保元解毒汤含药血清对肌细胞萎缩模型E3组分mRNA及蛋白表达的影响

Effect of Baoyuan Detoxofication Soup medicated serum on expression of E3 mRNA and protein of the muscle cell atrophy model

目的:研究保元解毒汤含药血清对小鼠成肌细胞(C2C12)萎缩模型中E3组分Atrogin-1、MuRF-1mRNA及蛋白表达的影响,探讨保元解毒汤干预肌肉蛋白降解的作用机制。方法:制备保元解毒汤含药血清,高效液相色谱法(HPLC)定性检测含药血清中人参皂苷Rb1、绿原酸、阿魏酸、乌头碱。建立TNF-α诱导的肌细胞萎缩模型,实验分为正常对照组,TNF-α干预组,含药血清组。TNF-α及含药血清干预6h后,实时荧光定量PCR法检测肌细胞中Atrogin-1、MuRF-1mRNA表达。TNF-α及含药血清干预72h后,Western blot法检测肌细胞中Atrogin-1、MuRF-1的蛋白表达。结果:HPLC结果显示含药血清中含有人参皂苷Rb1、绿原酸、阿魏酸、乌头碱。与正常对照组比较,水煎浓缩含生药量达2.76 g/ml时,TNF-α诱导组细胞中Atrogin-1、MuRF-1mRNA及蛋白表达水平明显升高(P<0.05),与TNF-α诱导组比较,含药血清组细胞中Atrogin-1、MuRF-1mRNA及蛋白表达水平明显降低(P<0.05)。结论:保元解毒汤含药血清能显著降低Atrogin-1、MuRF-1mRNA及蛋白表达水平,提示保元解毒汤可能通过UPP信号途径抑制肌细胞萎缩,改善恶病质肌肉萎缩状态,可为临症治疗癌性恶病质提供新的治疗思路和有效方药。

To investigate the effect of Bao Yuan detoxofication soup medicated serum on expression of Atrogin-1 ,MuRF-1 protein of the muscle cell atrophy model and the mechanism of muscle protein degradation. Methods： Drug-containing serum was prepared and analyzed by HPLC. Establishmentting the TNF-α induced muscle cell atrophy model Furthermore,C2C12 cells were divided into normal control group, TNF-α intervention group and medicated serum group. After TNF-α and medicated serum interventioned 6 hours, detecting the expression of Atrogin-1, MuRF-1 mRNA by real-time quantitative PCR. After TNF-α and medicated serum interventioned 72 hours, detecting the expres- sion of Atrogin-1, MuRF-1 protein by western-bloting. Results： There were gingsenoside Rbl, chlorogenic, acid, ferulic acid, ac.onitine. Compared to normal control group, when concentrated decoction containing crude drug amounted to 2.75 g/ml, the expression of Atrogin-1, MuRF-1 mRNA of TNF-ct intervention group were significantly increased（ P 〈0.05）. Compared to TNF-ct intervention group, the expression of Atrogin-1, MuRF-1 mRNA of medicated serum group were significantly reduced（ P 〈 0.05）. Compared to normal control group, the expres- sion of Atrogin-1, MuRF-1 protein of TNF-α intervention group were significantly increased （ P 〈 0.05 ）. Compared to TNF-α intervention group, the expression of Atrogin-1, MuRF-1 protein of medicated serum group were significantly reduced （P 〈 0.05 ）. Conclusion： Baoyuan detoxofication soup medicated serum could significantly reduced the expression of Atrogin-1, MuRF-1 protein. Promptting that Bao Yuan det- oxofication soup probably inhibited muscle cell atrophy through UPP signaling pathways, improving cachexia muscular atrophy status, provi- ding new treatment ideas and effective prescription for treatmenting cancer eaehexia of clinical disease.