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Title:   Author  item
                                            Lee, Sungitem
                                            CHA, SANG HO - Animal, Plant And Fisheries Quarantine And Inspection Agency (QIA)item
                                            Lillehoj, Hyunitem
                                            SONG, JAE-YOUNG - Animal, Plant And Fisheries Quarantine And Inspection Agency (QIA)item
                                            LEE, KYUNG-WOO - Animal, Plant And Fisheries Quarantine And Inspection Agency (QIA)
  Submitted to: Journal of Applied Animal Research
                                    Publication Type: Peer Reviewed Journal
                                    Publication Acceptance Date: 12/18/2013
                                    Publication  Date: 2/10/2014
                                    Citation: Lee, S.H., Cha, S., Lillehoj, H.S., Song, J., Lee, K. 2014. Enhanced adipogenic differentiation of bovine bone marrow-derived mesenchymal stem cells. Journal of Applied Animal Research. 43(1):15-21.Interpretive Summary: There is increasing understanding and need for developing new tools for basic research on how cells differentiate into specialized cell types of our body.  Cells which can develop into many different cell types have considerable therapeutic potential in human and veterinary science. In this study a ARS scientist collaborated in a study that showed bone-marrow derived cell differentiation into a fat-producing cell type.  Furthermore, this work showed various nutritional factors and tissue culture conditions that promotes cell differentiation of fat-producing cells. The results will provide useful information for clinicians and veterinarians to develop novel cell replacing strategy to repair damaged tissues.
                                Technical Abstract: Until now, the isolation and characterization of bovine bone marrow-derived mesenchymal stem cells (bBM-MSCs) have not been established, which prompted us to optimize the differentiation protocol for bBM-MSCs. In this study, bBM-MSCs were freshly isolated from three 6-month-old cattle and used for pluripotent or phenotypic characterization by RT-PCR and flow cytometer. Initially, the cells were adherent to plastic surfaces and exhibited spindle-like morphology. The cells expressed pluripotent as well as typical MSC markers. In addition, bBM-MSCs were differentiated into chondrocytes and osteocytes, but less efficiently into adipocytes. Accordingly, we conducted to establish the optimal adipogenic differentiation protocol under specific culture conditions. For this purpose, we formulated the basal differentiation medium using low-glucose DMEM media supplemented with 10% (v/v) foetal bovine serum, 1% (v/v) penicillin/streptomycin, 1μM dexamethasone, 0.5mM indomethacin and 0.5mM 3-isobutyl-l-methylxanthine, and added with three levels of insulin at 10, 15 and 20μg/mL as insulin is the key adipogenesis inducer. The level of differentiation was evaluated by Oil red O staining and by analyzing the expression of adipocyte-lineage specific genes by a quantitative real time RT-PCR. In this study, we found that the bBM-MSCs were effectively differentiated into adipocytes as manifested by the presence of Oil Red O stained lipid droplets. The mRNA levels of adipocyte-specific genes in the differentiated cells were highly expressed as compared with the non-differentiated bBM-MSCs. In conclusion, we successfully isolated and characterized bBM-MSCs with multipotent and differentiation potential. Additionally, enhanced in vitro adipogenic differentiation protocol for bBM-MSCs was established.


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