Yin S, Li W, Yang G, Cheng Y, Yi Q, Fan S, Ma Q, Zeng F
Department of Histoembryology, Genetics and Development, Shanghai Jiao Tong University School of Medicine, Shanghai, People's Republic of China.
Shanghai Institute of Medical Genetics, Shanghai Children's Hospital, Shanghai Jiao Tong University, Shanghai, People's Republic of China.
Balkan J Med Genet. 2020 Aug 26;23(1):5-13. doi: 10.2478/bjmg-2020-0003. eCollection 2020 Jun.
Induced pluripotent stem cells (iPSCs), generated from somatic cells, not only possess similar characteristics with embryonic stem cells (ESCs), but also present more advantages than ESCs in medical applications. The classical induction method that utilizes the integration of exogenous genes into chromosomes may raise the potential risk of the safety of iPSCs. To investigate the potential correlation between the integration sites of exogenous transcription factors (TFs) and iPSCs' pluripotency and safety, the integration of exogenous genes in three iPSC lines, which met the golden standard of murine developmental assay (tetraploid complementation), were analyzed. Twenty-two integration sites of exogenous TFs were identified by nested inverse polymerase chain reaction (iPCR) and 39 flanking genes' functions were analyzed by gene ontology (GO). In the 22 integrated sites, 17 (77.3%) were located in the intergenic regions and the remainder were located in introns far from the transcription start sites. Microarray analysis of the flanking genes in these cells showed that there was no distinct difference in expression levels between the iPSCs, ESCs and mouse embryonic fibroblast (MEF), suggesting that the integration of exogenous TFs has no significant influence on the expression of flanking genes. Gene ontology analysis showed that although most of the flanking genes were housekeeping genes, which were necessary for basic life activity, none of these 39 flanking genes have correlation with tumorigenesis or embryogenesis, suggesting that the integration sites hold low risk of tumorigenesis.
由体细胞产生的诱导多能干细胞（iPSC）不仅具有与胚胎干细胞（ESC）相似的特征，而且在医学应用中比ESC具有更多优势。利用外源基因整合到染色体中的经典诱导方法可能会增加iPSC安全性的潜在风险。为了研究外源转录因子（TF）的整合位点与iPSC的多能性和安全性之间的潜在相关性，分析了符合小鼠发育试验（四倍体互补）金标准的三个iPSC系中外源基因的整合情况。通过巢式反向聚合酶链反应（iPCR）鉴定了22个外源TF的整合位点，并通过基因本体论（GO）分析了39个侧翼基因的功能。在22个整合位点中，17个（77.3%）位于基因间区域，其余位于远离转录起始位点的内含子中。对这些细胞中侧翼基因的微阵列分析表明，iPSC、ESC和小鼠胚胎成纤维细胞（MEF）之间的表达水平没有明显差异，这表明外源TF的整合对侧翼基因的表达没有显著影响。基因本体论分析表明，虽然大多数侧翼基因是维持基本生命活动所必需的管家基因，但这39个侧翼基因中没有一个与肿瘤发生或胚胎发生相关，这表明整合位点的肿瘤发生风险较低。
