温针灸抑制NLRP3炎症小体激活改善兔膝骨关节炎的软骨损伤  

Warm-needling moxibusion inhibits NLRP3 inflammasome activation and improves cartilage injury in a rabbit model of knee osteoarthritis

作　　者：武永利[1,2] 李龙 刘君伟 刘娣[1] 王铎 Wu Yongli;Li Long;Liu Junwei;Liu Di;Wang Duo(Department of Orthopedics and Traumatology of Traditional Chinese Medicine,General Hospital of Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China;Key Laboratory of Ningxia Ethnomedicine Modernization,Ministry of Education,Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China;School of Traditional Chinese Medicine,Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China)

机构地区：[1]宁夏医科大学总医院中医骨伤科,宁夏回族自治区银川市750004 [2]宁夏医科大学,少数民族医药现代化教育部重点实验室,宁夏回族自治区银川市750004 [3]宁夏医科大学,中医学院,宁夏回族自治区银川市750004

出　　处：《中国组织工程研究》2023年第20期3202-3208,共7页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(82160939,81760891),项目负责人:武永利;宁夏回族自治区第二批科技领军人才项目(KJT201610),项目负责人:武永利。

摘　　要：背景:温针灸治疗膝骨关节炎疗效显著,但其作用机制尚不明确。目的:观察温针灸对兔膝骨关节炎软骨组织中NLRP3、基质金属蛋白酶3、基质金属蛋白酶13、Ⅰ型胶原蛋白表达的影响。方法:取6月龄新西兰雄兔40只,采用随机数字表法分成空白组、模型组、温针灸组、抑制剂组,每组10只:除空白组外,其他3组均采用石膏管型固定法建立右后肢膝骨关节炎模型。造模成功后,温针灸组进行温针灸治疗,1次/d,连续治疗4周;抑制剂组膝关节腔内注射NLRP3抑制剂MCC950,1次/周,注射4次。干预结束后,采用Lequesne MG评分评估兔行为学改变,苏木精-伊红染色观察软骨形态学结构,免疫组织化学染色及蛋白质印迹法检测软骨组织中NLRP3、基质金属蛋白酶3、基质金属蛋白酶13、Ⅰ型胶原蛋白的表达。结果与结论:(1)Lequesne MG评分:模型组Lequesne MG评分高于空白组(P<0.05),温针灸组、抑制剂组Lequesne MG评分低于模型组(P<0.05);(2)苏木精-伊红染色:与空白组比较,模型组软骨细胞数量减少,结构完整性被破坏,Mankin’s评分升高(P<0.05);与模型组比较,温针灸组、抑制剂组软骨细胞分布较均匀,软骨层结构相对清晰,Mankin’s评分降低(P<0.05);(3)免疫组织化学与蛋白质印迹检测:与空白组比较,模型组NLRP3、基质金属蛋白酶3、基质金属蛋白酶13表达升高(P<0.05),Ⅰ型胶原蛋白表达降低(P<0.05);与模型组比较,温针灸组与抑制剂组NLRP3、基质金属蛋白酶3、基质金属蛋白酶13表达降低(P<0.05),Ⅰ型胶原蛋白表达升高(P<0.05);(4)结果表明:温针灸可抑制NLRP3炎性启动因子的激活、下调炎性因子基质金属蛋白酶3,13的释放,降低炎性放大反应,减缓Ⅰ型胶原蛋白的降解,降低膝骨关节炎的关节软骨损伤及局部破坏。BACKGROUND:Warm-needling moxibusion has a significant clinical effect on knee osteoarthritis,but its therapeutic mechanism remains unclear.OBJECTIVE:To observe the effect of warm-needling moxibusion on the expression of NLRP3,matrix metalloproteinases 3,13,and typeⅠcollagen in cartilage tissue of rabbit knee osteoarthritis.METHODS:Forty 6-month-old male New Zealand rabbits were randomly divided into blank group,model group,warm-needling moxibusion group,and NLRP3 inhibitor group,with 10 rabbits in each group.Except for the blank group,osteoarthritic models of the right hind limb were established in the other three groups using plaster cast fixation.After modeling,intervention with warm-needling moxibusion in the warm-needling moxibusion group was performed once a day and lasted for 4 weeks;in the NLRP3 inhibitor group,MCC950,an NLRP3 inhibitor,was injected into the knee joint cavity,once a week,for four times in total.Lequesne MG score was used for behavioral evaluation,hematoxylin-eosin staining was used to observe the morphological structure of cartilage tissue,and the expression of NLRP3,matrix metalloproteinases 3,13,and typeⅠcollagen in chondrocytes was detected by immunohistochemistry and western blot.RESULTS AND CONCLUSION:Lequesne MG score:Compared with the blank group,Lequesne MG score increased in the other three groups after modeling(P<0.05).After treatment,the Lequesne MG scores were significantly decreased in the warm-needling moxibusion group and NLRP3 inhibitor group compared with the model group(P<0.05).Hematoxylin-eosin staining:compared with the blank group,the number of chondrocytes was decreased,the structural integrity was destroyed,and the Mankin’s score was significantly increased in the model group(P<0.05).Compared with the model group,the distribution of chondrocytes was more uniform,the structure of cartilage layer was relatively clear,and the Mankin’s score was decreased in the warm-needling moxibusion group and NLRP3 inhibitor group(P<0.05).Immunohistochemistry and western blot

关 键 词：温针灸 膝骨关节炎 NLRP3炎症小体 软骨损伤 基质金属蛋白酶 

分 类 号：R459.9[医药卫生—治疗学] R319[医药卫生—临床医学] R245