CAJ | 학술논문

目的：观察大鼠失血性休克(HS)+内毒素二次打击后膈肌组织内蛋白降解途径之一的泛素-蛋白酶体系统组成成分的变化。方法24只SD大鼠随机分为两组：假手术对照组(C组)和二次打击组(HS组)，每组12只。建立“失血性休克-内毒素”二次打击大鼠模型。蛋白免疫印记(Western blot)方法测定大鼠膈肌内E2-14k、MuRF1的蛋白表达；逆转录-聚合酶链式反应(RT-PCR)技术检测膈肌内泛素和蛋白酶体C2亚基的mRNA表达；采用苏木素-伊红(HE)染色在光镜下观察肺组织病理学改变；另外取膈肌标本固定后做电镜检测。结果蛋白免疫印记结果显示，HS组大鼠膈肌组织中E2-14k及MuRF1的蛋白表达分别为(1.65±0.38、1.88±0.32),与C组（分别为1.17±0.25,1.21±0.24）比较明显增加，差异有统计学意义，P<0.01。RT-PCR分析结果显示，HS组大鼠膈肌组织中泛素及蛋白酶体C2亚基的mRNA表达分别为(0.81±0.28、0.50±0.25，与C组（分别为0.89±0.20比0.50±0.15）比较明显增加，差异有统计学意义，P<0.05。HS组电镜下显示膈肌间线粒体肿胀、嵴减少，外膜模糊、变形，部分溶解破坏等超微结构的改变。结论二次打击大鼠膈肌组织内泛素-蛋白酶体降解途径被激活，可能是引起蛋白降解的重要因素之一。
목적：관찰대서실혈성휴극(HS)+내독소이차타격후격기조직내단백강해도경지일적범소-단백매체계통조성성분적변화。방법24지SD대서수궤분위량조：가수술대조조(C조)화이차타격조(HS조)，매조12지。건립“실혈성휴극-내독소”이차타격대서모형。단백면역인기(Western blot)방법측정대서격기내E2-14k、MuRF1적단백표체；역전록-취합매련식반응(RT-PCR)기술검측격기내범소화단백매체C2아기적mRNA표체；채용소목소-이홍(HE)염색재광경하관찰폐조직병이학개변；령외취격기표본고정후주전경검측。결과단백면역인기결과현시，HS조대서격기조직중E2-14k급MuRF1적단백표체분별위(1.65±0.38、1.88±0.32),여C조（분별위1.17±0.25,1.21±0.24）비교명현증가，차이유통계학의의，P<0.01。RT-PCR분석결과현시，HS조대서격기조직중범소급단백매체C2아기적mRNA표체분별위(0.81±0.28、0.50±0.25，여C조（분별위0.89±0.20비0.50±0.15）비교명현증가，차이유통계학의의，P<0.05。HS조전경하현시격기간선립체종창、척감소，외막모호、변형，부분용해파배등초미결구적개변。결론이차타격대서격기조직내범소-단백매체강해도경피격활，가능시인기단백강해적중요인소지일。
Objective To study the role of ubiquitin-proteasome pathway in a double-hit model (hemorrhagic shock followed by mimicked infection) rat diaphragmatic muscle. Methods 24 SD rats were randomly divided into two groups, control group with no surgery performed, and double-hit model group. A double-hit model of hemorrhagic shock with endotoxemia was utilized. The protein levels of E2-14k and MuRF1 in diaphragms were measured by Western blot. The mRNA levels of ubiquitin and proteasome subunit C2 in diaphragms were measured by means of real time polymerase chain reaction (RT-PCR). The lung Pathological changes of rats were observed, and the diaphragmatic ultrastructure was also measured by electron microscopy. Results The protein expression levels of E2-14k, MuRF1 of diaphragmatic tissues in HS group was 1.65 ± 0.38, 1.88±0.32, respectively, which were significantly higher than those 1.17±0.25, 1.21±0.24 in the group ,P<0.01. The mRNA expression of ubiquitin and C2 subunit of proteasome mRNA level of diaphragmatic tissues in HS group was 1.75 ± 0.36, 1.61 ± 0.41, respectively, which were significantly higher than those 1.22 ± 0.23,1.01 ± 0.25 in the control group ,P<0.01. Ultra structural examination showed mitochondria derangement in HS group, including swollen mitochondria with abnormal cristae, and disrupted external membrane of mitochondria. Conclusions The mRNA and protein expressions of ubiquitin-proteasome pathway in diaphragm increased significantly in HS rats, suggesting that the activity of ubiquitin-proteasome pathway increased which led to an increase of protein degradation.