益气活血方对大鼠放射性肺损伤组织MMP-2/TIMP-2表达的影响

Effect of Yiqi Huoxue Recipe to MMP-2 / TIMP-2 Expression in Radioactive Damaged Lung Tissue

目的:检测益气活血方干预前后大鼠放射性肺损伤组织基质金属蛋白酶2(matrix metalloproteinase-2,MMP2)和基质金属蛋白酶抑制剂2(tissue inhibitor of matrix metalloproteinase,TIMP2)表达的影响,了解该方剂的作用机制。方法:健康成年雌性SD大鼠120只,随机分为模型组30只、照射加益气活血方高剂量组30只、照射加益气活血方低剂量组30只、空白组30只。益气活血方高、低剂量组大鼠受照当天开始,用药量20,10 g·kg-1,每次ig益气活血方剂5 m L·kg-1,2次/d;模型组、空白组每次ig蒸馏水5 m L·kg-1,2次/d。4组大鼠分别于照射后第4,8,12,16,20周末随机抽取6只,处死并取其右肺组织保存。分别用反转录PCR技术(RT-PCR),蛋白质印迹法(Western-blotting)检测MMP-2/TIMP-2 mRNA及蛋白表达的动态变化。结果:模型组MMP-2 mRNA及蛋白表达于第12周达高峰,而TIMP-2 mRNA及蛋白表达在照射后第4周至20周期间出现逐渐升高的趋势,各时间点模型组与空白组相比均具有明显差异(P<0.01)。益气活血方高剂量组与模型组相比,各mRNA及蛋白表达趋势与模型组相似,MMP-2 mRNA及蛋白表达在12周前低于模型组,差异有统计学意义(P<0.05,P<0.01)。益气活血方低剂量组各项指标均有变化,趋势与模型组类似,差异没有统计学意义。结论:益气活血方高剂量在放射性肺损伤的各个时间段,从转录及翻译层面对MMP-2/TIMP-2的表达进行调整,使MMPs/TIMPs趋于平衡,从而减轻放射性肺损伤,这可能是该药防治放射性肺损伤的分子生物学机制。

Objective： To study the effect of the Yiqi Huoxue recipe on expression of the matrix metalloproteinase-2（MMP-2）/tissue inhibitor of matrix metalloproteinase（TIMP-2） in radiation-induced lung injury model rats, in order to know the mechanism of this recipe. Method： One hundred and twenty female rats were divided into four groups at random：namely simple exposure group（model group,n=30）, radiational Yiqi Huoxue recipe high-dose group and the radiational Yiqi Huoxue recipe low-dose group（n=30）,and blank control group（control group,n=30）. Six rats were selected randomly from the four groups each time at the 4^th, 6^th, 8^th, 12^th, 26^th weekend after first radiation, those rats were executed immediately after intraperitoneal anesthesia, took the right lung tissue of them and saved in right way. Reverse transcription PCR（RT- PCR） was used to detect the dynamic change of mRNA expression of MMP-2, TIMP-2 in each time point.Western-blotting was used to detect the dynamic change of protein expression of MMP-2, TIMP-2 in each time point. Result： The expression of MMP-2 mRNA and protein appeared a first increased and then decline trend from 4^th weekend to 20^th weekend after irradiation, and the peak was in 12^th.The expression of TIMP-2 mRNA and protein appeared a rising tend from 4^th weekend to 20^th weekend after irradiation. The mRNA and protein expression trend is similar to Yiqi Huoxue recipe high-dose group and model group,but they have different change in different time point. In Yiqi Huoxue recipe low-dose group the indicators changed with no statistical significance. Conclusion： High-dose Yiqi Huoxue recipe can adjust the expression of MMP-2/TIMP-2 from the transcription and translation level in each period of radioactive lung injury, making so as to reduce the radioactive lung injury. This may be the molecular biology mechanism of the drug in prevention and control of radioactive lung injury.