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Transplantation for retinal diseases

视网膜疾病移植

基本信息

批准号：
14370553
负责人：
ABE Toshiaki
金额：
$ 8.58万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2005
项目状态：
已结题

项目摘要

We report a subretinal iris pigment epithelial cell (IPE) transplantation with safely. When these cells were genetically engineered that expressing neurotrophic factors, the transplantation rescued the photoreceptor cells from phototoxicity. We used adeno-associate virus (AAV2) for delivering neurotrophic factors, such as brain-derived neurotrophic factor (BDNF) to the transplanted cells. If we used more than 1x107 capsides/ml AAV-BDNF for transfection, statistically significant photoreceptor protection was observed in the AAV-BDNF-IPE transplantation from phototoxicity. However, the rescue effect was not dose-dependent. Then, we examined the BDNF receptor TrkB expression, because the receptor may control the effect of BDNF. So far many isoforms of TrkB have reported and we examined two of the major isoforms of TrkB ; TrkB-FL which has tyrosine kinase activity in the cell and TrkB-T1 which has not. These isoforms showed not always completely same expression by immunohistochemistry. TrkB-FL was expressed mainly on nerve fiber layer, ganglion cells layer, and inner nuclear layer (INL), conversely TrkB-T1 was on INL and RPE. Both isoforms were not expressed in ONL and photoreceptor layer. From the results of double immunostaining with S100β, These isforms were expressed on Muller cells. Further these isoforms were expressed in the retina not only spatially but also temporally different way, although the expression was enhanced by the AAV-BDNF-IPE transplantation. When we performed in situ hybridization, same expression pattern were also confirmed. siRNA experiments showed significant less photoreceptor protection when we injected siRNA of TrkB-T1. When we examined differential gene expression by DNA microarray of Muller cell line, rMC-1 between BDNF stimulated and non-stimulated, neurotrophic factors that were so far reported were not many expressed. Another mechanism such as scavenging the ions in the surrounding circumstances may be important.

我们报道了一种安全的视网膜下虹膜色素上皮细胞移植术.当这些细胞被基因工程改造成表达神经营养因子时，移植拯救了感光细胞的光毒性。我们使用腺相关病毒（AAV 2）将神经营养因子，如脑源性神经营养因子（BDNF）输送到移植细胞。如果我们使用超过1 × 107个衣壳/ml AAV-BDNF进行转染，则在AAV-BDNF-IPE移植中观察到统计学显著的光感受器保护免受光毒性。然而，拯救作用不具有剂量依赖性。然后，我们检测了BDNF受体TrkB的表达，因为该受体可能控制BDNF的作用。到目前为止，已经报道了TrkB的许多同种型，并且我们检测了TrkB的两种主要同种型; TrkB-FL，其在细胞中具有酪氨酸激酶活性，而TrkB-T1则没有。免疫组化结果显示，这些亚型的表达并不总是完全相同。TrkB-FL主要表达于神经纤维层、神经节细胞层和内核层（INL），TrkB-T1主要表达于INL和RPE。这两种亚型在ONL和感光细胞层中均不表达。S100β免疫组化结果表明，这些异构体在Muller细胞上均有表达。此外，这些异构体在视网膜中的表达不仅在空间上而且在时间上不同，尽管AAV-BDNF-IPE移植增强了表达。当我们进行原位杂交时，同样的表达模式也得到了证实。当我们注射TrkB-T1的siRNA时，siRNA实验显示出显著更少的光感受器保护。当我们用DNA微阵列检测Muller细胞系rMC-1在BDNF刺激和未刺激之间的差异基因表达时，迄今报道的神经营养因子表达不多。另一种机制，如清除周围环境中的离子可能是重要的。