Inhibition of neuronal cell death in Parkinson's disease by transduction of neurotrophic factor gene.

色素上皮衍生因子(PEDF)是从培养的人视网膜色素上皮细胞条件培养液中首次发现的一种神经营养物质，可诱导Y-79视网膜母细胞瘤细胞向神经元分化，并对培养的小脑颗粒细胞具有神经营养存活作用。PEDF对谷氨酸毒性和诱导培养的颗粒细胞凋亡也有保护作用。在本项目中，我们首先研究了人重组PEDF(HrPEDF)对培养的黑质神经元增殖的影响。为了进行这项实验，我们使用了从胎鼠脑中分离的黑质神经元的培养。酪氨酸羟化酶免疫组织化学染色显示，PEDF组大鼠黑质神经元数量明显多于对照组。PEDF的作用呈剂量-反应关系。在第二项研究中，我们将PEDF基因导入小鼠的纹状体和黑质。为此，我们使用了足以包装和整合的重组腺相关(AAV)载体。将含有PEDF和GFP编码基因的AAV通过纹状体行纹状体内注射。在纹状体和黑质观察到GFP阳性细胞，它们通过纹状体途径运输。我们现在正在这些切片上进行PEDF的免疫组织化学研究，以确认PEDF的表达。我们的研究表明，PEDF基因转移到纹状体和黑质的可能性和对中性神经元存活的影响，以及进一步对帕金森病的基因治疗的潜在作用

Pigment epithelium-derived factor (PEDF), a neurotrophic agents first identified in conditioned medium from cultured human retinal pigment epithelial cells, induces neuronal differentiation with neuritic outgrowth in Y-79 retinoblastoma cells and has a neurotrophic survival effect on cerebellar granule cells in culture. PEDF also have protective effect against glutamate toxicity and induction of apoptotic cell death of cultured granular cells. In this project, first we investigated the effects of human recombinant PEDF (hrPEDF) on proliferation of cultured nigral neurones. To do this experiments, we used cultures of nigral neurones isolated from the fetal rat brains. The number of nigral neurones, which identified by immunohistochemistry for tyrosine hydroxilase, is larger in the presence of PEDF than that of controls. The effect of PEDF showed a dose-response relationship. In second studies, we have performed the transduction of PEDF gene to the striatum and nigra of mice. To this purpose, we used recombinent adeno-associated (AAV) vectors which are sufficient for packaging and integration. AAV containing cDNA encoding PEDF and GFP were injected to the striatum of the mice by streotaxic operation. GFP-positive cells were observed in the striatum and the substantia nigra, which were transported through strionigral pathway. We are now conducting immunohistochemistry for PEDF on these sections to confirm the expression of the PEDF.Our study suggest the possibility of PEDF gene transfer to the striatum and nigra and effect of survival of niral neurones, and furthermore potential role for the gene therapy of Parkinson's disease