Abstract:

BACKGROUND: Bone marrow mesenchymal stem cells are restricted in clinical application due to low contents in bone marrow, easy to lose stem cell potential during long-term in vitro culture, and unclear safety after in vivo transplantation.
OBJECTIVE: To explore an optimal isolation, purification and cytopreservation method of rat bone marrow mesenchymal stem cells and observe the tumorigenicity after implanting the in vitro cultured cells into nude mice.
METHODS: Bone marrow mesenchymal stem cells were obtained from bone marrow cavity, purified and cutured by three different methods. The culture-expanded bone marrow mesenchymal stem cells were cryopreserved with cytoprotectant solution containing different serum concentrations at -80 ℃. Following recovery from cryopreservation, cell growth capacity and adiogenic capacity were determined. Passage 3 and 15 bone marrow mesenchymal stem cells were injected into the muscle and liver of nude mice to observe the neoplasia in local site.
RESULTS AND CONCLUTION: The purity of bone marrow mesenchymal stem cells obtained by differential adhesion combined with 24-hour first exchange method was higher compared to other control groups, and there was no significant difference in cell proliferation capacity between groups. Bone marrow mesenchymal stem cells cryopreserved with 30% serum could ensure the viability, proliferation ability and stem cell characterisitics and multi-differentiation potential. Bone marrow mesenchymal stem cells might keep stem cell properties until to the 15th generation. After 45 days of transplantation to nude mice, no neoplasia was found in the local site. The muscle tissue had normal structure. Passage 15 bone marrow mesenchymal stem cells could survive and proliferate in the liver with no hetermorphism and invasion to the surrounding liver tissue.