Establishment and characterization of an ovarian cell line from half-smooth tongue sole Cynoglossus semilaevis
Establishment and characterization of an ovarian cell line from half-smooth tongue sole Cynoglossus semilaevis
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DOI：
10.1111/jfb.12535
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发表时间：
2015-01-01
影响因子：
2
通讯作者：
Chen, S. L.
中科院分区：
文献类型：
--
作者：
Sun, A.;Wang, T. Z.;Chen, S. L.
以半滑舌鳎为材料，建立了一个新的卵巢细胞系CSO。CSO细胞的原代培养起始于用胰蛋白酶溶液消化卵巢组织块，并在24 ℃下在补充有20%胎牛血清、碱性成纤维细胞生长因子（bFGF）、表皮生长因子（EGF）、胰岛素样生长因子-I（IGF-I）和人绒毛膜促性腺激素（HCG）的Dulbecco改良Eagle培养基-F12培养基（DMEM-F12，1：1）（pH 70）中培养。培养的CSO细胞，形态成纤维细胞，3天后增殖至100%汇合，并已传代至第80代。染色体分析表明，CSO细胞染色体非整倍性，众数为42，显示正常的二倍体核型。semilaevis [2n=42t，fundamental number（N-F）= 42].逆转录聚合酶链反应显示CSO细胞可表达卵巢体细胞功能基因p450 armo、foxl 2和sox 9a，但不表达卵巢生殖细胞标志基因vasa和雄性特异基因dmrt 1。转染实验表明，pEGFP-N3质粒转染的CSO细胞能表达绿色荧光蛋白（GFP），且转染效率较高。CSO细胞系可能为研究比目鱼性别决定和卵巢卵子发生机制提供有价值的工具。
A new ovarian cell line, CSO, was established from half-smooth tongue sole Cynoglossus semilaevis. Primary culture of CSO cells was initiated from digestion of ovarian tissues pieces by trypsin solution and cultured at 24 degrees C in Dulbecco's modified Eagle's medium-F12 medium (DMEM-F12, 1:1) (pH 70), supplemented with 20% foetal bovine serum, basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), insulin-like growth factor-I (IGF-I) and human chorionic gonadotropin (HCG). The cultured CSO cells, fibroblastic in morphology, proliferated to 100% confluency 3 days later and had been subcultured to passage 80. Chromosome analyses indicated that the CSO cells exhibited chromosomal aneuploidy with a modal chromosome number of 42 that displayed the normal diploid karyotype of C. semilaevis [2n=42t, fundamental number (N-F) = 42]. Reverse transcription polymerase chain reaction revealed that CSO cells could express ovarian somatic cell functional genes p450armo, foxl2 and sox9a but not ovary germ cell marker gene vasa and male-specific gene dmrt1. Transfection experiment demonstrated that CSO cells transfected with pEGFP-N3 plasmid could express green fluorescence protein (GFP) with higher transfection efficiency. The CSO cell line might serve as a valuable tool for studies on the mechanism of sex determination and oogenesis of ovary in flatfish.