Use of dezocine maleate in the preparation of a drug for treating peri-implant mucositis

Abstract

This invention belongs to the field of pharmaceutical technology and discloses the application of dezocampine maleate in the preparation of a drug for treating peri-implant mucositis. This invention is the first to propose that dezocampine maleate has a therapeutic effect on peri-implant mucositis. In in vivo experiments, a peri-implant mucositis model was established using C57BL / 6 mice. The study investigated that dezocampine maleate could improve the clinical manifestations of peri-implantitis in the model mice, reduce macrophage recruitment, regulate and decrease the polarization of pro-inflammatory M1 macrophages, increase the polarization of anti-inflammatory M2 macrophages, and enhance the expression of IL-1β, IL-6, TNF-α, IκB-α, and NF-κB. In in vitro experiments, the therapeutic effect of dezocampine maleate on peri-implant mucositis was investigated. Simultaneously, using a lipopolysaccharide-stimulated RAW264.7 cell model, the effect of dezocampine maleate on the Glu / NMDAR / NF-κB signaling pathway was verified.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to the application of dezocine maleate in the preparation of a drug for treating peri-implantitis. Background Technology

[0002] Peri-implant mucositis (PM) is defined as an inflammatory lesion of the mucosa surrounding the implant without persistent marginal bone loss. It is a common complication associated with implant support rehabilitation. PM is a precursor to peri-implantitis. At this stage, the lesion is confined to soft tissue and can be effectively treated and reversed with local sustained-release medications and non-surgical mechanical debridement. If left untreated, it may progress to peri-implantitis. The latter is complex to treat and carries a significant burden on the patient population; continued progression can lead to implant and implant-supported prosthesis loss. Therefore, early identification and aggressive treatment of PM are crucial to preventing disease progression to peri-implantitis and ensuring long-term implant survival.

[0003] When the microbial homeostasis of the periimplant tissue is disrupted, pathogens and secretions within the biofilm can trigger periimplantitis, inducing a host immune response. Dysbiosis of the periimplant microbiota triggers an inflammatory response mediated by the activation of innate immune cells such as macrophages, dendritic cells, mast cells, and neutrophils. This leads to inflammatory lesions around the implant, accelerating the production of cytokines, chemokines, prostaglandins, and proteases. Inflammation typically progresses in a non-linear, incremental manner, further exacerbating tissue damage and expanding the lesion area.

[0004] Although peri-implant mucositis (PM) is reversible with early detection and timely intervention, there is currently a lack of specific and effective treatments in clinical practice, aside from mechanical debridement. Early intervention to control the inflammatory process of PM holds promise for preventing its progression to irreversible peri-implantitis, ultimately ensuring and improving the long-term survival rate of the implant and superstructure. Therefore, elucidating the underlying pathogenesis of PM and developing safer treatment strategies is of significant clinical importance.

[0005] Dizocilpine maleate (MK-801) is a selective, non-competitive NMDA receptor antagonist that works by binding to a site within NMDA-associated ion channels, thereby inhibiting the absorption of calcium ions. 2+ The flow of [something unclear - possibly related to drug activity]. Recent studies have shown that dezocine maleate exhibits broad pharmacological activity in various diseases, including anti-inflammatory and anti-oxidative stress effects. However, its role in peri-implantitis mucositis remains unstudied. Summary of the Invention

[0006] The purpose of this invention is to provide new pharmaceutical uses for dezocine maleate.

[0007] To address the aforementioned technical problems, embodiments of the present invention provide the use of dezocampine maleate (MK-801) in the preparation of a medicament for treating peri-implant mucositis.

[0008] In the pharmaceutical applications provided by this invention, dezocypine maleate antagonizes the N-methyl-D-aspartate receptor (NMDAR) on the surface of peri-implant macrophages in mice, inhibiting peri-implant mucosal inflammation induced by Porphyromonas gingivalis in mice. The mice in question are C57BL / 6 mice.

[0009] In the pharmaceutical applications provided by this invention, the dezocypine maleate can improve the clinical manifestations of peri-implantitis in model mice, reduce macrophage recruitment, regulate the reduction of pro-inflammatory M1 macrophage polarization, increase the polarization of anti-inflammatory M2 macrophage, and the expression of IL-1β, IL-6, TNF-α, IκB-α, and NF-κB.

[0010] In the pharmaceutical applications provided by this invention, the dezocypine maleate was used in a RAW264.7 cell model stimulated by lipopolysaccharide extracted from Porphyromonas gingivalis to verify that dezocypine maleate has an inhibitory effect on the Glu / NMDAR / NF-κB signaling pathway.

[0011] Embodiments of the present invention also provide a medicament for treating peri-implant mucositis, the medicament comprising dezocine maleate and pharmaceutically acceptable excipients.

[0012] This invention proposes for the first time the application of dezocampine maleate in the preparation of a drug for treating peri-implant mucositis. The invention induces peri-implant mucositis in mice by ligating the implant neck with sutures soaked in *Porphyromonas gingivalis* supernatant. Dezocampine maleate is then administered to the mouse model of peri-implant mucositis via local infiltration. The therapeutic effect of dezocampine maleate on peri-implant mucositis is investigated. Peri-implant gingival tissue from mice was collected and subjected to the following tests: histopathological changes were observed by hematoxylin-eosin (HE) staining and Masson's trichrome staining; the expression and distribution of macrophage markers F4 / 80, CD86, and CD206 were detected by immunofluorescence staining; and the protein expression levels of inflammatory factors IL-1β, IL-6, TNF-α, IκB-α, and NF-κB were detected by Western blotting and / or enzyme-linked immunosorbent assay. In in vitro experiments, this invention used lipopolysaccharide (LPS) derived from *Porphyromonas gingivalis* to stimulate RAW264.7 macrophages to establish an inflammatory cell model. The cells were divided into a control group, an LPS model group, an MK-801 group, and an LPS + MK-801 treatment group. Before LPS stimulation, cells were pretreated with dezocampine maleate for 2 hours. The expression of M1 macrophage markers inducible nitric oxide synthase (IVO2) and CD86, and M2 macrophage markers arginase-1 and CD206 were detected by immunofluorescence staining, Western blotting, and flow cytometry. Extracellular glutamate concentration was measured using a glutamate assay kit. The expression levels of NMDA receptor subunits GRIN1 and GRIN2B, as well as the phosphorylation levels of NF-κB p65 and IκB-α, were detected by real-time quantitative reverse transcription polymerase chain reaction (RT-PCR) to verify the regulatory effect of dezocampine maleate on the Glu / NMDAR / NF-κB signaling pathway.

[0013] The above experimental results confirm that dezocampine maleate can improve peri-implant mucosal inflammation. In in vivo experiments, a peri-implant mucosal inflammation model was established using C57BL / 6 mice. The study investigated that dezocampine maleate could improve the clinical manifestations of peri-implant mucosal inflammation in the model mice, reduce macrophage recruitment, regulate and decrease the polarization of pro-inflammatory M1 macrophages, increase the polarization of anti-inflammatory M2 macrophages, and affect the expression of IL-1β, IL-6, TNF-α, IκB-α, and NF-κB. In in vitro experiments, the therapeutic effect of dezocampine maleate on peri-implant mucosal inflammation was studied. Simultaneously, a RAW264.7 cell model stimulated with lipopolysaccharide extracted from *Porphyromonas gingivalis* was used to verify the effect of dezocampine maleate on the Glu / NMDAR / NF-κB signaling pathway. In conclusion, it has a therapeutic effect on peri-implant mucosal inflammation under both in vivo and in vitro conditions. Attached Figure Description

[0014] Figure 1This is a diagram showing the clinical manifestations of the peri-implant tissues after implantation of an implant in the right maxillary first molar region of experimental mice in Example 1, and modeling of mice in blank group, peri-implant mucositis group, drug group, and treatment group. Figure 2 This is the result of HE staining of buccal and lingual sections of tissue surrounding the implant after implanting an implant in the right maxillary first molar region of experimental mice in Example 1, and modeling mice in blank group, peri-implant mucositis group, drug group, and treatment group. Figure 3 This is the result of masson staining of buccal and lingual sections of tissue surrounding the implant after implanting an implant in the right maxillary first molar region of experimental mice in Example 1, and modeling mice in blank group, peri-implant mucositis group, drug group, and treatment group. Figure 4 This is a graph showing the results of detecting the protein expression levels of IL-1β, IL-6, and TNF-α in the peri-implant tissue of experimental mice after implanting an implant in the right maxillary first molar region of experimental mice in Example 1, and establishing mouse models in the blank group, peri-implant mucositis group, drug group, and treatment group. Figure 5 In Example 2, an implant was placed in the right maxillary first molar region of experimental mice. After establishing mouse models in the blank group, peri-implant mucositis group, drug group, and treatment group, the fluorescence images of M1 polarization marker CD86 and M2 polarization marker CD206 were detected by immunofluorescence technology. Figure 6 This is a diagram showing the results of Western blotting analysis of the expression of CD86, CD206, iNOS, and Arg-1 in the peri-implant tissue of mice after implantation of an implant in the right maxillary first molar region of experimental mice in Example 2. The model mice were established in a blank group, a peri-implant mucositis group, a drug group, and a treatment group. Figure 7 This is a graph showing the results of detecting the glutamate concentration produced by RAW264.7 cells stimulated with LPS and dezocyclopine maleate using a glutamate concentration measurement kit in Example 3. Figure 8 This is a figure showing the results of using polymerase chain reaction and Western blotting in Example 3 to detect the effects of LPS and dezocine maleate on the mRNA and protein expression of IL-1β, IL-6, and TNF-α in RAW264.7 cells. Figure 9 This is a diagram showing the results of immunofluorescence detection of IL-10 protein expression in RAW264.7 cells stimulated with LPS and dezocyclopine maleate in Example 3. Figure 10This is a graph showing the results of Western blotting assays in Example 3 to detect the protein expression of CD86, CD206, iNOS, and Arg-1 in RAW264.7 cells stimulated with LPS and dezocyclopine maleate. Figure 11 This is a graph showing the effect of Western blot detection on the expression of NF-κB protein in the peri-implant tissue of experimental mice after implanting an implant in the right maxillary first molar region of experimental mice in Example 3, establishing mouse models in blank group, peri-implant mucositis group, drug group, and treatment group. Figure 12 This is a graph showing the results of Western blotting in Example 3, which detected the effect of LPS and dezocyclopine maleate on NF-κB protein expression in RAW264.7 cells. Figure 13 This is a diagram showing the results of immunofluorescence detection of NF-κB nuclear translocation in RAW264.7 cells stimulated by LPS and dezocyclopine maleate in Example 3. Figure 14 This is a schematic diagram illustrating the mechanism of action of dezocypine maleate in treating peri-implantitis. Detailed Implementation

[0015] To make the objectives, technical solutions, and advantages of this invention clearer, the various embodiments of this invention will be described in detail below. However, those skilled in the art will understand that many technical details have been provided in the various embodiments of this invention to facilitate a better understanding of this application. However, the technical solutions claimed in the claims of this application can be implemented even without these technical details and various variations and modifications based on the following embodiments.

[0016] Dezocliptin maleate (MK-801) is a selective, non-competitive NMDA receptor antagonist that works by binding to a site within NMDA-associated ion channels, thereby blocking Ca2+ receptor antagonism. 2+ The flow of NMDA receptors by MK-801 alleviates the pro-inflammatory changes induced by LPS, and has certain pharmaceutical potential.

[0017] This invention primarily explores the therapeutic effects of dezoclimine maleate on peri-implant mucositis under in vivo and in vitro conditions. In in vivo experiments, a peri-implant mucositis model was established using C57BL / 6 mice. The study investigated how dezoclimine maleate improved the clinical manifestations of peri-implantitis in the model mice, reduced macrophage recruitment, regulated decreased polarization of pro-inflammatory M1 macrophages, increased polarization of anti-inflammatory M2 macrophages, and its effects on the expression of IL-1β, IL-6, TNF-α, IκB-α, and NF-κB. In in vitro experiments, the therapeutic effect of dezoclimine maleate on peri-implant mucositis was investigated. Simultaneously, a RAW264.7 cell model stimulated with lipopolysaccharide extracted from *Porphyromonas gingivalis* was used to verify the effect of dezoclimine maleate on the Glu / NMDAR / NF-κB signaling pathway. A schematic diagram illustrating the mechanism of action of dezoclimine maleate in treating peri-implant mucositis is shown below. Figure 14 .

[0018] The experimental scheme and results established for the embodiments of the present invention are as follows: A mouse model of peri-implant mucositis was established using miniature implants. After extraction of the right maxillary first molar in C57BL / 6 mice and local hemostasis, a sterile miniature titanium implant was immediately inserted into the extraction socket using a miniature implant actuator. 5-0 sutures soaked in sterile saline or *Porphyromonas gingivalis* supernatant were ligated at the implant neck, achieving local drug administration through suture infiltration. The mice were divided into a control group, a peri-implant mucositis group, a drug group, and a treatment group. During the first week post-surgery, the peri-implant mucositis group and the treatment group received local injections of *Porphyromonas gingivalis* supernatant at the ligation site every other day, while the control group and the drug group received an equal volume of sterile saline. During the second week, the control group and the peri-implant mucositis group continued to receive saline injections, while the drug group and the treatment group received local injections of 1 mM MK-801 at the ligation site every other day. The modeling period was two weeks. After modeling, peri-implant tissue was collected, and the protein levels of inflammatory factors IL-1β, TNF-α, and IL-6 in the peri-implant tissue were detected by Western blotting. Hematoxylin and eosin (HE) staining and Masson's staining were used to detect inflammatory granulation tissue and macrophage infiltration in the peri-implant tissue. The results showed that, compared with the inflammation group, the peri-implant mucosal inflammation group treated with dezocine maleate showed reduced peri-implant inflammatory response, decreased macrophage infiltration and recruitment, and lower protein levels of related inflammatory factors.

[0019] The anti-inflammatory mechanism studied in vivo was then validated. C57BL / 6 mice were used to establish models of peri-implant mucositis, including a control group, a peri-implant mucositis group, a drug group, and a treatment group. Peri-implant tissue was harvested. Western blotting was used to detect the protein expression levels of phosphorylated IκB-α, phosphorylated NF-κB, and NF-κB in the peri-implant tissue. Immunofluorescence was used to detect the nuclear translocation of the key protein in the NF-κB p65 signaling pathway in the peri-implant tissue. Compared with the control group, the phosphorylation levels of the signaling pathway proteins IκB-α and NF-κB were decreased in the peri-implant mucositis group treated with dezocampine maleate.

[0020] Using C57BL / 6 mice, peri-implant mucosal inflammation (PMI) models were established in a blank control group, a peri-implant mucosal inflammation group, a drug group, and a treatment group. Peri-implant tissue was harvested. Immunofluorescence was used to detect macrophage recruitment in the PMI tissue, and the expression of macrophage polarization markers CD86 and CD206 was detected. Western blotting was used to detect the protein levels of macrophage polarization markers CD86, CD206, iNOS, and Arg-1 in the PMI tissue. Results showed that compared with the PMI group, the dezocine maleate-treated PMI group exhibited reduced macrophage infiltration and recruitment, decreased expression of M1 macrophage polarization markers CD86 and iNOS, and increased expression of M2 macrophage polarization markers CD206 and Arg-1.

[0021] In in vitro experiments, RAW264.7 cells were used to verify its specific anti-inflammatory mechanism. RAW264.7 cells in different groups were first treated with dimethyl sulfoxide or dezocampine maleate for 2 hours, followed by stimulation with lipopolysaccharide extracted from *Porphyromonas gingivalis*. Cells were collected after 8 hours. The mRNA expression of IL-1β, IL-6, and TNF-α in RAW264.7 cells was detected by polymerase chain reaction (PCR), and the protein levels of IL-1β, IL-6, TNF-α, IκB-α, and NF-κB in RAW264.7 cells were detected by Western blotting and immunofluorescence staining. The results were consistent with those of a mouse peri-implant mucosal inflammation model. Compared with the dimethyl sulfoxide control, dezocampine maleate intervention inhibited the expression of IL-1β, IL-6, TNF-α, IκB-α, and NF-κB in lipopolysaccharide-stimulated RAW264.7 cells.

[0022] Example 1: Effects of dezocampine maleate on inflammatory manifestations, macrophage infiltration and recruitment, and protein levels of related inflammatory factors in a mouse model of peri-implantitis mucositis. 1. Experimental Materials 1.1 Experimental reagents: Dezocyclopine maleate (MCE Company) 1.2 Experimental animals: C57BL / 6 mice (Jinan Pengyue Experimental Animal Breeding Co., Ltd.) 1.3 Experimental bacteria: Porphyromonas gingivalis 33277 (purchased from the American Center for Type Culture Collection) 2. Experimental Methods 2.1 Model Mice Eight-week-old healthy, clean-grade female C57BL / 6 mice were selected as research subjects. The oral cavity was examined before the experiment to rule out missing teeth, and the right side was selected as the experimental side. All procedures performed on the mice complied with the guidelines for humane treatment of laboratory animals issued by the Ministry of Science and Technology of China (vGKFCZ-2006-398).

[0023] 2.2 Effects of dezocine maleate on inflammatory manifestations, macrophage infiltration and recruitment, and protein levels of related inflammatory factors in a mouse model of peri-implantitis mucositis. C57BL / 6 mice were randomly divided into four groups: a control group treated with sterile saline, a group with peri-implant mucositis treated with *Porphyromonas gingivalis*, a group treated with dezocine maleate, and a group with peri-implant mucositis treated with dezocine maleate. Anesthesia was induced by intraperitoneal injection of sodium pentobarbital (50 mg / kg). The surgical area was disinfected with 0.5% povidone-iodine. After extraction of the right maxillary first molar and local hemostasis, a sterile miniature titanium implant was immediately inserted into the extraction socket using a miniature implant actuator. 5-0 sutures soaked in sterile saline or *Porphyromonas gingivalis* supernatant were ligated to the implant neck, achieving local drug administration through suture infiltration. Experimental mice were randomly divided into a control group, a peri-implant mucositis group (PM), a drug group (MK-801), and a treatment group (PM + MK-801). During the first week post-surgery, the PM and treatment groups received a local injection of 10 μL of *Porphyromonas gingivalis* supernatant at the ligation site every other day, while the control and drug groups received an equal volume of sterile saline. During the second week, the control and PM groups continued to receive saline injections, while the drug and treatment groups received a local injection of 1 mM MK-801 (10 μL) at the ligation site every other day. The modeling period was two weeks. Oral conditions were recorded and photographed at five time points: before tooth extraction, before implant placement, before suture ligation, after suture ligation, and immediately after euthanasia. The modeling period was two weeks. After modeling, mice were sacrificed, and peri-implant tissue was harvested for immunofluorescence experiments and Western blot analysis.

[0024] 3. Experimental Results 3.1 Effects of dezocyclopine maleate on inflammatory manifestations and inflammatory cell recruitment in a mouse model of peri-implantitis mucositis Figure 1This is a diagram showing the clinical manifestations of the peri-implant tissues after implantation of an implant in the right maxillary first molar region of experimental mice, and modeling of mice in the blank group, peri-implant mucositis group, drug group, and treatment group. Figure 2 This is a result of HE staining of buccal and lingual sections of tissue around the implant after implantation of an implant in the right maxillary first molar region of experimental mice, modeling mice in a blank group, a peri-implant mucositis group, a drug group, and a treatment group. In the figure, the blue box area represents the inflammatory cell infiltration in the tissue around the implant threads, and the red box area represents the inflammatory cell infiltration in the tissue at the base of the implant. Figure 3 This image shows the results of masson staining of buccal and lingual sections of tissue surrounding the implant after implantation of an implant in the right maxillary first molar region of experimental mice, modeling of mice in a blank group, peri-implant mucositis group, drug group, and treatment group.

[0025] As shown in Figures 1, 2, and 3, compared with the peri-implant mucositis group, the dezocyclopine maleate intervention group showed significantly improved inflammation in the peri-implant tissues of mice and reduced macrophage recruitment.

[0026] 3.2 Effects of dezocyclopine maleate on protein levels of related inflammatory factors in a mouse model of peri-implantitis mucositis Figure 4 The figure shows the results of Western blot analysis of the protein expression levels of IL-1β, IL-6, and TNF-α in the peri-implant tissue of mice after implantation of an implant in the right maxillary first molar region of experimental mice and establishment of mouse models in blank group, peri-implant mucositis group, drug group, and treatment group.

[0027] like Figure 4 As shown, Western blot experiments revealed that, compared with the peri-implant mucositis group, the protein levels of inflammatory factors such as IL-1β, IL-6, and TNF-α in the peri-implant tissue of mice treated with dezocyclopine maleate were significantly reduced, demonstrating the improvement of inflammatory effects.

[0028] Example 2: Dezocine maleate can regulate the polarization ratio of macrophages, reducing the polarization of pro-inflammatory M1 macrophages and increasing the polarization of anti-inflammatory M2 macrophages. 1. Experimental Materials 1.1 Experimental reagents: Dezocyclopine maleate (MCE Company) 1.2 Experimental animals: C57BL / 6 mice (Jinan Pengyue Experimental Animal Breeding Co., Ltd.) 1.3 Experimental bacteria: Porphyromonas gingivalis 33277 (purchased from the American Center for Type Culture Collection) 2. Experimental Methods 2.1 Model Mice Eight-week-old healthy, clean-grade female C57BL / 6 mice were selected as research subjects. The oral cavity was examined before the experiment to rule out missing teeth, and the right side was selected as the experimental side. All procedures performed on the mice complied with the guidelines for humane treatment of laboratory animals issued by the Ministry of Science and Technology of China (vGKFCZ-2006-398).

[0029] 2.2 Dezocyclopine maleate can regulate the polarization ratio of macrophages and inhibit the effects of peri-implant mucosal inflammation through the Glu / NMDAR / NF-κB signaling pathway. C57BL / 6 mice were randomly divided into four groups: a control group treated with sterile saline, a group with peri-implant mucositis treated with *Porphyromonas gingivalis*, a group treated with dezocine maleate, and a group with peri-implant mucositis treated with dezocine maleate. Anesthesia was induced by intraperitoneal injection of sodium pentobarbital (50 mg / kg). The surgical area was disinfected with 0.5% povidone-iodine. After extraction of the right maxillary first molar and local hemostasis, a sterile miniature titanium implant was immediately inserted into the extraction socket using a miniature implant actuator. 5-0 sutures soaked in sterile saline or *Porphyromonas gingivalis* supernatant were ligated to the implant neck, achieving local drug administration through suture infiltration. Experimental mice were randomly divided into a control group, a periimplantitis group, a drug group, and a treatment group. During the first week post-surgery, the periimplantitis group and the treatment group received a local injection of 10 μL of *Porphyromonas gingivalis* supernatant at the ligation site every other day, while the control group and the drug group received an equal volume of sterile saline. During the second week, the control group and the periimplantitis group continued to receive saline injections, while the drug group and the treatment group received a local injection of 1 mM MK-801 (10 μL) at the ligation site every other day. The modeling period was two weeks. Oral conditions were recorded and photographed at five time points: before tooth extraction, before implant placement, before suture ligation, after suture ligation, and immediately after euthanasia. The modeling period was two weeks. After modeling, mice were sacrificed, and periimplant tissue was harvested for immunofluorescence experiments and Western blot analysis.

[0030] 3. Experimental Results 3.1 Effects of dezocine maleate on macrophage infiltration and recruitment Figure 5 Implants were implanted in the right maxillary first molar region of experimental mice. After establishing mouse models of blank group, peri-implant mucositis group, drug group, and treatment group, the fluorescence images of M1 polarization marker CD86 and M2 polarization marker CD206 were detected by immunofluorescence technology. Figure 6This image shows the results of Western blotting analysis to detect the expression of CD86, CD206, iNOS, and Arg-1 proteins in the peri-implant tissue of experimental mice after implanting an implant in the right maxillary first molar region and establishing mouse models in a blank group, peri-implant mucositis group, drug group, and treatment group.

[0031] like Figure 5 , Figure 6 As shown, Western blotting and immunofluorescence experiments indicated that, compared with the peri-implant mucositis group, dezocyclopine maleate could reduce macrophage infiltration in the peri-implant mucositis group, regulate the reduction of pro-inflammatory M1 macrophage polarization, and increase the polarization of anti-inflammatory M2 macrophage.

[0032] Example 3: Dezocine maleate can affect the release of glutamate, and through the potential Glu / NMDAR / NF-κB pathway, influence the release of inflammatory factors and affect the polarization direction of macrophages. 1. Experimental Materials 1.1 Experimental reagents: Dezocyclopine maleate (MCE Company) 1.2 Experimental animals: C57BL / 6 mice (Jinan Pengyue Experimental Animal Breeding Co., Ltd.) 1.3 Experimental bacteria: Porphyromonas gingivalis 33277 (purchased from the American Center for Type Culture Collection) 1.4 Experimental Cells Mouse RAW264.7 cell line (purchased from Shanghai Cell Bank, Chinese Academy of Sciences) 2. Experimental Methods 2.1 Model Mice Eight-week-old healthy, clean-grade female C57BL / 6 mice were selected as research subjects. The oral cavity was examined before the experiment to rule out missing teeth, and the right side was selected as the experimental side. All procedures performed on the mice complied with the guidelines for humane treatment of laboratory animals issued by the Ministry of Science and Technology of China (vGKFCZ-2006-398).

[0033] 2.2 Dezocyclopine maleate can regulate the polarization ratio of macrophages and inhibit the effects of peri-implant mucosal inflammation through the Glu / NMDAR / NF-κB signaling pathway. C57BL / 6 mice were randomly divided into four groups: a control group treated with sterile saline, a group with peri-implant mucositis treated with *Porphyromonas gingivalis*, a group treated with dezocine maleate, and a group with peri-implant mucositis treated with dezocine maleate. Anesthesia was induced by intraperitoneal injection of sodium pentobarbital (50 mg / kg). The surgical area was disinfected with 0.5% povidone-iodine. After extraction of the right maxillary first molar and local hemostasis, a sterile miniature titanium implant was immediately inserted into the extraction socket using a miniature implant actuator. 5-0 sutures soaked in sterile saline or *Porphyromonas gingivalis* supernatant were ligated to the implant neck, achieving local drug administration through suture infiltration. Experimental mice were randomly divided into a control group, a periimplantitis group, a drug group, and a treatment group. During the first week post-surgery, the periimplantitis group and the treatment group received a local injection of 10 μL of *Porphyromonas gingivalis* supernatant at the ligation site every other day, while the control group and the drug group received an equal volume of sterile saline. During the second week, the control group and the periimplantitis group continued to receive saline injections, while the drug group and the treatment group received a local injection of 1 mM MK-801 (10 μL) at the ligation site every other day. The modeling period was two weeks. Oral conditions were recorded and photographed at five time points: before tooth extraction, before implant placement, before suture ligation, after suture ligation, and immediately after euthanasia. The modeling period was two weeks. After modeling, mice were sacrificed, and periimplant tissue was harvested for immunofluorescence experiments and Western blot analysis.

[0034] 2.3 Cell Model The mouse-derived RAW264.7 macrophage cell line was cultured in Dulbecco modified Eagle medium (DMEM) supplemented with 1% penicillin-streptomycin and 10% fetal bovine serum (FBS). Cells were maintained in a humidified incubator at 37°C and 5% CO2, with the medium changed periodically. Cells were passaged when confluence reached 80-90%. RAW264.7 cells were cultured in multi-well plates and used when confluence reached 70-80%. To synchronize the cell cycle, the basal medium was replaced and the cells were starved of serum for 12 hours before stimulation. Before LPS stimulation, cells were pretreated for 2 hours with 2 mL of DMEM basal medium containing 100 μM MK-801; the control group was pretreated for 2 hours with an equal volume of DMEM basal medium containing 0.1% DMSO (dimethyl sulfoxide, i.e., DMSO solvent control). After 8 hours of treatment, the medium was discarded, and cells were washed three times with pre-cooled PBS before collection for protein or mRNA extraction. The treatment groups were set as follows: 0.1% DMSO (control group), 1 μg / mL LPS, 100 μM MK-801, and 100 μM MK-801 + 1 μg / mL LPS. Cells after stimulation were used for immunofluorescence experiments, polymerase chain reaction (PCR), and Western blot analysis.

[0035] 3. Experimental Results 3.1 Dezocine maleate can affect the release of glutamate in macrophages. Figure 7 This is a graph showing the results of detecting the glutamate concentration in RAW264.7 cells stimulated with LPS and dezocyclopine maleate using a glutamate concentration measurement kit. like Figure 7 As shown in the figure, the glutamate concentration measurement experiment showed that, compared with the inflammation group, dezocyclopine maleate could reduce the release of glutamate in macrophages.

[0036] 3.2 Dezocyclopine maleate can affect the release of inflammatory factors and influence the polarization direction of macrophages. Figure 8 The figure shows the results of detecting the effects of LPS and dezocine maleate on the mRNA and protein expression of IL-1β, IL-6, and TNF-α in RAW264.7 cells using polymerase chain reaction and Western blotting. Figure 9 This is a graph showing the results of detecting IL-10 protein expression in RAW264.7 cells stimulated with LPS and dezocyclopine maleate using immunofluorescence technology. Figure 10 This figure shows the results of Western blotting assays to detect the protein expression of CD86, CD206, iNOS, and Arg-1 in RAW264.7 cells stimulated with LPS and dezocyclopine maleate.

[0037] like Figure 8 , Figure 9 , Figure 10 As shown, compared with the inflammation group, the expression levels of pro-inflammatory cytokines IL-1β, IL-6, and TNF-α decreased and the expression level of anti-inflammatory cytokines IL-10 increased in the treatment group treated with dezocine maleate intervention. Cell experiments also confirmed that dezocine maleate can regulate the reduction of polarization of pro-inflammatory M1 macrophages and the increase of polarization of anti-inflammatory M2 macrophages.

[0038] 3.3 Dezocyclopine maleate can affect the NF-κB pathway Figure 11 This figure shows the results of Western blot analysis of the effect of implantation on NF-κB protein expression in the peri-implant tissue of experimental mice after implantation of an implant in the right maxillary first molar region and establishment of mouse models in blank group, peri-implant mucositis group, drug group, and treatment group. Figure 12 The figure shows the results of Western blotting analysis to detect the effect of LPS and dezocine maleate on NF-κB protein expression in RAW264.7 cells. Figure 13 This image shows the results of detecting NF-κB nuclear translocation in RAW264.7 cells stimulated with LPS and dezocyclopine maleate using immunofluorescence technology.

[0039] like Figure 11 , Figure 12 , Figure 13 As shown, immunofluorescence and Western blotting experiments indicated that, compared with the peri-implant mucositis group, the expression levels of phosphorylated IκB-α and phosphorylated NF-κB proteins in the peri-implant tissue of mice in the dezocyclopine maleate intervention group were significantly reduced, and the establishment of the RAW264.7 cell model also verified this. Moreover, compared with the inflammation group, the nuclear translocation of NF-κB p65 in RAW264.7 cells in the dezocyclopine maleate intervention group was reduced, and the NF-κB pathway was inhibited.

[0040] Those skilled in the art will understand that the above embodiments are specific examples of implementing the present invention, and in practical applications, various changes in form and detail may be made without departing from the spirit and scope of the present invention.

Claims

1. The use of dezocyclopine maleate in the preparation of drugs for the treatment of peri-implantitis.

2. The application according to claim 1, characterized in that, The dezocyclopine maleate antagonizes the N-methyl-D-aspartate receptor on the surface of peri-implant macrophages in mice, inhibiting peri-implant mucosal inflammation induced by Porphyromonas gingivalis in mice.

3. The application according to claim 2, characterized in that, The mice in question were C57BL / 6 mice.

4. The application according to claim 2, characterized in that, Dezocyclopine maleate inhibits macrophage recruitment in mouse peri-implant tissues, regulates the reduction of pro-inflammatory M1 macrophage polarization, the increase of anti-inflammatory M2 macrophage polarization, and the expression of IL-1β, IL-6, TNF-α, IκB-α, and NF-κB.

5. The application according to claim 2, characterized in that, The dezocyclopine maleate has an inhibitory effect on the Glu / NMDAR / NF-κB signaling pathway in a RAW264.7 cell model stimulated by lipopolysaccharide extracted from Porphyromonas gingivalis.

6. A drug for treating peri-implant mucositis, characterized in that, It contains dezocine maleate and pharmaceutically acceptable excipients.

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