大麻二酚联合米诺环素对牙周炎治疗作用的实验研究

Therapeutic effect of cannabidiol combined with minocycline on periodontitis

目的通过体内外实验探索大麻二酚(CBD)联合米诺环素(MINO)对小鼠实验性牙周炎的治疗作用。方法丝线结扎法建立小鼠牙周炎模型,口腔局部分别给予CBD、MINO及二者联用,通过定量反转录聚合酶链反应(qRT-PCR)检测牙周炎小鼠用药后外周血炎症因子变化,Micro CT、免疫印迹(WB)及苏木精-伊红(HE)染色分析局部位点炎症改善情况;牙龈卟啉单胞菌抑菌实验检测联用药物体外抗菌性能;体外诱导巨噬细胞及牙龈成纤维细胞炎症,通过qRT-PCR检测炎症相关mRNA表达,划痕实验探索药物对细胞迁移的影响,并通过酶联免疫吸附(ELISA)测定探究药物对于胶原生成的作用。结果联用CBD与MINO后,与单独用药相比,小鼠全身炎症指标下降,局部附着丧失减轻、组织炎症水平降低、牙周软硬组织破坏得到改善;体外抑菌实验结果表明联用药物表现出良好的抗菌性能;体外细胞实验证明联用药物降低巨噬细胞多种炎症因子的表达水平,促进炎症状态下人牙龈成纤维细胞增殖、迁移,提升胶原形成功能。结论相较单一用药方式法,局部应用CBD联合MINO对小鼠实验性牙周炎有着显著疗效。

Objective The therapeutic effect of cannabidiol combined with minocycline on experimental periodontitis in mice was studied in vitro and in vivo.Methods The mouse periodontitis model was established by thread ligation.The oral area was treated with cannabidiol,minocycline,and both.Quantitative reverse transcription polymerase chain reaction(qRT-PCR)was used to detect the changes in inflammatory factors in peripheral blood of mice with periodontitis.Micro CT,western blot(WB),and hematoxylin-eosin(HE)staining were used to analyze the improvement in local inflammation.Porphyromonas gingivalis(P.gingivalis)bacteriostatic assay was used to detect the antibacterial properties of the combined drugs in vitro.Inflammation of macrophages and gingival fibroblasts was induced in vitro,and mRNA expression related to inflammation was detected by qRTPCR.The effect of drugs on cell migration was explored by wound-healing assay,and the effect of drugs on collagen production was explored by enzyme-linked immunosorbent assay.Results After combined administration of cannabidiol and minocycline,the systemic in‐flammation index of mice decreased,the local attachment loss was alleviated,the tissue inflammation level was reduced,and the destruction of periodontal soft and hard tissues improved compared with single administration.The results of the antibacterial experiment in vitro showed that the combined drugs had good antibacterial properties.In vitro cell experiments proved that the combination of drugs reduced the expression levels of various inflammatory factors in macrophages,promoted the proliferation and migration of human gingival fibroblasts in inflammatory states,and improved the collagen formation function.Conclusion Compared with the single medication method,the local application of cannabidiol combined with minocycline has a significant effect on experimental periodontitis in mice.