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The transplantation of muscle progenitor cells (MuPCs) differentiated from human induced pluripotent stem cells (hiPSCs) is a promising approach for treating skeletal muscle diseases such as Duchenne muscular dystrophy (DMD). However, proper purification of the MuPCs before transplantation is essential for clinical application. Here, by using MYF5 hiPSC reporter lines, we identified two markers for myogenic cell purification: CDH13, which purified most of the myogenic cells, and FGFR4, which purified a subset of MuPCs. Cells purified with each of the markers showed high efficiency for regeneration after transplantation and contributed to the restoration of dystrophin expression in DMD-immunodeficient model mice. Moreover, we found that MYF5 regulates CDH13 expression by binding to the promoter regions. These findings suggest that FGFR4 and CDH13 are strong candidates for the purification of hiPSC-derived MuPCs for therapeutical application., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Cardiac progenitor cells (CPCs) are a crucial source of cells in cardiac development and regeneration. However, reported CPCs are heterogeneous, and no gene has been identified to transiently mark undifferentiated CPCs throughout heart development. Here we show that Spalt-like gene 1 (Sall1), a zing-finger transcription factor, is expressed in undifferentiated CPCs giving rise to both left and right ventricles. Sall1 was transiently expressed in precardiac mesoderm contributing to the first heart field (left ventricle precursors) but not in the field itself. Similarly, Sall1 expression was maintained in the second heart field (outflow tract/right ventricle precursors) but not in cardiac cells. In vitro, high levels of Sall1 at mesodermal stages enhanced cardiomyogenesis, whereas its continued expression suppressed cardiac differentiation. Our study demonstrates that Sall1 marks CPCs in an undifferentiated state and regulates cardiac differentiation. These findings provide fundamental insights into CPC maintenance, which can be instrumental for CPC-based regenerative medicine., (Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objective: The pathogenesis of hypertension is dependent on tissue angiotensin (Ang) II, which induces cardiovascular and renal remodeling. The presence of label-retaining cells (LRCs) as renal stem cells has been reported in nephrotubulus. We examined effects of treatment with valsartan on endothelial progenitor cells (EPCs) and renal LRCs in stroke-prone spontaneously hypertensive rats (SHR-SP)., Methods: SHR-SP were salt-loaded and treated with hydralazine or valsartan. Peripheral blood mononuclear cells (MNCs) were cultured to assess EPC colony formation and migration. LRCs were labeled for 1 week with bromodeoxyuridine (BrdU) and were detected after a 2-week chase period. We measured expression of c-kit and Pax-2 mRNAs in renal medulla., Results: Colony formation and migration of EPCs were suppressed in salt-loaded SHR-SP. Treatment with valsartan markedly stimulated these EPC functions. There was no difference in the number of renal LRCs in normotensive Wistar-Kyoto rats and SHR-SP. Treatment with valsartan significantly improved renal tubular degeneration and increased the number of LRCs in renal medulla from salt-loaded SHR-SP. Treatment with valsartan significantly increased expression of c-kit and Pax-2 mRNAs in renal medulla from salt-loaded SHR-SP., Conclusion: These findings suggest that ARBs have cardiovascular and renal protective effects through an antioxidative action that stimulates ECP function and increases the number of the self-repairing renal LRCs.

Non-invasive evaluation of cell reprogramming by advanced image analysis is required to maintain the quality of cells intended for regenerative medicine. Here, we constructed living and unlabelled colony image libraries of various human induced pluripotent stem cell (iPSC) lines for supervised machine learning pattern recognition to accurately distinguish bona fide iPSCs from improperly reprogrammed cells. Furthermore, we found that image features for efficient discrimination reside in cellular components. In fact, extensive analysis of nuclear morphologies revealed dynamic and characteristic signatures, including the linear form of the promyelocytic leukaemia (PML)-defined structure in iPSCs, which was reversed to a regular sphere upon differentiation. Our data revealed that iPSCs have a markedly different overall nuclear architecture that may contribute to highly accurate discrimination based on the cell reprogramming status. [ABSTRACT FROM AUTHOR]

The article discusses the use of hypoxia to enhance induced pluripotent stem cells (iPSCs) generation. It mentions that mouse and human somatic cells can be reprogrammed to the iPSCs through transduction of four transcription factors. It notes that hypoxic conditions tend to improve the generation of iPSCs from mouse and human somatic cells.

Background: Although many studies have shown the feasibility of in vivo cardiac transplantation of human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) in animal experiments, nano-structural confirmation of the engrafted iPSC-CMs including electron microscopy (EM) has not been accomplished, partly because identification of engrafted cells in EM has proven to be difficult. However, with a new genetically encoded probe, the monomeric 28-kDa peroxidase reporter 2 (APEX2), which withstands strong EM fixation can resolve this problem. Moreover, immaturity of iPSC-CMs is a critical problem. Especially, excitation-contraction coupling is one of the fundamental properties of cardiomyocytes, the absence of dyad formed between T-tubule and junctional portion of the sarcoplasmic reticulum is one of the major reasons of arrhythmogenic risks after transplantation. Using the APEX2 system, we evaluated morphological alteration of the engrafted iPSC-CMs in EM in a mouse model of myocardial infarction. Methods: We established human iPSC lines which stably expressed histone H2B-APEX2 (APEX2 iPSCs). After differentiating APEX2 iPSCs into CMs in vitro , purified cells were transplanted into NOG mouse hearts with myocardial infarction by direct injections into the myocardium. We evaluated the ultrastructure of engrafted iPSC-CMs using EM at 3 and 6 months after transplantation. Results: APEX2 did not give significant influences on cardiac differentiation in vitro and stably expressed in iPSC-CMs over 6 months in vivo. APEX2 reaction observed in EM clearly identified engrafted APEX2 iPSC-CMs surrounded by host CMs. The maturation of sarcomeric structure and mitochondria were evident, and T-tubules and dyads started to emerge in engrafted iPSC-CMs at 6 months after transplantation. Conclusions: We demonstrated that APEX2 is a versatile genetic reporter to trace cell fates in living animals over many months. We unequivocally demonstrated that T-tubules and dyads can be formed in iPSC-CMs after a substantially long period of engraftment. This method should be useful to many studies of stem cell-based cell replacement therapy, as it allows direct nano-scale structural characterization of engrafted cells in EM. [ABSTRACT FROM AUTHOR]

Background: Although studies have demonstrated the feasibility of in vivo cardiac transplantation of pluripotent stem cell-derived cardiomyocytes (PSC-CMs) using large animals, it requires large quantities of purified PSC-CMs. Moreover, genetic modification and contamination of non-CMs are inappropriate for clinical application. Using antibodies on the surface of the transplanted cells is one of the useful methods, but can be immunogenic and cause local inflammation or graft failure. We have shown the synthetic mRNAs encoding a fluorescent protein tagged with sequences targeted by microRNAs (miRNAs) expressed in specific cell types can efficiently detect and purify the particular cell populations. Using a miRNA switch and magnetic-activated cell sorting (MACS), we evaluated the efficiency of purification of human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) in a large scale. Methods: We used CD4 as a selection marker for MACS and miR-208a as a specific mRNA of CMs. We synthesized CD4 mRNA and transfected it into differentiated cells from iPSCs to confirm CD4 expressing on the surface of the transfected cells. We also synthesized a miRNA switch encoding CD4 tagged with sequences targeted by miR-208a (CD4-208a switch) and transfected it into differentiated cells to demonstrate transfected non-CMs expressing CD4 and transfected CMs non-expressing CD4. Finally, we transfected the CD4-208a switch and puromycin resistance mRNA simultaneously, and purified iPSC-CMs by eliminating CD4+ cells using MACS and untransfected cells using puromycin. Purified cells were transplanted into NOG mouse hearts with myocardial infarction by direct injections into the myocardium. Results: After transfecting CD4 mRNA into differentiated cells from iPSCs, 78±5% expressed CD4 on the surface. We also confirmed that the CD4-208a switch separated CMs and non-CMs. Using MACS and puromycin selection, we purified iPS-CMs 69±5% to 97±2% assessed by troponin T. Purified cells were also engrafted as CMs in mouse hearts. Conclusions: We demonstrated that CD4-208a switch purifies iPSC-CMs efficiently in a large scale. Synthetic microRNA switches can apply for many studies of stem cell-based cell replacement therapy for clinical application. [ABSTRACT FROM AUTHOR]