TNF-INHIBITOR GENE THERAPY FOR NEOINTIMAL HYPERPLASIA

经皮冠状动脉介入治疗（PCI）和冠状动脉旁路移植术（CABG）在美国每年都有大约500，000例患者接受。这两种手术都因血管壁中细胞的活化、迁移和增殖而复杂化，导致新生内膜增生（NH）。大量的证据表明，这些细胞大多数是血管平滑肌起源。NH影响约30%的所有PCI患者，并且在两年内约30%的所有CABG病例中需要再次介入。每年仅治疗PCI再狭窄所需的费用就超过20亿美元。PCI和隐静脉CABG手术均导致下方血管壁的显著损伤，引起内皮功能障碍和血小板、中性粒细胞和单核细胞/巨噬细胞谱系的循环细胞的粘附。局部释放的细胞因子如肿瘤坏死因子（TNF）与生长因子（包括血小板衍生生长因子（PDGF）和碱性成纤维细胞生长因子（bFGF））协同作用，以激活受损血管段中的SMC。体外TNF是SMC的强促分裂剂，并且可以与PDGF-BB协同作用以刺激细胞生长。这项工作提出了检查的贡献，TNF的SMC的激活，单独和结合各种生长因子。为此，将在TNF处理的和未处理的SMC中检查丝裂原活化蛋白激酶（MAPK）活化、DNA合成、增殖、趋化性和凋亡。此外，该提案旨在测试抑制TNF可以降低SMC体外活化程度并可以在实验动物模型中减少体内新生内膜形成的大小的假设。为了抑制TNF，产生嵌合TNF受体胞外结构域-鼠免疫球蛋白重链构建体（TNF-I）并插入重组腺病毒（rec. AD-TNF-I）中。该病毒将用于感染离体兔颈静脉，其将在新生内膜增生的良好表征模型中跨兔颈动脉移植。旁路移植术后4周，将在组织学上测量rec.AD-TNF-I和rec. AD-对照感染静脉中新生内膜形成的程度。这将在接受正常饮食的家兔和接受高脂肪饮食的动物中进行，以检查高脂血症对这种情况下新生内膜增生发展的贡献。通过抑制TNF减少静脉移植物新生内膜增生可能对人类血管疾病具有重要的治疗意义。

Percutaneous coronary intervention (PCI) and coronary artery bypass graft surgery (CABG) are each performed on approximately 500,000 patients annually in the US. Both procedures are complicated by activation, migration and proliferation of cells in the vascular wall, leading to neointimal hyperplasia (NH). A preponderance of evidence suggests that the majority of these cells are of vascular smooth muscle origin. NH affects about 30 % of all PCI patients and necessitates reintervention in some 30 % of all CABG cases within two years. The annual cost needed to treat PCI restenosis alone exceeds $2 billion. PCI and saphenous vein CABG surgery both lead to significant injury of the underlying vessel wall, causing endothelial dysfunction and adherence of platelets, neutrophils and circulating cells of the monocyte / macrophage lineage. Locally released cytokines, such as tumor necrosis factor (TNF), synergize with growth factors, including platelet derived growth factor (PDGF) and basic fibroblast growth factor (bFGF), to activate SMCs in the injured vessel segment. In vitro TNF is a strong mitogen for SMCs and can act in concert with PDGF-BB, to stimulate cell growth. This work proposes to examine the contribution of TNF to the activation of SMCs, alone and in combination with various growth factors. To this end mitogen activated protein kinase (MAPK) activation, DNA synthesis, proliferation, chemotaxis and apoptosis will be examined in TNF treated and untreated SMCs. In addition, this proposal seeks to test the hypothesis that inhibition of TNF can decrease the degree of SMC activation in vitro and can reduce the size of neointima formation in vivo in an experimental animal model. To inhibit TNF, a chimeric TNF receptor extracellular domain - murine immunoglobulin heavy chain construct (TNF-I) was generated and inserted into a recombinant adenovirus (rec. AD-TNF-I). This virus will be used to infect rabbit jugular veins ex vivo that will be grafted across the rabbit's carotid artery in a well characterized model of neointima hyperplasia. Four weeks after bypass grafting the degree of neointima formation in rec.AD-TNF-I and rec.AD-control infected veins will be measured histologically. This will be done both in rabbits receiving a normal diet and also in animals given a high fat diet to examine the contribution of hyperlipidemia towards the development of neointimal hyperplasia in this setting. Reduction of vein graft neointimal hyperplasia by inhibition of TNF may have significant therapeutic implications for human vascular diseases.