Attardi B, Keeping H S, Winters S J, Kotsuji F, Maurer R A, Troen P
Department of Medicine, Montefiore Hospital, Pittsburgh, Pennsylvania 15213.
Mol Endocrinol. 1989 Feb;3(2):280-7. doi: 10.1210/mend-3-2-280.
We showed previously that inhibin, partially purified from cynomolgus monkey Sertoli cell culture medium (primate Sertoli cell inhibin referred to as pSCI), selectively suppressed basal FSH secretion from dispersed rat pituitary cells and decreased total cellular FSH, but not LH content, suggesting a decrease in FSH biosynthesis. In order to investigate the mechanism of action of inhibin at the molecular level, we have now examined the effects of pSCI on steady state levels of the subunit mRNAs encoding LH and FSH and correlated these with release and intracellular content of LH, FSH, and glycoprotein alpha-subunit. Dispersed pituitary cells from 7- to 8-week-old adult male rats were cultured in the presence of pSCI or control medium for 2-72 h. FSH secretion was reduced significantly by 6 h (P less than 0.05) and reached a nadir (38% of control) by 48 h. LH secretion was unchanged, while release of the alpha-subunit was decreased to 89% of control at 72 h (P less than 0.05). Also by 72 h, cell content of both FSH (73% of control) and alpha-subunit (81% of control) were significantly suppressed (P less than 0.001, P less than 0.01), while LH was slightly affected. Total RNA was extracted from the pituitary cell cultures, electrophoresed in 1.2% agarose-formaldehyde gels, transferred to nylon membranes, and hybridized with 32P-labeled cDNA probes for the rat alpha-, LH beta-, and FSH beta-subunits.(ABSTRACT TRUNCATED AT 250 WORDS)
我们之前发现,从食蟹猴支持细胞培养基中部分纯化得到的抑制素(灵长类支持细胞抑制素,简称pSCI)可选择性抑制分散的大鼠垂体细胞基础促卵泡激素(FSH)的分泌,并降低细胞内FSH总量,但不影响促黄体生成素(LH)含量,提示FSH生物合成减少。为了在分子水平上研究抑制素的作用机制,我们现在检测了pSCI对编码LH和FSH的亚基mRNA稳态水平的影响,并将其与LH、FSH和糖蛋白α亚基的释放及细胞内含量进行关联分析。将7至8周龄成年雄性大鼠的分散垂体细胞在pSCI或对照培养基中培养2至72小时。FSH分泌在6小时时显著降低(P<0.05),48小时时降至最低点(为对照的38%)。LH分泌未改变,而α亚基的释放在72小时时降至对照的89%(P<0.05)。同样在72小时时,FSH(为对照的73%)和α亚基(为对照的81%)的细胞含量均受到显著抑制(P<0.001,P<0.01),而LH受到的影响较小。从垂体细胞培养物中提取总RNA,在1.2%琼脂糖-甲醛凝胶中进行电泳,转移至尼龙膜上,并用针对大鼠α、LHβ和FSHβ亚基的32P标记cDNA探针进行杂交。(摘要截短于250字)