Immunoregulatory effect of neuronal-like cells in inducting differentiation of bone marrow mesenchymal stem cells

J. Xu, H. Lu, Z.-N. Miao, W.-J. Wu, Y.-Z. Jiang, F. Ge, W.-F. Fang, A.-H. Zhu, G. Chen, J.-H. Zhou, Y.-Z. Lu, Z.-F. Tang, Y. Wang

Wuxi Third People’s Hospital, Liangxi, Wuxi, Jiangsu, China. xq3053324wudaomu@163.com

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• Obstetrics

OBJECTIVE: To evaluate the immune activity of bone marrow mesenchymal stem cells (BMSCs), and explore the biological characteristics and capabilities of BMSCs and the potential to be differentiated into neuronal cells in vitro.

MATERIALS AND METHODS: The BMSCs were isolated and proliferated in vitro to generate the xenogeneic mixed lymphocyte reaction. Moreover, peripheral BMSCs (pBMSCs) were added according to different ratios, which methods were stated as follows: 1: Dulbecco’s Modified Eagle Medium (DMEM) + 10% Fetal Bovine Serum (FBS) + 1 μmol/L all-trans-retinoic acid (ATRA) + 20 μg/L basic fibroblast growth factor (bFGF) + 20 μg/L epidermal growth factor (EGF); 2: DMEM + 2% dimethyl sulfoxide (DMSO) + 100 μmol/L butylated hydroxyanisole (BHA). The immunofluorescence and immunohistochemical staining were finally used to evaluate the differentiation capabilities of human BMSCs (hBMSCs) induced in neuronal cells.

RESULTS: hBMSCs inhibited the lymphocyte proliferation in the mixed lymphocyte reaction (MLR) system at a proportional inhibition rate with additional numbers of stem cells. At hour 2 after culture with method 1, the plasma of hBMSCs shrank to nuclei and perinuclear bodies and was visualized under the light microscope. At hours 3-5, most of the hBMSCs formed neuron-like cells with total cell number unchanged. Afterward, the hBMSCs turned into bipolar or multipolar shaped cells and interconnected into a large network at Day 3. With immunofluorescence and immunohistochemical staining, 60-70% of the hBMSCs showed neurospecific enolase (NSE) positive and 45-50% glial fibrillary acidic protein (GFAP) positive while the Nestin-positive cells decreased to 3.4%. However, when cultured 2 hours with method 2, the most of the hBMSCs formed bipolar or multipolar shaped cells, then died after 48 hours. 40-50% NSE and 35-40% GFAP were positively expressed. Significantly, the rate of Nestin-positive cells decreased from 63% to 1.6% from hour 2 after culture to hour 48.

CONCLUSIONS: hBMSCs may be effective for cell therapy and tissue engineering for the capability of differentiating into neuronal-like cells, as well as the capability of inhibiting lymphocyte proliferation in MLR system.

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