Immune Response Of Allogeneic Tooth Bone Graft Material In Rat Tooth Extraction Model

Objective :Observe and compare the changes of immune response caused by the implantation of allogeneic and autogenous tooth bone as bone graft substitutes,so as to provide clinical guidance and theoretical basis for clinical bone defect repair substitutes.Methods: Forty-five SD male rats,which were SPF grade,6 weeks old and 180±20g,were selected.The mandibular central incisors were extracted,parodontium and pulp of central incisors were removed,the extraction model was established,and the tooth bone was prepared.Rats were randomly divided into A,B,C,D,E 5 groups.A group extraction socket implanted with fresh untreated autogenous tooth bone graft material,B group extraction socket implanted with fresh demineralized autogenous tooth bone graft material,C group extraction socket implanted with fresh untreated allogeneic tooth bone graft material,D group extraction socket implanted with fresh demineralized allogeneic tooth bone graft material and E group extraction socket without implanted bone graft material as blank control group.According to the observation time after operation,the rats were divided into three stages-first,second and fourth weeks.The T lymphocyte subsets in peripheral blood of rats were analyzed one week,two weeks and four weeks after operation,and the mandible was separated.The sections were made for hematoxylin-eosin(HE)staining.Three sections were randomly selected for histological observation and analysis under 50-fold,100-fold and 400-fold microscopesrespectively.Five visual fields were randomly selected for each sample according to cell nucleus shape under 400-fold microscopy.Inflammatory cells were counted artificially.The experimental data were analyzed statistically to evaluate the immune response after allogenic tooth bone transplantation.Image Pro Plus 6.0software was used for image processing analysis to calculate the trabecular area of tooth extraction fossa in each group,and to analyze the osteogenesis effect of each group.Result: 1.Gross observation: 1 day after operation,the face was slightly swollen and the amount of food intake and activity decreased.One week after operation: Tooth bone transplantation site healed well,no swelling and abscess,facial swelling was eliminated,diet of rats was normal,and activity returned to normal.2.Flow cytometry analysis of T lymphocyte subsets: comparison among different groups at the same time point:There was no significant difference in CD4+/CD8+ between fresh untreated allogeneic tooth bone group and other experimental groups at 1,2 and 4 weeks after operation(p>0.05),indicating that there was no significant difference in immunity between untreated allogeneic tooth bone group and other experimental groups.Comparisons at different time points between the same group: CD4+/CD8+ increased in each experimental group at 2 weeks after operation compared with 1 and 4 weeks.The difference of CD4+/CD8+ between autologous untreated group and allogeneic untreated group was significant,with p<0.05,indicating that the untreated group might reach the immune peak at 2 weeks after operation.At 4 weeks after operation,CD4+/CD8+ in each group decreased,and the difference was significant compared with 2 weeks after operation with p<0.05.3.HE staining observation and analysis: No significant inflammatory cell infiltration was found around the tooth bone implantation area in each group of 1,2 and 4 weeks after operation and there are only a few inflammatory cells.New irregular bone trabeculae could be seen in the extraction socket of each group at 2weeks after operation.Degradation and absorption of bone meal could be seen in different degrees in the experimental group at 4 weeks after operation.New bone trabeculae could be seen around bone meal and connected with bone meal closely.After demineralization,the degradation of bone meal particles was obvious,and the area of new bone trabeculae increased.According to the analysis of artificial inflammatory cell count of nuclear morphology.At the same time point: there was no significant difference between fresh untreated allogeneic tooth bone group and other experimental groups(p>0.05).After the analysis of bone trabecular area by Image Pro Plus 6.0software: At the same time point,comparisons were made among different groups: at 2weeks after operation,the difference between allogeneic treatment group,autologous treatment group and autologous untreated group were significant with p<0.05 compared with the blank control group.At 4 weeks after operation,trabecular area increased significantly in allogeneic treatment group and autogenous treatment group,with statistically significant differences from allogeneic untreated group,autogeneicuntreated group and blank control group(p<0.05),while there was no statistically significant difference between allogeneic treatment group and autogeneic treatment group(p>0.05).Comparison of different time points among the same group: irregular new bone trabeculae could be seen in tooth extraction fossa of each group 2 weeks after operation,and the difference of bone trabecular area was statistically significant compared with that at 1 week after operation(p<0.05).The trabecular area of each group increased significantly 4 weeks after operation,and the difference was statistically significant 1 and 2 weeks after operation(p<0.05).Conclusion: According to the experiment,the following conclusions are drawn:1.Allogeneic tooth bone graft material showing no difference with autogenous tooth bone graft material and has good osteogenic effect.2.Both allogeneic tooth bone graft material and autogenous tooth bone graft material after demineralization can induce new bone formation,which is better than untreated tooth bone.3.The immune status of allogeneic tooth bone graft material and autogenous tooth bone graft material after implantation was the same,and there was no significant difference in immune rejection.