氨甲喋呤(MTX)治疗类风湿性关节炎的免疫调节机制

目的:探讨MTX治疗类风湿性关节炎(RA)的免疫学机制,为临床MTX治疗RA提供理论依据。方法:在建立佐剂型关节炎(AA)动物模型基础上,通过关节炎指数评价,血清中细胞因子测定和关节切片的病理学检查,观察MTX对促炎症细胞因子如IL^-1β、TNF-α和抗炎症细胞因子IL-4、IL-10的调控作用。结果:MTX预防组和对照组关节炎指数和发病率的变化均无显著差异(P>0.05)。MTX肌肉注射治疗或外用治疗后,关节炎指数发生较显著的变化与对照组相比有显著性差异(P>0.05)。对照组与MTX预防组、MTX肌肉注射组、MTX外用组血清中细胞因子水平相比,对照组与各组TNF-α和IL-10的水平有明显差异,P<0.05 ,MTX肌肉注射组与MTX外用组的TNF-α水平相差很大,P>0.05。而IL-1β、IL-4的水平各组之间没有明显差异,P>0.05。病理组织学变化中炎症程度:对照组>MTX预防组>MTX肌肉注射组>MTX外用组。结论:本研究结果表明,MTX治疗(肌肉注射和外用)可以降低AA大鼠血清中TNF-α浓度,提高IL-10的浓度,从而抑制或控制RA的发生发展。但MTX治疗对IL-1β和IL-4没有明显作用,这方面的机制尚需进一步研究。     

集落培养及细胞因子检测在AA与MDS-RA鉴别诊断中的意义

目的 提高对再生障碍性贫血（AA）及骨髓增生异常综合征难治性贫血（MDS－RA）的鉴别诊断。方法 对28例AA及16例MDS－RA患者进行骨髓单个核细胞（MNC）体外集落培养（CFU－GM及CFU－C）及血浆中多种细胞因子（IL－3、IL－6、IL－2、IL－8、γ－IFN及TNF－α）的检测。结果 AA及MDS－RA患者CFU－GM及CFU－C集落数均低于正常对照组，AA患者CFU－C集簇数低于正常对照组，MDS－RA患者CFU－GM及CFU－C集簇数与正常对照组无显著差异。AA与MDS－RA相比较，CFU－GM及CFU－C集落和集簇数均较低。AA及MDS－RA患者血浆中IL－3及IL－6与正常组无显著差异，AA患者IL－2、IL－8、γ-IFN、TNF－α及MDS－RA患者TNF－α均高于正常对照组，MDS－RA患者IL－2及IL－8低于AA组。结论 AA及MDS－RA患者骨髓MNC的CFU－GM及CFU－C集落及集簇数量以及血浆中某些细胞因子含量存在差异，有助于两者的鉴别。     

血府逐瘀汤联合阿替普酶静脉溶栓对急性脑梗死患者神经功能及血清炎性因子水平的影响

目的：观察血府逐瘀汤联合阿替普酶静脉溶栓对急性脑梗死患者神经功能恢复及血清超敏C反应蛋白（hs－CRP）、白介素－1β（IL－1β）、肿瘤坏死因子－α（TNF－α）水平的影响。方法将120例急性脑梗死且具有溶栓适应证患者随机分为观察组与对照组各60例。所有患者入院后均进入脑卒中单元溶栓急诊路径管理,排除禁忌证后,给予阿替普酶50mg静脉溶栓治疗,溶栓开始后对照组依据2010年版中国急性缺血性脑卒中诊治指南分析患者危险因素给予规范药物治疗;观察组在对照组基础上加用我院自制中药合剂—血府逐瘀汤组方,2组均治疗2周。利用美国国立卫生研究院卒中量表（ NIHSS）比较2组神经功能缺损程度评分,运用Barthel指数评价患者日常生活能力。在入院溶栓前及溶栓后24h及治疗后第14天清晨检测患者hs－CRP、IL－1β、TNF－α水平。结果溶栓治疗前,2组患者NIHSS、Barthel指数评分差异无统计学意义（P＞0．05）。溶栓后24h,观察组NIHSS评分较对照组明显降低,Barthel指数评分较对照组明显升高,差异均有统计学意义（P＜0．05）。治疗14d后观察组NIHSS评分较对照组进一步降低, Barthel指数评分较对照组进一步升高,差异均有统计学意义（P＜0．05）。溶栓治疗前,2组患者血清hs－CRP、IL－1β、TNF－α水平差异无统计学意义（P＞0．05）。溶栓后24h,观察组血清hs－CRP、IL－1β、TNF－α水平较对照组明显下降,差异有统计学意义（P＜0．05）。治疗14d后,观察组血清hs－CRP、IL－1β、TNF－α水平较对照组进一步下降,差异均有统计学意义（P＜0．05）。结论血府逐瘀汤联合阿替普酶静脉溶栓治疗能明显改善急性脑梗死患者神经功能缺损程度,提高患者日常生活能力,其机制可能与抑制缺血后再灌注损伤造成的血清炎性因子反应失衡有关。     

醒脑通脉化瘀方对急性脑梗死患者血清 hs－CRP、IL－1β、TNF－α的影响

目的：观察醒脑通脉化瘀方对急性脑梗死患者血清超敏－C反应蛋白（ hs－CRP）、白介素－1β（ IL－1β）、肿瘤坏死因子－α（ TNF－α）水平的影响。方法将急性脑梗死患者160例随机分为研究组和对照组各80例。对照组依据2010年版中国急性缺血性脑卒中诊治指南分析患者危险因素给予规范药物治疗,研究组在对照组基础上加用中药合剂—醒脑通脉化瘀组方,2组均治疗2周。比较2组治疗前后患者hs－CRP、IL－1β、TNF－α水平变化。结果治疗前,2组患者血清hs－CRP、IL－1β、TNF－α水平比较,差异无统计学意义（P＞0．05）。治疗后,2组hs－CRP、IL－1β、TNF－α均较治疗前降低,且研究组降低幅度大于对照组,差异均有统计学意义（P＜0．05）。结论醒脑通脉化瘀方可以明显抑制急性脑梗死患者血清hs－CRP、IL－1β、TNF－α水平,抑制机体氧化应激反应,起到加强脑保护作用。     

甲氨蝶呤对炎性细胞因子及丝裂原活化蛋白激酶信号通路的影响

目的：探讨甲氨蝶呤（methotrexate,MTX）对类风湿关节炎（RA）发病起重要作用的炎性细胞因子IL-1β和TNF-α的作用及其可能的机制.方法：用Ⅱ型胶原建立类风湿关节炎（CIA）动物模型,以MTX（0.2 mg/kg/W）进行处理,6周后处死大鼠,取血清进行IL-1β和TNF-α检测;分离大鼠膝关节取关节滑膜细胞（FLS）进行培养、鉴定,检测脂多糖（LPS）诱导的FLS培养上清中IL-1β和TNF-α的含量,并观察MTX对FLS分泌IL-1β和TNF-α的影响;在FLS的培养中加入不同剂量的MTX,用Westen Blot方法检测FLS中ERK蛋白及磷酸化蛋白的表达.结果：关节炎指数评分、关节肿胀度测定及关节病理显示造模成功,分离关节滑膜细胞经流式细胞仪检测FLS的血管细胞黏附分子-1（VCAM-1）的表达为85.5％.造模后,CIA大鼠血清IL-1β和TNF-α明显增加,与空白对照组比较有统计学差异（P＜0.05）,MTX能有效减少CIA大鼠血清中IL-1β和TNF-α的含量（P＜0.05）,但未能恢复至正常水平.MTX能抑制LPS诱导的CIA大鼠关节FLS分泌IL-1β和TNF-α.Western Blot检测显示,不同浓度的MTX对CIA大鼠FLS中ERK蛋白的表达与模型对照组相比无统计学差异（P＞0.05）.CIA大鼠FLS中p-ERK蛋白的表达明显高于空白对照组（P＜0.01）,MTX干预后,低剂量组对p-ERK蛋白表达无明显影响,而中、高剂量组可降低p-ERK蛋白的表达（P＜0.05）.结论：MTX既有免疫抑制作用,同时还通过抑制炎性细胞因子IL-1β和TNF-α而具有抗炎作用,其机制可能部分与其抑制MAPK信号通路中的ERK蛋白磷酸化有关.     

5-氟脲嘧啶对急性胰腺炎病程中细胞因子的影响

目的：观察5－Fu对急性胰腺炎（AP）病程中TNF－α、IL－6和IL－10的影响。探讨其治疗价值。方法：对5％牛磺胆酸钠溶液（0．1ml/100g)逆行胰胆管注射建立急性死性胰腺炎（ANP）模型，雄性SD大鼠48只，随机平均分为三组：假手术组（Sham组）；ANP组；5－Fu组。ANP模型建立后经股静脉注射5－Fu4.5mg/100g。观察术后3小时、6小时（每时间段8只大鼠）血清TNF－α、IL－6、IL－10、血清淀粉酶和胰腺组织学病理改变。结果：（1）ANP组3h,6h血清淀粉酶明显升高（P＜0．01），同时血清TNF－α、IL－6和IL－10水平均明显高于Sham组（P＜0．001）。（2）5-Fu处理后3h血清淀粉酶及胰腺组织学评分均降低（P＜0．05）；而血清IL－6水平仅在3h时降低（P＜0．01）。结论：TNF－α、IL－6和IL－10在AP病程中起了重要的作用。5－Fu抑制了AP病程中细胞因子的产生，对其治疗有益，抑制细胞因子可能是它治疗急性胰腺炎的另一机制。     

消旋卡多曲联合甘草锌治疗轮状病毒性肠炎62例临床评价

目的：研究消旋卡多曲联合甘草锌对轮状病毒性肠炎患儿的肠道保护作用及炎性反应的调控。方法选择2014年6月至12月医院收治的轮状病毒性肠炎患儿124例,按随机数字表法分为治疗组和对照组,各62例。两组患儿均给予合理饮食、液体疗法、利巴韦林抗病毒治疗、微生态调节剂妈咪爱调节肠道菌群;治疗组患儿加用消旋卡多曲及甘草锌颗粒,疗程为7 d,出院后带甘草锌颗粒继续口服4周。结果治疗组总有效率为96.77%,明显高于对照组的82.26%（ P＜0.05）;治疗组患儿止泻、退热、止吐及脱水纠正时间均短于对照组（ P＜0.05）;两组患儿治疗后细胞因子肿瘤坏死因子－α（TNF－α）、白细胞介素（IL）－6,IL－8、IL－12及血清超敏C反应蛋白（hs－CRP）水平均较治疗前显著降低（ P＜0.05）,且治疗组降低更明显（ P＜0.05）;两组患儿不良反应发生率均较低,差异无统计学意义（ P＞0.05）。结论消旋卡多曲联合甘草锌治疗轮状病毒性肠炎疗效可靠,症状改善快,能有效降低TNF－α,IL－6,IL－8,IL－12水平,下调患儿血清hs－CRP含量,抑制炎性级联反应,值得临床推广。     

金乌健骨方对关节炎模型大鼠的抗炎作用研究

目的观察金乌健骨方对胶原诱导性关节炎（CIA）大鼠血清肿瘤坏死因子α（TNF α）、白细胞介素1（IL 1）及白细胞介素1β（IL 1β）水平的影响,探讨其作用机制。方法将60只Wistar大鼠随机分为空白对照组,模型对照组,金乌健骨方高、中、低剂量组和雷公藤多苷对照组,每组10只。除空白对照组外,其余各组制备CIA模型。给药组分别于造模后第3周开始连续灌胃给药4周,并于用药前及用药后每周观测CIA大鼠关节肿胀程度和关节炎指数,给药结束后检测血清TNF α、IL 1及IL 1β。结果金乌健骨方高、中剂量组关节肿胀程度和关节炎积分均明显降低,优于雷公藤多苷对照组（P＜0.01,P＜0.05）。与模型组比较,金乌健骨方高、中剂量组大鼠血清TNF α、IL 1、IL 1β表达水平显著下降（P＜0.01）,与雷公藤多苷对照组相当;高、中剂量金乌健骨方对IL 1β的影响优于雷公藤多苷（P＜0.01）。结论金乌健骨方能减轻CIA大鼠关节肿胀程度,降低关节炎积分,下调CIA大鼠血清TNF α、IL 1及IL 1β的表达。     

类风湿关节炎患者血清白介素-1β白介素-6 肿瘤死亡因子-α的水平测定及其临床意义

目的通过测定类风湿关节炎（RA）患者和正常对照组血清白介素-1β（IL-1β）、白介素-6（IL-6）、肿瘤死亡因子-α（TNF-α）的水平,探讨这些细胞因子在RA发病机制中的作用。方法收集RA患者20例随机分为两组和健康体检者10例为正常对照组,均采用酶联免疫吸附测定法（ELISA）检测血清IL-1β、IL-6和TNF-α的水平。结果MTX＋TGP＋LEF组与MTX组比较,前者治疗效果明显优于后者。治疗前RA患者血清中IL-1β、IL-6和TNF-α的水平均高于正常对照组（P〈0.01）,具有显著性统计学意义。两组RA患者治疗后血清中IL-1β、IL-6和TNF-α的水平与治疗前比较（P〈0.05）,具有显著性统计学意义。但与正常对照组比较除TNF-α外,IL-1β、IL-6均为P〉0.05,无统计学意义。结论IL-1β、IL-6和TNF-α在RA患者发病、发展中起着重要作用,定期检测其水平并采取对应治疗,对降低RA患者软骨破坏,提高疗效有着重要意义。     

急性心肌梗死再灌注中血浆白细胞介素8、肿瘤坏死因子α的动态变化及意义

目的：探讨急性心肌梗死（AMI）心肌缺血再灌注过程血浆白细胞介素8（IL－8）、肿瘤坏死因子α（TNF－α）的动态变化及其意义。方法：采用单抗夹心ELISA方法,观察比较27例经链激酶（SK）或重组组织型纤溶酶原激活剂（rt-PA),溶栓治疗的AMI患者溶栓前,溶栓后2h、4h、8h、10h、12h血浆IL－8和TNF－α动态变化,并进行统计学分析。结果：27例观察组AMI患者中,14例溶栓治疗后相关血管再通者（再通组）;13例溶栓治疗相关血管未通者（未通组）,再通组IL－8、TNF－α各时间点与正常对照组间有显著差异（F＝51．474、F＝21．322,均P＜0．01）,未能组IL－8、TNF－α各时间点与正常对照组间有显著差异（F＝43．694、F＝36．45,均P＜0．01）。再通组与未通组IL－8比较2h、4h(t=7.652、t=3.054,均P＜0．01）有显著性差异,再通组与未通组TNF－α比较2h、4h(t=1.667、t=19.389),（P＜0．05和P＜0．01）也有显著性差异。两组溶栓治疗前IL－8、TNF－α比较（q=2.498、q=0.326,均P＞0．05）无显著性差异。结论：IL－8、TNF－α在心肌缺血再灌注后炎症损伤中起关键作用,监测IL－8、TNF－α动态变化有助于临床对AMI再灌注损伤的认识,对防治AMI再灌注后炎症损伤的认识,对防治AMI再灌注后炎症损伤有一定实用价值。     

五灵胶囊对正常及免疫性肝损伤小鼠IL-2、IL-6的影响

目的:探讨五灵胶囊对正常小鼠及免疫性肝损伤小鼠体内白细胞介素2（IL-2）、白细胞介素6（IL-6）活性的影响。方法:BALB/C系小鼠经脂多糖（LPS）处理和未经LPS处理,灌胃12d后,取脾脏体外培养48h、72h,收集脾细胞培养上清作用于靶细胞,分别用MTT法测定IL-2活性,用放免法测定IL-6活性。结果:五灵胶囊有促进小鼠Th细胞分泌IL-2的作用,使肝损伤所致低水平的IL-2明显升高,对损伤或正常小鼠IL-6水平无影响。结论:五灵胶囊促进IL-2分泌,调节免疫作用。     

大鼠胶原诱导性关节炎模型的制备及评价

目的:制备型胶原诱导性关节炎大鼠模型(CIA模型)。方法:将40只大鼠随机分为正常组和模型组。以酶联免疫吸附测定法(ELISA法)检测模型组和正常组大鼠血清中炎性细胞因子肿瘤坏死因子-α(TNF-α)水平,同时采用组织病理学和X线摄片的方法显示关节炎大鼠的发病程度和病理学特征。结果:与正常组相比,模型组在免疫后第36天关节肿胀达到最高峰。ELISA法检测结果显示模型组TNF-α的水平均较正常组大鼠明显增高(P<0.05)。组织病理学和X射线摄片结果显示,关节软骨组织、骨组织和滑膜组织呈典型的关节炎病变。结论:胶原诱导的大鼠关节炎模型,其病理特征和人类类风湿关节炎(RA)极为相似,为进一步深入研究人类RA的发病机制及其临床治疗提供了有价值的实验材料。     

Visfatin as a therapeutic target for rheumatoid arthritis

Introduction: The rising prevalence of musculoskeletal pathologies in developed countries has caused a dramatic impact on social welfare. Amidst these musculoskeletal pathologies is Rheumatoid arthritis (RA), a chronic systemic autoimmune disease that mostly affects the synovium. RA metabolic-associated alterations, including distorted adipokine production, enhance RA inflammatory environment. Among the altered adipokines, visfatin is particularly involved in RA inflammation and catabolism and stands out as an essential enzyme linked to critical cell features.

Areas covered: We discuss the potential mechanism supporting the contribution of visfatin to RA and the association between RA and obesity. We discuss the repurposing of cancer-tested drugs to inhibit visfatin in the context of RA. Additionally, we address the possibility of combining these drugs with current RA therapy. Finally, we explore the future of visfatin as an RA biomarker or therapeutic target.

Expert opinion: Inhibition of visfatin has become an interesting therapeutic approach for RA pathology. Such a feat has already been attained in oncology using small molecule inhibitors, which suggest that a similar course of action would be worth pursuing in the RA context. Visfatin will become an important biomarker and therapeutic target for RA.     

RNAi Silencing of IL‐1β and TNF‐α in the Treatment of Post‐traumatic Arthritis in Rabbits

Post‐traumatic arthritis is a secondary complication to severe joint trauma. With the disease progression, it may eventually lead to osteoarthritis in patients whose age is considerably younger than patients with traditional bone arthritis. The main objective of this study was to explore the feasibility of using lentiviral‐mediated RNA interference silencing of IL‐1β and TNF‐α to treat post‐traumatic arthritis in rabbits. About 48 New Zealand rabbits underwent bilateral knee joint surgery to stimulate traumatic arthritis. They were then randomly divided into four groups of 12 rabbits each. The histopathology of the cartilage was observed, and the changes were assessed by Mankin scoring. ELISA was used to detect the expression of IL‐1β and TNF‐α in the synovial fluid. (i) Compared with the control group, the transfection and co‐transfected groups displayed reduced cartilage damage and speed of degeneration. The co‐transfected group showed the greatest alleviation of symptoms. The Mankin score was statistically different (p < 0.01). (ii) Compared with the control group, the expression of IL‐1β or TNF‐α was reduced in the respective transfection groups (p < 0.01 in both groups) and IL‐1β and TNF‐α were reduced in the co‐transfected group (p < 0.01). The co‐transfected group showed the lowest expression of the three experimental groups of both IL‐1β and TNF‐α (p < 0.01). Lentivirus‐mediated RNA interference can knock down the expression of IL‐1β and TNF‐α in joint fluids and, in a synergistic effect when two siRNAs are co‐transfected, ease cartilage degeneration.     

OPG-HSP70融合蛋白对CIA大鼠的保护性作用

目的:研究人骨保护素(OPG)-分枝杆菌热休克蛋白70(HSP70)融合蛋白对类风湿关节炎动物模型--胶原诱导的关节炎(CIA)大鼠的保护性作用.方法:将OPG-HSP70重组基因菌株诱导表达,收集蛋白并鉴定,大量提取,纯化,复性;实验大鼠分为OPG-HSP70组、OPC组、HSP70组、模型组和正常对照组,CIA大鼠模型制备后分别给予相应处理,观察对大鼠关节病变的影响.结果:与模型组相比,OPG-HSP70组关节炎指数明显降低(P<0.05);关节病理切片显示关节滑膜无明显增生;骨密度值增高(P<0.05).结论:OPGHSP70融合蛋白具有减轻CIA大鼠关节病变的作用.     

美洛昔康抗炎作用的实验研究

目的观察美洛昔康的抗炎作用。方法采用大鼠急性、亚急性及免疫性炎症模型,观察美洛昔康对角叉菜胶致大鼠足跖肿胀和大鼠棉球肉芽肿的影响以及美洛昔康对大鼠佐剂性关节炎的预防和治疗作用。结果美洛昔康对多种致炎剂引起的炎性水肿均具有明显的抑制作用,1.0和3.0mg     

类风湿性关节炎滑膜组织中VEGF表达及临床意义

目的：研究类风湿性关节炎患者滑膜组织中血管内皮生长因子（vascular endothel ialgrowth factor,VEGF）的表达情况。方法：采用免疫组化SP法,检测27例类风湿性关节炎活动期膝关节滑膜组织中VEGF蛋白表达,以15例骨性关节炎患者、15例正常人及该27例患者治疗后（稳定期）关节滑膜组织中VEGF蛋白作对照,进行对比分析。结果：VEGF在正常人、骨性关节炎患者滑膜组织中的表达明显低于类风湿性关节炎患者,在稳定期滑膜组织中的表达明显低于活动期,组间比较,差异均有统计学意义（P均〈0.05）。结论：VEGF与类风湿性关节炎的病情发展密切相关,针对血管内皮生长因子的靶点治疗有望成为类风湿性关节炎治疗的一种新的发展方向。     

Enantioselective kinetic disposition of fenoprofen in rats with experimental diabetes or adjuvant-induced arthritis

This study was aimed to investigate the influence of diabetes or arthritis on the enantioselective metabolism and kinetic disposition of fenoprofen in rats with streptozotocin-induced diabetes or Mycobacteriumtuberculosis adjuvant-induced arthritis. Animals received i.v. 10 mg/kg racemic fenoprofen and blood samples were collected up to 24 h thereafter, with 5 animals studied at each time point. Plasma concentrations of the fenoprofen enantiomers were determined by HPLC. Diabetic and arthritic animals showed significant differences when compared with respective controls for the following pharmacokinetic variables of the (+)-(S)-fenoprofen eutomer: area under the plasma concentration time curve, total clearance and volume of distribution. The results indicate that experimental diabetes and adjuvant-induced arthritis influence the fenoprofen enantioselective metabolism.     

Pharmacological blockade of CCR1 ameliorates murine arthritis and alters cytokine networks in vivo

BACKGROUND AND PURPOSE: The chemokine receptor CCR1 is a potential target for the treatment of rheumatoid arthritis. To explore the impact of CCR1 blockade in experimental arthritis and the underlying mechanisms, we used J-113863, a non-peptide antagonist of the mouse receptor. EXPERIMENTAL APPROACH: Compound J-113863 was tested in collagen-induced arthritis (CIA) and three models of acute inflammation; Staphylococcus enterotoxin B (SEB)-induced interleukin-2 (IL-2), delayed-type hypersensitivity (DTH) response, and lipopolysaccharide (LPS)-induced tumour necrosis factoralpha (TNFalpha) production. In the LPS model, CCR1 knockout, adrenalectomised, or IL-10-depleted mice were also used. Production of TNFalpha by mouse macrophages and human synovial membrane samples in vitro were also studied. KEY RESULTS: Treatment of arthritic mice with J-113863 improved paw inflammation and joint damage, and dramatically decreased cell infiltration into joints. The compound did not inhibit IL-2 or DTH, but reduced plasma TNFalpha levels in LPS-treated mice. Surprisingly, CCR1 knockout mice produced more TNFalpha than controls in response to LPS, and J-113863 decreased TNFalpha also in CCR1 null mice, indicating that its effect was unrelated to CCR1. Adrenalectomy or neutralisation of IL-10 did not prevent inhibition of TNFalpha production by J-113863. The compound did not inhibit mouse TNFalpha in vitro, but did induce a trend towards increased TNFalpha release in cells from synovial membranes of rheumatoid arthritis patients. CONCLUSIONS AND IMPLICATIONS: CCR1 blockade improves the development of CIA, probably via inhibition of inflammatory cell recruitment. However, results from both CCR1-deficient mice and human synovial membranes suggest that, in some experimental settings, blocking CCR1 could enhance TNF production.