探討誘導式多能性幹細胞於光誘導視網膜病變大鼠之效果

Abstract

在大鼠進行光誘導視網膜退化可以用來模仿視網膜退化疾病中視覺細胞細胞凋亡的狀況，近期研究中發現，在心肌梗塞的豬移植間質幹細胞可以經由幹細胞分泌的物質造成保護效果，進而達到心肌功能的回復，而非直接的心肌再生。因此本研究目的為了解Sprague-Dawley (SD)大鼠的視網膜退化模式中誘導式多能性幹細胞(induced-Pluripotent stem cells; iPS cells)是否具有視網膜保護效果。在經過12000Lux的白光照射2小時後，在30分鐘內，玻璃體內注射iPS cells (10000顆細胞/μl；100000顆細胞/μl；1000000顆細胞/μl；分別注射0.01ml)以及PBS 0.1mL (Vehicle組)。受試大鼠的視網膜功能以視網膜電波圖(Electroretinogram)分別在暴露前與光暴露後第24、第48、第96小時，以及第7天進行測量。光照後視網膜電波圖a波和b波的振幅(μV)除以光照前的振幅可以得到殘留百分比(Residual Persentage)，並且在7天後取大鼠眼睛作成組織切片觀察其視網膜型態，以及測量視網膜和視網膜外核層的厚度，統計以ANOVA來計算各時間點的視網膜電波圖a波和b波的殘留百分比，以及視網膜和視網膜外核層在7天後各組之間有無顯著差異(P<0.05)。結果發現誘導式多能性幹細胞治療組的殘留百分比和視網膜以及視網膜外核層厚度皆和未治療組有顯著差異，且發現使用100000顆細胞/μl的濃度可以得到最佳的效果，但在高濃度組會造成炎症反應。總結在本研究中發現，在光傷害的視網膜玻璃體內注射誘導式多能性幹細胞，具有保護的效果。

Light-induced retinal degeneration in a rat model was used to mimic the visual cell apoptosis of retinal degenerations. Recent reserchs found that transplantation of mesenchymal stem cells improves repair and functional recovery of myocardial infarction swine. The possible hyposis is that mesenchymal stem cells could achieve protection through release mediators rather than direct cardiac regeneration. Therefore, we aim to study whether induced-pluripotent stem cells (iPS cells) possess protect effects on degenerative retina in rats. iPS cells (10000 cells/μl, 100000 cells/μl, 1000000 cells/μl; 0.01ml) or vehicle (PBS; 0.01 ml) were injected intravitreally in Sprague-Dawley rats 30 minutes after exposure to 12000 lux of white light for two hours. The retinal function of the test rats were assessed by measuring the electroretinogram (ERG) before light damage and at 24 hours, 48 hours, and 96 hours, and seven days after light damage. The amplitude of a wave and b wave after light damage divided by the amplitude of a wave and b wave before light damage is equal to a wave and b wave residual percentage. The retinal sections were evaluated histologically and thickness of retina and outer nuclear layer (ONL) were measured. The residual percentage of retinal function and retinal thickness were statistically analysed by ANOVA. Results showed that the residual ratio of ERG were significantly higher in the iPS-cells-treated group (n=12) than the control (n=8) and vehicle-treated (n=8) group, respectively. Retinal sections revealed that the ONL of the retina were preserved in the iPS-cells-treated group, and the thickness of retina and ONL in iPS-cells-treated group (n=3) was thicker than the control (n=3) and vehicle-treated group (n=3). Thus, the iPS cells provided protective effect in light induced retinal degenerative rats.