利用患者特异iPS细胞模型研究低碱性磷酸脂酶症成骨发育不良的表观遗传学调控机制

Induced pluripotent stem cell (iPS) is a type of pluripotent stem cell artificially derived from adult somatic cell by by inducing a forced expression of specific genes, which are similar to natural pluripotent stem cells, such as embryonic stem cells. Modelling pathogenesis of the hereditary disease using patient-specific iPS has recently become a research focus. Hypophosphatasia (HPP) causes the hereditary developmental disorders of bone and teeth due to mutations in the liver/bone/kidney alkaline phosphatase gene (ALPL), and ALPL gene is always acknowleged as the biomarker of stem cells without any functions. Unfortunately, there are no related research reports that ALPL mutation could cause abnormal biological behaviors of stem cells resulting in osteogenesis imperfecta. Our previous research found that the decreasing osteogenic potentials of HPP messenchymal stem cells were related to the changes of genes patterning including epigenetic factors, then HPP-iPS cells also presented abnormal biological characteristics. However , its molecular and pathogenic mechanisms remain unraveled. In our project, we will plan to construct the modelling of the HPP patient-specific iPS, and characterize their biological behaviors change, and then examine the effects of ALPL mutations on cellular metabolisms, finally evaluate the epigenetic regulatory patterns in the osteogenic differentiation of HPP-iPS. We wish to explore the roles of ALPL gene in the differentiation of stem cells, and disclose the pathogenic mechanism of HPP phenotypes-osteogenesis imperfecta.

诱导多能干细胞iPS是由体细胞诱导而成的一类具有胚胎干细胞特性的多能干细胞, 建立遗传性疾病特异性iPS细胞模型进而探索致病基因功能和其致病机制是近期国际研究热点。低碱性磷酸酯酶症HPP是由于碱性磷酸酯酶基因ALPL突变导致的骨及牙齿发育缺陷的遗传性疾病， ALPL基因被公认是发育期干细胞生物学标志，其突变是否能影响发育期干细胞行为而导致成骨发育不良未见研究报道。我们前期研究发现HPP患者间充质干细胞成骨分化能力减弱与表观遗传学因子等表达异常相关，建立的HPP疾病iPS细胞也展示异常生物学行为，但其分子致病机制不甚清楚。本项目拟通过建立HPP疾病特异iPS细胞模型，检测疾病iPS细胞生物学行为，分析ALPL功能缺陷对iPS细胞相关生物分子代谢的影响，最后评估疾病iPS细胞成骨分化的表观遗传学调控模式，以期揭示ALPL基因在干细胞分化中的生物学作用，阐明HPP疾病成骨发育不良的分子致病机制。

课题组围绕本课题的主要研究内容及预期目标，主要开展了以下几方面的研究工作：.①收集的3例HPP患儿出现成骨发育不全及骨质疏松等表型；首次观察到HPP患儿机体出现细胞免疫异常现象；证实了HPP患者ALPL基因发生错义突变c.422C>A, p.T141N 和c.551G>A, p.R263Q。②HPP患者间充质干细胞的生物学活性明显降低，牙周膜干细胞的生物学行为缺陷是导致HPP患者牙齿过早脱落的致病因素，骨髓间充质干细胞的生物学性能异常是造成HPP患者成骨不全的主要原因，说明HPP患者BMMSC的生物活性降低以及免疫调控功能异常与HPP疾病的发生存在密切联系。③ALPL 基因突变明显地影响重编程细胞的生长速度和进程，结果提示我们ALPL 基因可能不仅仅是ES 和iPS标志物，而且在诱导体细胞重编程过程中发挥重要作用。④ADRB2交感神经信号通路具有抑制BMMSCs成骨分化的作用，调控ADRB2信号可以改善ALP缺陷所导致的成骨不全。⑤发表SCI 文章4 篇，中文源期刊文章2篇；⑥培养博士2名，硕士3名；⑦参加国际学术报告2人次，国内特邀学术报告2人次。

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