Nogami H, Inoue K, Moriya H, Ishida A, Kobayashi S, Hisano S, Katayama M, Kawamura K
Department of Anatomy, School of Medicine, Keio University, Tokyo, Japan.
Endocrinology. 1999 Jun;140(6):2763-70. doi: 10.1210/endo.140.6.6787.
Regulation of GH-releasing hormone receptor (GHRH-R) messenger RNA (mRNA) expression was studied, with the ribonuclease protection assay, in the fetal rat pituitary gland and in MtT-S clonal cells. GHRH-R mRNA was first detected on embryonic day (E)19 and increased rapidly thereafter, to reach a maximum at E21. Incubation of E17 or E18 pituitaries with 50 nM dexamethasone (DEX), a synthetic glucocorticoid, induced GHRH-R mRNA expression, suggesting that glucocorticoids play a pivotal role in the developmental expression of this mRNA. In E19 pituitaries, 24 h treatment with DEX increased GHRH-R mRNA by 60%, and GH mRNA by 76%, but did not affect pit-1 mRNA level, suggesting that the effect of DEX is specific for expressions of GH mRNA and GHRH-R mRNA. The accumulation of GHRH-R mRNA by DEX was time dependent, and it was slightly enhanced by the protein synthesis inhibitor, puromycin (100 microM). In MtT-S cells (a pituitary cell line established from an estrogen-induced tumor), DEX induced GHRH-R mRNA expression within 2 h in a dose-dependent manner. This induction was augmented by puromycin (100 microM) or cycloheximide (3.5 microM). However, the RNA synthesis inhibitor Actinomycin D (1 microM) completely inhibited GHRH-R mRNA accumulation in response to either DEX or DEX plus puromycin, suggesting that glucocorticoids induce GHRH-R mRNA mainly through stimulation of mRNA transcription. These results suggest: that GHRH-R mRNA accumulation in the fetal pituitary gland of rats normally occurs at E19, probably because of the direct action of glucocorticoids on the pituitary gland, to stimulate GHRH-R mRNA transcription; and that the expression of glucocorticoid receptors is an important event in GH cell development in rats. Accordingly, immunocytochemical results suggest an increase in glucocorticoid receptors in immature GH cells between E17 and E18. The present results also imply that MtT-S cells may be a good model in which to further study the molecular mechanisms of the regulation of GHRH-R gene expression.
采用核糖核酸酶保护分析法，对胎鼠垂体和MtT-S克隆细胞中生长激素释放激素受体（GHRH-R）信使核糖核酸（mRNA）的表达调控进行了研究。GHRH-R mRNA在胚胎第（E）19天首次被检测到，此后迅速增加，在E21达到最大值。用50 nM地塞米松（DEX，一种合成糖皮质激素）孵育E17或E18垂体，可诱导GHRH-R mRNA表达，这表明糖皮质激素在该mRNA的发育表达中起关键作用。在E19垂体中，用DEX处理24小时可使GHRH-R mRNA增加60%，生长激素（GH）mRNA增加76%，但不影响垂体特异性转录因子1（pit-1）mRNA水平，这表明DEX的作用对GH mRNA和GHRH-R mRNA的表达具有特异性。DEX诱导GHRH-R mRNA的积累具有时间依赖性，并且蛋白质合成抑制剂嘌呤霉素（100 microM）可使其略有增强。在MtT-S细胞（一种由雌激素诱导的肿瘤建立的垂体细胞系）中，DEX在2小时内以剂量依赖性方式诱导GHRH-R mRNA表达。嘌呤霉素（100 microM）或环己酰亚胺（3.5 microM）可增强这种诱导作用。然而，RNA合成抑制剂放线菌素D（1 microM）完全抑制了对DEX或DEX加嘌呤霉素反应的GHRH-R mRNA积累，这表明糖皮质激素主要通过刺激mRNA转录来诱导GHRH-R mRNA。这些结果表明：大鼠胎垂体中GHRH-R mRNA的积累通常发生在E19，可能是由于糖皮质激素对垂体的直接作用，刺激了GHRH-R mRNA转录；糖皮质激素受体的表达是大鼠GH细胞发育中的一个重要事件。因此，免疫细胞化学结果表明E17和E18之间未成熟GH细胞中糖皮质激素受体增加。目前的结果还表明，MtT-S细胞可能是进一步研究GHRH-R基因表达调控分子机制的良好模型。
