CAJ | 학술논문

目的观察叠氮胸苷对恶性胶质瘤细胞系TJ905细胞p33ING1b表达的影响及该影响与细胞凋亡和衰老的关系.方法将体外培养的TJ905细胞系分为对照组,50、100及200 μmol/L叠氮胸苷组,在后三组的培养液中分别给予相应终浓度的叠氮胸苷并继续传代培养;各组取第1、3、6代细胞以半定量逆转录聚合酶链反应(RT-PCR)和衰老相关β-半乳糖苷酶染色检测p33ING1bmRNA表达水平及衰老细胞;取第3、6代细胞以TUNEL法和单细胞凝胶电泳法检测凋亡细胞.结果从用药后第1代起叠氮胸苷即呈时间和剂量依赖性地诱导TJ905细胞p33ING1b mRNA表达和细胞衰老;200 μmol/L叠氮胸苷组第6代TJ905细胞的p33ING1b RT-PCR扩增条带相对灰度(1.44±0.23)显著高于同组第1代(0.95±0.13)、第3代(1.35±0.23)及同代50μmol/L组(0.85±0.24)和100 μmol/L组(1.23±0.34),其衰老相关β-半乳糖苷酶标记指数(45.62±6.74)亦显著高于同组第1代(7.82±2.40)、第3代(26.27±7.17)及同代50 μmol/L组(27.37±6.41)和100 μmol/L组(35.49±5.12),上述差异均有统计学意义(P＜0.01);而叠氮胸苷促凋亡作用出现在稍晚的第6代.结论叠氮胸苷可上调TJ905细胞p33ING1b的表达水平,这可能是叠氮胸苷诱导TJ905细胞衰老和凋亡的重要分子机制.
목적 관찰첩담흉감대악성효질류세포계TJ905세포p33ING1b표체적영향급해영향여세포조망화쇠로적관계.방법 장체외배양적TJ905세포계분위대조조,50、100급200 μmol/L첩담흉감조,재후삼조적배양액중분별급여상응종농도적첩담흉감병계속전대배양;각조취제1、3、6대세포이반정량역전록취합매련반응(RT-PCR)화쇠로상관β-반유당감매염색검측p33ING1bmRNA표체수평급쇠로세포;취제3、6대세포이TUNEL법화단세포응효전영법검측조망세포.결과 종용약후제1대기첩담흉감즉정시간화제량의뢰성지유도TJ905세포p33ING1b mRNA표체화세포쇠로;200 μmol/L첩담흉감조제6대TJ905세포적p33ING1b RT-PCR확증조대상대회도(1.44±0.23)현저고우동조제1대(0.95±0.13)、제3대(1.35±0.23)급동대50μmol/L조(0.85±0.24)화100 μmol/L조(1.23±0.34),기쇠로상관β-반유당감매표기지수(45.62±6.74)역현저고우동조제1대(7.82±2.40)、제3대(26.27±7.17)급동대50 μmol/L조(27.37±6.41)화100 μmol/L조(35.49±5.12),상술차이균유통계학의의(P＜0.01);이첩담흉감촉조망작용출현재초만적제6대.결론 첩담흉감가상조TJ905세포p33ING1b적표체수평,저가능시첩담흉감유도TJ905세포쇠로화조망적중요분자궤제.
Objectives To investigate the pharmacological effects of azidothymidine (AZT) on p33ING1b expression, senescence and apoptosis of TJ905 glioblastoma cells. Methods TJ905 cells were treated with AZT at a serial concentrations of 50, 100 and 200 μmol/L. Semi-quantitative RT-PCR and cytochemical staining of senescence related-galactosidase (sβ-Gal) were used to evaluate the expression of p33ING1b mRNA and to label the senescent cells at the 1st, 3rd and 6th generations, respectively. In situ cell death detection and single cell gel electrophoresis were used to detect the apoptosis at the 3rd and 6th generations. Results AZT induced the expression of p33ING1b mRNA and senescence of the tumor cells of the 1st generation in a dosage and time dependent manner. At the 6th generation, the relative amount of p33ING1b RT-PCR product (1.44 ±0. 23 ) and s3-Gal labeling index of 200 μ mol/L group (45. 62 ±6. 74)were significantly higher than those of the 1st( 0. 95 ± 0. 13 and 7. 82 ± 2. 40 ) and the 3rd generation cells (1.35 ± 0. 23, 26. 27 ± 7. 17 ) of the same group, and cells of the same generation in the 50 μmol/L(0.85 ±0.24, 27.37 ±6.41) and 100 μmol/L groups (1.23 ±0.34, 35.49 ±5.12, P＜0.01 ). Therewas a significant positive correlation between the p33ING1b mRNA expression and the labeling index of sβ-Gal. Pro-apoptotic effects of AZT became obvious at the 6th generation. Conclusion AZT upregulates the expression of p33ING1b, a possible mechanism in regulating senescence and apoptosis of the TJ905 cells.