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目的确定RNA合成抑制剂——放线菌素D(actinomycin D,ACTD)诱发V79细胞凋亡的最适作用浓度和时间,建立细胞凋亡模型。方法采用不同剂量(0～8.0 mg/L)的ACTD染毒V79细胞不同时间(0.5～4 h)。采用MTT法检测细胞活力;采用流式细胞仪法分析细胞凋亡和坏死率。结果各剂量ACTD染毒不同时间后V79细胞活力均下降(P<0.05)。与染毒0.5 h比较,在染毒1 h后仅0.25、1、2、8 mg/L ACTD染毒V79细胞活力下降,在染毒2 h后0.25、1、2、4、8 mg/L ACTD染毒V79细胞活力下降(P<0.05),在染毒4 h后各剂量ACTD染毒V79细胞活力均下降(P<0.05)。各剂量ACTD染毒不同时间后V79细胞凋亡率均升高(P<0.05);与染毒0.5 h比较,各剂量ACTD染毒1 h以及0.25、0.5、1、2 mg/L ACTD染毒2、4 h后V79细胞凋亡率均升高(P<0.05),4 mg/L ACTD染毒4 h后V79细胞凋亡率下降(P<0.05)。与对照组比较,0.25、2mg/L ACTD染毒0.5 h,1、2、4 mg/L ACTD染毒2 h以及各剂量ACTD染毒4 h后V79细胞坏死率均升高(P<0.05);与染毒0.5 h比较,2 mg/L ACTD染毒1 h后V79细胞坏死率下降(P<0.05),1、4 mg/L ACTD染毒2 h以及0.5、1、2、4 mg/L ACTD染毒4 h后V79细胞坏死率均升高(P>0.05)。结论 ACTD诱发V79细胞凋亡的最适剂量为4 mg/L,最佳作用时间为1 h。
Objective To determine the optimal concentration and time for the apoptosis of V79 cells induced by actinomycin D (ACTD), an inhibitor of RNA synthesis, and to establish a cell apoptosis model. Methods V79 cells were treated with different doses (0 ~ 8.0 mg / L) of ACTD for different times (0.5 ~ 4 h). Cell viability was detected by MTT assay. Apoptosis and necrosis rates were analyzed by flow cytometry. Results The viability of V79 cells in each dose of ACTD decreased at different time points (P <0.05). Compared with 0.5 h, the viability of V79 cells treated with ACTD at doses of 0.25, 1, 2, and 8 mg / L for 1 h decreased significantly, and at 0.25, 1, 2, 4 and 8 mg / L The viability of V79 cells in ACTD-treated V79 cells decreased (P <0.05), and the viability of V79 cells in ACTD-treated cells decreased after 4 h (P <0.05). The apoptotic rate of V79 cells was increased after each dose of ACTD exposure (P <0.05). Compared with the 0.5 hour exposure, ACTD exposure for 1 h and 0.25,0.5,1,2 mg / L ACTD After 2 and 4 h, the apoptosis rate of V79 cells increased (P <0.05). After 4 mg / L ACTD treatment, the apoptosis rate of V79 cells decreased (P <0.05). Compared with the control group, the necrosis rates of V79 cells in 0.25, 2 mg / L ACTD exposure 0.5 h, 1,2,4 mg / L ACTD exposure 2 h and ACTD 4 h (P <0.05) ; Compared with the 0.5 h exposure, the necrosis rate of V79 cells decreased after 1 mg / L ACTD exposure for 1 h (P <0.05), 2 h at 1 mg / L ACTD and 0.5, 1, 2, 4 mg / After 4 hours of LACTD treatment, V79 cell necrosis rate increased (P> 0.05). Conclusion The optimal dose of ACTD to induce apoptosis of V79 cells is 4 mg / L and the optimal time is 1 h.