Oride Aki, Kanasaki Haruhiko, Purwana Indri N, Mutiara Sandra, Miyazaki Kohji
Department of Obstetrics and Gynecology, Shimane University School of Medicine, Izumo 693-8501, Japan.
Regul Pept. 2009 Aug 7;156(1-3):65-71. doi: 10.1016/j.regpep.2009.05.001. Epub 2009 May 14.
Follistatin regulates FSHbeta gene expression by binding to and bioneutralizing activin effects. In this study, we found that thyrotropin-releasing hormone (TRH) increased follistatin gene expression in pituitary somatolactotroph GH3 cells. Treatment of GH3 with 100 nM TRH significantly increased follistatin mRNA expression as determined by real time PCR. TRH-induced follistatin expression was significantly abrogated in the presence of MEK inhibitor, U0126. Overexpression of constitutive active MEKK in GH3 cells dramatically increased follistatin expressions. Transfection of GH3 cells with follistatin siRNA reduced endogenous follistatin mRNA expression, but failed to modulate prolactin promoter activity. Prolactin mRNA levels were not affected by increasing the dose of follistatin, and TRH-induced prolactin promoter activity was not modulated in the presence of follistatin. In other experiments using pituitary gonadotroph LbetaT2 cells, activin increased FSHbeta promoter activity and mRNA expression, and follistatin completely inhibited this activin-increased FSHbeta gene expression. Treatment of GH3 cells with activin reduced the basal activity of prolactin promoter and follistatin prevented this effect. GH3 cells were co-cultured with LbetaT2 cells, which had been transfected with FSHbeta promoter-linked luciferase vectors and treated with activin in the presence of TRH. Activin-induced FSHbeta promoter activity was completely inhibited in the presence of TRH. In addition to that, FSHbeta mRNA was not detected from LbetaT2 cells which were co-cultured with GH3 cells. Our current results suggest the possibility that TRH increases follistatin gene expression in prolactin-producing cells in association with ERK pathways. Somatolactotroph-derived follistatin affects gonadotrophs by countering activin-induced FSHbeta gene expression in a paracrine fashion.
卵泡抑素通过结合并生物中和激活素的作用来调节促卵泡激素β（FSHβ）基因表达。在本研究中，我们发现促甲状腺激素释放激素（TRH）可增加垂体生长催乳素细胞系GH3细胞中卵泡抑素基因的表达。用100 nM TRH处理GH3细胞，通过实时定量PCR测定发现卵泡抑素mRNA表达显著增加。在MEK抑制剂U0126存在的情况下，TRH诱导的卵泡抑素表达显著消除。在GH3细胞中组成型活性MEKK的过表达显著增加了卵泡抑素的表达。用卵泡抑素小干扰RNA（siRNA）转染GH3细胞可降低内源性卵泡抑素mRNA表达，但未能调节催乳素启动子活性。增加卵泡抑素剂量对催乳素mRNA水平无影响，并且在卵泡抑素存在的情况下，TRH诱导的催乳素启动子活性未被调节。在使用垂体促性腺激素细胞系LβT2细胞的其他实验中，激活素增加了FSHβ启动子活性和mRNA表达，而卵泡抑素完全抑制了这种激活素诱导的FSHβ基因表达。用激活素处理GH3细胞可降低催乳素启动子的基础活性，而卵泡抑素可阻止这种作用。将GH3细胞与已用FSHβ启动子连接的荧光素酶载体转染并在TRH存在下用激活素处理的LβT2细胞共培养。在TRH存在的情况下，激活素诱导的FSHβ启动子活性被完全抑制。除此之外，与GH3细胞共培养的LβT2细胞中未检测到FSHβ mRNA。我们目前的结果提示，TRH可能通过细胞外信号调节激酶（ERK）途径增加催乳素分泌细胞中卵泡抑素基因的表达。生长催乳素细胞来源的卵泡抑素通过旁分泌方式对抗激活素诱导的FSHβ基因表达，从而影响促性腺激素细胞。
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