猪呼吸道上皮细胞的体外原代培养研究

Studies on primary culturing of porcine tracheal epithelial cells

为探索猪呼吸道上皮细胞体外的原代培养,对胶原酶Ⅳ+胰酶消化法、胰酶消化法、胶原酶Ⅳ和胰酶消化结合组织块培养法以及组织块培养法等4种分离细胞的方法进行了比较,同时比较了不同浓度胎牛血清的培养基所培养细胞的纯度及成活率,采用差速贴壁法纯化细胞并对所得的上皮细胞进行冻存复苏研究。结果表明:组织块培养法培养获得的细胞数量和纯度极显著高于胰酶消化法(P<0.01),胶原酶Ⅳ+胰酶消化法获得的细胞成活率显著高于胰酶消化法;含有5%血清的培养基培养的细胞成活率及纯度均高于用10%血清培养(P<0.01),冻存1个月后的细胞复苏率为85%。

To explore the method of culturing primary porcine respiratory tract epithelial cells, four methods were compared： collagenase Ⅳ digestion ＋trypsin digestion, trypsin digestion, collagenase Ⅳ digestion ＋trypsin digestion＋ tissue culture, tissue culture; Subsequently the cells were cultured in culture medium with 5% and 10% fetal bovine serum（FBS）.The quantity, purity and viability of cells isolated by three methods were compared. Purity cells with the differential adhesion method ,the cryopreservation and resuscitated to tracheal epithelial cells were studied. The result indicated that the quantity,purity and viability of cells obtained from tissue culture were significantly higher than that of other methods（P〈0.01）. The viability and purity of cells in culture medium with 5% FBS were significantly higher than that of the cells in culture medium with 10% FBS（P〈0.01）.After cryopreservation for one month, eighty percents cells were successfully resuscitated.