Characterisation of hiPSC lines derived from normal dermal human fibroblasts.

<p>A) Two hiPSC lines were selected for in-depth analyses, NHDF-1 and NHDF-2. These reprogrammed colonies showed hESC-like morphology, for example, tightly packed colonies with high nucleus to cytoplasm ratio (top row). These colonies expressed pluripotency markers hNanog, hOCT4, SSEA-4 and TRA-1-60. Scale bar: 100 µm. B) Genome integrity analysis was performed using both the Illumina Human CytoSNP-12 beadchip with analysis using Karyostudio and using M-FISH. The top panel shows the Illumina analysis of three derived hiPSC clones (iPS-NHDF-1, -2 and -6), using the parental fibroblasts as reference (NHDF). The two analyses indicated a normal chromosomal complement and no gross structural abnormalities for iPS-NHDF-1 and iPS-NHDF-2, but iPS-NHDF-6 showed a translocation and amplification of part of chromosome 2, t(X:2), and was not used in further experiments. GEO: GSE43904. C) iPS-NHDF-1 and iPS-NHDF-2 cell lines are competent at differentiation to three germ layers <i>in vitro</i>. Left column shows embryoid body (EB) formation 4 days after forced aggregation of hiPSCs using Aggrewells. Second column, directed differentiation to neuroectoderm, showing neurons stained for βIII tubulin, using a TujI antibody. Third column, directed differentiation to mesoderm, showing cells stained with antibody against α-sarcomeric actin (ASA). Fourth column, directed differentiation to endoderm, showing nuclei stained with an antibody against the transcription factor, FoxA2. Nuclei are counterstained with DAPI (blue). Scale bar:100 µm. D) PluriTest analysis of Illumina HT12v4 transcriptome array data shows iPS-NHDF-1 and iPS-NHDF-2 to cluster with pluripotent stem cells (red cloud) and not with partly- or differentiated cells (blue clouds). Each circle represents one iPSC line: 1 = iPS-NHDF-1; 2 = iPS-NHDF-2; C = control iPSC line iPS-DF19.9.11TH. GEO: GSE43904.