环孢素A对PAN损伤足细胞凋亡及p38通路的作用机制研究

发布时间：2018-09-08 18:12

【摘要】：目的：（1）建立氨基核苷嘌呤霉素足细胞损伤模型，观察氨基核苷嘌呤霉素（PAN）对小鼠肾脏足细胞p38、Bax蛋白表达的影响。（2）通过观察环孢素A（CsA）对PAN损伤足细胞p38、Bax蛋白表达的影响，来探讨环孢素A对足细胞凋亡p38MAPK信号通路影响以及其对肾脏的保护作用及其可能机制。方法: 本实验分两部分进行: （1）首先将同步化的足细胞分为11组，对照组用无血清1640培养基；后10组加入50μg/ml的PAN进行培养，其中前6组分别于培养10min、30min、1h、2h、4h、6h收集细胞，Westernblot法检测p-p38蛋白表达；后4组分别于培养12h、24h、36h、48h收集细胞，Western blot法检测Bax蛋白表达。（2）其次将足细胞分为：(a)对照组：无血清1640培养基；(b)PAN组：50μg/ml（c）CsA组：50μg/ml PAN+0.1μmol/L CsA；（d）SB203580组：10μmol/L SB203580预处理细胞1h后加入50μg/ml PAN。培养24h及48h后收集细胞观察细胞凋亡率，培养2h后Western blot法检测p38、p-p38蛋白表达，48h后Bax蛋白表达。结果：（1）PAN作用足细胞10分钟后p38表达开始增加，并随着时间的延长而逐步增加，2小时后活化最为显著，6小时接近基础水平。（2）PAN作用足细胞后，Bax蛋白的表达随时间持续增强，且与对照组相比，24小时后其表达有统计学意义。（3）24小时及48小时PAN组细胞凋亡率较对照组明显增加，差异有统计学意义（P0.05）。CsA组细胞凋亡率较PAN组均显著减少，差异均有统计学意义（P0.05）。（4）与对照组相比，PAN组细胞p-p38、Bax蛋白表达明显增加；而环孢素A组及SB203580组表达量较PAN组明显下降，差异有统计学意义（P0.05）。结论： CsA可减少PAN诱导的足细胞凋亡，起到肾脏保护作用，其机制可能是通过部分抑制p38MAPK活化，降低Bax的表达。
[Abstract]:Objective: (1) to establish the injury model of aminonucleoside purine mycin podocyte. To observe the effect of aminoglycoside purine mycin (PAN) on the expression of p38 (PAN) protein in mouse renal podocytes. (2) to observe the effect of cyclosporine A (CsA) on the expression of p38 Bax-Bax protein in podocytes injured by PAN. To investigate the effect of cyclosporine A on p38MAPK signaling pathway of podocyte apoptosis and its protective effect on kidney and its possible mechanism. Methods: the experiment was divided into two parts: (1) the synchronized podocytes were divided into 11 groups, the control group was cultured on serum-free 1640 medium, the latter 10 groups were cultured with 50 渭 g/ml PAN. In the first six groups, the expression of p-p38 protein was detected by Western blot at 10 min, 30 min, 1 h, 2 h and 4 h, respectively, and the last 4 groups were cultured for 12 h, 24 h, 36 h and 48 h, respectively. (2) podocytes were divided into: (a) control group (serum-free culture medium) and control group (: (a) control group). (b) PAN group: 50 渭 g/ml (c) CsA group: 0: 50 渭 g/ml PAN 0. 1 渭 mol/L CsA; (d) SB203580 group: 10 渭 mol/L SB203580 preconditioning cells for 1 h, then adding 50 渭 g/ml PAN. The apoptotic rate was observed at 24 h and 48 h after culture, and the expression of p38 p-p38 protein was detected by Western blot assay after 2 h culture, and the expression of Bax protein was detected after 48 h culture. Results: (1) the expression of p38 in podocytes began to increase after PAN treatment for 10 minutes. With the time prolonging, the activation reached the basic level at 6 hours after activation for 2 hours. (2) the expression of Bax protein in podocytes increased continuously with time after PAN treatment. (3) the apoptotic rate of PAN group was significantly higher than that of control group (P0.05). The apoptosis rate of CSA group was significantly lower than that of PAN group. The difference was statistically significant (P0.05). (4) compared with the control group, the expression of p-p38 Bax-Bax protein was significantly increased in the pan group, while the expression of p-p38 Bax protein in the cyclosporine A group and the SB203580 group was significantly lower than that in the PAN group (P0.05). Conclusion: CsA can reduce the apoptosis of podocyte induced by PAN and protect the kidney. The mechanism of CsA may be partly inhibiting the activation of p38MAPK and decreasing the expression of Bax.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R692

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