Oxidized Low-density Lipoprotein Impairs Bone Marrow Stem Cell Survival

氧化低密度脂蛋白损害骨髓干细胞的存活

• 批准号: 9128038
• 负责人: ZHENGUO LIU
• 金额: $ 52.08万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-01 至 2019-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9128038
• 关键词: Acute; Animal Model; Animals; Blood; Blood Circulation; Bone Marrow Stem Cell; Cardiac; Cardiac Myocytes; Cardiovascular Diseases; Cell Culture System; Cell Survival; Cell Therapy; Cell Transplantation; Cell membrane; Cells; Coronary Arteriosclerosis; Data; Doxycycline; Dyes; Endothelial Cells; Engineering; Ethics; Exposure to; Fibroblasts; Fluorescent Dyes; Health; Human; Hyperlipidemia; Hypoxia; Immune; Impairment; Implant; In Vitro; Individual; Injury; Intravenous; Knockout Mice; Lactate Dehydrogenase; Link; Lipids; Lipoprotein (a); Low Density Lipoprotein Receptor; Low-Density Lipoproteins; Mechanical Stress; Mechanics; Mediating; Membrane; Molecular; Mus; Muscle Cells; Myocardial Ischemia; Myocardium; Natural Products; Natural regeneration; Outcome; Patients; Principal Investigator; Production; Proteins; Recombinants; Role; Serum; Skeletal Muscle; Source; Stem cell transplant; System; Testing; Tissues; Transgenic Mice; Transplantation; Treatment Efficacy; Umbilical vein; Up-Regulation; base; body system; cardiac regeneration; cardiac repair; clinical application; design; extracellular; immunogenicity; in vivo; intravenous administration; mouse model; overexpression; oxidized low density lipoprotein; promoter; protective effect; repaired; research study; tissue regeneration; tissue repair

 DESCRIPTION (provided by applicant): Cell therapy with bone marrow stem cells (BMSCs) remains a viable option for tissue repair and regeneration. One major challenge for cell-based therapy is the poor survival of the cells after transplantation. The exact mechanism(s) for impaired survival of the implanted BMSCs remains to be defined. MG53 is an important cell membrane repair protein that is naturally produced in skeletal and cardiac muscles, and present in blood that mediates the tissue protective function for MG53. Oxidized low-density lipoprotein (ox-LDL) is a natural product in blood. We observed that ox-LDL at the concentrations compatible with the serum ox-LDL levels in patients with coronary artery disease produced significant membrane damage to BMSCs as reflected by the release of lactate dehydrogenase (LDH) from the cell and the entry of fluorescent dye FM1-43 into the cell. Treatment with recombinant human MG53 (rhMG53) protein could reduce LDH release and FM1-43 dye entry in BMSCs exposed to ox-LDL. Further in vivo studies showed that intravenous ox-LDL administration led to decreased serum MG53 level, and impaired survival of BMSCs implemented into normal C57B6 mice. The proposed study will test the hypothesis that "ox-LDL impairs the survival of BMSCs through direct damage to their cell membrane, and ox-LDL-mediated reduction of MG53 level in circulation is an important factor for the limited survival of BMSCs with compromised cell membrane repair capacity and reduced efficacy of cell-based therapy". The specific aims are: 1) to determine the effect of ox-LDL on the membrane integrity of BMSCs both in vitro and in vivo; and 2) to define the mechanism(s) for ox-LDL-mediated action on MG53 and the survival of BMSCs in cell transplantation. We will elucidate the mechanisms that underlie the ox-LDL-induced damage to BMSCs in culture system, and also conduct experiments to evaluate the protective effect of rhMG53 on BMSCs following mechanical or hypoxia-induced membrane injuries. We will further engineer BMSCs that express secretary MG53 to determine if sustained MG53 production has beneficial effects on the survival of BMSCs in vivo in ox-LDL-treated mice and in hyperlipidemic animals. Efforts will be made to determine the mechanism on how ox-LDL causes impairment in MG53-mediated membrane repair. We will determine whether ox-LDL causes decreased MG53 secretion from muscle cells or increased its clearance from circulation. We will further test the hypothesis with the use of MG53 knockout mice and transgenic mouse models where inducible and reversible MG53 expression in circulation can be tailored with a doxycycline-driven promoter system. These studies will reveal the role for MG53-mediated membrane repair in cell-based therapy, provide important information on the molecular mechanisms for the poor survival of implanted BMSCs, and help to explore new strategies like targeting ox- LDL and circulating MG53 to optimizing the survival of BMSCs and enhancing the efficacy of cell-based therapy for cardiovascular diseases. 1

 描述（由申请人提供）：使用骨髓干细胞（BMSC）的细胞疗法仍然是修复组织和再生的可行选择。基于细胞的疗法的一个主要挑战是移植后细胞的存活率较差。 ）对于植入的 BMSC 的存活受损仍有待确定。 MG53 是一种重要的细胞膜修复蛋白，在骨骼肌和心肌中自然产生，存在于血液中，介导 MG53 的组织保护功能。氧化低密度脂蛋白 (ox-LDL) 是血液中的天然产物，我们观察到，与冠状动脉疾病患者血清 ox-LDL 水平相匹配的 ox-LDL 对 BMSC 产生了显着的膜损伤。乳酸脱氢酶 (LDH) 从细胞中释放以及荧光染料 FM1-43 进入细胞内，用重组人 MG53 (rhMG53) 蛋白处理可以减少 LDH 释放和暴露于 ox-LDL 的 BMSC 中的 FM1-43 染料进入。进一步的体内研究表明，静脉注射 ox-LDL 会导致血清 MG53 水平降低，并损害正常 C57B6 小鼠中的 BMSC 存活率。 “ox-LDL 通过直接损伤细胞膜来损害 BMSC 的存活，ox-LDL 介导的循环中 MG53 水平的降低是 BMSC 存活有限的一个重要因素。细胞膜修复能力受损，细胞疗法疗效降低”。具体目标是：1) 确定 ox-LDL 在体外和体内对 BMSC 膜完整性的影响；2) 明确其机制(s) ox-LDL 介导的 MG53 作用和细胞移植中 BMSC 的存活 我们将阐明 ox-LDL 诱导培养系统中 BMSC 损伤的机制，并进行实验来评估其保护作用。 rhMG53 对机械或缺氧诱导的膜损伤后 BMSC 的影响我们将进一步改造表达秘书 MG53 的 BMSC，以确定持续产生 MG53 是否对 ox-LDL 处理的小鼠和高脂血症动物体内 BMSC 的存活产生有益影响。我们将努力确定 ox-LDL 如何导致 MG53 介导的膜修复受损的机制。我们将确定 ox-LDL 是否会导致 MG53 分泌减少。我们将使用 MG53 敲除小鼠和转基因小鼠模型进一步测试这一假设，其中循环中的可诱导和可逆 MG53 表达可以通过多西环素驱动的启动子系统进行定制。 MG53介导的膜修复在细胞治疗中的作用，提供有关植入的BMSCs存活不良的分子机制的重要信息，并有助于探索新的策略，例如靶向ox-LDL和循环MG53 可优化 BMSC 的存活并增强心血管疾病细胞疗法的功效 1。