吸入高浓度氢气对脓毒症小鼠心肌损伤及线粒体生物合成的影响

Effects of inhaling high concentration hydrogen on myocardial injury and mitochondrial biogenesis in septic mice

目的评价吸入高浓度氢气对脓毒症小鼠心肌损伤及线粒体生物合成的影响。方法清洁级健康雄性C57BL/6J小鼠128只,6~8周龄,体质量20~25 g,采用随机数字表法分为4组(n=32):假手术组(Sham组)、假手术+氢气组(Sham+H组)、脓毒症组(Sep组)和脓毒症+氢气组(Sep+H组)。采用盲肠结扎穿孔术建立小鼠脓毒症模型。Sham+H组和Sep+H组分别于术后1和6 h时吸入67%氢气1 h。每组随机取20只小鼠,观察术后7 d生存情况。剩余小鼠于术后24 h时取血液标本,采用ELISA法测定血清TNF-α、IL-1β、cTnI和CK-MB浓度;取心肌组织,HE染色后进行心肌病理评分,采用荧光分光光度法检测线粒体膜电位(MMP),萤光素酶法检测ATP含量,Western blot法测定过氧化物酶体增殖活化受体γ共激活因子-1α(PGC-1α)、核呼吸因子2(NRF2)、线粒体转录因子A(TFAM)的表达。结果与Sham组比较,Sep组生存率降低,血清TNF-α、IL-1β、cTnI和CK-MB浓度和心肌病理评分升高,MMP和ATP含量降低,心肌组织PGC-1α、NRF2和TFAM表达下调(P<0.05),Sham+H组上述指标差异无统计学意义(P>0.05);与Sep组比较,Sep+H组生存率升高,血清TNF-α、IL-1β、cTnI和CK-MB浓度和心肌病理评分降低,MMP和ATP含量升高,心肌组织PGC-1α、NRF2和TFAM表达上调(P<0.05)。结论吸入高浓度氢气可减轻脓毒症小鼠心肌损伤,机制可能与促进线粒体生物合成,改善线粒体功能有关。

Objective To evaluate the effects of inhaling high concentration hydrogen on myocardial injury and mitochondrial biogenesis in septic mice.Methods One hundred and twenty-eight clean-grade healthy male C57BL/6J mice,aged 6-8 weeks,weighing 20-25 g,were divided into 4 groups(n=32 each)using a random number table method:sham operation group(group Sham),sham operation+hydrogen group(group Sham+H),sepsis group(group Sep),and sepsis+hydrogen group(group Sep+H).The sepsis model was developed by cecal ligation and puncture in anesthetized animals.In Sham+H and Sep+H groups,67%H2 was inhaled for 1 h starting from 1 and 6 h after operation,respectively.Twenty mice in each group were randomly selected to observe the survival conditions at 7 days after operation.Blood samples were taken from the remaining mice at 24 h after operation for determination of tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),cardiac troponin I(cTnI)and creatine kinase isoenzyme(CK-MB)(by enzyme-linked immunosorbent assay),for examination of the pathological changes of myocardial tissues(by HE staining),and for determination of the mitochondrial membrane potential(MMP)(by fluorescence spectrophotometry),ATP content(by luciferase assay),and expression of myocardial peroxisome proliferator-activated receptor gamma coactivator-1α(PGC-1α),nuclear respiratory factor 2(NRF2)and mitochondrial transcription factor A(TFAM)(by Western blot).Results Compared with Sham group,the survival rate was significantly decreased,the serum concentrations of TNF-α,IL-1β,cTnI and CK-MB and pathological score were increased,the MMP and content of ATP in myocardial mitochondria were decreased,and the expression of PGC-1α,NRF2 and TFAM in myocardial tissues was down-regulated in Sep group(P<0.05),and no significant change was found in the parameters mentioned above in Sham+H group(P>0.05).Compared with group Sep,the survival rate was significantly increased,the serum concentrations of TNF-α,IL-1β,cTnI and CK-MB and pathological score were decreased,the MMP and content of ATP in myocardial mitochondria were increased,and the expression of PGC-1α,NRF2 and TFAM in myocardial tissues was up-regulated in group Sep+H(P<0.05).Conclusions Inhaling high concentration hydrogen can attenuate sepsis-induced myocardial injury in mice,and the mechanism may be related to promotion of mitochondrial biosynthesis and improvement in mitochondrial function.