【摘要】 目的:研究鬼臼乙叉甙(Etoposide,VP16)对表达β-1,4-半乳糖基转移酶V(beta-1,4-galactosyltransferaseV,β-1,4-GalTV)的SHG-44胶质瘤细胞死亡和凋亡的影响,为进一步探讨β-1,4-GalTV与化疗药物敏感性的关系奠定基础。方法:通过流式细胞仪(FCM)和Hoechst33258染色法,分析不同浓度的VP16处理转染了正义(GalTV-HA/SHG-44)、反义β-1,4-GalTV(GalTV-AS/SHG-44)和空载体(Mock)的SHG-44细胞后细胞的凋亡和死亡情况。结果:①低浓度的VP16(20μmoL/L)处理24h后,GalTV-HA/SHG-44组的死亡细胞加上凋亡细胞的比例明显低于Mock组(t=4.5506,P<0.01)和GalTV-AS/SHG-44组(t=5.0865,P<0.001)。GalTV-AS/SHG-44组的死亡细胞加上凋亡细胞比例明显高于Mock组(t=3.2873,P<0.01)。高浓度的VP16(120μmoL/L)处理48h和72h后,GalTV-HA/SHG-44组死细胞所占比例明显低于Mock组t=3.3664,(P<0.05;t=5.4953,P<0.01)和GalTV-AS/SHG-44组(t=3.1732,P<0.01;t=4.6035,P<0.001)。②MTT分析显示,VP16处理48,72h后,GalTV-HA/SHG-44组活细胞数明显高于Mock组(t=5.0783,P<0.05;t=3.8923,P<0.01)和GalTV-AS/SHG-44组(t=4.5667,P<0.01;t=4.7849,P<0.001)。③VP16(20μmoL/L)处理24h后,用Hoechst33258染色。镜下观察到死亡或者凋亡细胞中的细胞核更多还原

【Abstract】 AIM:To study the effect of etoposide (VP16) on the death and apoptosis of SHG 44 glioma cell which expresses beta 1, 4 galactosyltransferase Vβ 1,4 GalT V), so as to establish basis for further exploration of the relationship between β 1,4 GalT V and sensitivity of anti tumor drugs. METHODS: The effects of VP16 of different concentrations on the apoptosis and death of SHG 44 cells transfected with senseβ 1,4 GalT V (GalTV HA/SHG 44), antisense β 1,4 GalT V (GalTV AS/SHG 44) and empty vector (Mock) were detected by flow cytometry (FCM) and Hoechst 33258 staining. RESULTS: ①Twenty four hours after dealing with VP16 of low concentration (20 μmol/L), the ratio of dead cells and apoptosis cells in the GalTV HA/SHG 44 group was obviously higher than those in the Mock group (t=4.550 6, P< 0.01) and GalTV AS/SHG 44 group (t=5.086 5, P< 0.001), and that in the GalTV AS/SHG 44 group was obviously higher than that in the Mock group (t=3.287 3, P >0.01). After dealing with VP16 of high concentration (120 μmoL/L) for 48 and 72 hours, the proportion of death cells in the GalTV HA/SHG 44 group was obviously lower than that in the Mock group (t=3.366 4, P< 0.05;t=5.495 3, P< 0.01) and the GalTV AS/SHG 44 group (t=3.173 2, P< 0.01;t=4.603 5, P< 0.001). ② MTT analysis showed that after dealing with VP16 for 48 and 72 hours, the number of live cells in the GalTV HA/SHG 44 group was obviously high than that in the Mock group (t=5.078 3, P< 0.05;t=3.892 3, P< 0.01) and GalTV AS/SHG 44 group (t=4.566 7, P< 0.01;t=4.784 9, P< 0.001). ③ After dealing with VP16 for 24 hours, Hoechst 33258 staining was performed. Observation under fluorescence microscope showed that the cell nucleolus and disrupted segment of nucleus were showed crescent shape, or karyopyknosis and deeply dyed. The mean positive rate of death and apoptosis in the GalTV AS/SHG 44 group was 41%, significantly higher than Mock and GalTV HA/SHG 44 dealt with the same method. CONCLUSION: β 1,4 GalT V can influence the killing and injuring effect of VP16 on SHG 44 cells, and overexpression of β 1,4 GalT V can antagonize the effect of VP16, which provides new evidence for the explanation of the anti tumor tolerance of glioma.更多还原