Abstract:

Objective

To evaluate the effect of extensive subcultivation on osteoblast of rabbit marrow derived mesenchymal stem cells in vitro.

Methods

Mesenchymal stem cells were isolated from the thighbone marrow of new born rabbit， cultured and subcultured. The subcultured cells were assayed for both the growing rate and osteogenic potential in respose to an osteoinductive medium containing dexamethasone（10 nmol/L）， ascorbic acid（50 mg/L） and β-glycerophosphate（10 mmol/L）at 2，6 and 10 passage（P₂，P₆ and P₁₀） respectively. The osteogenic potential of cells was identified by the activity of alkaline phosphates（ALP） and formation of mineralized nodular aggregates.

Results

The growing rate， the activity of ALP and number of mineralized node showed no statistical different between the P₂ and P₆ cells at low passages. Compared with the values at low passages，the growing rate of P₁₀ at high passage was decreased （P＜0.05）， the activity of ALP and number of mineralized node also showed a decreasing tendency， but there were no statistical differences （P ＞0.05）.

Conclusion

Within 10 passages process of extensive subcultivation， the cells keep the capability of progressing through the osteogenic lineage and can be provided as seeding cells for bone tissue engineering. However， the growing rate and the osteogenic potential of P₁₀ at high passage decreased with various rates.

Key words: Bone marrow, Mesenchymal stem cell, Osteoblast/differentiation, Tissue engineering