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.

ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
    Stem cells culture
    Isolation and culture of swine umbilical veinendothelial cells (SUVECs)
    In vitro differentiation of stem cells into endothelialcells (ECs)
    Magnetic-activated cells sorting (MACS)
    Immunocytochemistry
    Quantitative real-time polymerase chain reaction PCR(qRT-PCR)
    Flow cytometry (FACS)
    Capillary-like structure formation assay
    Three-dimensional spheroid sprouting assay
    Statistical analysis
RESULTS
    Culture of pEpiSCs, pBM-MSCs and pA-MSCs fordifferentiation
    In vitro differentiation of pEpiSCs, pBM-MSCs andpA-MSCs into ECs
    Morphology of cells differentiated from pEpiSCs,pBM-MSCs and pA-MSCs
    Characterization of ECs differentiated from pEpiSCs,pBM-MSCs and pA-MSCs
    Expression of pluripotency markers in ECsdifferentiated from pEpiSCs, pBM-MSCs and pAMSCs
    Capillary-like structure formation of ECs differentiatedfrom pEpiSCs, pBM-MSCs and pA-MSCs
    Three-dimensional spheroids sprouting of ECsdifferentiated from pEpiSCs, pBM-MSCs and pAMSCs
DISCUSSION
CONCLUSION
REFERENCES

• Yeon-Ji Lee(Department of Animal Bioscience, College of Agriculture & Life Sciences, Gyeongsang National University, Jinju 52828, Korea, Division of Applied Life Science Gyeongsang National University, Jinju 52828, Korea)
• In-Won Lee(Department of Animal Bioscience, College of Agriculture & Life Sciences, Gyeongsang National University, Jinju 52828, Korea, Division of Applied Life Science Gyeongsang National University, Jinju 52828, Korea)
• Tae-Suk Kim(Department of Animal Bioscience, College of Agriculture & Life Sciences, Gyeongsang National University, Jinju 52828, Korea)
• Joon-Hee Lee(Department of Animal Bioscience, College of Agriculture & Life Sciences, Gyeongsang National University, Jinju 52828, Korea, Institute of Agriculture & Life Science, College of Agriculture & Life Sciences, Gyeongsang National University, Jinju 52828, Korea) Corresponding author
• Bo-Gyeong Seo(Division of Life Science, College of Natural Sciences, Gyeongsang National University, Jinju 52828, Korea, Division of Applied Life Science Gyeongsang National University, Jinju 52828, Korea)
• Cheol Hwangbo(Division of Life Science, College of Natural Sciences, Gyeongsang National University, Jinju 52828, Korea)
• Sang-Ki Baek(Gyeongsangnamdo Livestock Experiment Station, Sancheong 52263, Korea)