Effects of basic fibroblast growth factor in culture of mesenchymal stem cells derived from Wharton's jelly of human umbilical cord

BACKGROUND: During culture of mesenchymal stem cells (MSCs) derived from Wharton's jelly of human umbilical cord. MSC morphology tends to become hypertrophic and irregular. MSCs were found to have higher rate of death and not be adapt to be adherent after passage. It is necessary to find a method to maintain its stability. OBJECTIVE: To isolate MSCs from human umbilical cord wharton's jelly by tissue block method, and to investigate basic fibroblast growth factor (bFGF) effects on biological characteristics of MSCs. METHODS: Human umbilical cord mesenchymal stem cells (hUCMSCs) were separated by tissue block method. Tissue fragments in control group were cultured in growth medium consisting of Dulbecco's Modified Essential /F12 Media (DMEM/F12) and 10% volume fraction of fetal bovine serum. Tissue fragments in experiment group were cultured in growth medium including 20 ug/L bFGF besides these in control group. Time of tissue blocks emigrating from cells and morphology of cells were observed. The medium was changed every 3-4 days. When the cells reached 80%-90% confluency, they were detached with 0.25% trypsin and were passaged at a ratio of 1: 2 or 1:3. RESULTS AND CONCLUSION: A few long spindle or flat fibroblast-like cells were presented firstly after 8-10 days of incubation in control group, while in experiment group it took 6-8 days. One week later, lots of long spindle or flat fibroblast-like cells like whirlpool around micro-mass were presented in two groups. In the first 3 passages, cell morphous were similar and cells were passaged at approximately equivalent time. After passage 3, cells in experiment group were easier to be adherent, lower rate of death and better proliferation ability (shown by growth curve) in comparison with these in control group. Flow cytometry revealed that cell cycle at the passages 3 and 6 showed the percentage of G₀/G₁ was more than 70% respectively. CD44 and CD29 were highly expressed on the surface of passages 3 and 6 cells, whereas the expression of HLA-ABC was less positive, but there was negative for CD34, CD45 and HLA-DR. Cells from Wharton's jelly of the human umbilical cords, which have the biological characteristics of MSCs, have been isolated by tissue culture method. bFGF can shorten the time that MSCs are presented firstly, improve its adherence and proliferation ability and maintain its morphological stability in some degree, moreover keep surface marker expression stability of MSCs.