【摘要】：誘導多能干細胞(induced Pluripotent Stem Cell or iPSC)技術的產(chǎn)生和發(fā)展為建立神經(jīng)系統(tǒng)疾病的體外細胞模型,研究個體特異的疾病發(fā)生機理提供了新的方法。然而,既往大多數(shù)關于神經(jīng)系統(tǒng)疾病模型的研究主要局限于神經(jīng)元的病變,通常忽略了膠質(zhì)細胞在疾病表型中的作用。為了模擬唐氏綜合癥發(fā)病過程中細胞和分子水平上的缺陷,我們通過基因轉(zhuǎn)染技術將人體皮膚成纖維細胞重新編程為誘導多能干細胞。其中有兩株細胞系來源于唐氏綜合癥患者,另外兩株細胞系來源于正常人作為對照。然后,經(jīng)過體外培養(yǎng),將誘導多能干細胞定向分化為神經(jīng)干細胞,神經(jīng)元及星形膠質(zhì)細胞,從而建立唐氏綜合癥疾病模型。研究結果表明,相對于來源正常對照iPSC的星形膠質(zhì)細胞,來源于患者iPSC的星形膠質(zhì)細胞表達更多的病理學指標鈣結合蛋白S100B和淀粉樣肽前提蛋白(APP)。另外,患者來源的星形膠質(zhì)細胞內(nèi)活性氧物質(zhì)的水平也較正常對照組明顯增高。在神經(jīng)分化過程中,患者iPSC與對照組iPSC相比,兩者在神經(jīng)元及星形膠質(zhì)細胞的分化效率方面差別并不顯著。而運用不同來源的星形膠質(zhì)細胞條件培養(yǎng)基與的患者iPSC神經(jīng)干細胞共培養(yǎng)后,則發(fā)現(xiàn)神經(jīng)干細胞向神經(jīng)元和星形膠質(zhì)細胞的比例明顯不同。研究結果表明,在患者iPSC來源的星形膠質(zhì)細胞的條件培養(yǎng)基中,神經(jīng)干細胞分化為星形膠質(zhì)細胞的比例明顯增多,分化為神經(jīng)元的比例明顯減少。另外,在共培養(yǎng)體系中,生長在患者iPSC來源星形膠質(zhì)細胞上的神經(jīng)元的神經(jīng)突起較生長在正常iPSC來源星型膠質(zhì)細胞上的神經(jīng)元突起短。隨著人們越來越重視對星形膠質(zhì)細胞功能障礙在神經(jīng)系統(tǒng)疾病中重要作用,本研究認不同類型細胞(神經(jīng)元及星形膠質(zhì)細胞)間的相互聯(lián)系,相互作用對唐氏綜合癥的發(fā)病起重要的作用,因為21號染色體上基因的過度表達而導致的基因劑量效應不僅僅局限在神經(jīng)元上。
[Abstract]:The production and development of induced pluripotent stem cells (induced Pluripotent Stem Cell or iPSC) provides a new method for the establishment of in vitro cell models of nervous system diseases and for the study of individual specific disease mechanisms. However, most previous studies on neuropathic models were mainly confined to neuronal lesions, often ignoring the role of glial cells in disease phenotypes. In order to simulate the cellular and molecular defects in the pathogenesis of Down syndrome, we reprogrammed human skin fibroblasts to induce pluripotent stem cells by gene transfection. Two of the cell lines were from patients with Down syndrome and the other two were from normal controls. After in vitro culture, the induced pluripotent stem cells were differentiated into neural stem cells, neurons and astrocytes, and the disease model of Down's syndrome was established. The results showed that astrocytes from patients with iPSC expressed more calcium-binding protein S100B and amyloid precursor protein (APP). Than astrocytes derived from normal control iPSC. In addition, the level of reactive oxygen species in astrocytes from patients was significantly higher than that of normal controls. There was no significant difference in the differentiation efficiency of neurons and astrocytes between patients with iPSC and control group with iPSC during neural differentiation. However, the proportion of neural stem cells to neurons and astrocytes was significantly different after co-culture of iPSC neural stem cells with different astrocyte conditioned medium. The results showed that the proportion of neural stem cells differentiated into astrocytes increased and the proportion of neurons decreased significantly in the conditioned medium of astrocytes derived from patients with iPSC. In addition, in the co-culture system, the neurites of the neurons growing on the astrocytes derived from iPSC were shorter than those on the astrocytes derived from normal iPSC. As more and more attention has been paid to the important role of astrocytic dysfunction in nervous system diseases, this study recognizes the interrelationship between different types of cells (neurons and astrocytes). Interaction plays an important role in the pathogenesis of Down's syndrome, because the gene dose effect caused by over-expression of gene on chromosome 21 is not limited to neurons.
【學位授予單位】：天津醫(yī)科大學
【學位級別】：博士
【學位授予年份】：2012
【分類號】：R329

【共引文獻】

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本文編號：2365330