收稿日期: 2023-07-25
网络出版日期: 2024-03-26
基金资助
贵州医科大学国家自然科学基金培育项目(gyfynsfc-2021-54);贵州省卫生健康委科学技术基金项目(gzwjkj2020-1-026);贵州省黔西南州科技局科技计划项目(2022-2-08)
Inhibition of TRAF6 ameliorates myocardial inflammatory injury and cardiac dysfunction via regulating cardiomyocyte inflammation in sepsis mice
Received date: 2023-07-25
Online published: 2024-03-26
目的 探索抑制TRAF6后对脓毒症小鼠心肌细胞自噬、炎症反应和心功能的改善作用。 方法 （1）昆明雄性小鼠24只，随机分为4组，分别为假手术（sham）组、假手术+C25-140（sham+C）组、盲肠结扎穿刺术（CLP）组、盲肠结扎穿刺术+C25-140（CLP+C）组。sham+C组、CLP+C组术后腹腔注射C25-140。术后24 h超声评估LVEF、LVFS；ELISA测血清TNF-α、IL1-β；HE染色评估心肌炎症反应；透射电镜观察心肌细胞自噬小体、线粒体微结构；qPCR检测心肌组织TRAF6 mRNA；WB检测TRAF6、P62、Beclin-1、LC3B蛋白的表达；（2）予3-MA抑制自噬观察C25-140对脓毒症小鼠心肌损伤的影响。 结果 （1）与sham组比，CLP组心肌组织TRAF6mRNA及TRAF6蛋白显著升高（P < 0.05）；血清 TNF-α、IL1-β浓度显著升高（P < 0.05）；心肌组织HE染色见炎症细胞浸润；LVEF、LVFS显著降低（P < 0.05）；（2）与CLP组比，CLP+C组TRAF6 mRNA及TRAF6蛋白表达减少（P < 0.05）；血清TNF-α、IL1-β下降（P < 0.05）；心肌组织病理心肌炎症细胞浸润减少；LVEF、LVFS升高（P < 0.05）；电镜见线粒体肿胀减轻、自噬小体增多；Beclin-1、LC3Ⅱ/Ⅰ表达增加，P62表达减少（P < 0.05）；予3-MA抑制自噬后，与CLP+C组比，CLP+C+3-MA组小鼠心肌病理见炎症细胞浸润明显，LVEF、LVFS降低（P < 0.05）。 结论 抑制TRAF6改善脓毒症小鼠心肌炎症损伤和心功能障碍；抑制TRAF6促进心肌细胞自噬参与保护脓毒症诱导的心肌损伤。
关键词： 肿瘤坏死因子受体相关因子6; 自噬; 脓毒症; 心肌炎症; 心功能
周颖 , 蒋大军 , 田勇 , 古雍翔 , 杨国辉 . 抑制TRAF6调节炎症和自噬改善脓毒症小鼠的心肌损伤和心功能[J]. 实用医学杂志, 2024 , 40(5) : 608 -614 . DOI: 10.3969/j.issn.1006-5725.2024.05.004
Objective To explore the effects of TRAF6 inhibition on autophagy， myocardial inflammation and cardiac function in septic mice. Methods Twenty-four male Kunming mice were randomly divided into 4 groups： sham， sham + C25-140 （sham+C）， cecal ligation and puncture （CLP）， and cecal ligation and puncture+C25-140 （CLP+C） group. Sham+C group and CLP+C group were intraperitoneally injected with C25-140 after operation. LVEF and LVFS were evaluated by ultrasound 24 hours after operation. Serum TNF-α and IL1-β were measured by ELISA. HE staining was used to evaluate myocardial inflammatory response. Autophagosomes and mitochondrial microstructure of cardiomyocytes were observed by transmission electron microscopy. TRAF6 mRNA in myocardial tissue was detected by qPCR. The expression of TRAF6， P62， Beclin-1 and LC3B protein was detected by W-B. The effect of C25-140 on myocardial injury in the septic mice was observed by inhibiting autophagy with 3-MA. Results Compared with the sham group， the levels of TRAF6 mRNA and TRAF6 in the myocardial tissue in the CLP group were significantly increased （P < 0.05） and the serum TNF-α and IL1-β concentrations were significantly increased （P < 0.05）. Meanwhile， the myocardial tissue HE staining showed inflammatory cell infiltration and the LVEF and LVFS levels were significantly decreased in the CLP group （P < 0.05）. Compared with CLP group， the CLP+C group showed that the expression of TRAF6 mRNA and TRAF6 protein decreased （P < 0.05）， serum TNF-α and IL1-β decreased （P < 0.05）， myocardial histopathological myocardial inflammatory cell infiltration decreased， the LVEF and LVFS levels increased （P < 0.05）. Electron microscopy showed that the mitochondrial swelling decreased， autophagosomes increased， expression of Beclin-1 and LC3II/I increased， and P62 expression decreased （P < 0.05）. As compared with CLP+C group， the CLP+C+3-MA group showed that obvious inflammatory cell infiltration in the myocardial pathology and the LVEF and LVFS levels decreased after 3-MA inhibited autophagy （P < 0.05）. Conclusion Inhibition of TRAF6 can not only ameliorate myocardial inflammatory injury and cardiac dysfunction in septic mice， but promote the involvment of cardiomyocyte autophagy in provention from sepsis-induced myocardial injury.
Key words： TRAF6; autophagy; sepsis; myocardial inflammation; cardiac dysfunction
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