Development of a Polymeric Delivery System for Efficient in vivo Cellular Reprogramming for Cardiac Regeneration

开发用于心脏再生的有效体内细胞重编程的聚合物递送系统

• 批准号: 10063433
• 负责人: Laura Elisabeth Saunders
• 金额: $ 4.55万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-29 至 2021-09-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10063433
• 关键词: Acute myocardial infarction; Aftercare; Alternative Therapies; Automobile Driving; Bone Tissue; Cardiac; Cardiac Myocytes; Cardiovascular Diseases; Catheters; Cause of Death; Cell Line; Cell Nucleus; Cessation of life; Chemicals; Chronic; Complex; DNA delivery; Data; Development; Disease; EFRAC; Fibroblasts; Fibrosis; GATA4 gene; Gene Delivery; Gene Expression; Goals; Gold; Heart; Hydrogels; Immunohistochemistry; Impairment; In Vitro; Injectable; Injections; Kinetics; Lipofectamine; Magnetic Resonance Imaging; Measures; Mechanics; Modeling; Modulus; Myocardial Infarction; Myocardial tissue; Natural regeneration; Nuclear; Nuclear Localization Signal; Obesity; Operative Surgical Procedures; Organ; Outcome; Patients; Peptide Fragments; Phenotype; Plasmids; Polyethyleneimine; Polymers; Rattus; Recovery of Function; Research; Research Personnel; Risk Factors; Saline; Severities; Signaling Protein; Stains; Stroke Volume; System; Technology; Testing; Time; Tissues; Transfection; Traumatic injury; Treatment Efficacy; Viral; Work; World Health Organization; aging population; base; body system; cardiac regeneration; clinical application; clinically relevant; coronary fibrosis; crosslink; density; disability; gene delivery system; heart function; immunogenic; improved; in vivo; in vivo regeneration; minimally invasive; mouse model; non-viral gene delivery; novel; off-target mutation; plasmid DNA; regenerative; scaffold; tissue regeneration; transcription factor; treatment strategy; tumorigenic; uptake

PROJECT SUMMARY The goal of this study is to develop a targeted, injectable plasmid delivery system that is capable of efficient direct reprogramming of cardiac fibroblasts to cardiomyocytes (CMCs) in vivo, aiding in cardiac tissue regeneration following a myocardial infarction (MI) or ‘heart attack’. Cardiovascular disease (CVD) is the leading cause of death worldwide and resulted in 17.9 million deaths in 2016 alone. Cardiac fibrosis, a major contributor to CVD is which results from traumatic injury such as MI. A promising treatment avenue is direct reprogramming of cardiac fibroblasts into CMCs to regenerate the myocardial tissue and regain functional post-infarct heart tissue. Several recent studies have demonstrated that a combination of transcription factors, Gata4, Mef2c, and Tbx5 delivered simultaneously can directly reprogram cardiac fibroblasts to CMCs and induce CMC phenotype in vitro and in vivo. Unfortunately, while cardiac fibroblast to CMC differentiation is now a possibility, implementation of this approach as a CVD treatment has not been viable. The gene delivery vehicles (polyplexes) developed in the Ma Lab have been shown to drive bone tissue regeneration in vivo, setting an important precedent on which this study is based. Preliminary data using an improved polyplex has thus far driven in vitro reprogramming of cardiac fibroblasts into CMCs, a promising proof-of-concept result. In the proposed study, researchers will be further improving the recently developed system by improving the gene expression through nuclear targeting using a Nuclear Localization Signal (NLS). Preliminary results indicate this improves gene expression compared to polyplexes alone. Further, researchers will be driving direct reprogramming in vivo using a newly developed hydrogel scaffold to implement sustained delivery of the polyplexes. The hypothesis is that through addition of a NLS and sustained delivery of plasmid DNA using an injectable hydrogel, the nuclear localization and amount of plasmid delivered will be increased, resulting in an increase in in vivo reprogramming of fibroblasts into CMCs. This will be evaluated using PCR and immunohistochemistry to compare gene expression and later MRI to evaluate functional changes in the heart. This research aims to establish the first regenerative treatment option for patients who have suffered a heart attack. This work may likely lay the framework for an effective non-viral gene delivery system for use in treatment of other lost or diseased tissues, especially the fibrosis of other tissue/organ systems.

项目总结 这项研究的目标是开发一种靶向的、可注射的、能够有效地 心脏成纤维细胞在体内直接重编程为心肌细胞(CMC)，有助于心脏组织 心肌梗死(MI)或心脏病发作后的再生。心血管疾病(CVD)是 全球主要死因，仅在2016年就造成1790万人死亡。心脏纤维化，一个主要的 心血管疾病的诱因是由创伤造成的，如心肌梗死。一种有希望的治疗途径是直接的 将心脏成纤维细胞重新编程为CMCs以再生心肌组织并恢复功能 梗死后的心脏组织。最近的几项研究表明，转录因子的组合， 同时传递的Gata4、MEF2C和Tbx5可以直接将心脏成纤维细胞重新编程为CMCs和 体外和体内诱导CMC表型。不幸的是，虽然心脏成纤维细胞现在正在向CMC分化 有一种可能性，将这种方法作为一种心血管疾病的治疗方法是不可行的。基因传递 在Ma实验室中开发的载体(复合体)已被证明可以在体内驱动骨组织再生， 开创了这项研究的重要先例。使用改进的Polyplex的初步数据具有 到目前为止，心脏成纤维细胞在体外被重新编程为CMCs，这是一个有希望的概念验证结果。在……里面 在这项拟议的研究中，研究人员将通过改进基因来进一步完善最近开发的系统 通过使用核定位信号(NLS)进行核靶向表达。初步结果显示 与单独的多链相比，这提高了基因的表达。此外，研究人员将直接驾驶 使用新开发的水凝胶支架在体内重新编程以实现持续递送 复式影院。假设是通过添加NLS和使用AFP持续递送质粒DNA 注射水凝胶后，核的定位和运送的质粒量将增加，从而导致 成纤维细胞体内重编程为巨噬细胞的增加。这将使用聚合酶链式反应和 用免疫组织化学方法比较基因表达，然后用MRI来评估心脏的功能变化。 这项研究旨在为心脏病患者建立第一种再生治疗方案 进攻。这项工作可能会为有效的非病毒基因递送系统奠定基础，用于 治疗其他丢失或病变的组织，尤指其他组织/器官系统的纤维化。