【摘要】： 間充質(zhì)干細(xì)胞(mesenchymal stem cell,MSC)是具有自我復(fù)制和多向分化潛能的一類多能干細(xì)胞。目前成人骨髓是MSC的主要來(lái)源。骨髓間充質(zhì)干細(xì)胞的獲取須行骨髓穿刺術(shù),給供者造成一定痛苦,且骨髓間充質(zhì)干細(xì)胞的增殖及分化潛能隨供者年齡的增大而下降,因此,尋找新的MSC來(lái)源成為國(guó)內(nèi)外干細(xì)胞研究領(lǐng)域的熱點(diǎn)。本研究從足月胎兒臍帶中分離出MSC,對(duì)其生物學(xué)特征、免疫學(xué)特征及抗急性移植物抗宿主病(acute graft versus host disease, aGVHD)的作用進(jìn)行系統(tǒng)研究,旨在尋找MSC的新來(lái)源。 本實(shí)驗(yàn)第一部分建立從臍帶中分離MSC的方法,并與成人骨髓源MSC對(duì)比,研究臍帶源MSC的生物學(xué)特征。結(jié)果顯示,臍帶源MSC的分離成功率達(dá)100%;與骨髓源MSC相比,臍帶源MSC成纖維細(xì)胞集落形成單位(CFU-F)比例、增殖能力和神經(jīng)細(xì)胞誘導(dǎo)分化能力均高于骨髓源MSC,HLA-I和CD106分子表達(dá)低于骨髓源MSC(P0.05)。臍帶源MSC形態(tài)、大多數(shù)分子表型(CD13、CD29、CD44、CD105、CD73、CD166陽(yáng)性,CD14、CD34、CD38和CD45陰性,CD31陰性)、細(xì)胞周期狀態(tài)(超過(guò)80%的細(xì)胞處于G_0/G_1期)、脂肪和骨誘導(dǎo)分化能力、細(xì)胞因子分泌(表達(dá)SCF、TPO、FL、IL-6、M-CSF、LIF、SDF-1和VEGF,不表達(dá)IL-3)和長(zhǎng)期支持造血的能力與骨髓源MSC相似。 本實(shí)驗(yàn)第二部分研究臍帶源MSC的免疫學(xué)特征。免疫表型分析顯示,臍帶源MSC表達(dá)HLA-I,不表達(dá)HLA-DR,不表達(dá)協(xié)同刺激分子CD80、CD86和CD40。3H-胸腺嘧啶核苷(3H-Tdr)摻入法檢測(cè)淋巴細(xì)胞增殖結(jié)果顯示,臍帶源MSC不刺激小鼠淋巴細(xì)胞增殖,且可顯著抑制小鼠混合淋巴細(xì)胞反應(yīng)。 本實(shí)驗(yàn)第三部分通過(guò)GVHD小鼠模型初步研究了臍帶源MSC對(duì)aGVHD的影響。對(duì)GVHD鼠生存時(shí)間、GVHD靶器官(肝臟、小腸和皮膚)病理改變、淋巴細(xì)胞亞群和細(xì)胞因子水平的研究結(jié)果證明,臍帶源MSC靜脈輸入可顯著減輕小鼠半相合移植模型aGVHD的嚴(yán)重程度。其機(jī)制可能與增高Th2細(xì)胞因子
[Abstract]:Mesenchymal stem cells (mesenchymal stem) are multipotent stem cells with self-replication and multi-differentiation potential. Adult bone marrow is currently the main source of MSC. Bone marrow mesenchymal stem cells must be obtained by bone marrow puncture, causing some pain to the donor, and the proliferation and differentiation potential of bone marrow mesenchymal stem cells decrease with the age of the donor. Finding new sources of MSC has become a hot spot in stem cell research at home and abroad. In this study, MSCs were isolated from the umbilical cord of full-term fetus. The biological, immunological and antigraft versus host disease (acute graft versus host disease, aGVHD) effects of MSCs were systematically studied in order to find a new source of MSC. In the first part of the experiment, we established a method to isolate MSC from umbilical cord and compared with adult bone marrow derived MSC to study the biological characteristics of umbilical cord derived MSC. The results showed that the isolation rate of umbilical cord derived MSC was 100%, compared with bone marrow derived MSC, the proportion of umbilical cord derived MSC fibroblast colony forming unit (CFU-F) was higher than that of bone marrow derived MSC. The expression of HLA-I and CD106 in MSCs was lower than that in MSC derived from bone marrow (P0.05). Umbilical cord derived MSC morphology, most molecular phenotypes (CD13C CD29, CD44, CD105, CD73, CD166 positive, CD14, CD34, CD38 and CD45 negative, CD31 negative), cell cycle state (more than 80% of cells in G_0/G_1 phase), adipose and osteogenic differentiation ability, The ability of cytokine secretion (expression of IL-6 TPO-FLIL-6 and IL-3) and long-term hematopoietic support was similar to that of bone marrow-derived MSC. The second part of this experiment was to study the immunological characteristics of umbilical cord-derived MSC. Immunophenotypic analysis showed that MSC from umbilical cord source expressed HLA-I, did not express HLA-DRR, and did not express co-stimulatory molecules CD80 CD86 and CD40.3H-thymidine (3H-Tdr) incorporation assay. The results showed that cord derived MSC did not stimulate lymphocyte proliferation in mice. In addition, the mixed lymphocyte reaction in mice was significantly inhibited. In the third part of the experiment, the effect of umbilical cord MSC on aGVHD was studied by GVHD mouse model. The pathological changes of GVHD target organs (liver, small intestine and skin), lymphocyte subsets and cytokine levels in GVHD mice were studied. The results showed that umbilical cord derived MSC intravenous infusion could significantly reduce the severity of aGVHD in mice model of haploidentical transplantation. Its mechanism may be related to the increase of Th2 cytokines.
【學(xué)位授予單位】：福建醫(yī)科大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2006
【分類號(hào)】：R329.2

【引證文獻(xiàn)】

相關(guān)期刊論文 前2條

1 鄭志娟;莊文欣;付文玉;;人臍帶間充質(zhì)干細(xì)胞的研究進(jìn)展[J];解剖科學(xué)進(jìn)展;2008年01期

2 康鵬云;周鑫磊;張宇晶;平超強(qiáng);李亮;洪珞珈;;臍帶間充質(zhì)干細(xì)胞與再生障礙性貧血[J];現(xiàn)代生物醫(yī)學(xué)進(jìn)展;2012年10期

相關(guān)博士學(xué)位論文 前1條

1 胡勝利;人臍帶間充質(zhì)干細(xì)胞治療脊髓損傷的實(shí)驗(yàn)研究[D];第三軍醫(yī)大學(xué);2010年

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本文編號(hào)：2185761