Generation of induced pluripotent stem cells from renal tubular cells of a patient with Alport syndrome
Generation of induced pluripotent stem cells from renal tubular cells of a patient with Alport syndrome
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DOI：
10.2147/ijnrd.s85733
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发表时间：
2015-01-01
影响因子：
2
通讯作者：
Dai, Yong
中科院分区：
文献类型：
--
作者：
Chen, Wenbiao;Huang, Jianrong;Dai, Yong
Alport综合征（AS）是一种导致肾衰竭的遗传性疾病，由COL 4A 3、COL 4A 4和COL 4A 5基因突变引起，导致成熟肾小球基底膜中缺乏胶原蛋白α 3 α 4 α 5（IV）网络。大约80%的AS是X连锁的，因为编码IV型胶原α 5链的基因COL 4A 5突变。为了在遗传水平上研究AS的发病机制，我们从AS患者的肾小管细胞中产生了诱导多能干细胞（iPSC）。iPSC的成功产生为掌握COL 4A 5基因的修复和评估iPSC向支持细胞的分化以及每个阶段伴随的表观遗传变化奠定了基础。从AS患者中产生iPSC不仅证实了iPSC可以从肾小管细胞中产生，而且为AS患者提供了一种新型的基因治疗方法。在这项研究中，我们通过异位表达四种转录因子（Oct 4，Sox 2，c-myc和Klf 4）从肾小管细胞中产生iPSCs。根据人胚胎干细胞（human embryonic stem cell，hESC）的特性，通过集落形态学、免疫组化、qRT-PCR、流式细胞术、基因表达谱和核型分析等方法比较分析hESC的标志物，证实iPSC的形成。我们的研究结果表明，iPSCs的形态，增殖，hESC特异性表面标志物的表达，并分化成三个胚层的细胞类型方面的hESC相似。从AS患者的肾小管细胞中有效产生iPSC将为研究AS的潜在机制和开发新的AS治疗方法提供新的模型。
Alport syndrome (AS) is a hereditary disease that leads to kidney failure and is caused by mutations in the COL4A3, COL4A4, and COL4A5 genes that lead to the absence of collagen alpha 3 alpha 4 alpha 5 (IV) networks in the mature kidney glomerular basement membrane. Approximately 80% of AS is X-linked because of mutations in COL4A5, the gene encoding the alpha 5 chain of type IV collagen. To investigate the pathogenesis of AS at the genetic level, we generated induced pluripotent stem cells (iPSCs) from renal tubular cells of a patient with AS. The successful iPSC generation laid the foundation to master the repair of the COL4A5 gene and to evaluate the differentiation of iPSC into Sertoli cells and the accompanying epigenetic changes at each stage. The generation of iPSCs from AS patients not only confirms that iPSCs could be generated from renal tubular cells, but also provides a novel type of genetic therapy for AS patients. In this study, we generated iPSCs from renal tubular cells via ectopic expression of four transcription factors (Oct4, Sox2, c-myc, and Klf4). According to the human embryonic stem cell (hESC) charter, iPSC formation was confirmed by comparatively analyzing hESC markers via colony morphology, immunohistochemistry, qRT-PCR, flow cytometry, gene expression profiling of the three germ layers, and karyotyping. Our results demonstrated that iPSCs were similar to hESCs with regard to morphology, proliferation, hESC-specific surface marker expression, and differentiation into the cell types of the three germ layers. The efficient generation of iPSCs from the renal tubular cells of an AS patient would provide a novel model to investigate the mechanisms underlying AS and to develop new treatments for AS.