Recombinant human bone morphogenetic protein 2/porous calcium phosphate cement/autologous bone and combination of platelet-rich plasma in repairing of large bone defects in rabbits by Masquelet technique

Abstract

To investigate the optimal mixing ratio of recombinant human bone morphogenetic protein 2 (rhBMP-2) with porous calcium phosphate cement (PCPC) and autologous bone as bone grafting material for the repair of large bone defects using Masquelet technique. The effect of platelet-rich plasma (PRP) on the healing of bone defects was evaluated under the optimal ratio of mixed bone. Fifty-four New Zealand White rabbits were taken to establish a 2 cm long bone defect model of the ulna and treated using the Masquelet technique. Two parts of the experiment were performed in the second phase of the Masquelet technique. First, 36 modeled experimental animals were randomly divided into 4 groups ( n=9) according to the mass ratio of autologous bone and rhBMP-2/PCPC. Group A: autologous bone (100%); group B: 25% autologous bone+75% rhBMP-2/PCPC; group C: 50% autologous bone+50% rhBMP-2/PCPC; group D: 75% autologous bone+25% rhBMP-2/PCPC. The animals were executed at 4, 8, and 12 weeks postoperatively for general observation, imaging observation, histological observation (HE staining), alkaline phosphatase (ALP) activity assay, and biomechanical assay (three-point bending test) were performed to assess the osteogenic ability and to determine the optimal mixing ratio. Then, 18 modeled experimental animals were randomly divided into 2 groups ( n=9). The control group was implanted with the optimal mixture ratio of autologous bone+rhBMP-2/PCPC, and the experimental group was implanted with the optimal mixture ratio of autologous bone+rhBMP-2/PCPC+autologous PRP. The same method was used to observe the above indexes at 4, 8, and 12 weeks postoperatively. The bone healing process from callus formation to the cortical connection at the defected gap could be observed in each group after operation; new bone formation, bridging with the host bone, and bone remodeling to normal bone density were observed on imaging observation; new woven bone, new capillaries, bone marrow cavity, and other structures were observed on histological observation. The ALP activity of each group gradually increased with time ( P<0.05); the ALP activity of group A was significantly higher than that of the other 3 groups at each time point after operation, and of groups C and D than group B ( P<0.05); there was no significant difference between groups C and D ( P>0.05). Biomechanical assay showed that the maximum load in three-point bending test of each group increased gradually with time ( P<0.05), and the maximum loads of groups A and D were significantly higher than that of groups B and C at each time point after operation ( P<0.05), but there was no significant difference between groups A and D ( P>0.05). According to the above tests, the optimal mixing ratio was 75% autogenous bone+25% rhBMP-2/PCPC. The process of new bone formation in the experimental group and the control group was observed by gross observation, imaging examination, and histological observation, and the ability of bone formation in the experimental group was better than that in the control group. The ALP activity and maximum load increased gradually with time in both groups ( P<0.05); the ALP activity and maximum load in the experimental group were significantly higher than those in the control group at each time point after operation ( P<0.05), and the maximum load in the experimental group was also significantly higher than that in group A at 12 weeks after operation ( P<0.05). In the second phase of Masquelet technique, rhBMP-2/PCPC mixed with autologous bone to fill the bone defect can treat large bone defect of rabbit ulna, and it has the best osteogenic ability when the mixing ratio is 75% autologous bone+25% rhBMP-2/PCPC. The combination of PRP can improve the osteogenic ability of rhBMP-2/PCPC and autologous bone mixture.