Cytological character of mini pig mesenchymal stromal cells from various tissues and the attempt of cell sheet formation

INTRODUCTION: Large animal experiments are important for translational research in regenerative medicine. Recently, mini pigs have been used in large animal studies and surgical training. The use of multipotent mesenchymal stromal cell (MSC) sheets for the treatment of many diseases is increasing. The purpose of the present study was to establish optimal methods for generating mini pig MSC sheets from various tissues and to compare the properties of MSCs in these sheets. METHODS: MSCs were isolated from the bone marrow, adipose, periodontal ligament, gingiva, or periosteum of mini pigs. The proliferation, markers, and mRNA expression of these MSCs were examined. Colony-forming and differentiation assays were performed. MSCs were seeded onto temperature-responsive culture dishes to develop MSC sheets. RESULTS: MSCs derived from bone marrow (BMSCs), adipose (ASCs), periodontal ligament (PDLCs), gingiva (GMSCs), and periosteum (PSCs) were positive for MSC-related markers. BMSCs and PSCs showed increased proliferation compared with other MSCs. The osteogenic potential of PDLCs and the adipogenic potential of PSCs were the highest among these MSCs. The expression levels of COL1A1 and COL3A1 in BMSCs and PSCs were significantly higher than those in other MSCs. The expression levels of FGF2, VEGFA, ICAM-1, and TIE-1 in GMSCs were significantly higher than those in other MSCs. PSCs showed the highest levels of TGF-β1 and ANG-1 expression among all MSC types. We succeeded in developing MSC sheets from BMSCs, ASCs, and PSCs. CONCLUSIONS: We developed methods to generate MSC sheets from various tissues of mini pigs, and these methods are useful to pursue regenerative translational research using mini pigs.