Pluripotent stem cells (PSCs) are undifferentiated cells with the potential to develop into all cell types in the body. They have the potential to replenish cells in tissues and organs, and have unique properties that make them a powerful tool for regenerative therapy. Embryonic stem cells (ESCs) derived from the inner cell mass of the blastocyst of pre-implantation embryo and epiblast stem cells (EpiSCs) derived from the epiblast layer of post-implantation embryo are the well-known PSCs. These stem cells can differentiate into any of three germ layers of germ cells (endoderm, mesoderm and ectoderm). Additionally, induced pluripotent stem cells (iPSCs) refer to adult somatic cells reprogrammed to return to the pluripotent state by introducing specific factors. This is a breakthrough in stem cell research because ethical concerns such as fertilized embryo destruction can be avoided. PSCs have tremendous potential in treating degenerative cells by generating the cells needed to replace damaged cells, which can also allow to generate specific cell types to study the mechanisms of the disease and create disease models that screen for potential drugs. However, if the proliferative capacity of PSCs is not controlled, there is a risk that tumors will form, as this can lead to uncontrolled growth in their proliferative capacity. In addition, when PSCs are used for therapeutic purposes, there is a risk that the body’s immune system rejects the transplanted cells when the transplanted cells do not originate from the patient’s own tissue. Taken together, PSC is the foundation of stem cell research and regenerative medicine, providing disease treatment and animal development understanding. We would like to explain the classification of PSCs based on their developmental potential, the types of PSCs (ESCs, EpiSCs and iPSCs), their pluripotent status (naïve vs. primed) and alkaline phosphatase (AP) in PSCs and PSCs in domestic animals.

Background: Pluripotent stem cells (PSCs) are capable of differencing into various cell types in the body, providing them valuable for therapy of degenerative diseases. Patientspecific treatments using PSCs, such as mesenchymal stem cells in patient’s own body, may reduce the risk of immune rejection. Inducing the differentiation of PSCs into vascular endothelial cells (ECs) altering culture conditions or using specific growth factors is able to applied to the treatment of vascular diseases. The purpose of this study was to induce the differentiation of porcine epiblast stem cells (pEpiSCs), bone marrow-derived mesenchymal stem cells (pBM-MSCs) and adipose-derived mesenchymal stem cells (pAMSCs) into ECs and then examine the functionality of vascular ECs. Methods: Porcine pEpiSCs, pBM-MSCs and pA-MSCs were induced to differentiate into ECs on matrigel-coated plates in differentiation medium (EBM-2 + 50 ng/mL of VEGF) for 8 days. Cells differentiated from these stem cells were isolated using CD-31 positive (+) magnetic-activated cell sorting (MACS) and then proliferated in M199 medium. Evaluation of ECs differentiated from these stem cells was treated with capillary-like structure formation and three-dimensional spheroid sprouting assay. Results: Porcine pEpiSCs, pBM-MSCs and pA-MSCs showed similar expression of pluripotency-related genes (OCT-3/4. NANOG, SOX2). These stem cells were differentiated into vascular ECs, but showed different morphologies after the differentiation. Cells differentiated from pEpiSCs showed an elongated spindle-like morphology, whereas cells differentiated from pBM-MSCs showed a round pebble-like morphology. In the case of pA-MSCs, these two morphologies were mixed with each other. Additionally, vascular ECs differentiated from these stem cells showed different formation of capillary-like structure formation and three-dimensional spheroid sprouting assay. Conclusions: Cells differentiated from pEpiSCs, pBM-MSCs and pA-MSCs presented the functionality of different vascular ECs, demonstrating the potential of the excellent ECs differentiated from pEpiSCs.

Background: Porcine pluripotent stem cells (pPSCs) would provide enormous potential for agriculture and biomedicine. However, authentic pPSCs have not established yet because standards for pPSCs-specific markers and culture conditions are not clear. Therefore, the present study reports comparative pluripotency characteristics in porcine induced pluripotent stem cells (piPSCs) derived from different viral transduction and reprogramming factors [Lenti-iPSCs (OSKM), Lenti-iPSCs (OSKMNL) and Sev-iPSCs (OSKM)]. Methods: Porcine fibroblasts were induced into Lenti-iPSCs (OSKM) and Lenti-iPSCs (OSKMNL) by using Lentiviral vector and Sev-iPSCs (OSKM) by using Sendaiviral vector. Expressions of endogenous or exogenous pluripotency-associated genes, surface marker and in vitro differentiation in between Lenti-piPSCs (OSKM), Lenti-iPSCs (OSKMNL) and Sev-piPSCs (OSKM) were compared. Results: Colonial morphology of Lenti-iPSCs (OSKMNL) closely resembles the naïve mouse embryonic stem cells colony for culture, whereas Sev-iPSCs (OSKM) colony is similar to the primed hESCs. Also, the activity of AP shows a distinct different in piPSCs (AP-positive (+) Lenti-iPSCs (OSKMNL) and Sev-iPSCs (OSKM), but AP-negative (-) LentiiPSCs (OSKM)). mRNAs expression of several marker genes (OCT-3/4, NANOG and SOX2) for pluripotency was increased in Lenti-iPSCs (OSKMNL) and Sev-iPSCs (OSKM), but Sev-iPSCs (OSKM). Interestingly, SSEA-1 of surface markers was expressed only in Sev-iPSCs (OSKM), whereas SSEA-4, Tra-1-60 and Tra-1-81 were positively expressed in Lenti-iPSCs (OSKMNL). Exogenous reprogramming factors continuously expressed in Lenti-iPSCs (OSKMNL) for passage 20, whereas Sev-iPSCs (OSKM) did not express any exogenous transcription factors. Finally, only Lenti-iPSCs (OSKMNL) express the three germ layers and primordial germ cells markers in aggregated EBs. Conclusions: These results indicate that the viral transduction system of reprograming factors into porcine differentiated cells display different pluripotency characteristics in piPSCs.

Background: Pluripotent stem cells (PSCs) including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) offer the immense therapeutic potential in stem cell-based therapy of degenerative disorders. However, clinical trials of human ESCs cause heavy ethical concerns. With the derivation of iPSCs established by reprogramming from adult somatic cells through the transgenic expression of transcription factors, this problems would be able to overcome. In the present study, we tried to differentiate porcine iPSCs (piPSCs) into endothelial cells (ECs) for stem cell-based therapy of vascular diseases. Methods: piPSCs (OSKMNL) were induced to differentiation into ECs in four differentiation media (APEL-2, APEL-2 + 50 ng/mL of VEGF, EBM-2, EBM-2 + 50 ng/ mL of VEGF) on cultured plates coated with matrigel® (1:40 dilution with DMEM/F-12 medium) for 8 days. Differentiation efficiency of these cells were exanimated using qRT-PCR, Immunocytochemistry, Western blotting and FACS. Results: As results, expressions of pluripotency-associated markers (OCT-3/4, SOX2 and NANOG) were higher observed in all porcine differentiated cells derived from piPSCs (OSKMNL) cultured in four differentiation media than piPSCs as the control, whereas endothelial-associated marker (CD-31) in the differentiated cells was not expressed. Conclusions: It can be seen that piPSCs (OSKMNL) were not suitable to differentiate into ECs in the four differentiation media unlike porcine epiblast stem cells (pEpiSCs). Therefore, it would be required to establish a suitable PSCs for differentiating into ECs for the treatment of cardiovascular diseases.