白花丹素对人舌癌Tca细胞增殖及凋亡作用的探讨

Effect of Plumbapin on Cell Proliferation and Apoptosis in Tongue Cancer Cell Line

目的探讨白花丹抑制舌癌细胞Tca的增殖及对细胞凋亡的影响。方法将舌癌细胞Tca按5×103个/孔的浓度接种于96孔细胞培养板,置于37℃,5%CO2的培养箱中预培养24 h,待细胞80%长满孔底,24 h后将白花丹分别按0.025,0.1,0.2,0.3 mg/ml和0.4 mg/ml分别加入各培养组中,每组5复孔。置37℃、5%CO2培养箱中培养。分别培养48,72,96,120 h后,用酶标仪在490 nm下,MTT染色法测定细胞增殖情况,计算白花丹对肿瘤细胞的抑制率。同时,Hoe-chest33342荧光染色法测定细胞核变化情况,观察细胞凋亡的形态学变化情况。结果随着白花丹浓度的增大和作用时间的延长,白花丹对人舌癌细胞增殖的抑制作用越明显。当药物浓度达0.4 mg/ml时,培养48 h时与其他各组比较有统计学意义(P<0.05或P<0.01),当药物浓度在0.3 mg/ml细胞培养达96 h时,与小于0.3 mg/ml和小于96 h培养时间的其他各组比较均有显著性差异(P<0.05),说明此点是白花丹有效成分抑制肿瘤细胞的一个关键点。细胞培养72 h后Hoechst33342染色发现,随着药物浓度的不断增加,凋亡细胞的数量越来越多。当药物浓度达到0.4 mg/ml时,镜下几乎很少见到完整的细胞核出现,相反出现了大量的细胞和碎片和不完整的细胞核。结论白花丹提取物可以有效抑制肿瘤细胞增长,导致肿瘤细胞大量地细胞凋亡,说明其在肿瘤治疗中具有很大的应用价值。

Objective To investigate the effects of Plumbapin on the proliferation and apoptosis of tongue cancer cell line Tca.Methods The tongue cancer cells were put into 96-well plates（5×104 cells /well） and cultivated at 37℃,5% CO2 for 48 h.When cells grew to 80% confluence of the plate,the Plumbapin was added into each group respectively（with 0.025,0.1,0.2,0.3 and 0.4 mg/ml）.After the treated cell cultured for 48,72,96 and 120 h,cell proliferation was evaluated by ELISA Reader at 490 nm with MTT assay.At mean time,apoptotic cell was detected by immunofluorescence technique with Hoechst33342 staining.Results Inhibition rate of tongue cancer cell was obviously enhanced with the increase of Plumbapin concentration.When the cells were treated with Plumbapin（0.4 mg/ml） for 48 h,statistical significance was shown compared with the other treated groups（P0.05）.When the cells were cultured with Plumbapin（0.3 mg/ml）for 96h,cell apoptosis was significantly decreased compared with the groups of less than 0.3 mg/ml（P0.05）.Hoechst33342 staining revealed that with the concentration increased,cell apoptosis rate was raised after 72 h.When the concentration of Plumbapin over 0.4 g/ml,complete nucleus was seldom seen under Fluorescence microscope instead of a large number of incomplete cell nucleus,nucleus debris and apoptotic body.Conclusion The Plumbapin can effectively inhibit tumor cell growth and induce tumor cell apoptosis.