Abstract:

To evaluate the regulatory effect of hydrogen-rich saline （HRS） on mitochondrial autophagy in myocardial cells of septic mice and its therapeutic effect on cardiac dysfunction， the seventy two male C57BL/6J mice were selected as the research objects and randomly divided into sham operation group （Sham group）， sham operation+hydrogen-rich saline group （Sham+HRS group）， sepsis （CLP group） and CLP+hydrogen-rich saline group （CLP+HRS group）， with 18 mice in each group. CLP model in mice was established by cecal ligation and puncture. Sham+HRS group and CLP+HRS group were intraperitoneally injected with 10 mL·kg^-1 hydrogen rich solution at 1 and 6 hours after modeling， respectively. Six mice in each group were randomly selected， and the mouse carotid blood sample was collected 24 hours after modeling， and the levels of tumor necrosis factor-α （TNF-α）， interleukin-1β （IL-1β）， cardiac troponin I （cTnI） and creatine kinase MB （CK-MB） in blood were determined by ELISA. Myocardial tissue was taken at 24 hours after modeling， ATP was detected by fluorescein fluorescent enzyme luminescence method， and mitochondrial membrane potential （MMP） was detected by fluorescence spectrophotometry. The expression of microtubule-associated protein 1 light/protein 3 light （LC3Ⅱ/LC3Ⅰ） and protein 62 （P62） in myocardial tissue was measured by Western blot at 24 hours after modeling. Results showed that， compared with Sham group， the levels of serum TNF-α， IL-1β， cTnI and CK-MB in CLP group were increased， the levels of myocardial ATP and MMP were decreased， the expression of myocardial LC3 Ⅱ/LC3 Ⅰ was increased， and the expression of P62 was decreased， the difference was statistically significant （P<0.05）. Compared with CLP group， the levels of serum TNF-α， IL-1β， cTnI and CK-MB in CLP+HRS group decreased， the levels of ATP and MMP in myocardial tissue increased， the expression of LC3Ⅱ/ LC3Ⅰ increased， and the expression of P62 decreased （P<0.05）. The results showed that the therapeutic effect of hydrogen-rich saline on cardiac dysfunction in sepsis mice may be achieved by regulating mitochondrial autophagy of cardiomyocytes. The study amied to explore the therapeutic effect and mechanism of hydrogen-rich saline on cardiac dysfunction in septic mice， and provide a theoretical basis for the clinical transformation of hydrogen-rich saline.

Key words: hydrogen-rich saline, sepsis, cardiac dysfunction, mitochondrial autophagy

摘要：

为评价富氢液（hydrogen-rich saline，HRS）对脓毒症小鼠心肌细胞线粒体自噬的调节及其对心功能障碍的治疗作用，选取72只雄性C57BL/6J小鼠作为研究对象，采用随机数字表法分为假手术组（Sham组）、假手术+富氢液组（Sham+HRS组）、脓毒症组（CLP组）、CLP+富氢液组（CLP+HRS组），每组18只。采用盲肠结扎穿孔法建立小鼠CLP模型。Sham+HRS组和CLP+HRS组分别于造模后1、6 h时腹腔注射富氢液10 mL·kg^-1。每组随机取6只小鼠，于造模后24 h时收集小鼠颈动脉血样，采用ELISA法测定血液肿瘤坏死因子α（tumor necrosis factor-α，TNF-α）、白细胞介素（interleukin-1β，IL-1β）、肌钙蛋白I（cardiac troponin I，cTnI）和肌酸激酶同工酶（creatine kinase MB，CK-MB）水平；于造模后24 h时取心肌组织，采用荧光素-荧光酶发光法检测ATP，荧光分光光度法检测线粒体膜电位（mitochondrial membrane potential，MMP）。造模后24 h采用Western blot法测定心肌组织线粒体自噬相关蛋白微管关联蛋白轻链3Ⅱ/轻链3Ⅰ（microtubule-associated protein 1 light/protein 3 light，LC3Ⅱ/LC3Ⅰ）和蛋白62（protein 62，P62）的表达水平。结果显示，与Sham组比较，CLP组血清TNF-α、IL-1β、cTnI和CK-MB水平升高，心肌ATP、MMP水平下降，心肌LC3Ⅱ/LC3Ⅰ表达水平上调，P62表达水平下调，差异有统计学意义（P<0.05）；与CLP组比较，CLP+HRS组血清TNF-α、IL-1β、cTnI和CK-MB含量下降，心肌组织ATP、MMP水平升高，LC3Ⅱ/LC3Ⅰ表达水平进一步上调，P62表达进一步下调（P<0.05）。结果表明，富氢液对脓毒症小鼠心功能障碍的治疗作用可能是通过调节心肌细胞线粒体自噬实现的。研究旨在探讨富氢液对脓毒症小鼠心功能障碍的治疗作用及机制，以期为富氢液的临床转化提供理论依据。

关键词: 富氢液, 脓毒症, 心功能障碍, 线粒体自噬