褪黑素对大鼠胸腺细胞凋亡及线粒体途径凋亡调控蛋白的影响

发布时间：2018-04-30 01:01

本文选题：褪黑素 + 胸腺细胞　；参考：《福建医科大学》2008年硕士论文

【摘要】： 目的通过研究体外培养大鼠胸腺细胞的自然凋亡和褪黑素对其凋亡的影响及机制,探索褪黑素影响胸腺细胞凋亡的机制,进一步揭示褪黑素与胸腺发育及细胞免疫之间的关系,从而丰富对神经-内分泌-免疫网络的认识。方法①体外培养大鼠胸腺细胞,分为空白对照组(Ctrl)和褪黑素干预组M1、M2、M3、M4、M5(药物浓度分别为10~(-3)、10~(-4)、10~(-6)、10~(-8)、10~(-10)mol/L)。②在培养的第4、8小时以Annexin-V/PI双标法染色及流式细胞仪检测,观察不同浓度褪黑素干预下胸腺细胞凋亡率的变化。③用RT-PCR法及凝胶成像分析系统观察分析培养的第4、8小时,不同浓度褪黑素干预下胸腺细胞Bax、Bcl-xL、Bim mRNA量的变化。④运用western blot法观察培养的第4、8小时,不同浓度褪黑素干预下胸腺细胞Bax、Bcl-xL和Bim蛋白表达量的变化。结果①流式细胞检测结果显示,培养4小时,FITC~+/ PI~-的大鼠胸腺细胞高于8小时,相同培养时间的M1、M2、M3组FITC~+/ PI~-细胞明显下降。②半定量PT-PCR及Western blot结果显示,Bax和Bim的表达随时间延长而增高,mRNA与蛋白水平一致,但对照组与褪黑素干预组之间无明显差异;相同培养时间的M1、M2、M3组与对照组相比,Bcl-xL mRNA与蛋白水平均出现升高,但同一处理组的4小时与8小时比较无明显变化。结论体外培养的大鼠胸腺细胞随着时间延长而凋亡增加,褪黑素能降低胸腺细胞的早期凋亡率,并呈剂量依赖性。Bax和Bim的表达随培养时间延长而增高,褪黑素干预对其无明显影响;褪黑素干预后Bcl-x_L的表达水平呈剂量依赖性增高,但不随培养时间延长而变化。因此,褪黑素可通过影响线粒体途径从而调控大鼠胸腺细胞的凋亡。
[Abstract]:Objective to study the natural apoptosis of rat thymocytes in vitro and the effect and mechanism of melatonin on thymocyte apoptosis, and to explore the mechanism of melatonin affecting thymocyte apoptosis, and to further reveal the relationship between melatonin and thymus development and cellular immunity. So as to enrich the understanding of neural-endocrine-immune network. Methods 1 Rat thymocytes were cultured in vitro and divided into two groups: control group (Ctrl) and melatonin intervention group (M1OM2M3OM4M5). The concentrations of M1OM2M3OM4M5 (the drug concentrations were 10OTr / L5) were determined by Annexin-V/PI double labeling method and flow cytometry at 48 hours after culture. The apoptosis rate of thymocytes was observed at different concentrations of melatonin. 3 the culture was observed by RT-PCR method and gel imaging analysis system at the 4th hour of culture. Changes of Bax-xLnBim mRNA in thymocytes treated with melatonin at different concentrations. The expression of Bax-xL and Bim protein in thymocytes was observed by western blot method at the 8th hour of culture and by different concentrations of melatonin. Results 1 the results of flow cytometry showed that the thymocytes cultured for 4 hours in FITC- / PII- were higher than 8 hours. The expression of Bax and Bim in the FITC- / PII-M3 group decreased significantly with the increase of time, but there was no significant difference between the control group and the melatonin intervention group. The levels of Bcl-xL mRNA and protein in M1M2M3 group were higher than those in control group, but there was no significant change in the same treatment group at 4 hours and 8 hours. Conclusion the apoptosis of rat thymocytes in vitro increases with time. Melatonin can decrease the early apoptosis rate of thymocytes and increase the expression of .Bax and Bim in a dose-dependent manner. The level of Bcl-x_L expression was increased in a dose-dependent manner, but did not change with the prolongation of culture time. Therefore, melatonin can regulate the apoptosis of rat thymocytes by affecting the mitochondrial pathway.
【学位授予单位】：福建医科大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R392

【参考文献】