动物多克隆抗血清，通过抗生物素蛋白-生物素-过氧化物酶复合物法对胆固醇侧链裂解细胞色素P450（P450scc）、3β-羟基类固醇脱氢酶（3βHSD）、17α-羟化酶细胞色素P450（P450c17）和芳香化酶细胞色素P450（P450arom）进行定位。整个孕期，P450scc和3βHSD定位于所有黄体细胞。P450c17存在于少数黄体细胞中，尤其是整个孕期黄体的外层区域，但着床后期阳性免疫染色细胞数量减少。P450c17阳性免疫染色的细胞明显小于阴性免疫染色的细胞（P < 0.01）。整个孕期，P450arom偶尔存在于少数黄体细胞中，但着床后期阳性免疫染色细胞数量减少。P450arom阳性免疫染色细胞的大小与阴性免疫染色细胞的大小无显著差异。整个胎盘对P450scc、3βHSD和P450c17呈阴性免疫染色，但P450arom存在于合体滋养层细胞和母体血管内皮细胞中。这些结果表明，在日本黑熊中，黄体是孕酮的来源，孕酮可能在维持孕期胚胎延迟着床和胎儿发育中起重要作用。孕期，黄体在小黄体细胞中具有最低限度的合成雄激素的能力，在正常大小的黄体细胞中具有合成雌激素的能力，胎盘在妊娠后期具有合成雌激素的能力。
