Endogenous GD3 ganglioside induces apoptosis in U-1242 MG glioma cells.
Endogenous GD3 ganglioside induces apoptosis in U-1242 MG glioma cells.
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内源性 GD3 神经节苷脂诱导 U-1242 MG 神经胶质瘤细胞凋亡。
DOI：
10.1111/j.1471-4159.2005.03640.x
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发表时间：
2006
期刊：
影响因子：
--
通讯作者：
Yates,AJ
中科院分区：
文献类型：
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作者：
Saqr,HE;Omran,O;Dasgupta,S;Yu,RK;Oblinger,JL;Yates,AJ
GD 3神经节苷脂在几种细胞类型中诱导凋亡，但通过其发生的分子事件在很大程度上是未知的。我们使用U-1242 MG胶质母细胞瘤细胞研究了GD 3表达的凋亡效应，因为这些细胞几乎只合成GM 3和GM 2，而不合成GD 3。为了在TetOn系统的控制下以最小的泄漏表达GD 3，我们通过构建含有由两个内部核糖体进入位点分开的三个基因的单个三顺反子逆转录病毒载体来修改系统：（a）转录沉默子，tTS;（B）逆转录激活子的突变体，rtTA 2（S）-M2（由H. Bujard，Heidelberg，德国）;和（c）增强的绿色荧光蛋白（EGFP），作为三顺反子基因表达的指示剂。使用流式细胞术，我们选择了响应于多西环素而表达高水平GD 3的胶质瘤细胞（U1242 MG-GD 3克隆）。GD 3的表达与细胞凋亡相关，如通过膜联蛋白V结合、TdT介导的dUTPnick end labeling assay（TUNEL）和EGFP降解所证实的。GD 3诱导的细胞凋亡通过caspase-8激活发生，因为GD 3导致caspase-8裂解，zlETD-favor对caspase-8激活的抑制使GD 3诱导的细胞凋亡最小化。
GD3 ganglioside induces apoptosis in several cell types, but the molecular events through which this occurs are largely unknown. We investigated the apoptotic effects of GD3 expression using U‐1242 MG glioblastoma cells, as these cells synthesize almost exclusively GM3 and GM2 but not GD3. To express GD3 under the control of the TetOn system with minimum leakage, we modified the system by constructing a single tri‐cistronic retrovirus vector containing three genes separated by two internal ribosome entry sites: (a) transcriptional silencer, tTS; (b) mutant of reverse transcriptional activator, rtTA2(S)‐M2 (provided by H. Bujard, Heidelberg, Germany); and (c) enhanced green fluorescent protein (EGFP), as an indicator of the tri‐cistronic gene expression. Using flow cytometry, we selected glioma cells (U1242MG‐GD3 clone) that express high levels of GD3 in response to doxycycline. Expression of GD3 was associated with apoptosis as verified by annexin‐V binding, TdT‐mediated dUTPnick end‐labelling assay (TUNEL), and EGFP degradation. GD3‐induced apoptosis occurred via caspase‐8 activation, as GD3 caused cleavage of caspase‐8 and inhibition of caspase‐8 activation by zlETD‐fmk minimized GD3‐induced apoptosis.