中国组织工程研究

中国组织工程研究 ›› 2021, Vol. 25 ›› Issue (17): 2676-2680.doi: 10.3969/j.issn.2095-4344.3157

• 组织构建实验造模 experimental modeling in tissue construction • 上一篇    下一篇

脉络宁注射液干预挤压伤综合征模型猪线粒体自噬及PINK1/Parkin通路的变化

陈天贵，高  磊，李天博，王江宁   

  1. 首都医科大学附属北京世纪坛医院骨科，北京市   100038
  • 收稿日期:2020-06-24 修回日期:2020-07-01 接受日期:2020-08-19 出版日期:2021-06-18 发布日期:2021-01-08
  • 通讯作者: 王江宁，博士，主任医师，首都医科大学附属北京世纪坛医院骨科，北京市 100038
  • 作者简介:陈天贵，男，1981年生，云南省永善县人，汉族，硕士，主治医师，主要从事挤压伤综合征基础与临床研究。
  • 基金资助:
    国家自然科学基金面上项目(81473502)，项目负责人：王江宁

Effects of Mailuoning Injection on mitochondrial autophagy and PINK1/Parkin pathway in a pig model of crush injury syndrome

Chen Tiangui, Gao Lei, Li Tianbo, Wang Jiangning   

  1. Department of Orthopaedics, Beijing Shijitan Hospital affiliated to Capital Medical University, Beijing 100038, China
  • Received:2020-06-24 Revised:2020-07-01 Accepted:2020-08-19 Online:2021-06-18 Published:2021-01-08
  • Contact: Wang Jiangning, MD, Chief physician, Department of Orthopaedics, Beijing Shijitan Hospital affiliated to Capital Medical University, Beijing 100038, China
  • About author:Chen Tiangui, Master, Attending physician, Department of Orthopaedics, Beijing Shijitan Hospital affiliated to Capital Medical University, Beijing 100038, China
  • Supported by:
    the National Natural Science Foundation of China (General Project), No. 81473502 (to WJN)

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摘要：

文题释义：
挤压伤综合征：其病理特征为局部缺血性骨骼肌再灌注后引起的损伤，不仅影响局部组织活性及功能，还引起全身炎症反应造成休克、肾衰竭等症状，严重危害患者的生命健康。
线粒体自噬：指通过选择性清除损伤或不需要的线粒体来维持细胞稳态，在能量代谢和组织稳态中起重要作用。

背景：挤压伤综合征常引起实质性脏器损害，且其病情发展较快、死亡率高，探究其病理机制，缓解缺血再灌注损伤，可改善挤压伤病理症状。
目的：模拟体内生理环境体外灌注脉络宁注射液对挤压伤综合征模型猪线粒体自噬及PINK1/Parkin通路的影响。
方法：将巴马小型猪随机分为4组：对照组、模型组、灌注组、脉络宁组，除对照组外，其余3组制备挤压伤模型，灌注组解除挤压后先进行体外循环灌注再恢复自体血供，灌注过程中以50 mL/h的速度滴入生理盐水50 mL，脉络宁组在灌注过程中以50 mL/h的速度滴入脉络宁注射液50 mL，自体供血恢复6 h后，用全自动生化分析仪检测血清肌酐、尿素氮、肌酸激酶、丙氨酸氨基转移酶、天冬氨酸氨基转移酶、钾离子生化指标；ELISA检测血清白细胞介素1β、白细胞介素6、肿瘤坏死因子α水平；苏木精-伊红染色观察胫前肌组织病理变化；透射电镜观察胫前肌组织线粒体变化及自噬小体数量；Western blot检测胫前肌组织LC3-Ⅰ和LC3-Ⅱ、PINK1、Parkin蛋白表达。
结果与结论：①与对照组相比，模型组、灌注组小猪胫前肌组织肌丝紊乱，炎症细胞浸润，血清各项生化指标和炎症指标均显著升高(P均 < 0.001)，胫前肌组织自噬小体数目、LC3-Ⅱ/LC3-Ⅰ、PINK1、Parkin蛋白表达显著降低(P均 < 0.05)；②与模型组、灌注组相比，脉络宁组胫前肌组织上述病理症状明显减轻，血清各项生化指标和炎症指标均显著降低(P均 < 0.001)，胫前肌组织自噬小体数目、LC3-Ⅱ/LC3-Ⅰ、PINK1、Parkin蛋白表达显著增加(P均 < 0.05)；③结果表明，脉络宁注射液可能通过PINK1/Parkin通路激活线粒体自噬，改善挤压伤病理症状。

关键词: 脉络宁注射液, 挤压伤综合征, 体外循环灌注, 胫前肌, 线粒体, 自噬, 通路

Abstract: BACKGROUND: Crush injury syndrome often causes substantial organ damage, and the disease progresses rapidly with a high mortality. To explore its pathological mechanism can relieve ischemia-reperfusion injury, and thereby improve the pathological symptoms of crush injury.
OBJECTIVE: To study the effects of Mailuoning Injection on mitochondrial autophagy and PINK1/Parkin pathway in the crush injury syndrome model pigs. 
METHODS: Bama miniature pigs were randomly divided into four groups: control group, model group, perfusion group, and Mailuoning group. A crush injury model was prepared in all groups except for the control group. The perfusion group was subjected to cardiopulmonary bypass with 50 mL of saline at a speed of 50 mL/h first and then restoring autologous blood supply. The Mailuoning group was subjected to cardiopulmonary bypass with 50 mL of Mailuoning injection at a speed of 50 mL/h. The model group was not treated. Six hours after the recovery of autogenous blood supply, the biochemical indexes, including creatinine, urea nitrogen, creatine kinase, alanine aminotransferase, aspartate aminotransferase, and potassium ion, were measured by automatic biochemical analyzer. The levels of serum interleukin-1β, interleukin-6 and tumor necrosis factor α were detected by ELISA. Hematoxylin-eosin staining was used to observe the pathological changes of tibialis anterior muscle tissues; the changes of mitochondria and autophagosomes were observed by transmission electron microscopy. Western blot was used to detect the expression of microtubule associated protein 1 light chain 3 (LC3)-I and II, PINK1 and Parkin proteins. 
RESULTS AND CONCLUSION: Compared with the control group, in the model group and perfusion group, the myofilaments of tibialis anterior muscle tissues were disordered and inflammatory cells infiltrated, the levels of creatinine, urea nitrogen, creatine kinase, alanine aminotransferase, aspartate aminotransferase, potassium ion, interleukin-1β, interleukin-6 and tumor necrosis factor α in serum were significantly higher (P < 0.001), the number of autophagosomes and the expressions of LC3-II/LC3-I, PINK1 and Parkin proteins in tibialis anterior muscle tissues were significantly lower (P < 0.05). Compared with the model group and the perfusion group, the above pathological symptoms of tibialis anterior muscle tissues were significantly reduced in the Mailuoning group, the levels of serum biochemical indexes and inflammatory indexes were significantly lower (P < 0.001), and the number of autophagosomes and the expressions of LC3-II/LC3-I, PINK1 and Parkin proteins in tibialis anterior muscle tissues were significantly higher (P < 0.05). To conclude, Mailuoning Injection may activate mitochondrial autophagy through PINK1/Parkin pathway and improve the pathological symptoms of crush injury.

Key words: Mailuoning Injection, crush injury syndrome, cardiopulmonary bypass perfusion, tibialis anterior muscle, mitochondria, authophagy, pathway

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