Establishment Of Mice Model Of Peri-implantitis And Study Of NLRP3 Expression In The Model

Objective: To establish mice model of peri-implantitis and investigate the expression of NLRP3 in gingival tissue of the mice model of peri-implantitis,and to clarify its pathogenic effect on peri-implantitis.Methods: 36 4-week-old C57 BL / 6J mal e mice were used in this study.The upper and lower first molar on the right side were extracted,and custom-made implants were implanted immediately.After 4 weeks of healing,osseointegration was obtained.Then the mice were randomly divided into blank group,control group and model group with 12 mice in each group.Mice in the blank group were killed on day 0 after ligation(i.e.before ligation treatment)to obtain specimens;The model group was treated with ligation for 2 weeks.The 5-0 silk thread was firmly placed on the subgingival neck of the model group mice implant,and the mice were killed after 2 weeks of ligation treatment to obtain specimens.The mice in the control group were not ligated,and they were killed after 2 weeks of observation to obtain specimens.Six mice in each group were used for Micro-CT and gene expression analysis,and the other six mice in each group were used for histological analysis.During the experiment,morphological observation was used to evaluate the signs of inflammation of the gingival tissue.Micro-CT was used to scan the bone tissue of mice to evaluate the bone height and bone density difference around implants.Histological analysis was carried out on specimens of each group by HE staining and TRAP staining methods.mRNA expression levels of NLRP3 and IL-1? in gingival tissue of each group were detected by RT-PCR method.Results: The results of morphological observation showed that the gingival tissue around the implant in the model group showed obvious redness,edema,texture softening and exudation,while there were no visible signs of inflammation in the blank group and the control group.After Micro-CT scanning and analysis,the bone height data around the implant of each group of mice are as follows: 876.6 ± 50.03?m(mesial side of blank group),834.3 ± 37.56?m(mesial side of control group),576.2 ± 55.81?m(mesial side of model group),906.2 ± 87.58?m(distal side of blank group),864.5 ± 58.70?m(distal side of control group),555.8 ± 26.61?m(distal side of model group),1019 ± 86.59?m(buccal side of blank group),954.4 ± 42.86?m(buccal side of control group),584.3 ± 75.00?m(buccal side of model group),819.9 ± 42.17?m(palate side of blank group),792.8 ± 78.99?m(palate side of control group),556.8 ± 60.59 ?m(palate side of model group).Compared with the control group,the bone height of the mesial,distal,buccal,and palatal sides of the model group was significantly reduced(P <0.05),while the bone heights of the mesial,distal,buccal,and palatal sides of the blank group had no statistical difference(P>0.05).For the residual bone mineral density in the region of interest(VOI)around the implant,there was a statistical difference between the model group[(964.6±6.586)mg/HA ccm] and the control group[(1034±44.05)mg/HA ccm](P<0.05),and there was no significant difference between the control group [(1034±44.05)mg/HA ccm] and the blank group [(993.5±10.02)mg/HA ccm](P>0.05).The histological sections demonstrated that the bone levels of buccal and palatal parts of the implant decreased significantly.the number of TRAP positive cells in the ROI of the model group(26.67±10.02)was significantly different from that of the control group(5.00±2.00)(P<0.05),while the number of TRAP positive cells in the ROI of the blank group(3.33±1.53)was not significantly different from that of the control group(P>0.05).At high magnification(200 times),a large number of inflammatory cells(including plasma cells,macrophages and polymorphonuclear leukocytes)infiltrated into the connective tissue around the implant were observed in the ROI of the model group,while there was almost no infiltration of inflammatory cells in the blank group and the control group.The number of inflammatory cells in ROI of model group(36.67±9.074)was significantly higher than that of control group(14.67±3.215)(P<0.05),while the number of inflammatory cells in ROI of blank group(10.67±2.082)had no statistical difference compared with control group(P>0.05).The relative level of NLRP3 mRNA expressed in gingival tissue of model group(4.23±0.51)was significantly higher than that of control group(1.29±0.41)(P<0.05).When comparing the blank group(1.04 ± 0.05)with the control group(1.29 ± 0.41),there was no significant difference in the relative expression level of NLRP3 mRNA between the two groups(P > 0.05).The relative level of IL-1 ? mRNA in gingival tissue of model group(2.84 ± 0.59)was significantly higher than that of control group(1.10 ± 0.33)(P < 0.05).When comparing the blank group(1.07 ± 0.07)with the control group(1.10 ± 0.33),there was no significant difference in the relative expression level of IL-1 ? m RNA between the two groups(P > 0.05).Conclusion: The mice model of peri-implantitis was successfully established by ligation induction method.NLRP3 and IL-1? were highly expressed in gingival tissue of the model mice,which can be preliminarily determined to promote the pathogenesis of peri-implantitis.