Experimental Studies Of The Role Of Children Bone Marrow Mesenchymal Stem Cells For T Lymphocytes And Hematopoietic Stem Cells In Vitro And Transplantation In Hematopoietic Damage

Objective: Mesenchymal stem cells (MSC) isolated from children bone marrow are cultured to examine their roles for T lymphocytes and hematopoietic stem cells in vitro and we study the feasibility of their promoting the hematopoietic covery of irradiafed-damage mice. Method:(1) The MNC from children bone marrow were cultured. The suspicious cells were obtained after adherent-culture for about a month in vitro,these cells were identified by flow cytometric analysis (FACS). Doses of 1.0×103,1.0×10~4 BMSC per recipient respectively were cocultured with actived T lymphocytes(1.0×10~5), which the proliferation rate was evaluated by incorporating 3H-TdR. T lymphocytes and cord blood CD34+ cells were chemoattracted respectively by cultured super natants of doses of 1.0×10~3,1.0×10~4,1.0×10~5BMSC respectively. Using doses of 1.0×10~3,1.0×10~4per well established UCB MNC ex vivo expansion system, The quantity of hematopoietic stem/progenifor cells and the expansion efficiencies of CFU-GM,CFU-E,CFU-GEMM colony formation capability were analyzed. SCID mice underwent 2.5Gy of irradiation followed by bone marrow reconstitution with human UCB MNC alone or cotransplantation with different doses of BMSC marked by hoechst 33258, mice life rate and the amount of white blood cell and platelet were observed by day 21 after transplantation. Result: (1)Children BMSC is easy for culture. BMSC shows surface markers of CD29,CD44,CD105,none of them express CD19,CD34,CD45; (2)During 200ul culture system, 1.0×103 BMSC promote T-cell proliferation, while 1.0×104 BMSC inhibit T-cell proliferation; (3) All levels of BMSC cultured supernatants can chemoattract T lymphocytes and cord blood CD34+ cells ,but the efficiency of 1.0×105 super natants is the highest; (4) During 200ul culture system, UCB MNC ex vivo expansion system of 1.0×104 BMSC per well was more capable of proliferation for hematopoietic stem/progenifor cells and formation CFU-GM,CFU-E,CFU-GEMM colony。During the earlier period(day 8), expansion system containing BMSC is more effective on expansion CD34+ cells. During the later period(day15),HSC was promoted to differentiate into hematopoietic progenifor cells; (5)The higher BMSC dosage co-transplant may be equal with the ten times cells of MNC needed in transfusion of MNC alone for the same expected effects and improve the longevity of mice. Conclusion: children Mesenchymal stem cell has rich resoure ,convenient availability,proliferation ability.Adjustment to the dose of BMSC has different effect for actived T cell. BMSC ex vivo culture system can promote the proliferation of hematopoietic stem/progenifor cell proliferation and the cultured supernatant of BMSC can chemoattract T lymphocytes and cord blood CD34+ cells. BMSC co-transplantation can promote the hematopoietic covery of irradiated-damage mice.children BMSC meets the need of transplantation treatment for malignant haematological diseases, It is expected to become a new treatment to haematological system diseases.