连翘昔对ACLT诱导的骨关节炎大鼠模型软骨组织损伤和TGF-β活性的调节作用

Regulatory Effect of Forsythin on Cartilage Tissue Damage and TGF-β Protein Activity in Rats with ACLT-induced Osteoarthritis

目的以前交叉韧带切断术(anterior cruciate ligament transection, ACLT)诱导的骨关节炎(osteoarthritis,0A)大鼠为模型,研究连翘昔(forsythin )对软组织损伤和转化生长因子β( transforminggrowth factorβ, TGF-β)活性的影响、方法将20只SD雄性大鼠随机分成4组:健康对照组(Ctrl)、连翘昔单独加药组(Forsythin)、模型组(OA model)和模型加药组(OA + Forsythin),每组5只。膝前交叉韧带切断建立骨关节炎模型。按50mg/kg的剂量,腹腔注射连翘昔,1次/d,共4周。苏木伊红染色(hematoxylinand eosin, HE)观察软骨损伤;番红ο染色和马森三色染色(MASSON)观察软组织损伤和纤维增生情况蛋白质印迹检测基质金属蛋白酶13 (matrix metalloproteinase 13, MMP-13)、Ki67和半胱氨酸天冬氨酸蛋白酶 3(cysteinyl aspartate specific proteinase 3 , Caspase-3)的表达。酶联免疫吸附法(enzymelinked immunosorbent assay, ELISA)检测炎性因子肿瘤坏死因子 a ( tumor necrosis factor α, TNF-α、IL-6、IL-10和诱导性一氧化氮舎酶(induced-Nitric oxide synthase, iNOS)的表达。蛋白质印迹检测TGF-β1、SMAD2和P-SMAD2的表达.结果模型组与对照组相比,软骨组织出现明显损伤,纤维增多;MMP-13和Caspase-3的表达显著上调,Ki67的表达显著下调;TNF-α、IL-6和iNOS的含量显著上升,11,-10的含量没有明显变化;TGF-β1表达显著上调,p-Smad2/Smad2比率明显上升。模型加药组与模型组相比,软骨组织损伤明显减轻,纤维化明显改善;MMP-13和Caspase-3的表达显著下调,Ki67的表达显著上调;TNF-α,IL-6和iNOS的含量显著降低,IL-10的含量没有明显变化;TGF-β1表达显著下调,p-Smad2/Smad2比率明显降低结论连翘昔缓解ACLT诱导的骨关节炎大鼠模型软骨组织损伤,调节TGF-β的活性。

Objective To study the effect of forsythin on the soft tissue damage and the activity of transforming growth factor beta in rats with osteoarthritis induced by anterior cruciate ligament transaction ( ACLT). Methods 20 male SD rats were randomly divided into 4 groups: healthy control group ( Ctrl), forsythia solo alone group ( Forsythin ), model group ( OA model ) and model treatment group ( OA + Forsythin), with 5 rats in each group. Osteoarthritis model was established by cutting off the anterior cruciate ligament. Intraperitoneal injection of 50mg/kg BW forsythin was given once a clay for 4 weeks. The cartilage damage was observed by using hematoxylin and eosin staining, and the damage of soft tissue and fibrous hyperplasia by safranine 0 staining and MASON staining. The expression of matrix metalloproteinase 13 , Ki67 and cysteine aspartic proteinase 3 was detected by Western blotting. Enzyme linked immunosorbent assay ( ELISA ) was used to detect tumor necrosis factor alpha, interleukin 6 , interleukin 10 and inducible nitric oxide synthase expression. Western blotting was used to detect the expression of TGF-beta1 , SMAD2 AND PSMAD2. Results Compared with the control group, the cartilage tissue was obviously damaged and the fiber number increased in the OA group. The expression of MMP-13 and Caspase-3 was up significantly;the expression of Ki67 was significantly down-regulated;the content of TNF-alpha , IL-6 and iNOS were increased significantly, the content of IL-10 was not significantly changed;the expression of TGF-beta1 was significantly up-regulated , and the p-Smad2/Smad2 ratio was significantly increased. Compared with the OA model group, in the OA + Forsythin group, the injury of cartilage tissue was obviously reduced and the fibrosis was obviously improved;the expression of MMP-13 and Caspase-3 was significantly down-regulated;the expression of Ki67 was significantly up-regulated;the content of TNF-alpha, IL-6 and iNOS decreased significantly;the content of IL-10 was not significantly changed;the expression of TGF-beta1 was significantly down-regulated, and the p-Smad2/Smad2 ratio was decreased significantly. Conclusion Forsythin alleviates carti lage damage and regulates TGF-beta activity in ACLT-induced osteoarthritis rats.