發(fā)布時(shí)間：2018-01-18 05:26

  本文關(guān)鍵詞：克服多能干細(xì)胞移植免疫排斥反應(yīng)的實(shí)驗(yàn)研究　出處：《第四軍醫(yī)大學(xué)》2009年博士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 多能干細(xì)胞 胚胎干細(xì)胞 人白細(xì)胞抗原G 免疫排斥 重編程

【摘要】： 多能干細(xì)胞(Pluripotent Stem Cells)主要包括胚胎干細(xì)胞和誘導(dǎo)多潛能干細(xì)胞(induced pluripotent stem cell, iPS),均具有自我復(fù)制及多潛能分化的能力。理論上,它們能夠?yàn)榧?xì)胞移植治療提供無限的供體細(xì)胞來源,因此是理想的“種子細(xì)胞”。然而,免疫排斥反應(yīng)是細(xì)胞移植治療面臨的共同問題,當(dāng)然是多能干細(xì)胞移植治療首先需要考慮的問題。本課題根據(jù)兩種多能干細(xì)胞的不同特點(diǎn),為解決免疫排斥進(jìn)行了以下兩個(gè)方面嘗試: 一、通過在人胚胎干細(xì)胞上穩(wěn)定表達(dá)人白細(xì)胞抗原G(Human Leucocyte Antigen G,HLA-G),獲得具有免疫耐受能力的通用的種子細(xì)胞。HLA-G是母胎免疫及器官移植免疫耐受的重要分子。本研究建立了穩(wěn)定表達(dá)HLA-G1的hES細(xì)胞系。該細(xì)胞系核型正常,仍具有自我復(fù)制和多潛能性(表達(dá)干細(xì)胞特異性的轉(zhuǎn)錄因子Oct4和Sox2,接種SCID小鼠后能夠形成畸胎瘤,其中有三胚層的分化細(xì)胞)。由HLA-G1+hESCs分化為神經(jīng)前體細(xì)胞(Neural Progenitor cells,NPCs)的過程中,HLA-G1持續(xù)表達(dá),通過細(xì)胞毒實(shí)驗(yàn)及混合淋巴細(xì)胞反應(yīng)證實(shí)HLA-G1對(duì)由hESCs分化獲得的NPCs具有明顯的免疫保護(hù)作用。 二、提高建立iPS細(xì)胞(induced pluripotent stem cells,iPS細(xì)胞)的效率,同時(shí)探討iPS細(xì)胞形成的機(jī)制。iPS細(xì)胞是個(gè)體特異性多能干細(xì)胞,可完全避免個(gè)體的免疫排斥反應(yīng)。本研究采用人胚胎神經(jīng)干細(xì)胞作為重編程對(duì)象,與傳統(tǒng)的四分子轉(zhuǎn)染體系不同,我們用Oct4進(jìn)行單分子轉(zhuǎn)染。轉(zhuǎn)染1周后出現(xiàn)iPS細(xì)胞克隆,2周統(tǒng)計(jì)iPS細(xì)胞的形成效率為0.015±0.001,比成纖維細(xì)胞效率高10倍。對(duì)獲得的iPS細(xì)胞與hES細(xì)胞進(jìn)行比較鑒定后發(fā)現(xiàn),二者在表達(dá)譜、重要標(biāo)志物以及DNA甲基化方式和多潛能分化能力等方面都非常相似。由此,本研究?jī)H采用了一個(gè)分子就獲得了人iPS細(xì)胞,同時(shí)神經(jīng)干細(xì)胞重編程獲得iPS細(xì)胞的效率遠(yuǎn)高于成纖維細(xì)胞。
[Abstract]:The pluripotent stem cells (Pluripotent Stem cells) mainly include embryonic stem cells and induced multipotential stem cells. Induced pluripotent stem cell. IPSs are capable of self-replication and multipotential differentiation. In theory, they provide an unlimited source of donor cells for cell transplantation therapy and are therefore ideal "seed cells". Immune rejection is a common problem in cell transplantation therapy, of course, it is the first problem to be considered in the treatment of pluripotent stem cell transplantation. According to the different characteristics of two kinds of pluripotent stem cells. In order to solve the immune rejection, the following two aspects have been attempted: Firstly, human Leucocyte Antigen HLA-Gs were stably expressed on human embryonic stem cells. HLA-G is an important molecule in maternal and fetal immunity and organ transplantation tolerance. A stable hES cell line expressing HLA-G1 was established in this study. The karyotype was normal. Still self-replicating and multipotent (expressing stem cell-specific transcription factors Oct4 and Sox2), teratoma can be formed after inoculation with SCID mice. In the process of differentiation from HLA-G1 hESCs to neural Progenitor cells (NPCs). The expression of HLA-G1 was sustained. The results of cytotoxicity assay and mixed lymphocyte reaction confirmed that HLA-G1 had a significant protective effect on NPCs derived from hESCs differentiation. Second, to improve the efficiency of establishing iPS cells induced pluripotent stem cells. The mechanism of iPS cell formation. IPS cells are individual specific pluripotent stem cells, which can completely avoid individual immune rejection. In this study, human embryonic neural stem cells were used as reprogramming objects. Different from the traditional four-molecule transfection system, we used Oct4 for monolayer transfection. After one week of transfection, iPS cell clones appeared. At 2 weeks, the efficiency of iPS cells was 0.015 鹵0.001, which was 10 times higher than that of fibroblasts. The iPS cells were compared with hES cells. They are very similar in expression profile, important markers, DNA methylation mode and multipotential differentiation ability. Therefore, human iPS cells were obtained by using only one molecule in this study. At the same time, neural stem cells were reprogrammed to obtain iPS cells more efficiently than fibroblasts.
【學(xué)位授予單位】：第四軍醫(yī)大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2009
【分類號(hào)】：R329

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 Martin Oudega;Bas Blits;;GENETIC ENGINEERING NEURAL STEM CELL MODIFIED BY LENTIVIRUS FOR REPAIR OF SPINAL CORD INJURY IN RATS[J];Chinese Medical Sciences Journal;2006年02期

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本文編號(hào)：1439650