大泷六线鱼鳍、吻端和肾脏组织原代培养的研究

Primary culture of fin,lip and kidney cells in fat greenling Hexagrammos otakii

应用组织块培养法,研究了大泷六线鱼Hexagrammos otakii鳍、吻端和肾脏细胞在含有体积分数为20%胎牛血清(FBS)的L-15、DMEM和DMEM/F12 3种培养基中的生长情况,以及人碱性成纤维细胞(bFGF)、硫酸软骨素、Ⅰ型胰岛素样(IGF-Ⅰ)3种生长因子对细胞贴壁率和迁出率的影响。结果表明:大泷六线鱼鳍、吻端和肾脏细胞适宜的体外培养体系为DMEM/F12培养基+20%FBS,pH为7.2,温度为25℃;在最适培养基中分别添加5.0 ng/mL bFGF、20μg/mL硫酸软骨素、40 ng/mL IGF-Ⅰ时,3种组织的贴壁率和细胞迁出率均最高;鳍、吻端和肾脏组织分别经培养21、15、18 d后长满单层,主要为成纤维样细胞;采用低渗滴片法制备染色体,3种组织原代细胞的染色体数目为48条。本研究为大泷六线鱼鳍、吻端和肾脏组织细胞系的构建和生物学特性的研究奠定了基础。

In vitro culture performance of fin,lip and kidney cells was compared in fat greenling Hexagrammos otakii by explant method with three media（L-15,DMEM and DMEM/F12） and three growth factors（bFGF,chondroitin sulfate and IGF-Ⅰ）.Depending on the migrating rate,anchorage-dependent rate and morphology,the optimal culture system for fat greenling fin,lip and kidney cells was found to be DMEM/F12 medium supplemented with 20% FBS,at pH 7.2 and 25 ℃,while the maximal attachment rate and cell emigration were observed in the cell culture media supplemented with 5.0 ng/mL bFGF,20 μg/mL chondroitin sulfate or 40 ng/mL IGF-Ⅰ.The cells in primary culture were all in fibroblastic morphology and proliferated to confluence for the fin in 21 d,lip in 15 d and kidney in 18 days,with 2n=48 chromosomes.The findings provide a useful foundation with the establishment of in vitro cell-line and the functional investigation of fin,lip and kidney cells in fat greenling.