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赵宇含, 曹国栋, 郭立春, 等. 硫化氢通过抑制ROS介导的内质网应激改善脓毒症心肌损伤的研究[J]. 四川大学学报(医学版), 2022, 53(5): 798-804. DOI: 10.12182/20220960106
引用本文: 赵宇含, 曹国栋, 郭立春, 等. 硫化氢通过抑制ROS介导的内质网应激改善脓毒症心肌损伤的研究[J]. 四川大学学报(医学版), 2022, 53(5): 798-804. DOI: 10.12182/20220960106
ZHAO Yu-han, CAO Guo-dong, GUO Li-chun, et al. Hydrogen Sulfide Ameliorates Myocardial Injury Caused by Sepsis Through Suppressing ROS-Mediated Endoplasmic Reticulum Stress[J]. Journal of Sichuan University (Medical Sciences), 2022, 53(5): 798-804. DOI: 10.12182/20220960106
Citation: ZHAO Yu-han, CAO Guo-dong, GUO Li-chun, et al. Hydrogen Sulfide Ameliorates Myocardial Injury Caused by Sepsis Through Suppressing ROS-Mediated Endoplasmic Reticulum Stress[J]. Journal of Sichuan University (Medical Sciences), 2022, 53(5): 798-804. DOI: 10.12182/20220960106

硫化氢通过抑制ROS介导的内质网应激改善脓毒症心肌损伤的研究

Hydrogen Sulfide Ameliorates Myocardial Injury Caused by Sepsis Through Suppressing ROS-Mediated Endoplasmic Reticulum Stress

  • 摘要:
      目的  探讨硫化氢(H2S)对脓毒症心肌损伤中活性氧(ROS)介导的内质网应激的影响。
      方法  采用盲肠结扎穿刺术(CLP)诱导SD大鼠脓毒症模型,将大鼠随机分成假手术(sham)组、脓毒症(CLP)组、脓毒症+硫氢化钠(NaHS)(CLP+NaHS)组。超声心动图观察各组大鼠左心室功能,检测大鼠血浆H2S的水平;通过乳酸脱氢酶(LDH)、丙二醛(MDA)、谷胱甘肽(GSH)水平检测以及ROS染色评估大鼠心肌氧化应激水平;HE染色观察大鼠心肌组织形态变化,透射电子显微镜观察心肌细胞线粒体超微结构;Western blot检测细胞内源性硫化氢合成酶半胱硫氨酸-γ-裂解酶(CSE)、3-巯基丙酮酸硫转移酶(3-MST)表达变化,内质网应激标志性蛋白:磷酸化(p)-激酶蛋白激酶R样内质网激酶(PERK)、p-细胞翻译启始因子2α(eIF2α)、p-肌醇需要酶1α(IRE1α)、活化转录因子(ATF)4和C/EBP同源蛋白(CHOP)的表达变化;TUNEL染色观察大鼠心肌细胞凋亡变化。
      结果  脓毒症大鼠左室射血分数(LVEF)、左心室缩短分数(LVFS)降低(P<0.05),血浆H2S降低(P<0.05),血浆H2S与LVEF和LVFS呈线性相关(r2=0.62、r2=0.64,P均<0.05)。脓毒症组大鼠ROS水平明显增强,LDH水平增高(P<0.05),MDA表达增强(P<0.05),GSH表达下降(P<0.05);在HE染色中可见炎性细胞浸润,心肌细胞水肿,透射电子显微镜可观察到线粒体明显损伤,可见线粒体水肿,嵴结构溶解等情况;Western blot结果显示,CSE、3-MST表达水平有所下降(P<0.05),内质网应激标志性蛋白p-PERK、p-eIF2α、IRE1α、ATF4和CHOP蛋白表达水平升高(P<0.05);TUNEL染色结果显示心肌细胞凋亡水平明显增加(P<0.05)。NaHS处理后,LVEF和LVFS升高(P<0.05),血浆H2S升高(P<0.05)。心肌氧化应激水平降低,HE与透射电镜显示心肌形态学有改善,线粒体损伤减轻,CSE、3-MST水平升高(P<0.05),p-PERK、p-eIF2α、IRE1α、CHOP蛋白表达下降(P<0.05),心肌细胞凋亡水平有所下降(P<0.05)。
      结论  H2S通过抑制ROS介导的内质网应激,减少脓毒症心肌细胞凋亡,从而改善脓毒症心肌功能障碍。

     

    Abstract:
      Objective  To investigate the effect of hydrogen sulfide (H2S) on reactive oxygen species (ROS)-mediated endoplasmic reticulum stress in myocardial injury caused by sepsis.
      Methods  A sepsis model was induced in Sprague-Dawley (SD) rats by cecal ligation and puncture (CLP). The rats were randomly divided into sham operation (sham) group, sepsis (CLP) group, and sepsis+sodium hydrosulfide (NaHS) (CLP+NaHS) group. The left ventricular function of the rats was observed with echocardiography and their plasma H2S levels were measured. Lactate dehydrogenase (LDH), malondialdehyde (MDA), glutathione (GSH) levels were measured and HE staining was done to evaluate the level of myocardial oxidative stress in rats. HE staining was done to observe the morphological changes of rat myocardium, and transmission electron microscope was used to observe the ultrastructure of myocardial mitochondria. Western blot was done to examine changes in the expression of two endogenous hydrogen sulfide synthases, cystathionine γ-lyase (CSE) and 3-mercaptopyruvate sulfur transferase (3-MST), and changes in the expression of endoplasmic reticulum stress (ERS) marker proteins, including phosphorylated (p) protein kinase R-like endoplasmic reticulum kinase (p-PERK), p-eukaryotic translation initiation factor 2α (p-eIF2α), p-inositol requires enzyme 1α (IRE1α), recombinant activating transcription factor 4 (ATF4), and C/EBP homologous protein (CHOP). TUNEL staining was performed to observe the changes of cardiomyocyte apoptosis in rats.
      Results  Left ventricular ejection fraction (LVEF), left ventricular shortening fraction (LVFS) and plasma H2S decreased in septic rats (P<0.05). Plasma H2S exhibited linear correlation with LVEF and LVFS (r2=0.62 and r2=0.64, all P<0.05). The ROS levels were significantly elevated in rats of the CLP group. In addition, these rats showed increased level of LDH (P<0.05), increased expression of MDA (P<0.05), and decreased expression of GSH (P<0.05). Inflammatory cell infiltration and cardiomyocyte edema were observed in HE staining. Transmission electron microscopic observation revealed significant mitochondrial damage, observable mitochondrial edema, and cristae structure dissolution. The Western blot results showed that the expression levels of CSE and 3-MST decreased (P<0.05), while the ERS marker proteins, including p-PERK, p-eIF2, IRE1α, ATF4, and CHOP, were expressed at increased levels (P<0.05). TUNEL staining showed significant increase of apoptosis in cardiomyocytes (P<0.05). After NaHS treatment, LVEF and LVFS increased (P<0.05) and plasma H2S increased in septic rats (P<0.05). Myocardial oxidative stress levels decreased. HE staining and transmission electron microscopy showed improved myocardial morphology. Mitochondrial damage was reduced and CSE and 3-MST levels were significantly increased (P<0.05). The expression of p-PERK, p-eIF2α, p-IRE1α, and CHOP proteins decreased (P<0.05). A decrease in cardiomyocyte apoptosis levels was observed by TUNEL staining (P<0.05).
      Conclusion  H2S reduces septic cardiomyocyte apoptosis by inhibiting ROS-mediated ERS, thereby improving myocardial dysfunction in sepsis.

     

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