Transcriptomics of adherent endothelial cells for improved endothelialization of small-diameter vascular grafts

项目摘要 本研究的长期目标是在血液接触表面建立自体内皮 植入式心血管设备这对于提供血液相容性和最小化 血栓形成、栓塞和其他不良后果的风险，以提高患者安全性。 这一目标直接支持NHLBI促进心脏病治疗的使命， 增强所有人的健康，使他们能够活得更长，更有成效。许多心血管 疾病是用可植入的装置来治疗的。在拯救生命的同时，这些设备也存在固有的风险， 血栓形成和栓塞，需要抗凝治疗。 这项研究解决了在小直径血管内建立内皮细胞的迫切临床需求。 血管移植物需要搭桥手术的冠心病患者目前接受自体 血管收获，因为合成移植物在小直径应用中显示出较差的通畅性。自体 血管获取与额外的成本和显著的供体部位发病率相关。此外，一个大型和 越来越多的患者由于包括预先存在的血管 疾病，静脉剥离，和以前的收获。自体小直径血管内皮细胞的建立 旁路移植物对于实现可接受的通畅率是必要的，而无需自体血管收获。 本研究的目的是通过以下方法改善血管移植生物材料上内皮细胞的保留 为促进细胞粘附的分子调节策略鉴定新的分子信号传导靶标 实力这是由我们的假设驱动的，即参与细胞凋亡的分子信号通路的差异调节。 细胞粘附是内皮细胞亚群在植入时抵抗分离的原因。 具体目标1将确定负责允许内皮细胞亚群的分子信号传导途径， 细胞在暴露于生理剪切应力时保持粘附。RNA测序将用于比较 所有细胞和贴壁细胞的转录组。转录组学数据将用于鉴定分子 在贴壁细胞亚群中高度差异调节的信号。 具体目标2将开发和测试改善血管移植物上内皮细胞保留的策略 生物材料将开发一种分子调节策略，通过上调或抑制细胞粘附来增强细胞粘附。 适当下调关键信号分子。将在以下条件下比较细胞保留： 生理剪应力 特定目标3将评估接种内皮细胞的小直径血管移植物的生物学性能 使用临床前模型的细胞。最有前途的细胞粘附增强策略将用于 内皮化移植物，将其植入猪的颈动脉循环中并进行临床分析。 相关结果包括细胞滞留、通畅性、血栓形成、新生内膜增生和炎症。

Project Summary The long-term objective of this study is to establish an autologous endothelium on the blood-contacting surface of implantable cardiovascular devices. This is critically important for providing hemocompatibility and minimizing the risk of thrombosis, embolism, and other adverse outcomes in order to improve patient safety. This objective directly supports the mission of the NHLBI to promote the treatment of heart diseases and enhance the health of all individuals so that they can live longer and more productive lives. Many cardiovascular diseases are treated with implantable devices. While lifesaving, these devices also carry an inherent risk of thrombosis and embolism that requires administration of anticoagulation therapy. The proposed study addresses the dire clinical need for establishing an endothelium within small-diameter vascular grafts. Patients with coronary heart disease who require bypass surgery currently undergo autologous vessel harvest because synthetic grafts demonstrate poor patency in small-diameter applications. Autologous vessel harvest is associated with additional cost and significant donor site morbidity. In addition, a large and growing number of patients do not possess suitable autologous vessel for reasons including preexisting vascular disease, vein stripping, and previous harvest. Establishment of an autologous endothelium on small-diameter bypass grafts is necessary for achieving acceptable patency rates without autologous vessel harvest. The goal of the proposed research is to improve endothelial cell retention on vascular graft biomaterials by identifying novel molecular signaling targets for molecular modulation strategies to promote cellular adhesion strength. This is driven by our hypothesis that differential regulation of molecular signaling pathways involved in cellular adhesion is responsible for a subpopulation of endothelial cells resisting detachment upon implantation. Specific Aim 1 will identify molecular signaling pathways responsible for allowing a subpopulation of endothelial cells to remain adherent upon exposure to physiological shear stress. RNA-sequencing will be used to compare the transcriptome of all cells and adherent cells. The transcriptomics data will be used to identify molecular signals that are highly differentially regulated in adherent cell subpopulations. Specific Aim 2 will develop and test strategies to improve endothelial cell retention on vascular graft biomaterials. A molecular modulation strategy will be developed to enhance cell adhesion by upregulating or downregulating critical signaling molecules as appropriate. Cell retention will be compared under conditions of physiological shear stress. Specific Aim 3 will assess the biological performance of small-diameter vascular grafts seeded with endothelial cells using a preclinical model. The most promising cell adhesion enhancement strategy will be used to endothelialize grafts, which will be implanted into the carotid circulation of swine and analyzed for clinically relevant outcomes including cell retention, patency, thrombosis, neointimal hyperplasia, and inflammation.