摘要
目的:探究携带miR-122a的间充质干细胞来源外泌体对糖尿病性心肌病(DCM)的作用及其相关分子机制。方法:将miR-122a和miR-NC转染至分离的小鼠骨髓来源间充质干细胞(BMSC),提取BMSC外泌体(BMSC-Exos)并通过Western blot鉴定。将30只C57/B6小鼠平均分为:对照组、miR-122a组和miR-NC组。除对照组外,miR-122a组和miR-NC组通过高脂高糖饮食联合STZ诱导DCM模型,随后miR-122a组和miR-NC组分别尾静脉注射BMSC-Exos-miR-122a和BMSC-ExosmiR-NC,而对照组腹腔注射等量枸橼酸钠缓冲液。结束实验后检测各组小鼠心功能指标,通过HE检测心肌组织病理水平,ELISA检测心肌组织ROS、IL-1β、IL-6和TNF-α水平,western blot检测小鼠心肌组织LC3Ⅱ、LC3Ⅰ、Beclin1和γH2AX蛋白表达水平。结果:细胞转染后,与BMSC-miR-NC比较,BMSC-miR-122a中miR-122a表达升高(P<0.05)。提取的BMSC-Exos有典型的Exo形态,粒径大约为100nm,外泌体标志蛋白CD63和CD81高表达。与对照组比较,miR-NC组舒张末期室间隔厚度(IVSd)、舒张末期后壁厚度(PWd)、左室短轴缩短率(LVFS)、每搏输出量(SV)、心排出量(CO)降低(P<0.05),左室舒张末期内径(LVDd)、左室收缩末期内径(LVDs)、左室质量(LVM)升高(P<0.05);心肌结构紊乱,炎性细胞浸润增加;心肌组织ROS、IL-1β、IL-6和TNF-α水平升高(P<0.05),LC3Ⅱ/LC3Ⅰ、Beclin1蛋白表达水平降低(P<0.05),γH2AX蛋白表达水平升高(P<0.05)。与miR-NC组比较,miR-122a组IVSd、PWd、LVFS、SV、CO升高(P<0.05),LVDd、LVDs、LVM降低(P<0.05);心肌结构恢复,炎性细胞浸润减少;心肌组织ROS、IL-1β、IL-6和TNF-α水平降低(P<0.05),LC3Ⅱ/LC3Ⅰ、Bec-lin1蛋白表达水平升高(P<0.05),γH2AX蛋白表达水平降低(P<0.05)。结论:DCM会引发心肌细胞自噬水平大幅度降低。携带miR-122a的BMSC-Exos可通过恢复心肌细胞自噬水平,降低炎性水平和细胞损伤,改善DCM的疾病症状。
Objective:To explore the effects of mesenchymal stem cell derived exosomes carrying miR-122a on diabetic cardiomyopathy(DCM)and the related molecular mechanisms.Methods:miR-122a and miR-NC were transfected into isolated mouse bone marrow derived mesenchymal stem cells(BMSC),and BMSC exosomes(BMSC-Exos)were extracted and identified by western blot.Thirty C57/B6 mice were divided into control group,miR-122a group and miR-NC group.In addition to the control group,the miR-122a group and the miR-NC group were treated with high-fat and high-sugar diet combined with STZ to induced DCM model,and then injected with BMSC-Exos-miR-122a and BMSC-Exos-miR-NC through caudal vein,respectively.The control group was intraperitoneally injected with the same amount of sodium citrate buffer.At the end of the experiment,cardiac function indexes of mice in each group were detected.The pathological level of myocardial tissue was detected by HE,the levels of ROS,IL-1β,IL-6 and TNF-αwere detected by ELISA,and the protein expression levels of LC3Ⅱ,LC3Ⅰ,Beclin1 andγH2AX in myocardial tissue were detected by western blot.Results:After transfection,compared with BMSC-miR-NC,the expression of miR-122a in BMSC-miR-122a was increased(P<0.05).The extracted BMSC-Exos showed a typical Exo morphology with a particle size of about 100 nm and high expression of exosome marker proteins CD63 and CD81.Compared with the control group,end-diastolic ventricular septal thickness(IVSd),end-diastolic posterior wall thickness(PWd),left ventricular short-axis shortening rate(LVFS),stroke output(SV)and cardiac output(CO)were decreased(P<0.05),while left ventricular end-diastolic diameter(LVDd),left ventricular end-systolic diameter(LVDs)and left ventricular mass(LVM)were increased(P<0.05),the myocardial structure was disturbed and inflammatory cell infiltration increased,and the levels of ROS,IL-1β,IL-6 and TNF-αin myocardial tissue were increased(P<0.05),the expression levels of LC3Ⅱ/LC3Ⅰand Beclin1 protein were decreased(P<0.05),while the expression levels ofγH2AX protein were increased in miR-NC group(P<0.05).Compared with miR-NC group,IVSd,PWd,LVFS,SV and CO were increased(P<0.05),while LVDd,LVDs and LVM were decreased(P<0.05),the myocardial structure recovered and inflammatory cell infiltration decreased,and the levels of ROS,IL-1β,IL-6 and TNF-αwere decreased(P<0.05),LC3Ⅱ/LC3Ⅰand Beclin1 protein expression levels were increased(P<0.05),whileγH2AX protein expression levels were decreased in miR-122a group(P<0.05).Conclusion:DCM can sig-nificantly reduce the autophagy level of cardiomyocytes.BMSC-Exos carrying miR-122a can improve disease symptoms in DCM by restoring autophagy levels of cardiomyocytes,reduce inflammatory levels and cell dam-age.
作者
亚森江·买买提
郭自同
买迪娜依·斯热吉丁
余小林
刘小方
程慧
YASENJIANG·Maimaiti;GUO Zitong;MAIDINAYI·Sirejiding(The People's Hospital of Xinjiang Uygur Autonomous Region,Xinjiang Urumqi 830001,China)
出处
《河北医学》
CAS
2023年第10期1593-1600,共8页
Hebei Medicine
基金
新疆维吾尔自治区自然科学基金项目,(编号:2022D01C133)。
