Lentiviral and Moloney retroviral expression of green fluorescent protein in somatotrophs in vivo.
Lentiviral and Moloney retroviral expression of green fluorescent protein in somatotrophs in vivo.
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DOI：
10.1371/journal.pone.0054437
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Okimura Y
中科院分区：
文献类型：
--
作者：
Okada M;Matsuda H;Okimura Y
已有研究表明，生长激素基因的基因座控制区（locus control region， LCR）和启动子可以调控生长激素的表达。因此，含有这些元素的慢病毒和逆转录病毒载体可能有助于监测生长激素基因的激活和新生儿生长发育不良的发生。为了验证这一点，我们首先构建了一个慢病毒载体，在这些元件的控制下表达绿色荧光蛋白（GFP），并将它们注射到大鼠垂体的原位和体内。慢病毒载体在前叶特异性表达GFP，几乎所有GFP阳性细胞都具有抗gh免疫反应。生长激素释放激素和IGF-1受体阻滞剂可上调GFP的表达。此外，社会隔离应激降低了生长激素分泌，降低了GFP的表达。其次，我们将逆转录病毒载体注射到新生大鼠垂体内。注射后30天，几乎所有gfp阳性细胞均呈抗gh阳性，抗催乳素阴性。然而，GFP在8和16 DPI时通过培养乳养菌短暂表达，这表明我们的载体缺乏抑制表达的元件。同时，逆转录病毒标记的细胞倾向于与相同类型的细胞聚集。对每簇细胞数量的分析揭示了细胞增殖的一些特征。这些病毒载体被证明是监测生长激素基因激活和生长发育的有用工具。
Previous studies have shown that the locus control region (LCR) and the promoter of the growth hormone (GH) gene can control the expression of GH. Therefore, lenti- and retro-viral vectors with these elements might be useful to monitor the activation of the GH gene and the development of newborn somatotrophs. To test this, we first constructed a lentiviral vector, which expresses green fluorescent protein (GFP) under the control of these elements, and injected them into rat pituitaries in situ and in vivo. The lentiviral vector expressed GFP specifically in the anterior lobe, and nearly all GFP-positive cells were anti-GH immunoreactive. The GFP expression was upregulated by the administration of growth hormone releasing hormone and an IGF-1 receptor blocker. Furthermore, the social isolation stress, which was shown to decrease the GH secretion, decreased the GFP expression. Second, we injected the retroviral vector into neonatal rat pituitaries in vivo. At 30 days postinjection (DPI), almost all GFP-positive cells were anti-GH positive and anti-prolactin negative as the lentiviral expression. However, GFP was transiently expressed by developing lactotrophs at 8 and 16 DPI, suggesting that our vector lacks an element(s) which suppresses the expression. Meanwhile, the retrovirally labeled cells tended to cluster with the cells of same type. An analysis of cell numbers in each cluster revealed some features of cell proliferation. These viral vectors are shown to be useful tools to monitor the activation of the GH gene and the development of somatotrophs.
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