| 罗刚,喻坚柏,陈涛,等.异柠檬酸脱氢酶 1基因突变对替莫唑胺干预下脑胶质瘤 U87细胞凋亡的影响[J].安徽医药,2021,25(8):1492-1496. |
| 异柠檬酸脱氢酶 1基因突变对替莫唑胺干预下脑胶质瘤 U87细胞凋亡的影响 |
| Mechanisms of mIDH1 enhancing the therapeutic sensitivity of temozolomide by interfering with U87 cell apoptosis |
| |
| DOI：10.3969/j.issn.1009-6469.2021.08.003 |
| 中文关键词: 神经胶质瘤 人异柠檬酸脱氢酶 1 替莫唑胺 胶质瘤细胞 凋亡率 |
| 英文关键词: Glioma Human isocitrate dehydrogenase 1 Temozolomide Glioma cells Apoptotic rate |
| 基金项目:湖南省自然科学基金( 2018JJ3404) |
|
| 摘要点击次数: 1696 |
| 全文下载次数: 450 |
| 中文摘要: |
| 目的研究异柠檬酸脱氢酶 1(isocitrate dehydrogenase 1，IDH1)基因突变对替莫唑胺干预下脑胶质瘤 U87细胞凋亡的影响，以期深入了解其生物学特性及其化疗敏感性。方法通过 DNA重组技术构建携带野生 IDH1基因(wIDH1)或突变的 IDH1基因(mIDH1)的真核表达载体；通过 CCK-8检测三种稳定细胞系的增殖能力和细胞活力；利用流式细胞仪技术检测稳转细胞系细胞凋亡率及替莫唑胺干预后细胞凋亡率的变化；构建裸鼠移植瘤模型，替莫唑胺干预， ELISA技术和蛋白质免疫印迹检测细胞相关凋亡蛋白［半胱氨酸天冬氨酸蛋白酶 -3(Caspase-3)、半胱氨酸天冬氨酸蛋白酶 -9(Caspase-9)、B淋巴细胞瘤 -2相关蛋白(Bax)和B淋巴细胞瘤 -2(Bcl-2)］的表达情况；测量肿瘤大小为宽和长度，计算肿瘤大小；统计不同处理组肿瘤形成抑制率和总有效率。结果 wIDH1组(100.37±10.24)对照组(102.45±13.57)和mIDH1组(100.63±12.42)之间细胞增殖差异无统计学意义(P>0.05)。mIDH1组U87细胞在 CCK-8测定示出对替莫唑胺的敏感性， mIDH1组( 36.84±3.55)细胞增殖和细胞活力较 wIDH1组( 98.17±8.54)和对照组中显，(73.26±5.37)低(P<0.05)； mIDH1组(62.18±3.47)%细胞的凋亡率高于对照组(33.16±6.54)%和 wIDH1组(15.36±2.33)%；蛋白质免疫印迹和酶联免疫吸附测定(ELISA)结果显示，三组不同处理中， mIDH1组在 Caspase-3，Caspase-9和Bax中的蛋白表达水平最高， Bcl-2的蛋白表达水平最低(P<0.05)，wIDH1的异位过表达诱导 Bcl-2蛋白表达水平的上调及 Caspase-3，Caspase-9和Bax蛋白水平的下调(P<0.05)；与其余两组相比， mIDH1组(220.17±12.15)mm3的肿瘤体积最小(P<0.05)。wIDH1组(624.36±33.54)mm3与对照组(601.17±36.18)mm3差异无统计学意义(P>0.05)；在替莫唑胺治疗组中， mIDH1组的总有效率为(73.15±8.46)%，wIDH1组的总有效率为(22.16±3.96)%。对照组的结果为(42.25±3.17)%。结论 mIDH1通过上调替莫唑胺后胶质瘤细胞 Caspase-3、Caspase-9、Bax表达并下调 Bcl-2表达提高胶质瘤细胞凋亡率，增强治疗敏感性。 |
| 英文摘要: |
| Objective To study the effect of isocitrate dehydrogenase 1 (IDH1) gene mutation on the apoptosis of glioma U87 cellstreated with temozolomide, in order to understand its biological characteristics and chemosensitivity.Methods The eukaryotic expression vector carrying the wild IDH1 gene (wIDH1) or the mutant IDH1 gene (mIDH1) was constructed by DNA recombination technology.The proliferation ability and cell viability of three stable cell lines were detected by CCK-8. Flow cytometry was used to detect the apoptosis rate of stable cell lines and the change in cell apoptosis rate after the intervention of temozolomide. The nude mice xenograft modelwas constructed and temozolomide was adopted for intervention. ELISA and Western blotting were used to detect the expressions of cell-associated apoptosis proteins (Caspase-3, Caspase-9, Bax and Bcl-2). The tumor size was measured as width and length, and the tumorsize was calculated. The tumor formation inhibition rate and the total effective rate of different treatment groups were figured out.Re? sults There was no significant difference in cell proliferation among wIDH1 group (100.37±10.24), control group (102.45±13.57) andmIDH1 group (100.63±12.42) (P>0.05). U87 cells in mIDH1 group showed sensitivity to temozolomide in CCK-8 assay. Cell proliferation and cell viability in mIDH1 group (36.84±3.55) were lower than those in wIDH1 group (98.17±8.54) and control group (73.26±5.37)(P<0.05). The apoptotic rate of cells in mIDH1 group (62.18±3.47)% was higher than that in control group (33.16±6.54)% and wIDH1group (15.36±2.33)%. The protein immunoblotting and enzyme-linked immunosorbent assay (ELISA) results showed that among thethree different treatments, the protein expression levels of Caspase-3, Caspase-9 and Bax were the highest in mIDH1 group, while the protein expression level of Bcl-2 was the lowest in mIDH1 group (P<0.05). The ectopic overexpression of wIDH1 induced the upregulation of Bcl-2 protein expression and the downregulation of Caspase-3, Caspase-9 and Bax protein expresssions (P<0.05). Compared with the other two groups, the mIDH1 group (220.17±12.15) mm3 had the smallest tumor volume (P<0.05). There was no significant difference in tumor volume between wIDH1 group (624.36±33.54) mm3 and control group (601.17±36.18) mm3 (P>0.05). In temozolomide treatmentgroup, the total effective rate of mIDH1 group was (73.15±8.46) %, and the total effective rate of wIDH1 group was (22.16±3.96) %. Theresult of the control group was (42.25±3.17) %.Conclusion mIDH1 can increase the apoptosis rate and the sensitivity of glioma cells by up-regulating the expressions of Caspase-3, Caspase-9 and Bax in glioma cells and down-regulating the expression of Bcl-2. |
|
查看全文
查看/发表评论 下载PDF阅读器 |
| 关闭 |
