2018-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/652371摘要：視神經相關病變與後續可能之視神經萎縮，目前影響全球超過六千萬人，是不可逆失明之主因之一。人體神經細胞再生性低，治療困難，在急性期減少受傷害的視神經細胞主體 – 視網膜神經節細胞 (retinal ganglion cells, RGCs) 凋亡的程度，甚而發展出促使視網膜神經節細胞或軸突再生之療法，乃臨床當務之急。近年誘導性多功能幹細胞 (induced pluripotent stem cells, iPSCs) 的發展，為眼底相關神經退化性疾病，提供了許多契機。首先，幹細胞可分泌神經滋養因子，有助於神經保護；再者，幹細胞分化技術的進步，使得各種視網膜神經元的細胞替代補充變得可能；更重要的，從個人自身細胞重新分化培養之幹細胞，若將來應用於臨床治療，可免除免疫排斥與醫療倫理爭議。精準的細胞分化與足量的細胞移植，是幹細胞療法邁向成功的基本要件。選擇適當材質的生物相容性支架，具有輔助細胞分化生長及增進手術移植效率之功能。幹細胞應用於視網膜神經節細胞的治療，比起其他幾種視網膜神經元，困難度相對較高。但本實驗室近年資料顯示，人類誘導性多功能幹細胞株可順利分化出類視網膜神經節細胞 (iPSC-derived RGC-like cells)，並且在聚苯基麩胺酸(polybenzyl glutamate, PBG) 支架上，能夠展現更好之軸突生長與分化標記表現。因此，在此兩年期研究中，我們將根據上述基礎，第一年首先研發並測試此類細胞在不同電紡纖維模式之聚苯基麩胺酸支架上的表現，觀察其細胞生理表現與細胞-支架接觸介面之特質，尋求最佳細胞培養模式與材料配方，同時進行基因轉殖小鼠之育種，準備動物實驗。第二年之動物實驗，將利用視神經損傷之SPIG1::GFP 基因轉殖小鼠為動物實驗模型，利用其標記輔助光學系統 (accessory opticsystem) 中特定方向選擇性節細胞 (direction sensitive ganglion cells) 之特性，方便追蹤視神經損傷前後以及接受不同幹細胞-支架複合物配方移植治療下，在細胞存活，軸突再生，與幹細胞融合替代的治療成效；加以電生理檢查及視動反射 (Optokinetic reflex, OKR) 之行為測量，完整評估幹細胞移植療法對於視神經傷害，組織學上與功能表現上之治療成效關聯性。希望藉本次之研究，能夠更了解此類衍生幹細胞搭配生物相容性支架之產物，未來提供臨床應用之潛力。<br> Abstract: Optic neuropathy influences more than 60 million people and is one of the leading causes of irreversibleblindness worldwide. Developing an effective approach to rescue injured or dying RGCs and to promotecellular or axon regeneration is an urgent need for these patients. Currently, there is no clinically effectivetreatment for RGC degeneration. The development of induced pluripotent stem cell (iPSC)-based therapyprovides us a window for this field. Stem cells have been proved to increase retinal ganglion cell (RGC)survival after optic nerve injury through the secretion of neurotrophic factors. It is also possible for stem cellsto differentiate into retinal neurons. Furthermore, iPSCs can be derived from the patient’s somatic cells toavoid potential immune rejection.Adequate differentiation and effective transplantation are the basis of a successful stem cell-basedtherapy. It has been known that appropriate scaffold can influence, or induce, the differentiation and growthof neural stem cells. Furthermore, with electrospinning process, the nanofibrous scaffold itself was proved tofacilitate neuronal differentiation in vitro.In our preliminary data, we have proved that iPSCs can differentiate into RGC-like cells. We furtherfound that these iPSC-derived RGC-like cells demonstrated better axonal growth and RGC-likecharacteristics, such as Brn3, when cultured on the polybenzyl glutamate (PBG) scaffold.Based on the preliminary results stated above, this ongoing project will be performed in two years. Inthe first year, we will keep designing the PBG-based scaffold with different electrospun techniques. Then wewill investigate the physiological properties and cell-scaffold interactions among these groups.Simultaneously, we will breed the SPIG1::GFP knockin transgenic mice for the optic nerve crush animalmodel. In the second year, we will perform the subretinal transplantation of iPSC-derived RGC-like cells tomouse eyes. ON DSGCs which are involved in vertical optokinetic reflex (OKR) will be labeled with greenfluorescence in SPIG1::GFP knockin mice, so we can easily access both the anatomical and functionalchange before and after the transplantation of iPSC-derived RGC-PBG complex. We hope that, through ourintensive study, we can know more about the potentials of iPSC-based cell therapy in treating optic neuropathy.