TRANSPLANT OF TRANSDUCED ENDOTHELIUM INTO CAPILLARIES

这项研究项目的长期目标是开发一种技术 在体细胞基因治疗中，内皮细胞经历了基因 体外移植到骨骼的毛细血管床中 肌肉。这种体细胞基因治疗技术可以用来实现 通过共同或重新填充血管床而产生的局部或系统效应 构成或诱导表达保护性蛋白的内皮细胞 重组蛋白，如组织型纤溶酶原激活剂或因子IX。 因此，这种体细胞基因治疗技术的发展可能 对外科和遗传性人类死亡都有广泛的应用。这个 该项目的具体目标是： 目的I：鉴定和表征介导血管紧张素转换酶的分子机制。 种植于静止、融合内皮细胞上的内皮细胞的黏附 细胞单层。 目的II：优化基因工程的黏附和整合 转导内皮细胞移植到血管内皮细胞床 骨骼肌。 目的III：证明血管内皮细胞长期表达重组基因 将细胞移植到骨骼肌的毛细血管床中。 研究人员在体外模型方面有相当多的经验 研究内皮细胞-内皮细胞黏附以及体内实验 RT模型用于转基因内皮细胞移植的研究 进入骨骼肌毛细血管床。建议的实验以实现 上述目标包括识别和表征特定的 黏附分子介导内皮细胞与内皮细胞的黏附。 这些实验将揭示关于 内皮细胞与内皮细胞的相互作用。这一知识还可以 用于增强转导内皮细胞的黏附和掺入 进入毛细管壁。在大鼠模型中，黏附和整合 将基因转导的内皮细胞移植到毛细血管床将 通过注射碱性成纤维细胞诱导血管生成来优化 生长因子或刺激骨骼肌再生。最后， 转导内皮细胞移植到宿主体内的最佳技术 在上述研究中确定的毛细管床将用于 记录这种传递重组蛋白的技术的有效性 进入血液循环。在这些研究完成后，这项技术 体细胞基因疗法将准备好在灵长类动物模型中进行评估 为它在人类身上的使用做准备。

The long-term objective of this research project is to develop a technique of somatic gene therapy in which endothelial cells that have undergone gene transfer ex vivo are transplanted into the capillary bed of skeletal muscle. This technique of somatic gene therapy could be used to achieve a regional or systemic effect by co-or repopulating a vascular bed with endothelial cells which constitutively or inducibly express a protective recombinant protein such as tissue plasminogen activator or Factor IX. Thus, the development of such a technique of somatic gene therapy could have a wide application to both surgical and genetic human deceases. The Specific Aims of the project are: Aim I: To identify and characterize the molecular mechanism mediating the adhesion of endothelial cells seeded onto quiescent, confluent endothelial cell monolayers. Aim II; To optimize the adhesion and incorporation of genetically transduced endothelial cells transplanted into the capillary bed of skeletal muscle. Aim III; To document long-term recombinant gene expression by endothelial cells transplanted into the capillary bed of skeletal muscle. The investigators have considerable experience with an in vitro model to study endothelial cell-endothelial cell adhesion as well as with an in vivo rt model to study transplantation of genetically modified endothelial cells into skeletal muscle capillary beds. The proposed experiments to achieve th above Aims include identification and characterization of specific adhesion molecules mediating endothelial cell-endothelial cell adhesion. These experiments would reveal fundamental new knowledge concerning endothelial cell-endothelial cell interactions. This knowledge could also be used to enhance adhesion and incorporation of transduced endothelium into the capillary wall. In a rat model the adhesion and incorporation of genetically transduced endothelium transplanted into the capillary bed will be optimized by angiogenesis induced with injection of basic fibroblast growth factor or stimulation f skeletal muscle regeneration. Finally, the optimal techniques of transplantation of transduced endothelium into host capillary beds, determined in the aforenoted studies, will be used to document the efficacy of this technique to deliver recombinant proteins into the circulation. At the completion of these studies, this technique of somatic gene therapy will be ready for evaluation in a primate model in preparations for its use in humans.