大泷六线鱼鳍、吻端和肾脏组织原代培养的研究

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、20 μg/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.

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

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.