防风石油醚提取物调节中性粒细胞胞外诱捕网对类风湿性关节炎大鼠的改善作用及机制

Improvement effect and mechanism of petroleum ether extract of Saposhnikovia divaricata on rheumatoid arthritis rats by regulating neutrophil extracellular traps

目的 探究防风石油醚提取物通过调节中性粒细胞胞外诱捕网(NETs)对类风湿性关节炎(RA)大鼠的改善作用及机制。方法 采用牛Ⅱ型胶原与弗氏完全佐剂建立大鼠RA模型。将造模成功的大鼠随机分为模型组和防风石油醚提取物低、中、高剂量组(55、110、220 mg/kg),另设不造模的正常组,每组10只。各组大鼠灌胃相应药液或等体积2%吐温-80溶液,每天1次,连续28 d。观察大鼠足趾肿胀度并进行关节炎指数(AI)评分,检测大鼠血清中白细胞介素1β(IL-1β)、IL-10、IL-17、肿瘤坏死因子α(TNF-α)、25-羟基维生素D_3[25(OH)D_3]、髓过氧化物酶(MPO)、中性粒细胞弹性蛋白酶(NE)和NETs的水平,观察大鼠踝关节病理学形态变化,检测大鼠踝关节中瓜氨酸化组蛋白H3(CitH3)以及滑膜组织中细胞色素P450 24A1(CYP24A1)、细胞色素P45027B1(CYP27B1)、维生素D受体(VDR)、肽基精氨酸脱亚胺酶4(PAD4)的表达水平。结果 与模型组比较,防风石油醚提取物中、高剂量组大鼠给药后第14天至第28天的足趾肿胀度和AI评分均显著降低(P<0.05或P<0.01);血清中IL-1β、IL-17、TNF-α、MPO、NE和NETs水平均显著降低,IL-10、25(OH)D_3水平均显著升高(P<0.05或P<0.01);踝关节间隙变宽,结构明显改善;踝关节中CitH3以及滑膜组织中CYP24A1、PAD4表达水平均显著降低,滑膜组织中CYP27B1、VDR表达水平均显著升高(P<0.05或P<0.01)。结论 防风石油醚提取物可能通过调控维生素D系统,抑制NETs生成,进而改善RA症状。

OBJECTIVE To explore the improvement effect and mechanism of petroleum ether extract of Saposhnikovia divaricata on rheumatoid arthritis(RA)rats by regulating neutrophil extracellular traps(NETs).METHODS Establishment of rat RA model using bovine typeⅡcollagen and Freund’s complete adjuvant.The model rats were randomly divided into model group and low-dose,middle-dose and high-dose groups(55,110,220 mg/kg)of petroleum ether extract of S.divaricata;the normal group without modeling was also established,with 10 rats in each group.Each group was given corresponding drugs or constant volume of 2%Tween-80 solution intragastrically,once a day,for consecutive 28 days.The toe swelling degree in rats was observed,and the arthritis index(AI)was scored.The serum levels of interleukin-1β(IL-1β),IL-10,IL-17,tumor necrosis factor-α(TNF-α),25-hydroxyvitamin D3[25(OH)D3],myeloperoxidase(MPO),neutrophil elastase(NE)and NETs in rats were detected.The histopathological changes in ankle joint were observed.The expression of citrullinated histone H3(CitH3)in ankle joint as well as the expressions of cytochrome P45024A1(CYP24A1),cytochrome P45027B1(CYP27B1),vitamin D receptor(VDR)and peptidyl arginine deiminase 4(PAD4)in synovium were all determined.RESULTS Compared with the model group,the toe swelling degree and AI score in the middle-dose and high-dose groups of petroleum ether extract of S.divaricata decreased significantly from day 14 to day 28 after administration(P<0.05 or P<0.01).The serum levels of IL-1β,IL-17,TNF-α,MPO,NE and NETs decreased significantly,while the levels of IL-10 and 25(OH)D3 increased significantly(P<0.05 or P<0.01).The widened ankle joint space and the improved structure were found;the expression of CitH3 in ankle joint,and the expressions of CYP24A1 and PAD4 in synovium were down-regulated significantly,while the expressions of CYP27B1 and VDR were up-regulated significantly in synovium(P<0.05 or P<0.01).CONCLUSIONS The petroleum ether extract of S.divaricata may inhibit the production of NETs and improve the symptoms of RA by regulating the vitamin D system.