金丝桃苷抑制Toll样受体4/髓样分化因子88/核因子-κB 信号通路减轻牙周炎大鼠牙周组织损伤实验研究

Hyperoside inhibits TLR4/MyD88/NF-κB signaling pathway to reduce periodontal tissue damage in rats with periodontitis

目的:探究金丝桃苷调控Toll样受体4(TLR4)/髓样分化因子88(MyD88)/核因子-κB(NF-κB)信号通路对牙周炎大鼠牙周组织损伤的影响。方法:构建牙周炎大鼠模型,建模成功大鼠随机分为模型组、金丝桃苷(30 mg/kg)组、金丝桃苷(30 mg/kg)+TLR4激活剂(脂多糖,0.4 mg/kg)组,每组15只,另取15只健康大鼠作为对照组,建模结束后,各组大鼠按对应方式给药,1次/d,连续4周。HE染色及抗酒石酸酸性磷酸酶染色分别观察各组大鼠牙周组织病理变化及破骨细胞数量;ELISA法检测各组大鼠血清骨保护素(OPG)、NF-κB受体活化因子配体(RANKL)、肿瘤坏死因子-α(TNF-α)、白介素-6(IL-6)水平;荧光定量PCR法检测各组大鼠牙周组织TLR4、MyD88、NF-κB mRNA表达水平;蛋白印迹法检测各组大鼠牙周组织TLR4、MyD88、NF-κB蛋白表达水平。结果:对照组大鼠牙周组织结构正常;与对照组相比,模型组大鼠牙周组织炎性细胞浸润明显,伴随骨吸收陷窝数量增多,牙周组织破骨细胞数量、TLR4、MyD88、NF-κB mRNA和蛋白表达水平、血清RANKL、TNF-α、IL-6水平显著升高(均P<0.05),血清OPG水平显著降低(P<0.05);与模型组相比,金丝桃苷组大鼠牙龈上皮较完整,炎性细胞浸润程度较轻,骨吸收陷窝减少,牙周组织破骨细胞数量、TLR4、MyD88、NF-κB mRNA和蛋白表达水平、血清RANKL、TNF-α、IL-6水平显著降低(均P<0.05),血清OPG水平显著升高(P<0.05);与金丝桃苷组相比,金丝桃苷+TLR4激活剂组大鼠牙周组织炎性细胞浸润程度加深,骨吸收陷窝增多,牙周组织破骨细胞数量、TLR4、MyD88、NF-κB mRNA和蛋白表达水平、血清RANKL、TNF-α、IL-6水平显著升高(均P<0.05),血清OPG水平显著降低(P<0.05)。结论:金丝桃苷可缓解牙周炎大鼠牙周组织损伤,抑制大鼠炎性反应,其机制与抑制TLR4/MyD88/NF-κB信号通路有关。

Objective:To investigate the effect of hyperoside on periodontal tissue damage in rats with periodontitis by regulating Toll-like receptor 4(TLR4)/myeloid differentiation factor 88(MyD88)/nuclear factor-κB(NF-κB)signaling pathway.Methods:The rat model of periodontitis was established,and the successful model rats were randomly divided into model group,hyperoside(30 mg/kg)group,and hyperoside(30 mg/kg)+TLR4 activator(lipopolysaccharide,0.4 mg/kg)group,with 15 in each group.Another 15 healthy rats were selected as control group.Following the modeling procedure,the corresponding method was administered to each group of rats once daily for a duration of 4 weeks.HE staining and tartrate-resistant acid phosphatase staining were used to observe the pathological changes of periodontal tissue and the number of osteoclasts in each group.The serum levels of osteoprotegerin(OPG),receptor activator of NF-κB ligand(RANKL),tumor necrosis factor-α(TNF-α)and interleukin-6(IL-6)were detected by ELISA.The mRNA levels of TLR4,MyD88 and NF-κB in periodontal tissues were detected by fluorescence quantitative PCR method.Western blot analysis was employed to determine the protein expression levels of TLR4,MyD88,and NF-κB in periodontal tissues of rats.Results:The periodontal tissue architecture of the control group rats exhibited normalcy.Compared with control group,the model group exhibited obvious inflammatory cell infiltration in periodontal tissue,accompanied by an increase in the number of bone resorption lacunae,and the number of osteoclasts,the mRNA and protein levels of TLR4,MyD88,and NF-κB in the periodontal tissue,and the serum levels of RANKL,TNF-α,and IL-6 were significant increased,and the serum OPG level was significantly decreased(all P<0.05).Compared with model group,the gingival epithelium of the hyperoside group was more intact,the degree of inflammatory cell infiltration was lighter,and the bone resorption lacunae were reduced,the number of osteoclasts in periodontal tissue,the mRNA and protein levels of TLR4,MyD88,and NF-κB,and the serum levels of RANKL,TNF-α,and IL-6 were significantly decreased,while the serum level of OPG was significantly increased(all P<0.05).Compared with the hyperoside group,the hyperoside+TLR4 activator group had inflammatory cell infiltration in periodontal tissue,bone resorption lacunae,the number of osteoclasts,the mRNA and protein levels of TLR4,MyD88,and NF-κB in periodontal tissue,and serum RANKL,TNF-α,and IL-6 levels were significantly increased,and the serum OPG level was significantly decreased(all P<0.05).Conclusion:Hyperoside can alleviate periodontal tissue damage and inhibit inflammatory response in rats with periodontitis,and the mechanism is related to the inhibition of TLR4/MyD88/NF-κB signaling pathway.