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， 将离体移植物移植到骨骼肌的毛细血管床中， 肌肉. 这种体细胞基因治疗技术可以用来实现 通过与血管床共同或重新填充血管床产生的局部或全身效应 内皮细胞组成性或诱导性表达一种保护性 重组蛋白，如组织纤溶酶原激活物或因子IX。 因此，这种体细胞基因治疗技术的发展可以 广泛应用于外科和遗传性人类疾病。 的 该项目的具体目标是： 目的I：鉴定和表征介导细胞凋亡的分子机制。 内皮细胞在静止的融合的内皮细胞上的粘附 细胞单层。 目的II：优化基因工程药物的粘附和掺入， 转导的内皮细胞移植到毛细血管床， 骨骼肌。 目的III：记录内皮细胞的长期重组基因表达 细胞移植到骨骼肌的毛细血管床中。 研究者在体外模型方面有相当丰富的经验， 研究内皮细胞-内皮细胞粘附以及体内 研究转基因内皮细胞移植的rt模型 进入骨骼肌毛细血管床。 所提出的实验，以实现 上述目的包括鉴定和表征特定 粘附分子介导内皮细胞-内皮细胞粘附。 这些实验将揭示关于 内皮细胞-内皮细胞相互作用。 这些知识也可以 用于增强转导的内皮细胞的粘附和结合 进入毛细血管壁。 在大鼠模型中， 移植到毛细血管床中的基因转导的内皮细胞将 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 转基因内皮细胞移植的最佳技术 在上述研究中确定的毛细管床将用于 证明了该技术递送重组蛋白质的功效 进入循环。 在这些研究完成后，这项技术 体细胞基因治疗的研究将准备在灵长类动物模型中进行评估， 准备用于人类。

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.