可溶性糖基化终末产物受体对缺血再灌注泛素-26S蛋白酶体系统活性的影响

Effect of soluble form of receptor for advanced glycation end products on 26 S proteasome activity in the animal model of ischemia reperfusion

【目的】建立动物心脏缺血再灌注模型,观察缺血再灌注模型中泛素-26 S蛋白酶体系统活性及总泛素蛋白表达的变化,检测s RAGE对缺血再灌注泛素-26 S蛋白酶体系统活性及总泛素蛋白表达的影响。【方法】复制C57BL/6J小鼠心脏缺血再灌注模型,Evans blue/2,3,5-triphenyltetrazolium chloride(TTC)双染色检测心肌缺血范围;检测泛素-26 S蛋白酶体系统活性:糜蛋白酶样、半胱天冬酶样、胰蛋白酶样;通过Western blotting检测总泛素蛋白的表达。【结果】与假手术组比较,缺血再灌注组梗死区面积/危险区面积%明显增加(I/R:36.3%±2.3%vs.Sham:0%),泛素-26 S蛋白酶体系统活性明显降低(半胱天冬酶样:0.74±0.04 vs.1.00±0.07;胰蛋白酶样:0.61±0.05 vs.1.00±0.02;糜蛋白酶样:0.63±0.06 vs.1.00±0.07),总泛素蛋白表达水平明显升高(2.73±0.14 vs.1.00±0.05);与缺血再灌注组比较,s RAGE预处理能够明显降低梗死区面积/危险区面积%(I/R+s RAGE:18.0%±1.1%vs.I/R:36.3%±2.3%),明显升高缺血再灌注的泛素-26 S蛋白酶体系统活性(半胱天冬酶样:1.25±0.09vs.0.74±0.04;胰蛋白酶样:1.05±0.06 vs.0.61±0.05;糜蛋白酶样:1.12±0.06 vs.0.63±0.06),同时明显抑制缺血再灌注诱导的总泛素蛋白表达水平(2.17±0.09 vs.2.73±0.14)。【结论】s RAGE能够抑制缺血再灌注诱导的泛素-26 S蛋白酶体系统活性降低及总泛素蛋白表达的增加。

【Objective】To test the change of the 26 S proteasome activity and total ubiquitinated proteins expression in the animal model of ischemia-reperfusion（I/R）. To detect the effect of s RAGE on 26 S proteasome activity and total ubiquitinated proteins expression.【Methods】C57BL/6J mice exposured to left anterior descending coronary artery ligation were used as in vivo models. At the end of reperfusion, the myocardial infarct size was determined by the Evans blue/2, 3, 5-triphenyltetrazolium chloride（TTC） double staining, the26 S proteasome activity was detected with Chymotrypsin-like kit, trypsin-like kit, and caspase-like kit, and then total ubiquitinated proteins expression was evaluated by western blot assay.【Results】Compared to Sham group, the myocardial infarct area was markedly increased（I/R：36.3% ±2.3% of area at risk vs. sham：0% of area at risk）, the 26 S proteasome activity was significantly decreased（caspase-like,0.74±0.04 vs. 1.00±0.07. trypsin-like, 0.61±0.05 vs. 1.00±0.02.chymotrypsin-like, 0.63±0.06 vs. 1.00±0.07）, and the level of total ubiquitinated proteins was significantly increased（2.73±0.14 vs. 1.00±0.05）. Compared to I/R group, s RAGE pretrentment significantly lower the infarct area（I/R＋s RAGE：18.0%±1.1% of area at risk vs. I/R：36.3%±2.3% of area at risk）, elevated 26 S proteasome activity（caspase-like, 1.25 ± 0.09 vs. 0.74 ± 0.04. trypsinlike 1.05 ±0.06 vs. 0.61±0.05;chymotrypsin-like, 1.12±0.06 vs. 0.63±0.06）,and inhibited total ubiquitinated proteins expression（2.17 ± 0.09 vs.2.73±0.14）.【Conclusion】s RAGE inhibits 26 S proteasome functional insufficiency and the increased ubiquitinated proteins expression induced by ischemia-reperfusion in vivo.