摘要
背景:脱细胞软骨细胞外基质治疗骨关节炎具有一定的效果,但其单独应用时的治疗效果有限。氧化苦参碱可缓解白细胞介素1β诱导的软骨细胞凋亡和细胞外基质降解。目的:观察不同剂量氧化苦参碱与脱细胞软骨细胞外基质联合膝关节腔注射对大鼠骨关节炎的治疗作用。方法:获取SD大鼠股骨软骨,采用物理、化学与酶相结合的方法制备脱细胞软骨细胞外基质。取50只SD大鼠,利用随机数字表法分为假手术组、骨关节炎组、单纯材料组、低剂量药物组、高剂量药物组,每组10只,后4组采用改良Hulth法建立大鼠骨关节炎模型。造模成功后,单纯材料组大鼠膝关节腔注射脱细胞软骨细胞外基质匀浆,低剂量药物组、高剂量药物组膝关节腔分别注射含50,100μg氧化苦参碱的脱细胞软骨细胞外基质匀浆。注射4周后取材进行相关检测。结果与结论:①与假手术组比较,骨关节炎组关节腔积液中白细胞介素1β、肿瘤坏死因子ɑ、丙二醛水平升高(P<0.05),超氧化物歧化酶水平降低(P<0.05);与骨关节炎组比较,低剂量药物组、高剂量药物组关节腔积液中白细胞介素1β、肿瘤坏死因子ɑ、丙二醛水平降低(P<0.05),超氧化物歧化酶水平升高(P<0.05),其中高剂量药物组变化更显著。②Tunel染色显示,与假手术组相比,骨关节炎组凋亡软骨细胞增多;与骨关节炎组相比,单纯材料组、低剂量药物组、高剂量药物组凋亡软骨细胞减少,其中高剂量药物组减少更显著。③苏木精-伊红与免疫组化染色显示,骨关节炎组大鼠膝关节软骨严重退变,软骨组织中Ⅱ型胶原表达降低(P<0.05)、基质金属蛋白酶9表达升高(P<0.05);与骨关节炎组相比,单纯材料组、低剂量药物组、高剂量药物组大鼠膝关节软骨退变明显改善,软骨组织中Ⅱ型胶原表达升高(P<0.05)、基质金属蛋白酶9表达降低(P<0.05),其中高剂量药物组改善最显著。④Western blot检测显示,与假手术组相比,骨关节炎组软骨组织中Nrf2、HO-1、Bcl-2蛋白表达降低(P<0.05),Cleaved-caspase-3、Cleaved-caspase-9、Bax蛋白表达升高(P<0.05);与骨关节炎组相比,低剂量药物组、高剂量药物组Nrf2、HO-1、Bcl-2蛋白表达升高(P<0.05),Cleaved-caspase-3、Cleaved-caspase-9、Bax蛋白表达降低(P<0.05),其中高剂量药物组改善更显著。⑤结果表明,膝关节腔内联合注射脱细胞软骨细胞外基质与氧化苦参碱可通过促进细胞外基质的合成、抑制炎症与氧化应激反应、抑制软骨细胞凋亡发挥骨关节炎治疗作用,该治疗作用可能与激活Nrf2/HO-1通路有关。
BACKGROUND:Acellular cartilage extracellular matrix is effective in the treatment of osteoarthritis,but its efficacy is limited when applied alone.Oxymatrine alleviates interleukin-1β-induced chondrocyte apoptosis and extracellular matrix degradation.OBJECTIVE:To observe the effect of different doses of oxymatrine combined with acellular cartilage extracellular matrix injection in knee cavity on osteoarthritis in rats.METHODS:The femoral cartilage of SD rats was obtained.The acellular cartilage extracellular matrix was prepared by physical,chemical and enzyme methods.Fifty SD rats were selected and divided into sham operation group,osteoarthritis group,simple material group,low-dose drug group,and high-dose drug group by random number table method,with 10 rats in each group.The latter four groups were treated with modified Hulth method to establish the rat model of osteoarthritis.After successful modeling,acellular cartilage extracellular matrix homogenate was injected into the knee cavity of rats in the simple material group.Acellular cartilage extracellular matrix homogenate containing 50,100μg oxymatrine was injected into the knee cavity of rats in the low-dose drug group and the high-dose drug group,respectively.Samples were taken 4 weeks after injection for relevant detection.RESULTS AND CONCLUSION:(1)Compared with the sham operation group,the concentrations of interleukin 1β,tumor necrosis factorɑ,and malondialdehyde in the joint effusion were increased(P<0.05),and the concentration of superoxide dismutase in the joint effusion was decreased(P<0.05)in the osteoarthritis group.Compared with osteoarthritis group,the levels of interleukin 1β,tumor necrosis factorɑ,and malondialdehyde in joint effusion were decreased(P<0.05),while the level of superoxide dismutase was increased(P<0.05)in the low-dose drug group and high-dose drug group,and the changes were more significant in high-dose drug group.(2)TUNEL staining showed that compared with sham operation group,apoptotic chondrocytes increased in osteoarthritis group.Compared with the osteoarthritis group,the apoptotic chondrocytes decreased in the simple material group,the low-dose drug group,and the high-dose drug group,and the decrease was more significant in the high-dose drug group.(3)Hematoxylin-eosin staining and immunohistochemical staining showed that the knee cartilage was seriously degraded,the expression of type II collagen was decreased(P<0.05),and the expression of matrix metalloproteinase-9 was increased(P<0.05)in the osteoarthritis group.Compared with the osteoarthritis group,the knee cartilage degeneration of rats in the simple material group,the low-dose drug group,and the high-dose drug group was significantly improved;the expression of type II collagen was increased(P<0.05)and the expression of matrix metalloproteinase 9 was decreased(P<0.05),and the improvement was most significant in the high-dose drug group.(4)Western blot assay showed that compared with sham operation group,the expression of Nrf2,HO-1,and Bcl-2 protein in cartilage tissue decreased(P<0.05);expression levels of Cleaved-caspase-3,Cleaved-caspase-9,and Bax were increased(P<0.05)in the osteoarthritis group.Compared with the osteoarthritis group,the protein expression levels of Nrf2,HO-1,and Bcl-2 were increased(P<0.05),and the protein expressions of Cleaved-caspase-3,Cleaved-caspase-9,and Bax were decreased(P<0.05)in the low-dose and high-dose drug groups.The improvement was more significant in the high-dose drug group.(5)In conclusion,intracavicular injection of acellular cartilage extracellular matrix and oxymatrine can promote the synthesis of extracellular matrix,inhibit inflammation and oxidative stress,and suppress chondrocyte apoptosis,and play a therapeutic role in osteoarthritis,which may be related to the activation of Nrf2/HO-1 pathway.
作者
吴富章
张鹏礼
章振华
何永兵
孙和炎
Wu Fuzhang;Zhang Pengli;Zhang Zhenhua;He Yongbing;Sun Heyan(Department of Orthopedics,Anhui Provincial Corps Hospital of Chinese Peoples Armed Police Force,Hefei 230041,Anhui Province,China;Department of Orthopedics,First Affiliated Hospital of Anhui Medical University,Hefei 230001,Anhui Province,China)
出处
《中国组织工程研究》
CAS
北大核心
2025年第13期2675-2682,共8页
Chinese Journal of Tissue Engineering Research
基金
安徽省“十三五”医疗卫生重点专科建设项目(皖卫科教秘[2021]230号),项目负责人:吴富章
安徽省自然科学基金项目计划(2008085MH291),项目负责人:孙和炎。
