塞来昔布抑制凝血酶诱导的大鼠软骨细胞退变  

Celecoxib inhibits thrombin-induced chondrocyte degeneration in rats

作　　者：朱志恒 丁佳滢 葛杨硕 黄春萌 沈军 王学宗[4] 郑昱新[4] 丁道芳 Zhu Zhiheng;Ding Jiaying;Ge Yangshuo;Huang Chunmeng;Shen Jun;Wang Xuezong;Zheng Yuxin;Ding Daofang(School of Rehabilitation Science,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China;Institute of Rehabilitation Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China;Shang Guanghua Hospital of Integrated Traditional Chinese and Western Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 200052,China;Shi’s Medical Center for Injuries,Shuguang Hospital,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China)

机构地区：[1]上海中医药大学康复医学院,上海市201203 [2]上海中医药大学康复医学研究所,上海市201203 [3]上海中医药大学附属光华医院,上海市200052 [4]上海中医药大学附属曙光医院石氏伤科医学中心,上海市201203

出　　处：《中国组织工程研究》2024年第34期5446-5451,共6页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(82174406),项目负责人:王学宗;国家自然科学基金项目(81902306),项目负责人:丁道芳;上海中医药大学科技发展项目(23KFL023),项目负责人:丁道芳。

摘　　要：背景:骨关节炎患者血清中凝血酶含量显著高于正常人,且凝血酶可以诱导大鼠软骨细胞退变,提示抑制凝血酶功能可能成为治疗骨关节炎的一种方法。塞来昔布为临床治疗骨关节炎的常见药物,是否可以抑制凝血酶的促炎作用而改善大鼠软骨细胞退变尚未可知。目的:探讨塞来昔布对凝血酶诱导软骨细胞退变的影响。方法:使用ELISA试剂盒检测骨关节炎患者与正常人血清中凝血酶含量。体外提取、培养SD乳鼠关节软骨细胞,使用第一代细胞进行实验,将软骨细胞分对照组、凝血酶组和塞来昔布组。显微镜下观察3组细胞形态,并使用Edu试剂盒检测细胞增殖情况;qRT-PCR检测软骨细胞外基质成分(聚集蛋白聚糖、弹性蛋白、软骨寡聚基质蛋白)、炎症因子(白细胞介素1、白细胞介素6、肿瘤坏死因子α)、炎症趋化因子(单核细胞趋化蛋白2、单核细胞趋化蛋白7、粒细胞趋化蛋白6)的表达;免疫荧光法检测Ⅱ型胶原α1的表达;Western Blot检测分解代谢基因如基质金属蛋白酶9、基质金属蛋白酶13和环氧化酶2等蛋白的表达。结果与结论:(1)与正常人相比,骨关节炎患者血清中凝血酶含量显著增高;(2)塞来昔布可明显抑制凝血酶诱导的软骨细胞纤维样形态改变;(3)与凝血酶组相比,塞来昔布抑制了软骨细胞的增殖;(4)凝血酶组Ⅱ型胶原α1等软骨细胞外基质基因表达的下调被塞来昔布抑制(P <0.05);(5)凝血酶可促进软骨细胞炎症因子(白细胞介素1、白细胞介素6、肿瘤坏死因子α)、炎症趋化因子(单核细胞趋化蛋白2、单核细胞趋化蛋白7、粒细胞趋化蛋白6)以及分解代谢基因(基质金属蛋白酶9、基质金属蛋白酶13和环氧化酶2)的表达,在塞来昔布干预下,上述基因的表达下调(P <0.05);(6)结果提示,塞来昔布可以抑制凝血酶的促炎作用,从而改善大鼠软骨细胞退变。BACKGROUND:The content of serum thrombin in patients with osteoarthritis is significantly higher than that in normal individuals,and thrombin can induce inflammatory degeneration of rat chondrocytes,suggesting that inhibiting the function of thrombin may become a method for treating osteoarthritis.Celecoxib is a common therapeutic drug for the clinical treatment of osteoarthritis.It is not yet known whether it improves chondrocyte degeneration by inhibiting the activity of thrombin.OBJECTIVE:To investigate the effect of celecoxib on thrombin-induced degeneration of rat chondrocytes.METHODS:Thrombin levels in the serum of osteoarthritis patients and normal individuals were detected by an ELISA kit.Primary chondrocytes of neonatal Sprague-Dawley rats were isolated,and all experiments were performed with cells from passage one.Chondrocytes were randomly divided into three groups:control group,thrombin group,and celecoxib group.The cell morphology of the three groups was observed under an inverted microscope,and an Edu kit was used to detect the cell proliferation.qRT-PCR was used to detect the expression of extracellular matrix components(aggrecan,elastin,cartilage oligomeric matrix proteins),inflammatory factors(interleukin-1,interleukin-6,and tumor necrosis factor-α),and chemokines(monocyte chemotactic protein 2,monocyte chemotactic protein 7,granulocyte chemotactic protein 6).The expression of type 2 collagenα1 was detected by immunofluorescence.Western blot method was used to detect the expression of catabolic metabolism genes,such as matrix metalloproteinase 9,matrix metalloproteinase 13,and cyclooxygenase 2.RESULTS AND CONCLUSION:Patients with osteoarthritis had higher levels of thrombin in the serum compared with normal individuals.Under the microscope,celecoxib was found to significantly inhibit fibroid changes in chondrocytes.Compared with the thrombin group,celecoxib inhibited the proliferation of chondrocytes.The downregulation of extracellular matrix gene expression,such as type II collagenα1,in the t

关 键 词：软骨细胞 凝血酶 塞来昔布 细胞增殖 细胞退变 

分 类 号：R459.9[医药卫生—治疗学] R318[医药卫生—临床医学] R496