The Experimental Study Of The Repairing Of Segmental Bone Defects Using MSCs Transferred With BFGF Gene And Combined With Nhac

ObjectivesBone defects were common conditions caused by trauma, sarcoma or infection. Small bone defects can be repaired automatically by bone induction, conduction or regeneration. Large bone defects will not heal without bone transplantation.The current bone defects repairing materials include: Autogenous bone, allogenic bone, artificial materials, composite materiel etc. All of these materials were not ideal. It is clinical desirable to find ideal materials to repair larger bone defects. Bone tissue engineering may provide an alternative for bone defects repairing. The aim of this study was to investigate the bone defects repairing ability of the combined material, which with MSCs transduced with bFGF gene as its seeding cells and with nHAC as its scaffolds. Materials and methods1. 20 young New Zealand white rabbits about 2~3 months old were used to harvest bone marrow. The bone marrow was inspired from two femoral trochanter of the rabbits. MSCs were isolated from the bone marrow cells and anchoring at one hour and physically curetle and cultured in vitro. Small samples of MSCs were cultured with conditioned culture medium to assess the alkaline phosphates and the mineralization ability.2. Half of the MSCs were transferred with bFGF gene and cultured. Small samples were used to assess the expression of the bFGF with RT-PCR methods andimmuohistological methods.3. MSCs or MSCs transferred with bFGF gene were combined with nHAC and cultured for 5~7 days respectively. Specimens were obsevered under microscope and electron microscope.4. 62 New Zealand white rabbits which were about 3~4 months old were divided into A, B, D group. 10 mm segmental defect were made on the bilateral radius of the rabbits. A group (n=30): the defect areas on the left radius were transplanted with MSCs combined with nHAC. B group (n=30): MSCs transduced with bFGF gene combined with nHAC were transplanted into the defect areas on the left radius. C group (n=60): All the right radiuses of A and B group were divided into this group, the defect areas were implanted with nHAC scaffolds without cells. D group: 2 other rabbits were divided into this group, the same bone defect were made on the bilateral radius without implanting materials. After 2 weeks, 4 weeks, 6 weeks, 12 weeks, 24 weeks from operation, 6 rabbits were killed from A or B group and X ray radio graphical, histological and immuohistological studies were performed. Specimen from C group were harvest and assessed consistent with A and B group, Specimen from D group were harvest at 24 weeks after operation and were assessed with radio graphical and histological method.Results1. The characters of MSCs: the shape of MSC is fusiform or angiogram, with multiple processes, the plasma is pale blue, the nucleus is round with multi-differentiation. MSCs survived and proliferated and differentiated well, the biochemical indexes were stable, MSCs showed the expression of alkaline phosphates, and localized regions of mineralization were found at about 2 weeks with conditioned culture medium.2. bFGF gene transferred MSCs: It didn't change the morphological characteristics of MSCs when MSCs were transferred with bFGF gene. The proliferation and differation of MSCs with bFGF gene transferred were active. The MSCs with bFGF gene transferred can express bFGF at 24h, can express bFGF notably at about 2 weeks and can still express bFGF at 5 weeks after gene transferring.