Background: In healthy dentin conditions, odontoblasts have an important role such as protection from invasion of pathogens. In mammalian teeth, progenitors such as mesenchymal stem cells (MSCs) can migrate and differentiate into odontoblast-like cells, leading to the formation of reparative dentin. For differentiation using stem cells, it is crucial to provide conditions similar to the complex and intricate in vivo environment. The purpose of this study was to evaluate the potential of differentiation into odonto/ osteoblasts, and compare co-culture with/without epithelial cells. Methods: MSCs and epithelial cells were successfully isolated from dental tissues. We investigated the influences of epithelial cells on the differentiation process of dental pulp stem cells into odonto/osteoblasts using co-culture systems. The differentiation potential with/without epithelial cells was analyzed for the expression of specific markers and calcium contents. Results: Differentiated odonto/osteoblast derived from dental pulp tissue-derived mesenchymal stem cells with/without epithelial cells were evaluated by qRT-PCR, immunostaining, calcium content, and ALP staining. The expression of odonto/ osteoblast-specific markers, calcium content, and ALP staining intensity were significantly increased in differentiated cells. Moreover, the odonto/osteogenic differentiation capacity with epithelial cells co-culture was significantly higher than without epithelial cells co-culture. Conclusions: These results suggest that odonto/osteogenic differentiation co-cultured with epithelial cells has a more efficient application.

Current studies have revealed the capacity of mesenchymal stem cells (MSCs) in term of immunomodulatory properties, and this distinct potential is downgraded according to the disease duration of patients-derived MSCs. In order to enhance the immunomodulatory and anti-tumorigenic properties of the rheumatoid arthritis (RA) joints-derived MSCs, we aggregate synovial fluid-derived MSCs from RA joints (RA-hMSCs) into 3D-spheroids by the use of hanging drop culture method. Cells were isolated from synovial fluids of RA joints with longstanding active status over 13 years. For aggregation of RA-hMSCs into 3D-spheroids, cells were plated in hanging drops in 30 μL of advanced DMEM (ADMEM) containing 25,000-30,000 cells/ drop and cultured for 48 h. To analyze the comparative immunomodulatory effects of 3D-spheroid and 2D monolayer cultured RA-hMSCs and then cells were cultured in ADMEM supplemented with 20% of synovial fluids of RA patients for 48 h and were evaluated by qRT-PCR for their expression of mRNA levels of inflammatory and antiinflammatory markers. Cellular aggregation of RA-hMSCs was observed and cells were aggregate into a single sphere. Following treatment of RA patient’s synovial fluids into the RA-hMSCs, spheroids formed RA-hMSCs showed significantly (p < 0.05) higher expression of TNFα stimulated gene/protein 6 (TSG-6) than the monolayer cultured RAhMSCs. Therefore, the 3D-spheroid culture methods of RA-hMSCs were more effective than 2D monolayer cultures in suppressing inflammatory response treated with 20% of RA-synovial fluids by expression of TNFα (TSG-6) according to the immune response and enhanced secretion of inflammatory factors.

Abnormal epigenetic reprogramming of donor nuclei is supposed to be one of the factors that causes low development efficiency of mammalian somatic cell nuclear transfer (SCNT). Trichostatin A (TSA) is an inhibitor of histone acetylase, and so development of SCNT embryos could be increased by treatment with TSA. In the present study, we examined the effect of TSA on in vitro development of porcine embryos derived from NT (nuclear transfer) by investigating the status of histone acetylation in TSA-treated and control NT embryos and the expression of developmental related genes. In this study, we found that incubating　NT embryos with 40nM TSA for 24h after activation could improved the blastocyst formation rate from 13.7% to 32.5%. Thechange in histone acetylation level as a reslut of TSA treatment were validated using immunofluorescence and confocal microscopy. Immunofluorescence results indicated that the level of aetylation at histone 3 lysine 18 (AcH3K18) was increased at early embryo development stage after TSA treatment. furthermore, we compared the expression patterns of several genes (developmental related genes; Oct4, Sox2, Nanog, Cdx2, the imprinting genes; igf2r). TSA treatment improved the expression of development related genes such as Oct4, Cdx2, Nanog as well as the imprinted genes like igf2r. In conclusion, our results demonstrated that TSA treatment improves the in vitro development of porcine NT embryos, increased the global histone acetylation (AcH3K18) and enhances the expression of some developmentally important genes (Oct4, Cdx2, Nanog) at blastocyst stages.

Due to their anatomical, physiological and genetic similarities, pig is attractive animal model in biomedical research. In the recent stem cell research era, porcine derived stem cells also gain attention due to its use for the preclinical application of human. Mesenchymal stem cells (MSCs) have been studied by many researchers over decade, and their prospect for clinical application is recognized. Although porcine derived MSCs (pMSCs) have confirmed to be differentiated into various types of cells, such as osteocyte, chondrocyte, neuronal cell, cardiomyocyte and pancreatic β cell, few report has been studied regarding hepatocyte differentiation in vitro. The present study was therefore aimed for bone marrow MSCs derived from pig femur to differentiate into hepatocyte. The cells were confirmed as MSCs by characterizing their morphology, lineage differentiation capacity and surface phenotype. They showed spindle like morphology and adipocytic, osteoblastic, and chondrocytic differentiation potentials and displayed positive expression of mesenchymal markers CD29, CD44 and CD90 while lacked the expression of hematopoietic marker CD45. Under appropriate differentiation conditions, MSCs displayed hepatocyte-like morphology depending on duration of differentiation. The differentiated MSCs into hepatocyte expressed hepatocyte-specific genes including hepatocyte nuclear factor 4 (HNF4), albumin (ALB), alpha fetoprotein (AFP), alpha-1-anti trypsin (A1AT). They also showed hepatocyte-like function, glycogen storage which is identified by PAS staining. Taken together, it concluded that the bone marrow MSCs have the potential to differentiate into hepatocyte. Further studies are needed on additional hepatocytic functional assays, such as low density lipoprotein (LDL) uptake and urea synthesis of differentiated MSC.