人類多能幹細胞在不同塗佈細胞外間質上分化視網膜色素上皮細胞;Differentiation of human pluripotent stem cells into retinal pigmented epithelial cells on different ECM-coated surface

NCUIR > Engineering College > National Central University Department of Chemical & Materials Engineering > Electronic Thesis & Dissertation > Item 987654321/92066

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Title:	人類多能幹細胞在不同塗佈細胞外間質上分化視網膜色素上皮細胞;Differentiation of human pluripotent stem cells into retinal pigmented epithelial cells on different ECM-coated surface
Authors:	林蕙瑜;Lin, Hui-Yu
Contributors:	化學工程與材料工程學系
Keywords:	細胞外基質;生醫材料;人類多能幹細胞;細胞分化;視網膜色素上皮細胞;extracellular matrix;biomaterials;human pluripotent stem cells;cell differentiation;retinal pigment epithelium
Date:	2023-08-14
Issue Date:	2024-09-19 14:48:50 (UTC+8)
Publisher:	國立中央大學
Abstract:	老年性黃斑部病變是不可逆視力損害的主要原因，與感光細胞及視網膜色素上皮細胞的功能損害有關。人類多能幹細胞衍生的視網膜色素上皮細胞的移植被認為為治療老年性黃斑部病變的方法。人類多能幹細胞可區分為人類胚胎幹細胞和人類誘導多能幹細胞，因為其具有無限的自我更新特性，所以可以提供作為分化視網膜色素上皮細胞的來源。此外，微環境在人類多能幹細胞的分化中起非常重要的作用。因此，我們研究了不同的培養方案及細胞培養生物材料對人類多能幹細胞向視網膜色素上皮細胞分化的影響，本研究使用修正後的NIC84 和 Activin A 製程去提升人類多能幹細胞分化為成熟視網膜色素上皮的效率，並選擇 Matrigel-、Laminin-521-、Laminin-511-、Synthemax II- 和Recombinant vitronectin (rVN)塗層表面作為細胞培養生物材料，以研究視網膜色素上皮細胞的最佳分化條件。我們觀察到人類誘導多能幹細胞 (HPS0077) 衍生的視網膜色素上皮細胞表現多邊形形態，並且經由流式細胞儀測試和免疫染色分析發現其表達成熟視網膜色素上皮細胞的標記（ZO-1 和 RPE65）。因視網膜色素上皮細胞有色素沉澱之功能，細胞呈棕色，而本研究之人類誘導多能幹細胞衍生的視網膜色素上皮細胞也呈棕色細胞。在不同培養基質中，與Synthemax II 和rVN表面相較之下，Matrigel和Laminin-521塗層表面更可以支持人類誘導多能幹細胞有效分化為視網膜色素上皮細胞。;Age-related macular degeneration (AMD), which is the leading cause of irreversible visual impairment, is associated with the progressive dysfunction and death of photoreceptor cells and their supportive retinal pigment epithelial (RPE) cells. Transplantation of human pluripotent stem cell (hPSC)-derived RPE cells is considered as a promising approach to regenerate cell function and cure AMD. Human PSCs, human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), can provide unlimited source of RPE cells because of indefinite self-renewal characteristics. Moreover, microenvironment plays an important role in differentiation of hPSCs. Therefore, I investigated the effect of cell culture biomaterials on the differentiation of hPSCs into RPE cells where Matrigel-, Laminin-521-, Laminin-511-, Synthemax II- and Recombinant vitronectin-coated surface were selected as the cell culture biomaterials in this study. Human PSCs were differentiated into RPE cells on different extracellular matrix (ECM) protein-coated surface using some different protocols (i.e., NIC84 and Activin A protocols) in order to investigate the optimal differentiation conditions into RPE cells. I observed the polygonal morphologies of hiPSCs (HPS0077)-derived RPE cells, which expressed RPE specific markers (ZO-1 and RPE65) by flow cytometry and immunostaining analysis using both protocols. Finally, hiPSCs-derived RPE cells showed brown color (pigmented) cells. Matrigel-, Laminin-521- and Laminin-511-coated surfaces could support the differentiation of hPSCs into RPE cells efficiently, which were compared with Synthemax II- and Recombinant vitronectin-coated surfaces. This is explained that RPE cells are ectodermal lineage of the cells where Laminin preferably supports ectodermal cells via integrin α6β1 rather than integrin αVβ5, which is the main binding site of vitronectin.
Appears in Collections:	[National Central University Department of Chemical & Materials Engineering] Electronic Thesis & Dissertation

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