孕酮对原代培养小胶质细胞及BV-2小胶质瘤细胞增殖效应的影响

发布时间：2018-03-23 21:36

本文选题：孕酮　切入点：小胶质细胞　出处：《兰州大学》2014年硕士论文

【摘要】：孕酮是人体内的一种类固醇激素类物质,其对于神经系统类疾病的治疗作用已经为广大的学者所认可。近年来,孕酮已经被应用于临床实验中对神经元及神经胶质细胞的损伤进行治疗。但到目前为止,虽然发现孕酮对治疗中风类疾病具有一定的效果,但是其具体的作用机理还缺乏详细的研究。由于孕酮的作用涉及的过程比较复杂,除了对神经元具有保护作用外,还对神经胶质细胞起到一定的作用。但是,关于孕酮对小胶质细胞进行处理后所发挥的效果的研究还很有限。实验采用小胶质细胞带有GFP荧光标记的转基因小鼠新生鼠作为实验材料,进行原代小胶质细胞培养,同时采用BV-2小胶质瘤细胞进行传代培养。对两种细胞分别进行氧糖剥夺模型(oxygen and glucose deprivation, OGD)的实验处理后,再使用适量的孕酮对其进行处理,探索孕酮对小胶质细胞的作用。本研究主要采用了Hoechst33258荧光染色,Iba-1和BrdU免疫细胞化学染色,MTT细胞活性检测及流式细胞术等实验技术。原代培养的小胶质细胞在OGD处理24小时后,对其使用40μM的孕酮溶液处理后再进行检测,结果发现OGD模型处理可以使小胶质细胞总数降低,凋亡增加。发现孕酮可以促进经过OGD处理的小胶质细胞的增殖,同时还可以降低经过OGD处理的小胶质细胞的凋亡比率。对BV-2小胶质瘤细胞进行OGD处理6小时后,再用10μM的孕酮进行处理。发现OGD模型处理可以使BV-2细胞细胞活力降低,细胞凋亡比例增加,细胞增殖降低。发现经过OGD联合孕酮处理后的BV-2细胞活性上升,细胞周期基本恢复正常,并且细胞的凋亡比例下降,细胞发生增殖。综览上述结果,可以发现OGD模型分别处理原代培养的小胶质细胞和传代培养的BV-2细胞,都可以产生明显的效果。之后再用适宜剂量的孕酮对经过OGD处理后的原代培养的小胶质细胞和BV-2细胞进行处理,发现孕酮促进了细胞活力的恢复,同时促进了细胞的增殖。因此,孕酮对于经过OGD处理的小胶质细胞的确会产生一定的保护作用,为临床中孕酮的使用提供了一定的理论依据。
[Abstract]:Progesterone is a steroid hormone in human body, and its therapeutic effect on nervous system diseases has been recognized by many scholars in recent years. Progesterone has been used in clinical trials to treat injuries to neurons and glial cells. But so far, progesterone has been found to be effective in the treatment of apoplectic diseases. However, the specific mechanism of the action is still lack of detailed study. As the process involved in progesterone is relatively complex, in addition to the protection of neurons, it also plays a certain role in neuroglial cells. Studies on the effects of progesterone on microglia are limited. The primary microglia of transgenic mice with GFP fluorescent labeling were used as experimental materials. At the same time, BV-2 small glioma cells were subcultured. The two kinds of cells were treated with oxygen and glucose privacy, and then treated with appropriate amount of progesterone. In order to explore the effect of progesterone on microglia, Hoechst33258 fluorescence staining and BrdU immunocytochemical staining were used to detect the cell activity and flow cytometry. The primary cultured microglial cells were treated with OGD for 24 hours, then treated with 40 渭 M progesterone solution. The results showed that OGD model treatment could reduce the total number of microglia cells. Apoptosis increased. It was found that progesterone could promote the proliferation of microglia treated with OGD and decrease the apoptosis rate of microglia treated with OGD. Then treated with 10 渭 M progesterone, it was found that OGD model treatment could decrease the viability, increase the proportion of apoptosis and decrease the proliferation of BV-2 cells. The activity of BV-2 cells treated with OGD combined with progesterone was increased. The cell cycle returned to normal, and the proportion of apoptosis decreased and cell proliferation occurred. After reviewing the above results, we can find that the OGD model deals with the primary cultured microglia and the subcultured BV-2 cells, respectively. Then the primary cultured microglia and BV-2 cells treated with OGD were treated with the appropriate dose of progesterone, and it was found that progesterone promoted the recovery of cell viability. Therefore, progesterone does have a protective effect on microglia treated with OGD, which provides a theoretical basis for the use of progesterone in clinic.
【学位授予单位】：兰州大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R739.41

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