Experimental Study On Repairing Bone Defect With Mononuclear Cells Derived From Bone Tissue Hemorrhage

Objective: The purpose of this study was to simulate the collection of mononuclear cells derived from bone tissue hemorrhage during the operation, and to observe the regenerative potential of mononuclear cells derived from bone tissue hemorrhage in a rabit’s critical size segmental defect model. To provide the experimental basis for clinical application of mononuclear cells derived from bone tissue hemorrhage for treatment of bone defects.Methods: The 22 New Zealand white rabbits were randomly divided into two groups based on "random number table". Experimental group,named group A, n1=13, simulated the collection of bone tissue hemorrhage during the operation. Mononuclear cells were isolated from the bone tissue bleeding in the rabbit by means of "density gradient centrifugation". After PKH26 fluorescence labeled, cells were mixed with allogeneic bone particles, and migrated to the ulna defects. Control group,named group B, n2=9, just only implanted allogeneic bone into rabbits’ ulna bone defect. After 8 weeks, the bone graft was evaluated by general observation of bone tissue. Ulna bone defect was examed by X-ray. And the healing of bone defect was evaluated by Lane-Sandhu’s X-ray grading standard. The histological characteristics of the newly formed bone tissue were observed, and the healing of bone defect was evaluated by Lane-Sandhu′s histological grading standard. Fluorescence microscopy was used to observe the distribution of PKH26 labeled cells in the newborn bone tissue.Results: In group A, the number of mononuclear cells isolated and mixed with allogeneic bone was(6.32 ±2.38)×107. Eight weeks after bone grafting, in group A, bone defect was healing through general observation. However, in group B, bone defect was still evident.( 9.92 ± 2.22) and( 5.11 ± 1.17) were evaluated by Lane-Sandhu’s X-ray grading standard, respectively in group A and group B. There was a significant statistic difference between the two groups(t=6.21,P<0.05). In group A, the new bone tissue sections stained with H-E showed normal bone tissue adjacent to os cartilaginea tissue. However, in group B, H-E staining section was implanted scaffold and less bone formation.(8.69±1.89)and(3.44±1.59)were evaluated by Lane-Sandhu′s histological grading standard, respectively in group A and group B. There was a significant statistic difference between the two groups(t=6.32,P<0.05). In the experimental group, the PKH26 labeled cells were widely distributed in the new bone tissue.Conclusion: Mononuclear cells derived from bone tissue hemorrhage combined with allogeneic bone in the treatment of bone defect could promote the healing of bone defect better than simply in the application of allograft bone. The greatest potential for bone regeneration medicated by mononuclear cells derived from bone tissue hemorrhage was found in the experiment. And it is the ideal choice for tissue engineering treating bone defect.