| 宋雨萱,王佳丽,郑智礼,魏伟莉,张卫平,封 婷,钟相根.模拟外湿环境探讨外湿对正常及类风湿关节炎大鼠线粒体自噬的影响[J].现代生物医学进展英文版,2023,(3):417-422. |
| 模拟外湿环境探讨外湿对正常及类风湿关节炎大鼠线粒体自噬的影响 |
| The Effect of External Humidity on Mitochondrial Autophagy in Normal and Rheumatoid Arthritis Rats - Discussed by Simulating the External Wet Environment |
| Received:June 28, 2022 Revised:July 24, 2022 |
| DOI：10.13241/j.cnki.pmb.2023.03.004 |
| 中文关键词: 外湿干预 类风湿关节炎 痹证 线粒体自噬 |
| 英文关键词: Dampness intervention Rheumatoid arthritis Arthromyodynia Mitochondrial autophagy |
| 基金项目:北京中医药新奥基金奖励课题（2019-XAJLJJ-004）；北京中医药大学校级科研纵向发展基金课题（2019-ZXFZJJ-024） |
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| 中文摘要: |
| 摘要 目的：观察外湿干预下，正常及胶原诱导性类风湿关节炎（CIA）大鼠骨骼肌线粒体自噬水平的变化，研究外湿对线粒体自噬的影响。方法：将24只SPF级雄性SD大鼠随机分为4组：正常大鼠组（Con组）、湿邪干预正常大鼠组（Damp组）、CIA大鼠组（CIA组）、湿邪干预CIA大鼠组（CIA+Damp组）。采用二次免疫法建立牛Ⅱ型胶原乳化剂诱导性关节炎模型，自造模次日起，将Damp组和CIA+Damp组大鼠置于人工气候箱内进行湿邪干预。60 d后取骨骼肌组织，透射电镜观察线粒体形态，ELISA法检测骨骼肌三磷酸腺苷（ATP）和反应性氧簇（ROS）含量，RT-PCR和Western blot检测骨骼肌组织中腺苷酸活化蛋白激酶（AMPK）、微管相关蛋白轻链3B（LC3B）、线粒体外膜转位酶20（TOMM20）的mRNA表达量，过氧化物酶体增殖激活受体γ共激活因子1α（PGC-1α）的表达，并计算LC3-II/LC3-I比值。结果：随着湿邪干预天数增加，与Con组相比，各组大鼠关节指数均上升。Damp组关节骨质破坏较轻；线粒体轻微变形；骨骼肌ATP含量减少，ROS含量上升；在mRNA水平上，AMPK、LC3B表达升高，TOMM20表达降低；在蛋白水平上，PGC-1α、TOMM20表达下调，LC3-II/LC3-I比值上升。与CIA组相比，CIA+Damp组关节指数上升，关节骨质破坏加重；线粒体数量增多、形态损伤严重；骨骼肌ATP含量减少，ROS含量上升；在mRNA水平上，AMPK、LC3B的mRNA表达降低，TOMM20的mRNA表达升高；在蛋白水平上，PGC-1α表达下调，TOMM20表达升高，LC3-II/LC3-I比值上升。结论：外湿会引起细胞中的线粒体损伤，导致正常大鼠线粒体自噬代偿性上升，CIA大鼠线粒体自噬障碍、损伤线粒体堆积。 |
| 英文摘要: |
| ABSTRACT Objective: To observe the changes of mitophagy level in skeletal muscle of normal and collagen-induced rheumatoid arthritis (CIA) rats under external wetness intervention, and to study the effects of external wetness on mitochondrial autophagy. Methods: Twenty-four SPF male SD rats were divided into 4 groups randomly: normal rat group (Con group), damp pathogen intervention normal rat group (Damp group), CIA rat group (CIA group), damp pathogen intervention CIA rat group (CIA+Damp group). The bovine type Ⅱ collagen emulsifier-induced arthritis model was established by the method of secondary immunization. From the day after the model was established, the rats in Damp group and CIA+Damp group were placed in an artificial climate incubator, carry out dampness intervention. After 60 days, the skeletal muscle tissue was collected, and the morphology of mitochondria was observed by transmission electron microscope. The contents of adenosine triphosphate (ATP) and reactive oxygen species (ROS) in skeletal muscle were detected by ELISA. RT-PCR and western blot detected the expression of AMP-activated protein kinase (AMPK), microtubule-associated protein 1 light chain 3 (LC3B), translocase of outer mitochondrial membrane 20 (TOMM20) and PGC-1α in skeletal muscle tissue, and the LC3-II/LC3-I ratio was calculated. Results: With the increase of dampness intervention, the joint index of rats in each group was increased compared with the Con group. In the Damp group, the joint bone injuries and the mitochondrial deformation were slight. The ATP levels were decreased, while the ROS levels were rose in the skeletal muscle. In terms of mRNA level, the expression of AMPK and LC3B were increased, while the TOMM20 was decreased. In terms of protein expression levels, TOMM20 and PGC-1α were decreased, LC3-II/LC3-I ratio was increased. Compared with the CIA group, the joint index of rats in CIA+Damp group increased, the joint bone destruction was aggravated and the morphological deformation were serious. The ATP levels were decreased, while the ROS levels were rose in the skeletal muscle. At the mRNA level, the expression of AMPK and LC3B were decreased , the TOMM20 was increased. At the protein level, the PGC-1α was declined, the TOMM20 was increased, and the ratio of LC3-II/LC3-I was increased. Conclusion: The intervention of dampness could cause mitochondrial damage, lead to increasement of compensatory mitochondrial autophagy in normal rats, as well as develop mitochondrial autophagy disorders and damage mitochondrial accumulation in CIA rats. |
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