新风胶囊对强直性脊柱炎活动期患者血液高凝状态影响及其机制探讨

Impact of Xinfeng Capsules on Hypercoagulable State in Active Ankylosing Spondylitis and its Mechanism

目的观察新风胶囊(XFC)对强直性脊柱炎(AS)活动期患者血液高凝状态相关指标外周血细胞因子(TNF-α、IL-4、IL-10、IL-17)、NF-κB信号通路等相关指标表达变化,探讨新风胶囊(XFC)对强直性脊柱炎患者活动期患者血液高凝状态影响的机制。方法采用随机数字表的方法将80例AS活动期患者分为新风胶囊组(XFC组)和柳氮磺吡啶组(SASP组),XFC组服用新风胶囊,3次/d,3粒/次,4周为1个疗程,连服3个疗程;SASP组服用柳氮磺吡啶,4片/次,2次/d,服用天数及疗程同XFC组。另设30名健康志愿者为健康对照组。采用酶联免疫吸附法(ELISA)检测AS活动期患者和健康对照者外周血血清中血栓素A2(TXA2)、前列环素I2(PGI2)、P-选择素(GMP140)、血小板活化因子(PAF)、纤溶酶原激活剂抑制剂(PAI-2)、肿瘤坏死因子(TNF)-α、白介素(IL)-4、IL-10、IL-17、核因子激活剂(Act1)、NF-κB抑制蛋白(IκBα)、IκB激酶(IKKβ)、核转录因子(NF-κB)/P65、NF-κB/P50。采用实时荧光定量(real time-PCR,RT-PCR)检测Act1、IκBα、IKKβ、NF-κB/P65、NF-κB/P50 mRNA变化,采用蛋白免疫印迹法(Western blot)检测NF-κB/P65、NF-κB/P50表达。采用Sysmex K-4500型全自动血液分析仪检测血常规、凝血常规、D-D,采用魏氏法测定血沉(ESR),采用日立7060型全自动生化分析仪检测C反应蛋白(CRP),同时观察AS患者临床症状变化。结果与健康对照组比较,AS患者PLT、FBG、D-D、GMP140、PAF、PAI-2、TXA2、PGI2、TNF-α、IL-17、Act1、IκBα、IKKβ、NF-κB/P65、NF-κB/P50明显升高(P<0.05,P<0.01),IL-4、IL-10明显降低(P<0.05,P<0.01)。与本组治疗前比较,XFC组和SASP组治疗后ESR、CRP、VAS、BASDAI、BASFI、BAS-G、晨僵时间、枕墙距、指地距、Act1、IKKβ、P50、P65明显下降(P<0.05,P<0.01),PF、RP、BP、VT明显升高(P<0.05,P<0.01)。XFC组总有效率显著高于对照组(P<0.05);与SASP组治疗后比较,XFC组在上调IL-4、IL-10,下调TNF-α、IL-17,纠正凝血-纤溶系统(PLT、FBG、D-D、TXA2/PGI2、GMP140、PAF、PAI-2)失衡,抑制IKKβ、IκBα、P50、P65等过度活化方面,差异有统计学意义(P<0.05,P<0.01)。结论 XFC能有效改善AS活动期患者的血液高凝状态,其机制可能通过上调抑炎因子(IL-4、IL-10),下调致炎因子(TNF-α、IL-17)、疾病活动指标(ESR、CRP),纠正凝血-纤溶系统(PLT、FBG、D-D、TXA2/PGI2、GMP140、PAF、PAI-2)失衡,抑制NF-κB信号通路(Act1、IKKβ、IκBα、P65、P50)过度活化,从而提高患者的生活质量。

Objective To observe the impacts of xinfeng capsule( XFC) on the relevant indexes of hypercoagulable state,the expressions of relevant indexes of peripheral blood cytokines( TNF- α、IL- 4、IL- 10,IL- 17) and NF- κB signaling pathway in the patients of AS at active stage and explore the mechanism of XFC on the hypercoagulable state. Methods The randomized number table was used to divide 80 cases of active AS into a XFC group and a sulfasalazine( SASP) group. In the XFC group,XFC was taken orally,three times a day,3 capsules each time,the treatment for 4 weeks as 1 session,continuously for 3 sessions. In the SASP group,SASP was taken orally,twice a day,4 tablets each time,the duration and session same as XFC group. Additionally,a health control group was set up,30 cases. Using enzyme- linked immunosorbent method( ELISA),serum thromboxane A2( TXA2),prostacyclin I2( PGI2),P- selection element( GMP140 or CD62p),platelet activation factor( PAF),plasminogen activator inhibitor( PAI- 2),tumor necrosis factor-alpha( TNF- α),interleukin( IL)- 4,IL- 10,IL- 17,nuclear factor activating agent- 1( Act1),nuclear factor kappa B inhibitor protein( IκB- alpha,IκBα),nuclear factor kappa B kinase( IKK beta,IKKβ),and nuclear factor protein( NF- kappa B65 /50,NF- κB- P65 / P50) were determined in the patients of active AS and the subjects in the control group. The Real time- PCR( RT- PCR) was used to determine mRNA changes of Act1,IκBα,IKKβ,NF- κB / P65,and NF- κB / P50. Western blot was used to determine the expressions of NF- κB / P65,and NF- κB / P50. Automatic blood analyzer,Sysmex K- 4500 was used to determine blood routine,coagulation routine,and D- D. Using the Westergren method,erythrocyte sedimentation rate( ESR) was determined. Hitachi 7060 automatic biochemistry analyzer was used to detect C reactive protein( CRP) and observe the changes of clinical symptoms in AS patients. Results Compared with the health control group,the levels of PLT,FBG,D- D,GMP140,PAF,PAI- 2,TXA2,PGI2,TNF- α,IL- 17,Act1,IκBα,IKKβ,NF- κB / P65 and NF- κB / P50 were significantly increased in AS patients( P < 0. 05,P <0. 01),and the levels of IL- 4 and IL- 10 were reduced apparently( P < 0. 05,P < 0. 01). As compared with those before treatment,ESR,CRP,VAS,BASDAI,BASFI,BAS- G,morning stiffness time,occiput to wall distance,finger to floor distance,Act,IκBα,IKKβ,NF- κB / P65 and NF- κB / P50 were decreased significantly( P < 0. 05,P < 0. 01),and PF,RP,BP,and VT were increased significantly( P < 0. 05,P < 0. 01) in the XFC group and the SASP group. The total effective rate of the XFC group was significantly higher than that of the control group( P < 0. 05). Compared with the SASP group after treatment,the significant differences presented in the up- regulation of IL- 4 and IL- 10,the down- regulation of TNF- α and IL- 17,the rectification of the imbalance of coagulation fibrinolytic system( PLT,FBG,D- D,TXA2 / PGI2,GMP140,PAF and PAI- 2) and the inhibition of the excessive activation of Act1,IκBα,P65 and P50( P < 0. 05,P <0. 01) in the XFC group. Conclusion XFC effectively improves in the hypercoagulable state in the patients of active AS. The potential mechanism may be related to the up- regulation of anti- inflammatory factors( IL- 4 and IL- 10) and the down- regulation of inflammatory factors( TNF- α and IL- 17),active indexes( ESR and CRP),the rectification of the imbalance in the coagulation fibrinolytic system( PLT,FBG,D- D,TXA2 / PGI2,GMP140,PAF and PAI- 2) and the inhibition of the excessive activation of NF- κB signaling pathway( Act1,IκBα,P65 and P50). As a result,the quality of life is improved in the patients.