ABSTRACT

We aimed to generate human induced pluripotent stem cell (iPSC) lines from New Zealand donors. These lines are the first to be generated in New Zealand. Human dermal fibroblasts were collected from two individual donors and reprogrammed with the human OSKM transcription factors using the Sendai virus system. Emerging iPSC colonies were picked, expanded and karyotyped. Clones with normal karyotype were characterised for pluripotency marker expression, p53 mutational status and trilineage differentiation potential. The MANZ-2-2 and MANZ-4-37 iPSC lines showed normal karyotype and expressed pluripotency markers at RNA and protein levels without detectable transgene expression. Both lines differentiated into the three germ layers in vitro and passed the hPSC Scorecard assay for pluripotency and trilineage differentiation. Furthermore, both lines were susceptible to cell apoptosis mediated by nutlin-3a indicative of their wildtype p53 status. This study presents the successful derivation and characterisation of iPSC lines derived from New Zealand donors. These lines will facilitate iPSC-based research in New Zealand and beyond.

摘要

我们的目标是从新西兰捐赠者那里产生人类诱导多能干细胞 (iPSC) 系。这些线是在新西兰产生的第一条线。人类真皮成纤维细胞是从两个个体供体收集的，并使用仙台病毒系统用人类 OSKM 转录因子重新编程。挑选、扩增和核型分析新兴的 iPSC 集落。具有正常核型的克隆的特征在于多能性标记表达、p53 突变状态和三系分化潜能。MANZ-2-2 和 MANZ-4-37 iPSC 系显示出正常的核型，并在 RNA 和蛋白质水平表达多能性标记，而没有可检测的转基因表达。两个品系在体外均分化为三个胚层并通过了多能性和三系分化的 hPSC 记分卡检测。此外，这两个品系都容易受到由 nutlin-3a 介导的细胞凋亡的影响，这表明它们的野生型 p53 状态。本研究展示了来自新西兰捐赠者的 iPSC 系的成功推导和表征。这些线路将促进新西兰及其他地区基于 iPSC 的研究。