硫化氢对实验性病毒性心肌炎小鼠的保护作用及其机制

Protective effect of hydrogen sulfide on mice with experimental viral myocarditis and its mechanism

目的探讨硫化氢(H2S)对病毒性心肌炎(VMC)小鼠的保护作用及其机制。方法将70只BALB/c小鼠随机分为正常对照组(n=10)、心肌炎组(n=20)、炔丙基甘氨酸(PAG)组(n=20)、硫氢化钠(NaHS)组(n=20),后3组小鼠腹腔接种0.1 mL内含柯萨奇病毒B3(CVB3)的Eagle液建立VMC模型,对照组仅接种Eagle液。接种后当日,PAG、NaHS组分别腹腔注射40 mg/kg PAG、50μmol/kg NaHS,其余2组腹腔注射PBS,1次/d,连续2周。第15天称体质量(BM)后处死全部小鼠,比较各组死亡率,分离血清,通过ELISA测定血清心肌肌钙蛋白I(cTnI)水平,取心脏,称心脏质量(HM),计算HM/BM,HE染色计算心肌病理积分,测定心肌匀浆液中丙二醛(MDA)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)、过氧化氢酶(CAT)活性,ELISA检测心肌组织中白介素1β(IL-1β)、IL-6、肿瘤坏死因子-α(TNF-α)含量,Western blot法检测心肌细胞核内核因子-κB(NF-κB)p65表达水平。结果心肌炎组HM/BM、血清cTnI水平、心肌MDA活性、胞核NF-κB p65表达水平及心肌IL-1β、IL-6、TNF-α含量较对照组增加(P<0.05或0.01),而心肌SOD、GSH-Px和CAT活性较对照组减少。与心肌炎组比较,PAG组HM/BM、血清cTnI水平、心肌病理积分、心肌MDA活性、胞核NF-κB p65表达水平及心肌IL-1β、IL-6、TNF-α含量增加,心肌SOD、GSH-Px和CAT活性则减少(P<0.05),但2组死亡率比较差异无显著性(P>0.05)。与心肌炎组相比,NaHS组死亡率、HM/BM、血清cTnI水平、心肌病理积分、心肌MDA活性、胞核NF-κB p65表达水平及心肌IL-1β、IL-6、TNF-α含量降低(P<0.05),而SOD、GSH-Px、CAT活性则升高(P<0.05)。结论 H2S对VMC小鼠产生良好的保护作用,其机制可能与增强抗氧化能力及抑制炎症反应有关。

Objective To explore the protective effect of hydrogen sulfide （H2S） on viral myocarditis （VMC） mice and its mechanism. Methods Seventy BALB/c mice were randomly divided into 4 groups： normal control group （n=10）, myocarditis group （ n = 20）, DL-propargylglycine （PAG） group （ n =20） and sodium hydrosulfide （NariS） group （ n = 20）. Mice in the latter three groups were inoculated with 0. 1 mL Eagle＇s solution containing Coxsackievirus 133 （CVB3） intraperitoneally; whereas those in normal control group were treated with 0.1 mL Eagle＇s solution. On the day of inoculation, mice in PAG and NariS groups received intraperitoneal administration with 40 mg/kg PAG and 50 μmol/kg NariS, respectively; however, those in the other two groups were treated with PBS instead. PAG, NariS or PBS were given one time for one day and persisted for 14 days. On day 15, all mice were killed after weighing body mass （BM）. The mortality was compared among groups. Serum was separated and serum cardiac troponin I （cTnl） levels were detected using ELISA. The heart was removed and weighed to calculate heart mass （HM）. Histological cross sections of heart were stained with hematoxylin-eosin, and myocardial histopathologic scores were counted under optical microscope. The activities of malondialdehyde （MDA）, superoxide dismutase （SOD）, glutathione-peroxidase （GSH-Px） and catalase （CAT） in the homogenate of myocardium were examined. Myocardial interleukin （IL）-1β, IL-6 and tumor necrosis factor-α （TNF-α） contents were detected by ELISA. The expression level of myocardial nuclear factor-KB （NF-KB） p65 in the nucleus was examined using Western blotting. Results Compared with normal control group, the HM/BM, serum cTnl levels, myocardial MDA activity, NF-κB p65 expression level in the nucleus and myocardial contents of IL-1β, IL-6 and TNF-α were elevated, but myocardial SOD, GSH-Px and CAT activities were reduced in myocarditis group （ P 〈 0.05 or P 〈 0.01 ）. In comparison with myocarditis group, HM/BM, serum cTnl levels, myocardial histopathologic scores, myocardial MDA activity, NF-κB p65 expression level in the nucleus and myocardial contents of IL-1β, IL-6 and TNF-α significantly increased while myocardial SOD, GSH-Px and CAT activities significantly decreased in PAG groups （ P 〈 0. 05 ）. There was no significant difference in the mortality between PAG group and myocarditis group （P 〉 0. 05）. The mortality, HM/BM, serum cTnl levels, myocardial histopathologic scores, myocardial MDA activity, NF-KB p65 expression level in the nucleus and myocardial contents of IL-1β, IL-6 and TNF-α were lower, while myocardial SOD, GSH-Px and CAT activities were higher in NariS group than in myocarditis group （P〈0. 05）. Conclusion H2S can produce effective protection against VMC in mice. The mechanism may be associated with enhancing antioxidative ability and inhibiting inflammatory response.