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目的 在培养的心脏成纤维细胞模型上建立检测增殖的方法。 方法 (1)差速贴壁分离法培养乳鼠心脏成纤维细胞 ,用光镜、电镜及免疫细胞化学染色进行鉴定。 (2 )同步测定心脏成纤维细胞在不同浓度胎牛血清作用下 Brd U掺入和 WST- 1代谢活性。 结果 (1)培养的细胞经形态学观察和波形蛋白 SP染色 ,鉴定为成纤维细胞。 (2 )在 0 ,1% ,2 .5 % ,5 %和 10 %的胎牛血清作用下 ,心脏成纤维细胞 Brd U ,WST- 1的 OD值分别是 0 .186± 0 .0 2 5 ,0 .30 0± 0 .0 73;0 .46 6± 0 .0 49,0 .5 31± 0 .0 72 ;0 .76 1± 0 .0 39,0 .716± 0 .0 6 4;0 .834± 0 .0 48,0 .833±0 .0 73和 0 .90 5± 0 .0 6 4,0 .935± 0 .0 5 3(P<0 .0 1)。 结论 (1) WST- 1可在活的心脏成纤维细胞代谢 ,WST- 1法适用于该细胞的增殖。 (2 ) Brd U、WST- 1EL ISA可同步测定心脏成纤维细胞的 DNA合成及其代谢活性 ,是检测细胞增殖的新方法。
Objective To establish a method for detecting proliferation in cultured cardiac fibroblast models. Methods (1) The fibroblasts of neonatal rat cardiac fibroblasts were cultured by differential adherence method and identified by light microscope, electron microscope and immunocytochemical staining. (2) Simultaneous determination of Brd U incorporation and WST-1 metabolic activity of cardiac fibroblasts under different concentrations of fetal bovine serum. Results (1) The cultured cells were identified as fibroblasts by morphological observation and vimentin SP staining. (2) Under the action of 0, 1%, 2.5%, 5% and 10% fetal calf serum, the OD values of BrdU and WST-1 in cardiac fibroblasts were 0.186 ± 0.255 , 0 .30 0 ± 0 .0 73; 0 .46 6 ± 0 .0 49,0 .5 31 ± 0 .0 72; 0 .76 1 ± 0 .0 39,0 .716 ± 0 .0 6 4 ; 0 .834 ± 0 .0 48,0 .833 ± 0 .0 73 and 0 .90 5 ± 0 .0 6 4,0 .935 ± 0 .0 5 3 (P <0.01). Conclusion (1) WST-1 can be metabolized in live cardiac fibroblasts, WST-1 method is suitable for the proliferation of the cells. (2) BrdU and WST-1EL ISA can simultaneously determine DNA synthesis and metabolic activity of cardiac fibroblasts, which is a new method to detect cell proliferation.