摘要目的 研究O连接的N-乙酰葡萄糖胺(O-GlcNAc)糖基化及乙酰氨基葡萄糖转移酶(OGT)对Nalm-6细胞生物学行为及对依托泊苷(Vp16)诱导凋亡的影响.方法 利用OGT抑制剂Alloxan构建低O-GlcNAc修饰的Nalm-6细胞模型,CCK-8法检测Alloxan对细胞增殖的影响,流式细胞术检测Alloxan对细胞凋亡及细胞周期的影响;以不同浓度的Vp16处理Nalm-6细胞12 h,Western blot检测O-GlcNAc糖基化修饰程度及OGT表达量的变化;以Alloxan处理Nalm-6细胞24 h后再加入Vp16(5μg/ml)处理12 h,应用流式细胞术检测不同组别细胞凋亡,Western blot检测凋亡相关蛋白Bax和Bcl-2的表达情况.结果 随着Vp16浓度的增加,Nalm-6细胞O-GlcNAc修饰及OGT表达均逐渐上调(P<0.05);Alloxan可抑制Nalm-6细胞增殖,诱发Nalm-6细胞凋亡[(15.190±2.539)%对(21.910±4.105)%,P=0.007],阻滞细胞周期[G1期:(43.534±4.453)%对(57.322±6.091)%,P=0.003;S期:(50.747±5.937)%对(37.201±4.661)%,P=0.001];Alloxan可抑制Vp16诱导的Nalm-6细胞凋亡[(75.195±13.845)%对(52.741±10.815)%,P=0.011],并伴随Bax表达下调(5.496±1.998对2.950±0.703,P=0.015)、Bcl-2表达上调(0.454±0.125对0.803±0.223,P=0.013).结论 通过抑制OGT而改变Nalm-6细胞O-GlcNAc修饰程度可影响其增殖、凋亡、改变细胞周期并抑制Vp16诱导的细胞凋亡.

abstractsObjective To explore the effects of O-GlcNAc glycosylation and its key enzyme OGT on the biological behaviors and etoposide(Vp16)- induced apoptosis of Nalm-6 cells. Methods Low O-GlcNAc modified Nalm-6 cells model was established with Alloxan, an inhibitor of OGT. The influence of Alloxan on Nalm-6 cells proliferation was checked by CCK-8 assay, apoptosis and cell cycle by flow cytometry. Nalm-6 cells were treated with different concentrations of Vp16 for 12 h, and then the O-GlcNAc level and the expressions of OGT were examined by Western blot. After treating Nalm-6 with Alloxan for 24 h and then 5μg/ml of Vp16 for 12 h, the apoptosis of different groups were measured with flow cytometry, and the expression of apoptosis-associated proteins Bax and Bcl-2 were examined by Western blot. Results With the concentration of Vp16 increasing, the O-GlcNAc modified levels of total protein and the expression of OGT were up regulated (P<0.05, n=6); Alloxan could slow down the proliferation capacity, induce apoptosis[(15.190±2.539)%vs(21.910±4.105)%,P=0.007], arrest cell cycle [G1 phase:(43.534 ± 4.453)% vs(57.322 ± 6.091)%, P=0.003; S phase:(50.747 ± 5.937)% vs(37.201 ± 4.661)%, P=0.001]. Alloxan could inhibit the apoptosis caused by Vp16[(75.195±13.845)%vs(52.741± 10.815)%, P=0.011] along with Bax decreasing (5.496 ± 1.998 vs 2.950 ± 0.703, P=0.015) and Bcl-2 increasing(0.454 ± 0.125 vs 0.803 ± 0.223, P=0.013). Conclusion Changes of O-GlcNAc modified level of Nalm-6 cells along with the inhibition of OGT could influence the biological behaviors and inhibit apoptosis induced by Vp16.