Abstract:

BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) are widely utilized as seed cells or carriers in bone tissue engineering and gene therapy. Thus, how to obtain BMSCs with high purity arose more attentions of researchers.

OBJECTIVE: To observe the effects of different separation methods and culture conditions on biological characteristics of rabbit BMSCs.

METHODS: BMSCs were obtained by whole bone marrow culture, density grand centrifugal and red blood cell lysis. At 48 hours after culture, the cell numbers were counted, the time of passage was recorded, in addition, the cell morphology was observed by phase contrast microscope, and the CD44 antigen expression was identified using flow cytometry. The 3^rd and the 7^th generation aging cells were cultured with DMEM-LG, MEM-HG, and DMEM/F12 culture medium. MTT and count cell plat were used to evaluate the growth of BMSCs. Phase contrast microscope was used to observe the morphological changes of aging cells.

RESULTS AND CONCLUSION: BMSCs could be separated by each method. The adherent cells showed shuttle or multiple angle shapes, with rich cytoplasm, and positive for CD44 antigen. The more cell number and shortest primary culture time was presented in red blood cell lysis group (P < 0.05). DMEM/F12 could promote the proliferation of quiescent cells. And the cells prevented the better viability. The method of read blood cell lysis improving the efficiency of BMSCs adherent is an effective method of extraction of BMSCs. DMEM/F12 could promote the proliferation Maybe, DMEM/F12 is more suitable for BMSCs.