Establishment and evaluation of a stable steroidogenic caprine luteal cell line
Establishment and evaluation of a stable steroidogenic caprine luteal cell line
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稳定的类固醇生成山羊黄体细胞系的建立和评价
DOI：
10.1016/j.theriogenology.2012.01.009
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发表时间：
2012-07-15
期刊：
影响因子：
2.8
通讯作者：
Tong, Dewen
中科院分区：
文献类型：
--
作者：
Li, Wei;Xu, Xingang;Tong, Dewen
许多生理学、生物学、药理学和毒理学事件和化合物影响萨宁奶山羊黄体细胞的功能，导致着床失败或早期胚胎丢失。虽然原代黄体细胞培养已被使用，其有限的寿命排除了长期影响的评估。在本研究中，原代山羊黄体细胞（CLC）通过转染含有人端粒酶逆转录酶（hTERT）基因的质粒永生化。在转导的CLC（hTERT-CLC）中检测hTERT的表达和端粒酶活性。在这项研究中，这些细胞稳定表达hTERT基因，并在第30代和第50代表现出较高的端粒酶活性。第30代和第50代的hTERT-CLC表达编码正常黄体细胞固有的关键蛋白质、酶和受体的基因，例如，类固醇生成急性调节蛋白（星星）、细胞色素P450胆固醇侧链裂解酶（P450 SCC）、3 β-羟基类固醇脱氢酶（3 β-HSD）和LH-受体（LH-R）。此外，永生化山羊黄体细胞产生可检测数量的孕酮，以响应8-溴-cAMP（8-Br-cAMP）或22（R）-羟基胆固醇（22 R-HC）刺激。此外，该细胞系似乎比对照细胞增殖更快，尽管在体内或体外均未发生肿瘤转化。结论：hTERT永生化的CLC保留了其原有的特性，为研究黄体细胞功能提供了一个有用的模型。(c)2012 Elsevier Inc. All rights reserved.
Many physiological, biological, pharrnacologic, and toxicologic events and compounds affect the function of Saanen dairy goat luteal cells, resulting in implantation failure or early embryonic loss. Although primary luteal cell cultures have been used, their finite lifespan precludes assessment of long-term effects. In the present study, primary caprine luteal cells (CLCs) were immortalized through transfection of a plasmid containing the human telomerase reverse transcriptase (hTERT) gene. The expression of hTERT and telomerase activity were evaluated in transduced CLCs (hTERT-CLCs). In this study, these cells steadily expressed hTERT gene and exhibited higher telomerase activity at Passages 30 and 50. The hTERT-CLCs at Passages 30 and 50 expressed genes encoding key proteins, enzymes and receptors inherent to normal luteal cells, e.g., steroidogenic acute regulatory protein (StAR), cytochrome P450 cholesterol side-chain cleavage enzyme (P450scc), 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD), and LH-receptor (LH-R). In addition, immortalized caprine luteal cells produced detectable quantities of progesterone in response to 8-bromo-cAMP (8-Br-cAMP) or 22(R)-hydroxycholesterol (22R-HC) stimulation. Furthermore, this cell line appeared to proliferate more quickly than control cells, although no neoplastic transformation occurred either in vivo or in vitro. We concluded the immortalized CLCs by hTERT retained their original characteristics and may provide a useful model to study luteal cell functions. (c) 2012 Elsevier Inc. All rights reserved.