吉林大学学报(医学版) ›› 2024, Vol. 50 ›› Issue (5): 1348-1357.doi: 10.13481/j.1671-587X.20240519

• 基础研究 • 上一篇    

骨髓间充质干细胞来源外泌体对异丙肾上腺素诱导的大鼠心肌纤维化的影响及其作用机制

魏俊萍1,符达佳2,孟庆雯1(),林道飞1,林燕仔1   

  1. 1.海南医学院第一附属医院心血管内一科,海南 海口 570102
    2.海南医学院第一附属医院 心电图室,海南 海口 570102
  • 收稿日期:2023-09-05 出版日期:2024-09-28 发布日期:2024-10-28
  • 通讯作者: 孟庆雯 E-mail:443130019@qq.com
  • 作者简介:魏俊萍(1987-),女,海南省海口市人,主治医师,主要从事心力衰竭、高血压和冠心病的进展及心脏康复方面的研究。
  • 基金资助:
    海南省卫健委卫生健康行业科研项目(22A200151);海南省科技厅自然科学基金青年项目(821QN401);海南医学院第一附属医院青年培育基金(HYYFYPY202020)

Effect of bone marrow mesenchymal stem cell-derived exosomes on myocardial fibrosis in rats induced by isoproterenol and its mechanism

Junping WEI1,Dajia FU2,Qingwen MENG1(),Daofei LIN1,Yanzai LIN1   

  1. 1.Department of Cardiovascular,First Affiliated Hospital,Hainan Medical College,Haikou 570102,China
    2.Echocardiography Room,First Affiliated Hospital,Hainan Medical College,Haikou 570102,China
  • Received:2023-09-05 Online:2024-09-28 Published:2024-10-28
  • Contact: Qingwen MENG E-mail:443130019@qq.com

摘要:

目的 探讨骨髓间充质干细胞(BMSCs)来源外泌体(Exo)对异丙肾上腺素(ISO)诱导的大鼠心肌纤维化的影响,并初步阐明其作用机制。 方法 从BMSCs中分离提取Exo,采用透射电子显微镜、纳米颗粒跟踪分析仪和Western blotting 法对Exo进行分析鉴定。将40只SD大鼠分为对照组、模型组、BMSCs-Exo组和BMSCs-Exo+铁死亡激活剂(Erastin)组,每组10只。除对照组外,其余3组大鼠均皮下注射ISO建立心肌纤维化模型,BMSCs-Exo组和BMSCs-Exo+Erastin组大鼠尾静脉注射BMSCs-Exo,且BMSCs-Exo+Erastin组大鼠再腹腔注射Erastin。4周后,超声心动图检测各组大鼠左心室射血分数(LVEF)、左心室短轴缩短率(LVFS)、左心室舒张末期内径(LVEDD)和左心室收缩末期内径(LVESD),HE染色观察各组大鼠心肌组织病理形态表现,Masson染色观察各组大鼠心肌组织纤维化程度,免疫组织化学染色检测各组大鼠心肌组织中α-平滑肌肌动蛋白(α-SMA)、Ⅰ型胶原(COL?Ⅰ)和Ⅲ型胶原(COL?Ⅲ)阳性表达率,比色法测定各组大鼠心肌组织中铁离子(Fe2+)水平,Western blotting法检测各组大鼠心肌组织中酰基辅酶A合成酶长链家族成员4(ACSL4)、铁蛋白重链1(FTH1)、谷胱甘肽过氧化物酶4(GPX4)和溶质载体家族7成员11(SLC7A11)蛋白表达水平。 结果 从BMSCs中分离的颗粒物具有典型的脂质双分子层,粒径大多分布在波长100 nm处,CD63、CD9、肿瘤易感基因101(TSG101)和热休克蛋白70(HSP70)蛋白均高表达,由此判定为Exo。与对照组比较,模型组大鼠LVEF和LVFS明显降低(P<0.05),LVEDD和LVESD明显升高(P<0.05),心肌细胞排列紊乱,部分细胞核皱缩和坏死,胶原容积分数(CVF)明显升高(P<0.05),α-SMA、COL-Ⅰ和COL-Ⅲ阳性表达率明显升高(P<0.05),心肌组织中Fe2+水平明显升高(P<0.05),ACSL4蛋白表达水平明显升高(P<0.05),FTH1、GPX4和SLC7A11蛋白表达水平明显降低(P<0.05)。与模型组比较,BMSCs-Exo组大鼠LVEF和LVFS明显升高(P<0.05),LVEDD和LVESD明显降低(P<0.05),心肌组织损伤明显减轻,CVF明显降低(P<0.05),α-SMA、COL-Ⅰ和COL-Ⅲ阳性表达率明显降低(P<0.05),心肌组织中Fe2+水平明显降低(P<0.05),ACSL4蛋白表达水平明显降低(P<0.05),FTH1、GPX4和SLC7A11蛋白表达水平明显升高(P<0.05)。与BMSCs-Exo组比较,BMSCs-Exo+Erastin组大鼠LVEF和LVFS明显降低(P<0.05)、LVEDD和LVESD明显升高(P<0.05),心肌细胞出现水肿和坏死,CVF明显升高(P<0.05),α-SMA、COL-Ⅰ及COL-Ⅲ阳性表达率明显升高(P<0.05),心肌组织中Fe2+水平明显升高(P<0.05),ACSL4蛋白表达水平明显升高(P<0.05),FTH1、GPX4和SLC7A11蛋白表达水平明显降低(P<0.05)。 结论 BMSCs来源Exo能够改善ISO诱导的大鼠心肌纤维化,该机制可能与抑制铁死亡有关。

关键词: 心肌纤维化, 异丙肾上腺素, 骨髓间充质干细胞, 外泌体, 铁死亡

Abstract:

Objective To discuss the effect of bone marrow mesenchymal stem cells (BMSCs)-derived exosomes (Exo) on isoproterenol (ISO)-induced myocardial fibrosis in the rats, and to clarify its mechanism. Methods The Exo was isolated from the BMSCs and characterized by transmission electron microscope, nanoparticle tracking analysis, and Western blotting methods. Forty SD rats were divided into control group, model group, BMSCs-Exo group, and BMSCs-Exo+ferroptosis activator (Erastin) group, and there were 10 rats in each group. The myocardial fibrosis models were established by subcutaneous injection of ISO in all the rats except control group. The rats in BMSCs-Exo and BMSCs-Exo+Erastin groups were given BMSCs-Exo, and the rats in BMSCs-Exo+Erastin group were additionally injected with Erastin intraperitoneally. After 4 weeks, echocardiography was used to detect the left ventricular ejection fraction (LVEF), left ventricular fractional shortening (LVFS), left ventricular end diastolic diameter (LVEDD), and left ventricular end systolic diameter (LVESD) of the rats in various groups; HE staining was used to observe the pathomorphology of myocardium tissue of the rats in various groups; Masson staining was used to observe the fibrosis degrees of myocardium tissue of the rats in various groups; immunohistochemistry was used to detect the positive expression rates of α-smooth muscle actin (α-SMA), type Ⅰ collagen (COL-Ⅰ), and type Ⅲ collagen (COL-Ⅲ) in myocardium tissue of the rats in various groups; colorimetry was used to detect the Fe2+ level in myocardium tissue of the rats in various groups; Western blotting method was used to detect the expression levels of acyl-CoA synthetase long chain family member 4 (ACSL4), ferritin heavy chain 1 (FTH1), glutathione peroxidase 4 (GPX4), and solute carrier family 7 member 11 (SLC7A11) proteins in myocardium tissue of the rats in various groups. Results The particles isolated from BMSCs had a typical lipid bilayer structure, and most particle sizes distributed around 100 nm. High expression levels of CD63, CD9, tumor susceptibility gene 101 (TSG101), and heat shock protein 70 (HSP70) proteins confirmed the particles as Exo. Compared with control group, the LVEF and LVFS of the rats in model group were significantly decreased (P<0.05), LVEDD and LVESD were increased (P<0.05), the myocardial cell arrangement was disordered, some nuclear shrinkage and necrosis were seen, the collagen volume fraction (CVF) was significantly increased (P<0.05), the positive expression rates of α-SMA, COL-Ⅰ, and COL-Ⅲ were significantly increased (P<0.05), the Fe2+ level in myocardium tissue was significantly increased (P<0.05), the expression level of ACSL4 protein was significantly increased (P<0.05), and the expression levels of FTH1, GPX4, and SLC7A11 proteins were significantly decreased (P<0.05). Compared with model group, the LVEF and LVFS of the rats in BMSCs-Exo group were significantly increased (P<0.05), the LVEDD and LVESD were significantly decreased (P<0.05), the myocardial tissue damage was significantly alleviated, the CVF was significantly decreased (P<0.05), the positive expression rates of α-SMA, COL-Ⅰ, and COL-Ⅲ were significantly decreased (P<0.05), the Fe2+ level in myocardium tissue was significantly decreased (P<0.05), the expression level of ACSL4 protein was significantly decreased (P<0.05), and the expression levels of FTH1, GPX4, and SLC7A11 proteins were significantly increased (P<0.05). Compared with BMSCs-Exo group, the LVEF and LVFS of the rats in BMSCs-Exo+Erastin group were significantly decreased (P<0.05), the LVEDD and LVESD were significantly increased (P<0.05), the myocardial cell edema and necrosis were seen, the CVF was significantly increased (P<0.05), the positive expression rates of α-SMA, COL-Ⅰ, and COL-Ⅲ were significantly increased (P<0.05), the Fe2+ level in myocardium tissue was significantly increased (P<0.05), the expression level of ACSL4 protein was significantly increased (P<0.05), and the expression levels of FTH1, GPX4, and SLC7A11 proteins were significantly decreased (P<0.05). Conclusion BMSC-derived Exo can improve the myocardial fibrosis in the rats induced by ISO, and its mechanism may be related to the inhibition of ferroptosis.

Key words: Myocardial fibrosis, Isoproterenol, Bone marrow mesenchymal stem cell, Exosome, Ferroptosis

中图分类号: 

  • R542.2