| Background and purposeWith the improvement of People’s living standard and the rapid development of dental implant technology,dental implant has gradually become the first choice for patients with dentition defects.Insufficient alveolar bone is a common problem in implant repair,and suitable bone graft material is more and more important in clinical application,which has a significant impact on the success rate of implant repair.A wide variety of bone graft materials in clinical patients to provide implant bone defect repair,long-term results are uneven.The purpose of this study was to investigate the effect of human bone graft material demineralized by ultrasound on the adhesion and polarization of mouse macrophage RAW264.7 and to explore the Biocompatibility between human bone graft material and mouse macrophage RAW264.7.MethodsCollect human tooth and spray water with high-speed to remove tooth enamel,pigmentation,tartar and pulp tissue.The bulk human tooth bone graft materials of similar size were prepared and divided into two groups,namely untreated block human tooth bone graft material and Vacua Sonic vacuum ultrasound combined with hydrochloric acid treated block human tooth bone graft material,and the morphology of the two groups of materials was observed by scanning electron microscopy.The extract was prepared at a concentration of 1 mL of human tooth bone graft material,and the two groups of extract were cultured with RAW264.7 mouse macrophage for 1,3 and 5 days,respectively,and differentiation of RAW264.7 mouse mononuclear macrophage was observed.Two sets of extracts were co-cultured with RAW264.7 mononuclear macrophages,and cell proliferation was detected by MTT and CCK-8 for 1,3,and 5 days.ELISA measured IL-10 inflammatory factor expression levels secreted by RAW264.7 mouse mononuclear macrophages after leachate treatment.Raw264.7 macrophage was lysed,total macrophage RNA was extracted,reverse transcription,mRNA was turned into cDNA,PCR was performed by cDNA,mRNA level was detected by qPCR,and Western Blot was used to detect protein expression.ResultsElectron microscope showing,ultrasonic vacuum Sonic hydrochloric acid demineralized massive human dental bone grafts were fully opened and widened,and the contact area was increased compared to untreated massive human dental bone grafts.Compared with untreated massive human dental bone graft material,the extract of Vacua Sonic ultrasonic hydrochloric acid demineralized massive human dental bone graft material was co-cultured with RAW264.7 macrophage under a light microscope,and the differentiated morphology of RAW264.7 macrophage was closer and the pseudopod shorter.Macrophages cultured from extracts of untreated human dental bone graft material showed long spindle type and long pseudopodia.The light absorption values of raw human dental bone graft extract and Vacua Sonic ultrasonic hydrochloric acid demining extract cultured with macrophages for 1,3 and 5days,respectively,were detected by MTT and CCK-8 experiments.The light absorption values of the two groups of materials were different only after 3 days.The light absorption value of Vacua Sonic ultrasonic hydrochloric acid demineralized group was significantly higher than that of untreated group,which promoted cell proliferation.ELISA showed that the level of IL-10 secreted by macrophages after co-culture of Vacua Sonic hydrochloric acid demineralization extract with RAW264.7 macrophage was higher than that of untreated human dental bone graft extract.qPCR and Western Blot tests detected transcription and protein expression levels of M1 and M2 polarization markers in macrophages,suggesting high transcription and protein expression levels of M2 markers ARG1 and CD163 in the human dental bone graft material group Vacua Sonic ultrasonic demineralization.The transcription and protein expression levels of IL-1α and IL-1β markers of M1 were lower,and the differences were statistically significant(P<0.05).Conclusion1.Vacua Sonic hydrochloric acid demineralization of massive human dental graft material extract group promoted the differentiation of macrophage RAW264.7,which could transform macrophages from M1 to M2.2.The expression level of IL-10 secreted by macrophage RAW264.7cultured in the extract of Vacua Sonic ultrasonic hydrochloric acid demineralized massive human tooth transplantation material group was higher than that in the control group.3.Vacua Sonic ultrasonic hydrochloric acid demineralized massive human tooth bone graft material can promote macrophage proliferation and differentiation,and has good biocompatibility. |
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