CAJ | 학술논문

目的研究顺铂在增加人脑胶质瘤细胞U251对肿瘤坏死因子相关凋亡诱导配体(TRAIL)敏感性的作用,并探讨其分子机制.方法通过倒置荧光显微镜下观察重组腺病毒载体携带的绿色荧光蛋白的表达确定最适感染复数MOI;运用MTT法检测细胞增殖活性;倒置荧光显微镜下观察Hoechst33342染色细胞凋亡形态;PI染色后通过流式细胞术检测诱导凋亡作用,RT-PCR法检测凋亡相关基因.结果感染后TRAIL基因的表达明显上调.顺铂增敏TRAIL组与单独组相比,有显著抑制细胞增殖作用(P<0.05),Hoechst33342染色观察有明显的核固缩,核碎裂.流式细胞术检测细胞有凋亡峰的出现,增敏组与单独组有显著性差异(P<0.05).RT-PCR检测到TRAIL、DB5、caspase-3基因上调,survivin基因下调,DB4表达无明显变化.结论顺铂可以增加人脑胶质瘤细胞U251对TRAIL的敏感性,其分子机制可能与DR5、caspase-3基因上调,survivin基因下调有关.
목적 연구순박재증가인뇌효질류세포U251대종류배사인자상관조망유도배체(TRAIL)민감성적작용,병탐토기분자궤제.방법 통과도치형광현미경하관찰중조선병독재체휴대적록색형광단백적표체학정최괄감염복수MOI;운용MTT법검측세포증식활성;도치형광현미경하관찰Hoechst33342염색세포조망형태;PI염색후통과류식세포술검측유도조망작용,RT-PCR법검측조망상관기인.결과 감염후TRAIL기인적표체명현상조.순박증민TRAIL조여단독조상비,유현저억제세포증식작용(P<0.05),Hoechst33342염색관찰유명현적핵고축,핵쇄렬.류식세포술검측세포유조망봉적출현,증민조여단독조유현저성차이(P<0.05).RT-PCR검측도TRAIL、DB5、caspase-3기인상조,survivin기인하조,DB4표체무명현변화.결론 순박가이증가인뇌효질류세포U251대TRAIL적민감성,기분자궤제가능여DR5、caspase-3기인상조,survivin기인하조유관.
Objective To evaluate the positive effects of cisplatin on sensitivity of human glioma U251 to tumor necrosis factor-related apoptosis inducing ligand and to investigate the potential mechanism. Methods The expression of green fluorescent protein (GFP) in U251 which was transfected with pAdxsi-GFP-TRAIL was observed by inverted fluorescent microscope ×400) and to ascertain the MOI. The proliferation inhibition was studied by MTT method. Morphological change was detected through inverted florescent microscope and the Hoechst33342 staining assay was used to verify whether cell apoptosis could be induced or not. The cell apoptosis was also analyzed by flow cvtometry with propidium iodide staining. Semi-quantitative RT-PCR was introduced to detect the mRNA expression of apoptosis related gene.Results The expression of TRAIL mRN A was significantly upregulated after transfection. Compared with treatment group of cisplatin and TRAIL alone, the proliferation of U2S1 was significantly inhibited in the cisplatin sensitizing TRAIL group (P < 0.05 ). Nuclear shrinkage and pyknosis fragmentation were observed by Hoechst 33342 staining assay; Apoptotic peak was detected from the results of flow cvtometry and there were significant differences between the sensitizing group and the other two groups ( P < 0.05 ) ; Moreover, the relatively high expression of TRAIL, DR5, caspaseS and down - regulated survivin genes were also observed. There was no significant changes in DR4 expression. Conclusion Cisplatin could extremely enhance the sensitivity of U251 cells to TRAIL And the potential mechanism may related to the increase of TRAIL, DRS, caspaseS genes while the reduction of surivivin gene.