摘要
背景:Wnt信号转导的活化与膝骨关节炎的发病机制密切相关,当前对Wnt信号通路的研究侧重于Wnt/β-catenin信号通路及其激活或抑制蛋白的表达。目的:观察院内协定处方骨痹散对膝骨关节炎模型兔体内炎症因子与Notch3/Hes1及Wnt/β-catenin信号通路的影响,为临床治疗提供新的用药思路。方法:将40只新西兰大白兔随机分为假手术组、模型组、阳性组与骨痹散组,每组10只,使用改良Hulth法构建兔膝骨关节炎模型,持续造模8周,模型构建完毕后假手术组、模型组膝骨关节处涂抹生理盐水,阳性组涂抹双氯芬酸钾凝胶,骨痹散组涂抹骨痹散,每天敷8 h,连续1周。给药完毕后观察动物活动情况并进行行为学评分,ELISA法检测血清中前列腺素E2、环氧化酶2、白细胞介素1β与肿瘤坏死因子α水平,苏木精-伊红染色与番红O染色观察软骨组织病变情况,免疫组织化学法检测膝关节软骨组织中基质金属蛋白酶1,2,3,13蛋白的表达水平,Western blot法检测膝关节软骨组织中Notch3、Hes1、Wnt5a、β-catenin、Bcl-2、Bax、Caspase-3的蛋白表达水平。结果与结论:①与假手术组相比,模型组动物行走困难,膝关节软骨组织损伤严重,血清中前列腺素E2、环氧化酶2、白细胞介素1β与肿瘤坏死因子α水平升高(P<0.05),软骨组织中基质金属蛋白酶1,2,3,13和Bax、Caspase-3、Wnt5a、β-catenin蛋白表达升高(P<0.05),Notch3、Hes1、Bcl-2蛋白表达降低(P<0.05);②与模型组相比,骨痹散组动物行走情况良好,血清中前列腺素E2、环氧化酶2、白细胞介素1β与肿瘤坏死因子α水平降低(P<0.05),膝关节软骨组织损伤减轻,软骨组织中基质金属蛋白酶1,2,3,13和Bax、Caspase-3、Wnt5a、β-catenin蛋白表达降低(P<0.05),Notch3、Hes1、Bcl-2蛋白表达升高(P<0.05);③结果表明,院内协定处方骨痹散能减轻兔膝骨关节炎的症状,改善软骨组织病变,减少血清中炎症因子水平,作用机制可能与调控Notch3/Hes1、Wnt/β-catenin信号通路有关。
BACKGROUND:Activation of Wnt signaling is closely related to the pathogenesis of knee osteoarthritis.Current research on the Wnt signaling pathway has focused on the Wnt/β-catenin signaling pathway and the expression of its activation or inhibition proteins.OBJECTIVE:To observe the effect of Gubi Powder,an in-hospital prescription,on inflammatory factors and Notch3/Hes1 and Wnt/β-catenin signaling pathways in rabbits with knee osteoarthritis model,providing new drug ideas for clinical treatment.METHODS:Forty New Zealand rabbits were divided into sham operation group,model group,positive group(diclofenac potassium gel)and Gubi Powder group,with 10 rabbits in each group.The modified Hulth method was used to construct the rabbit knee osteoarthritis model,which lasted for 8 weeks.After modeling,normal saline was applied to the knee joints of the sham operation group and model group,diclofenac potassium gel was applied to the positive group,and Gubi Powder was applied to the Gubi Powder group,8 hours a day,for 1 week.After administration,animal activity and behaviors were evaluated and scored;serum levels of prostaglandin E2,cyclooxygenase-2,interleukin 1βand tumor necrosis factorαwere determined by ELISA;pathological changes of cartilage tissue were observed by hematoxylin-eosin staining and Safranin-O staining;protein levels of matrix metalloproteinases 1,2,3,and 13 in cartilage tissue of the knee joint were determined by immunohistochemistry;and protein levels of Notch3,Hes1,Wnt5a,β-catenin,Bcl-2,Bax,and Caspase-3 in cartilage tissue of the knee joint were determined by western blot assay.RESULTS AND CONCLUSION:Compared with the sham operation group,animals in the model group had difficulties in walking,serious cartilage injury in the knee joint,significantly elevated levels of serum prostaglandin E2,cyclooxygenase 2,interleukin 1βand tumor necrosis factorα(P<0.05),significantly increased protein expression levels of matrix metalloproteinases 1,2,3,and 13,Bax,Caspase-3,Wnt5a andβ-catenin in cartilage tissue(P<0.05),and decreased protein expression levels of Notch3,Hes1,and Bcl-2(P<0.05).Compared with the model group,the walking condition of animals in the positive group and Gubi Powder group was improved,the serum levels of prostaglandin E2,cyclooxygenase-2,interleukin 1βand tumor necrosis factorαwere decreased,cartilage damage of the knee was alleviated,the protein expression levels of matrix metalloproteinases 1,2,3,and 13,Bax,Caspase-3,Wnt5a andβ-catenin in cartilage tissue were reduced(P<0.05),and the protein expression levels of Notch3,Hes1,and Bcl-2 were increased(P<0.05).To conclude,Gubi Powder can alleviate the symptoms of knee osteoarthritis in rabbits,improve the pathological changes of the cartilage,and reduce inflammatory factor levels in serum.The mechanism of action may be related to the inhibition of Notch3/Hes1 and Wnt/β-catenin signaling pathways.
作者
李东东
陈光友
李晓明
吴雪莲
朱凯
雷杨
易露
Li Dongdong;Chen Guangyou;Li Xiaoming;Wu Xuelian;Zhu Kai;Lei Yang;Yi Lu(Department of Orthopedics and Traumatology,Affiliated Hospital of Traditional Chinese Medicine,Southwest Medical University,Luzhou 646000,Sichuan Province,China;Department of Rehabilitation Medicine,Southwest Medical University,Luzhou 646000,Sichuan Province,China)
出处
《中国组织工程研究》
CAS
北大核心
2023年第35期5622-5627,共6页
Chinese Journal of Tissue Engineering Research
基金
西南医科大学附属中医医院课题(2018XYLH-019),项目负责人:陈光友。
