Enhanced Generation of Human Induced Pluripotent Stem Cells from Peripheral Blood and Using Their Mesoderm Differentiation Ability to Regenerate Infarcted Myocardium

Figure 1

Generation of induced pluripotent stem cells (iPSCs) from human circulating multipotent adult stem (CiMS) cells. (a) Time schedule of CiMS-iPSC generation. (b) CiMS morphology before reprogramming factor transduction and human embryonic stem cell- (hESC-) like CiMS-iPSC colonies on feeder cells after transduction. (c) CiMS-iPSCs were positive for ALP staining and expressed pluripotency markers, such as OCT3/4, NANOG, and TRA-1-81. (d) Transduction efficiency was estimated using GFP retroviral transduction. Retroviral transduction efficiency was higher in CiMS than in human dermal fibroblasts (HDFs) and even 293T cells () (, , and : statistically significant, ns: statistically not significant). (e) Comparison of ALP-positive colony formation efficiency of CiMS cells and HDF () (f) Expression levels of CiMS-iPSC pluripotency genes were similar to those in hESCs, confirmed by reverse transcription PCR. (g) The global gene expression profiles compared CiMS, CiMS-iPSCs, and hESCs (H9) with the oligonucleotide microarray. (h) Bisulfite genomic sequencing of the promoter regions of OCT3/4 and NANOG. Open and closed circles indicate unmethylated and methylated CpGs. (i) Karyotyping analyses of the CiMS-iPSCs. (j) CiMS-iPSCs were spontaneously differentiated into three germ layers, and immunofluorescence staining showed positivity for each marker. The markers used were beta III tubulin and nestin for ectoderm, alpha-sarcomeric actin and smooth muscle actin for mesoderm, and AFP for endoderm. (k) Teratoma derived from CiMS-iPSCs. H&E staining of teratoma derived from CiMS-iPSCs. CiMS-iPSCs were transplanted subcutaneously on the back of a SCID mouse.

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