骨髓源性成体干细胞体外转化为心肌样细胞的研究

发布时间：2018-05-26 22:22

本文选题：骨髓间充质细胞 + 细胞分化　；参考：《大连医科大学》2006年博士论文

【摘要】：目的：急性心肌梗死(AMI)是危害人类健康的主要疾病之一。虽然成熟心肌细胞不是终末分化细胞，但也只有有限的分裂、增殖能力。AMI患者即使有幸渡过急性期，但可由于心肌梗死(MI)后梗死部位瘢痕形成及残留心肌代偿性肥厚，引起心室重构，进而促进心力衰竭发生，最终造成患者死亡。现今用于AMI治疗的方法通常为溶栓治疗、经皮冠状动脉成形术(PTCA)、冠状动脉旁路移植术(CABG)及心脏移植术等。前3项方法可改善心肌供血，但不能使坏死心肌复生，而心脏移植虽可替代受损心脏，但难以广泛的推广应用。近年来，随着分子生物学的发展，利用成体干细胞种植方法来增加梗死部位心肌细胞的数量，从而改善心功能及降低AMI后远期病死率，为AMI的治疗开辟了新的前景。目前，用于心脏疾病方面的成体干细胞来源主要有骨髓干细胞、骨骼肌卫星细胞、平滑肌干细胞、胚胎干细胞以及胎儿心肌细胞。骨髓干细胞在人体广泛分布而且容易获取，种植中无免疫排斥反应，也不存在伦理学的争论，因此它们在细胞性心肌整复的临床应用中很有优势。本研究探讨骨髓源性的成体干细胞体外在5-氮胞苷、中药三七皂甙的诱导下转化为心肌样细胞的可行性，并将骨髓干细胞与心肌细胞共培养，探讨“心肌样”微环境对这一转化过程的影响。方法：一．骨髓间充质干细胞(marrow mesenchymal stem cells，MSCs)体外诱导分化为心肌样细胞 1．体外分离、培养SD大鼠MSCs。 2．分别用不同浓度的5-氮胞苷(5-azacytidine，5-aza)对MSCs定向诱导，观察细胞形态学变化：在培养3周时，用免疫细胞化学染色的方法检测细胞中的心肌特异性肌钙蛋白T(cTnT)；用RT-PCR对心肌肌球蛋白重链(β-MHC)进行鉴定；及电镜下观察细胞内部结构。取SD大鼠心肌细胞作为阳性对照，，未经5-氮胞苷诱导正常培养的MSCs作为阴性对照。
[Abstract]:Objective: acute myocardial infarction (AMI) is one of the major diseases endangering human health. Although mature cardiomyocytes are not terminal differentiation cells, they are also limited in division. The proliferative ability of AMI patients may be due to scar formation and residual myocardial compensatory hypertrophy after myocardial infarction (MI), even though they are lucky enough to pass through the acute phase. It causes ventricular remodeling, which in turn promotes heart failure and ultimately leads to death. Nowadays, thrombolytic therapy, percutaneous transluminal coronary angioplasty (PTCA), coronary artery bypass grafting (CABG) and heart transplantation are commonly used in the treatment of AMI. The first three methods can improve the blood supply of myocardium, but can not regenerate the necrotic myocardium. Although heart transplantation can replace the damaged heart, it is difficult to be widely used. In recent years, with the development of molecular biology, the method of adult stem cell implantation is used to increase the number of myocardial cells in the infarct area, thus improve the cardiac function and reduce the long-term mortality after AMI, which opens up a new prospect for the treatment of AMI. At present, adult stem cells are mainly derived from bone marrow stem cells, skeletal muscle satellite cells, smooth muscle stem cells, embryonic stem cells and fetal cardiomyocytes. Bone marrow stem cells are widely distributed and easy to obtain in human body. There is no immune rejection in implantation and there is no ethical debate, so they have advantages in the clinical application of cellular myocardial repair. The aim of this study was to investigate the feasibility of transforming bone marrow derived adult stem cells into cardiomyocyte-like cells induced by 5-azacytidine and Panax notoginseng saponins in vitro, and co-cultured bone marrow stem cells with cardiomyocytes. To investigate the effect of myocardial microenvironment on this transformation process. Methods: I. Differentiation of bone marrow mesenchymal stem cells into cardiomyoid cells in vitro 1. MSCs of SD rats were isolated and cultured in vitro. 2. MSCs was induced by 5-azacytidine 5-azazacydine 5-aza at different concentrations, and the morphological changes were observed. The specific cardiac troponin TcTnTnTnTnTX was detected by immunocytochemical staining, the cardiac myosin heavy chain (尾 -MHCs) was identified by RT-PCR, and the structure of the cells was observed under electron microscope. SD rat cardiomyocytes were used as positive control and normal cultured MSCs without 5-azacytidine as negative control.
【学位授予单位】：大连医科大学
【学位级别】：博士
【学位授予年份】：2006
【分类号】：R542.22;R329

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