Genetic Engineering of Vein Bypass Grafts in Vascular and Cardiovascular Surgery

项目总结/摘要 自体静脉移植是最常见和最有效的旁路移植时，用于心脏治疗 冠状动脉疾病或下肢，以防止截肢。尽管如此， 静脉移植物中约有30%是由于内膜增生（IH）所致。反过来，IH是 当静脉从低压、血管依赖性血管移走时对植入损伤的反应 动脉系统的高压、管腔依赖性生理学。我们的目标是尽量减少 这种植入损伤，从而减少与IH相关的适应不良的病理性重塑。我们 实验设计总是以实际考虑为指导， 可以在手术室的临床限制内进行。我们的策略是首先 确定植入后1天至30天的时间依赖性基因表达反应（转录组）， 然后确定驱动病理性重塑的最重要的“枢纽”基因。我们的治疗 一种方法是沉默血管壁中的这些致病性枢纽基因。因此，我们击倒了， 测试了我们最有希望的两个靶点，血小板反应蛋白-2（TSP-2）和富含豆蔻酰丙氨酸的C-激酶 基质（MARCKS）在两种不同的可重复的大型动物模型中记录分子和组织学 结果。展望未来，我们将通过过度表达 动脉粥样硬化保护，抗炎基因，TNF-α诱导蛋白-3（TNFAIP 3或A20），临床使用 可应用的亲血管性基因治疗载体。一种多模式和双向（动脉粥样硬化和 动脉粥样硬化保护基因的过度表达）的治疗方案可能需要解决复杂的 IH的发病机制。首次使用最先进的集成单电池（Sc）， RNA测序我们将绘制VG对植入的异质性反应的转录组粒度图， 伤害和治疗。这可以更精确地确定血管和非血管的作用，但影响， 免疫细胞、肌成纤维细胞等细胞亚群参与VG植入损伤的发病机制。我们 对人类疾病具有高度生物逼真度的犬静脉移植模型的初步结果揭示了相关的新 信息，强调了迄今为止被低估的致病性T辅助1细胞（Th 1）对VG的贡献 重塑这些新的实验将使我们进入临床前准备阶段， 最先进的输送系统和分子技术，以减少静脉移植物的植入损伤， 减少其他血管疾病中的IH。我们的研究团队的结构是利用MPI格式， 整合使该项目取得成果所需的广泛专业知识，并保持对信息健康的持久关注 问题.

PROJECT SUMMARY/ABSTRACT Autologous vein grafts are the most common and most effective bypass grafts when used in the heart to treat coronary artery disease or in the lower extremity to prevent amputations. Nonetheless the delayed failure rate of vein grafts is about 30%, almost all due to intimal hyperpalsia (IH). IH, in turn, is an integral part of the response to implantation injury when the vein is moved from the low pressure, vasa vasorum dependent physiology, to the high pressure, luminal dependent physiology of the arterial system. Our goal is to minimize this implantation injury, and thereby diminish the maladaptive, pathologic remodeling associated with IH. Our experimental design has always been guided by practical considerations whereby the therapeutic intervention could be conducted within the clinical constraints of the operating room. Our strategy has been to first determine the time dependent gene expression response (transcriptome), 1 day to 30 days after implantation, followed by identifying the most important ‘Hub” genes that drive the pathologic remodeling. Our therapeutic approach is to silence these pathogenic hub genes in the vascular wall. Therefore, we knocked down and tested two of our most promising targets, Thrombospondin-2 (TSP-2) and Myristoylated alanine-rich C-kinase substrate (MARCKS) in two different reproducible large animal models to document molecular and histologic outcomes. Going forward we are going to supplement our therapeutic regimen by overexpressing the atheroprotective, anti-inflammatory gene, TNF-alpha induced protein-3 (TNFAIP3 or A20), using clinically applicable vasculotropic gene therapy vectors. A multimodal and bidirectional (knockdown of atherogenic and overexpression of atheroprotective genes) therapeutic regimen may well be required to address the complex pathogenesis of IH. Using, for the first time, the state-of-the-art integrated single cell (Sc), coupled with bulk RNA-sequencing we will map the transcriptomic granularity of the VG heterogeneous response to implantation injury and to therapy. This could determine more precisely the role of vascular and non-vascular, yet influential, cellular subsets such as immune cells, myofibroblasts etc. in the pathogenesis of VG implantation injury. Our preliminary results in a canine vein graft model with high biofidelity to human disease uncovers pertinent novel information, highlighting a so far underestimated contribution of pathogenic T helper 1 cells (Th1) to VG remodeling. These new experiments will bring us to preclinical readiness in a strategy designed to use the most advanced delivery systems and molecular technologies to reduce implantation injury to vein grafts and diminish IH in other vascular diseases. Our research team is structured to take advantage of the MPI format to coalesce the broad expertise required to bring this project to fruition and maintain an enduring focus on the IH problem.