丙酮酸乙酯对胶原诱导性关节炎大鼠的治疗作用

目的探讨丙酮酸乙酯（EP）在胶原诱导关节炎（CIA）大鼠模型中的治疗作用和安全性。方法利用牛Ⅱ型胶原建立Wistar大鼠CIA模型,随机分为CIA对照组和EP干预组,并设立正常对照组。每周观察各组大鼠的一般情况变化,并对双后足进行关节炎指数（AI）评分及测量后足容积;在实验第42、63天分别对各组大鼠的右后足进行放射学及病理学观察。结果 CIA对照组大鼠与正常对照组大鼠比较,有脱毛、精神萎靡,摄食及其他活动减少;EP干预组大鼠精神食欲尚可。体重变化,与正常对照组大鼠比较,CIA对照组大鼠从造模第28天开始,体重增加缓慢;EP干预组从造模第35天开始体重增加程度也有所下降,差异均有统计学意义（P〈0.05）;从造模第56天开始EP干预组体重增加明显,与正常组比较差异无统计学意义（P〉0.05）。关节炎症,EP干预组大鼠造模第28、42、63天AI值分别为3.83±0.71、3.42±0.79、1.50±0.54,明显低于同期CIA对照组AI值4.58±1.08、5.17±1.19、3.67±0.81,差异均有统计学意义（P〈0.05）。足肿胀度,EP干预组大鼠足肿胀度在造模第42、63天分别为（1.58±0.11）、（1.51±0.09）ml,低于CIA对照组足肿胀度（1.97±0.10）、（1.81±0.10）ml,差异均有统计学意义（P〈0.05）。影象学表现,CIA对照组X线示关节间隙消失,严重的可出现骨质破坏、融合;与CIA对照组相比,EP干预组仅可见部分关节有关节间隙模糊、骨质疏松,很少见关节破坏融合。病理学表现,CIA对照组滑膜肥厚,大量炎性细胞浸润,并有骨结构破坏;与CIA对照组相比,EP干预组滑膜细胞增生不明显,有少量炎性细胞浸润,关节软骨和骨组织破坏不明显。EP干预过程中未见明显不良反应。结论 EP能改善CIA大鼠关节炎症状、放射学及滑膜组织病理变化,起到抑制炎症的作用,且安全性好。     

胶体磷酸铬32P关节腔内注射对大鼠关节炎模型的影响

目的 探讨关节腔内注射胶体磷酸铬^32P对Ⅱ型胶原诱导大鼠关节炎模型的影响,为难治性类风湿关节炎（RA）的治疗探索一条新路。方法 利用Ⅱ型胶原免疫SD大鼠诱发的关节炎模型（CIA）,给予胶体磷酸铬^32P踝关节腔内注射,观察右踝关节左右径平均宽度、关节体积、关节指数及关节病理的变化。结果 与CIA对照组相比,注射后6周右踝关节左右径平均宽度、关节体积与CIA对照组比较差异有显著性（P＞0．05）。关节指数二者差异无显著性。组织病理学改变示滑膜组织增生及炎症细胞浸润程度相对较轻。结论 提示胶体磷酸铬^32P关节腔内注射治疗能减轻CIA关节滑膜的增生,对CIA有一定治疗作用,可用于难治性RA的局部治疗。     

α黑素细胞刺激素对大鼠胶原性关节炎的保护作用

目的 观察α黑素细胞刺激素(α—MSH)对大鼠胶原性关节炎(CIA)发病过程的影响。方法 经牛Ⅱ型胶原(bcⅡ)诱导建立大鼠关节炎动物模型，随机分为4组：正常对照组，CIA对照组，α—MSH低剂量组和α—MSH高剂量组，每组8只，于致敏当天连续腹腔注射不同剂量的α—MSH或PBS，观察比较4组大鼠体重、足爪容积、关节炎指数、病理评分、X线放射学表现等变化。结果α—MSH可缓解大鼠关节炎症状，减轻滑膜增生和炎性细胞浸润，抑制关节骨质破坏。结论 α—MSH可抑制大鼠胶原性关节炎发病，具有潜在的临床应用价值。     

羟基喜树碱与甲氨蝶呤治疗胶原诱导性关节炎模型大鼠的比较研究

目的 探讨羟基喜树碱(HCPT)在胶原诱导性关节炎(CIA)大鼠的治疗作用及临床治疗类风湿关节炎(RA)的潜在价值.方法 ①建立CIA大鼠模型:随机分为健康对照组、模型组、甲氨蝶呤(MTX)治疗组、低剂量HCPT治疗组、高剂量HCPT治疗组;②观察不同时间点大鼠一般情况、关节肿胀度;③测量造模15周后大鼠关节滑膜病理改变及原位末端标记技术(TUNEL)染色滑膜细胞凋亡率;④测定血清肿瘤坏死因子(TNF)-α和白细胞介素(IL)-1β浓度.结果 HCPT、MTX治疗组与模型组比较,各治疗组均可改善模型鼠一般情况、缓解关节炎指数及足肿胀度、减少炎性细胞浸润、滑膜细胞增生及减轻关节内软骨和骨破坏、提高滑膜细胞凋亡率、降低血清TNF-α、IL-1β浓度,差异有统计学意义(P<0.05),各治疗组间无统计学意义(P>0.05).结论 HCPT在临床治疗RA方面具有潜在价值.     

青藤碱对胶原诱导型关节大鼠滑膜细胞增殖及凋亡影响的研究

目的研究青藤碱对胶原诱导型关节炎(CIA)大鼠关节炎症,滑膜增生、凋亡及突变型p53的影响,揭示青藤碱抗类风湿关节炎(RA)可能的作用途径。方法采用牛Ⅱ型胶原混合弗氏不完全佐剂于W istar大鼠尾根部注射,足肿后随机分为造模加生理盐水组、甲氨蝶呤(M TX)治疗组和青藤碱(SIN O)治疗组,采用容积法和计分法分别评价后肢肿胀度和四肢关节炎症程度;左前肢和右后肢近端第3足趾关节苏木素-伊红(H E)染色,评价中性粒细胞、淋巴细胞、浆细胞浸润、滑膜细胞增生和纤维组织增生的情况;对足肿胀和病理各项指标进行回归性分析。碱性磷酸酶标记法检测大鼠滑膜细胞凋亡;免疫组织化学法检测突变型p53的表达。比较M TX和SINO灌胃治疗后各项指标的差异。结果CIA大鼠造模后,炎症迅速发展,1周内即可达到肿胀高峰,然后逐渐消退,在第21～25天后又出现第2个炎症高峰。药物治疗17d后抑制CIA肢炎症的作用逐渐显现出来。M TX和SIN O都可明显抑制第2个炎症高峰,后肢肿胀度和四肢肿胀积分在35d与造模加生理盐水组差异有统计学意义(P<0.01);病理结果中,CIA大鼠关节表现为单个核炎症细胞浸润(包括中性粒细胞、淋巴细胞和浆细胞的浸润),滑膜细胞增生、纤维组织增生;足肿与淋巴细胞浸润,滑膜增生呈直线回归关系,其中以滑膜增生为主(P<0.01     

缺氧诱导因子-1α及血管内皮生长因子在Ⅱ型胶原诱导性关节炎模型中的致病作用

目的 探讨低氧诱导因子-1α(HIF-1α)及血管内皮生长因子(VEGF)在Ⅱ型胶原诱导性关节炎(CIA)模型中的致病作用.方法 选用50只SD大鼠(雌雄各半),建立CIA模型,分别于造模第21、28、35、42天取踝关节作HE染色及HIF-1α免疫组化染色,分析其在CIA中与关节炎活动指标,滑膜病理学评分之间的关系.结果 CIA大鼠关节滑膜层和滑膜下层均表达HIF-1α,第21天阳性表达量最高,随病程进展,表达逐渐下降,与滑膜病理学评分、滑膜增生评分及血管生成评分呈显著正相关.结论 HIF-1α在RA致病机制中发挥关键作用,可能是重要的治疗靶点.     

比较磁共振成像和病理对大鼠Ⅱ型胶原诱导性关节炎早期关节病变的诊断价值

目的 建立Ⅱ型胶原诱导性大鼠关节炎模型(CIA),结合病理改变研究磁共振成像(MRI)检测对早期类风湿关节炎的诊断价值.方法 用牛Ⅱ型胶原和不完全弗氏佐剂联合皮内注射免疫Wistsr大鼠,制作关节炎模型;免疫后不同时期分别采用普通X线摄片、1.5T磁共振仪进行双踝关节平扫加增强扫描以及病理组织学等方法 进行大鼠踝关节病理学特征分析.结果 CIA实验组大鼠免疫14 d后,其关节指数(AI)显著增加,28 d AI增加达到最高(3.6±1.0),抗Ⅱ型胶原抗体水平较对照组明显高(P<0.001).根据病理学分析CIA成模率为93.3%.MRI平扫加增强显示有12只出现异常征象,敏感性85.7%,特异性100%,与病理诊断呈正相关(r=0.5345,P<0.05).X线有4只出现关节软组织肿胀,MRI较X线检出率高(P=0.008).结论 MRI对早期关节滑膜炎、滑膜增生、血管翳等病理学的诊断生成方面有明显优势.     

三氧化二砷对胶原诱导的关节炎大鼠凋亡及肿瘤坏死因子-α白细胞介素-1的作用

目的研究三氧化二砷（As2O3）对Ⅱ型胶原诱导的关节炎（CIA）模型大鼠滑膜组织凋亡的影响,并进一步探讨As2O3对细胞因子肿瘤坏死因子（TNF）-α、白细胞介素（IL）-1的作用.方法将72只DA大鼠随机分成正常对照组、CIA大鼠模型组及不同浓度As2O3治疗组,共6组.As2O3治疗后第15天各组取膝关节滑膜、软骨和骨组织,光、电镜观察,TUNEL法检测凋亡.酶联免疫吸附试验（ELISA）测定血清细胞因子IL-1和TNF-α的浓度.结果治疗组滑膜组织病理改变比模型组减轻,滑膜细胞凋亡数明显增多,滑膜细胞超微结构接近正常对照组.并且与模型对照组相比,治疗组大鼠血清IL-1、TNF-α浓度降低,以第Ⅴ组和第Ⅵ组更显著（P＜0.01）.结论As2O3可能通过促进CIA大鼠滑膜细胞及组织的凋亡,抑制炎性因子TNF-α及IL-1的释放,起到减少CIA损害的保护作用.     

99Tc-亚甲基二膦酸盐对胶原诱导性关节炎大鼠的治疗作用及其对滑膜MMP-3和TIMP-1的影响

目的观察~(99)Tc-亚甲基二膦酸盐(~(99)Tc-MDP)对Ⅱ型胶原诱导性关节炎(CIA)大鼠的治疗作用及其对滑膜基质金属蛋白酶-3(MMP-3)、基质金属蛋白酶组织抑制物-1(TIMP-1)的影响,探讨~(99)Tc-MDP治疗类风湿关节炎(RA)的机制。方法建立CIA大鼠关节炎模型,分为正常对照组、模型对照组、~(99)Tc-MDP组和甲氨蝶呤组。采用关节炎指数评分法评价其关节的炎症程度；观察关节病理组织形态学变化；反转录-聚合酶链反应(RT-PCR)法检测滑膜MMP-3和TIMP-1 mRNA水平。结果①模型组、甲氨蝶呤组、~(99)Tc-MDP组大鼠随着免疫时间延长关节炎指数(Al)评分增加。②模型组关节组织学评分均高于甲氨蝶呤组和~(99)Tc-MDP组(P＜0．05)；软骨破坏和骨质破坏评分,~(99)Tc-MDP组低于甲氨蝶呤组(P＜0．05)。③模型组、甲氨蝶呤组、~(99)Tc-MDP组MMP-3 mRNA水平显著高于正常组(P＜0．01)；模型组高于~(99)Tc-MDP组(P＜0．05)。各组滑膜TIMP-1 mRNA水平差异无统计学意义(P＞0．05)。结论~(99)Tc-MDP明显减轻CIA大鼠的关节炎症状,延缓关节破坏的进展；~(99)Tc-MDP降低滑膜MMP-3 mRNA水平,可能是其治疗RA的机制之一；在延缓CIA大鼠软骨和骨质破坏的病理学改变及降低滑膜MMP-3 mRNA水平,其近期疗效优于甲氨蝶呤。     

低氧诱导因子-1α在大鼠胶原诱导性关节炎中的表达及与血管生成的关系

目的制备牛Ⅱ型胶原诱导性大鼠关节炎(CIA)模型,探讨低氧诱导因子-1α(HIF-1α)、血管内皮生长因子(VEGF)在类风湿关节炎(RA)组织中的动态表达及其相关性作用。方法 50只SD大鼠,随机分为对照组与模型组。建立CIA模型,动态观察各项关节炎活动指标,于造模21、28、35、42d取踝关节行HE染色及HIF-1α、VEGF免疫组化染色,分析两者在CIA中的相关性及与关节炎活动指标、滑膜病理学评分之间的关系。结果 CIA大鼠关节滑膜层和滑膜下层均表达HIF-1α、VEGF,第21天阳性表达量最高,随病程进展,表达逐渐下降,与滑膜病理学评分、滑膜增生评分及血管生成评分呈显著正相关,与炎症浸润评分无明显相关。结论Ⅱ型胶原可成功诱导大鼠关节炎模型。HIF-1α、VEGF在RA组织中的表达,与炎症严重程度呈明显正相关。HIF-1α可能通过调控一系列下游靶基因,如VEGF等,促进滑膜增生及血管生成,影响RA的发生、发展。     

移植间充质干细胞在胶原诱导性关节炎大鼠关节的分布和作用

目的 探讨移植人的异基因骨髓间充质干细胞(BM-MSCs)在类风湿关节炎(RA)动物模型胶原诱导性关节炎(CIA)大鼠炎症关节局部的转归以及对关节局部组织损伤的修复作用.方法 分别选取5只Wistar幼年大鼠和30只Wistar成年雌性大鼠,5只Wistar幼年大鼠用以提取BM-MSCs,30只Wistar成年雌性大鼠随机分为CIA造模A组、B组和正常C组各10只.BM-MSCs分离、传代培养后用5-溴脲嘧啶脱氧核苷(5-BrdU)进行标记.CIA大鼠成功造模后,采取尾静脉注射移植于A组和C组大鼠,移植细胞数为1.0×107/kg.移植后1-4周观察各组动物一般状况和关节局部肿胀度的变化;于移植4周后用关节组织病理切片局部免疫组织化学抗5-BrdU和骨保护素的方法观察BM-MSCs向受体关节局部的趋化和聚集以及对关节组织中骨保护素表达的影响.2组间比较采用t检验.结果 BM-MSCs移植后,受试动物关节肿胀开始消退[(200±350)μl],移植后2、3、4周时与未移植BM-MSCs的B组动物[(320±110)μl]之间比较差异有统计学意义(P＜0.05);在受者关节滑膜、血管内膜及软骨组织中均可见5-BrdU阳性细胞,滑膜组织处的平均灰度值CIA造模A组(85±9)低于正常C组(110±6),差异有统计学意义(P＜0.05);BM-MSCs移植后CIA造模A组关节滑膜中骨保护素阳性率增加,平均灰度值为54±4,明显低于未移植的CIA造模B组(77±6),差异有统计学意义(P＜0.05).结论 通过外周静脉移植的BM-MSCs可以特异性趋化和聚集到炎症关节局部,并可以减轻关节损害,而这种局部组织损伤修复作用可能是通过增加局部炎症关节组织中骨保护素的含量而发挥的.

Abstract：

Objective To study the distribution of allogenic bone marrow-derived mesenchymal stem cells (BM-MSCs) on joints of collagen-induced arthritis (CIA) rats and to investigate their repair effects on joint damages. Methods Five Wistar rats were used for extraction of mesenchymal stem cells and 30 adult female Wistar rats were divided into 3 groups: the CIA rats group A (n=10), CIA rats group B (n=10) and normal rats control group C (n=10). BM-MSCs of Wistar rats were isolated, cultured in vitro routinely and the fourth passages was taken for identification of specific surface antigens by flow cytometry, then the cells were labeled with 5-bromodeoxyuridine (5-BrdU) in vitro. The models of CIA rats were established. 5-BrdU labeled BM-MSCs (1.0×107 cells/kg) were imfused from through tail vein to CIA rats group A and control group C. During the first 4 weeks after BM-MSCs transplantation, changes of general condition and left hind paw swelling were examined. At the fourth week, immunohistochemical examination of 5 -BrdU and osteoprotegerin (OPG) were performed to investigate BM-MSCs aggregation around the knee joints. The contribution of BM -MSCs to repairing of joint damages was identified. Comparisons between groups were performed by t-test. Results After BM-MSCs transplantation, left hindpaw swelling of group A were relieved compared with group B (P＜0.05) and the mobility of the joints was significantly improved. At the fourth week, much more implanted cells (5-BrdU positive cells.) were detected in the damaged knee joints than those in normal knee joints. The average grey scale values on synovium of knee joints in the CIA group A (85±9) was significantly lower than that of the normal group C (110±6, P＜0.05). At the same time, OPG expression was increased in damaged knee joints. The average grey scale values on synovium of knee joints in CIA group A (54±4) was significantly lower than that of the CIA group B (77±6, P＜0.05). Conclusion The transplanted allogeneic bone marrow mesenchy-mal stem cells can migrate to sites of damaged tissue in arthritis. They can prevent tissue damage and repair the damaged joints tissue by increasing OPG expression. This study has provided some evidence for developing effective therapy for rheumatoid arthritis.     

Beneficial effects of tempol, a membrane-permeable radical scavenger, in a rodent model of collagen-induced arthritis

OBJECTIVE: To investigate the effects of tempol, a membrane-permeable radical scavenger, in rats with collagen-induced arthritis (CIA). METHODS: CIA was induced in Lewis rats by intradermal injection of 100 microl of an emulsion of 100 microg of bovine type II collagen (CII) in complete Freund's adjuvant (FCA) at the base of the tail. On day 21, a second injection of CII in FCA was administered. RESULTS: Lewis rats developed an erosive arthritis of the hind paws when immunized with CII in FCA. Macroscopic evidence of CIA first appeared as periarticular erythema and edema in the hind paws. The incidence of CIA was 100% by day 27 in the CII-challenged rats, and the severity of CIA progressed over a 35-day period. Radiographs revealed focal resorption of bone, with osteophyte formation in the tibiotarsal joint, and soft tissue swelling. The histopathologic features included erosion of the cartilage at the joint margins. Treatment of rats with tempol (10 mg/kg/day intraperitoneally) starting at the onset of arthritis (day 23) delayed the development of the clinical signs on days 24-35 and improved the histologic status of the knee and paw. Immunohistochemical analysis for nitrotyrosine and poly(ADP-ribose) synthetase (PARS) revealed positive staining in the inflamed joints of CII-treated rats. The degree of nitrotyrosine and PARS staining was markedly reduced in tissue sections obtained from CII-treated rats that had received tempol. Furthermore, radiographs revealed protection against bone resorption and osteophyte formation in the joints of tempol-treated rats. CONCLUSION: This study is the first to provide evidence that tempol, a small molecule that permeates biologic membranes and scavenges reactive oxygen species, attenuates the degree of chronic inflammation and tissue damage associated with CIA in the rat.     

Amelioration of collagen-induced arthritis in rats by nanogold

OBJECTIVE: Angiogenesis plays a part in the pathogenesis of rheumatoid arthritis (RA), and nanogold inhibits the activity of an angiogenic factor, vascular endothelial growth factor (VEGF). We therefore investigated whether intraarticular delivery of nanogold ameliorates collagen-induced arthritis (CIA) in rats. METHODS: Binding of 13-nm nanogold to VEGF in human RA synovial fluid (SF) and its effects on RA SF-induced endothelial cell proliferation and migration were assessed. Nanogold was administered intraarticularly to rats with CIA before the onset of arthritis. Progression of CIA was monitored by measures of clinical, radiologic, and histologic changes. In addition, the microvessel density and extent of infiltrating macrophages as well as levels of tumor necrosis factor alpha (TNFalpha) and interleukin-1beta (IL-1beta) in the ankle joints were determined. RESULTS: Nanogold bound to VEGF in RA SF, resulting in inhibition of RA SF-induced endothelial cell proliferation and migration. Significant reductions in ankle circumference, articular index scores, and radiographic scores were observed in the nanogold-treated rats with CIA compared with their control counterparts. In addition, the histologic score (of synovial hyperplasia, cartilage erosion, and leukocyte infiltration), microvessel density, macrophage infiltration, and levels of TNFalpha and IL-1beta were also significantly reduced in the ankle joints of nanogold-treated rats. CONCLUSION: Our results are the first to demonstrate that intraarticular administration of nanogold ameliorates the clinical course of CIA in rats. Nanogold exerted antiangiogenic activities and subsequently reduced macrophage infiltration and inflammation, which resulted in attenuation of arthritis. These results demonstrate proof of principle for the use of nanogold as a novel therapeutic agent for the treatment of RA.     

Amelioration of joint disease in a rat model of collagen-induced arthritis by M40403, a superoxide dismutase mimetic.

OBJECTIVE: To investigate the effects of M40403, a synthetic mimetic of superoxide dismutase (SOD), on collagen-induced arthritis (CIA) in rats. METHODS: CIA was elicited in Lewis rats by intradermal injection of 100 microl of an emulsion of bovine type II collagen (CII) in Freund's incomplete adjuvant at the base of the tail. A second injection was given on day 21. RESULTS: Immunization induced an erosive arthritis of the hind paws. Macroscopic evidence of CIA first appeared as periarticular erythema and edema in the hind paws by days 24-26 after the first injection, with a 100% incidence by days 27. Severity progressed over a 35-day period. Radiography revealed soft tissue swelling and focal resorption of bone, together with osteophyte formation in the tibiotarsal joint. Histopathologic features included erosion of the articular cartilage at the joint margins and subchondral bone resorption associated with bone-derived multinucleated cell-containing granulomatous lesions. Treatment with M40403 (2-10 mg/kg/day) starting at the onset of arthritis (day 25) ameliorated the clinical signs on days 26-35 and improved the histologic findings in the joint and paw. Immunohistochemical analysis for nitrotyrosine (a marker of peroxynitrite formation) and poly(ADP-ribose) polymerase (PARP; a nuclear enzyme activated by DNA single-strand damage) revealed positive staining in the inflamed joints of CII-treated rats, suggestive of the formation of peroxynitrite and DNA damage, both of which were markedly reduced by M40403 treatment. Radiographic evidence of protection from bone resorption, osteophyte formation, and soft tissue swelling was apparent in the tibiotarsal joints of M40403-treated rats. Arthritic rats treated with M40403 gained weight at the same rate and to the same extent as normal, nonarthritic rats. CONCLUSION: This study shows that a low molecular weight mimetic of SOD, M40403, attenuates the degree of chronic inflammation, tissue damage, and bone damage associated with CIA in the rat, and supports the possible use of SOD mimetics as therapeutic agents for the management of chronic diseases such as rheumatoid arthritis.     

Prevention of the onset and progression of collagen-induced arthritis in rats by the potent p38 mitogen-activated protein kinase inhibitor FR167653

OBJECTIVE: FR167653 is a potent inhibitor of p38 mitogen-activated protein kinase (MAPK) and inhibits tumor necrosis factor alpha (TNFalpha) and interleukin-1 beta (IL-1 beta) production in inflammatory cells. In this study we investigated the effect of FR167653 on collagen-induced arthritis (CIA). METHODS: Rats with CIA were subcutaneously injected with FR167653 (32 mg/kg/day) starting on the day of the booster injection (day 7) in the prophylactic treatment group and after the onset of arthritis (day 21) in the therapeutic treatment group. Hind-paw swelling, body weight, radiographic and histologic scores, and osteoclast number were evaluated. Cytokine levels in the serum and tissue were assessed by enzyme-linked immunosorbent assays. Flow cytometric analysis of T lymphocytes from bone marrow was performed. The effect of FR167653 on in vitro osteoclast formation induced by soluble receptor activator of nuclear factor kappa B ligand (sRANKL) and TNFalpha was examined. RESULTS: Swelling of hind paws and loss of weight occurred in the CIA rats, but this was not evident in the prophylactic treatment group. Therapeutic treatment also significantly reduced paw swelling. The mean radiographic and histologic scores as well as the osteoclast numbers were significantly lower in the treatment group than in the CIA rats without treatment. FR167653 treatment reduced the serum levels of TNFalpha and IL-1 beta, lowered the IL-1 beta concentration in the ankle joints, and decreased the CD4-,CD8a+ T cell population in bone marrow. Furthermore, FR167653 inhibited the osteoclast-like cell differentiation induced by both sRANKL and TNFalpha in vitro. CONCLUSION: FR167653 prevents the onset of arthritis in a prophylactic treatment model and suppresses the progression of joint destruction in a therapeutic treatment model, suggesting that p38 MAPK is a potential therapeutic target for rheumatoid arthritis.     

Synergistic interaction between methotrexate and a superoxide dismutase mimetic: pharmacologic and potential clinical significance

OBJECTIVE: To investigate the effects of combination therapy with M40403 and methotrexate (MTX) on collagen-induced arthritis (CIA) in rats. METHODS: CIA was elicited in Lewis rats that had been assigned to different experimental groups, and the rats were treated daily, starting at the onset of arthritis (day 26), with M40403 2 mg/kg intraperitoneally, MTX 0.15 mg/kg orally, or combination therapy (M40403 2 mg/kg plus MTX 0.015 mg/kg). RESULTS: The histopathologic features of CIA in type II collagen-challenged rats included erosion of the articular cartilage and bone resorption. Treatment of rats with MTX 0.15 mg/kg orally delayed the development of clinical signs (days 26-35) and improved histologic status in the knee and paw, as clearly demonstrated by a significant reduction in erosion of the articular cartilage at the joint margins and subchondral bone resorption. Furthermore, radiographic evidence of protection against bone resorption and soft tissue swelling was apparent in the tibiotarsal joints of rats treated with MTX 0.15 mg/kg daily. Furthermore, combination therapy with M40403 2 mg/kg plus MTX 0.015 mg/kg exerted significant protection against the development of arthritis, similar to that observed with MTX alone at a dose of 0.15 mg/kg. In contrast, no significant protection was observed in animals treated with M40403 2 mg/kg alone or with MTX 0.015 mg/kg alone. CONCLUSION: This study provides the first evidence that M40403, a potent superoxide dismutase mimetic, exerts a significant synergistic effect with MTX in rats with CIA.     

The effect of apoptosis signal-regulating kinase 1 gene transfer on rat collagen induced arthritis

OBJECTIVE: To examine the apoptosis-inducing effect of apoptosis signal-regulating kinase 1 (ASK1) gene transfer into synovial cells in vitro and in vivo. METHODS: An adenovirus vector was constructed so that a constitutively active form of ASK1 gene (ASK1DeltaN) was expressed in the presence of the Cre recombinase. The ASK1DeltaN and Cre adenovirus vectors were cotransduced into cultured synoviocytes derived from patients with rheumatoid arthritis (RA), and apoptosis was evaluated by TUNEL and Hoechst staining. Collagen induced arthritis (CIA) was induced in 8-week-old male DA rats, and 10 days later the 2 adenovirus vectors were coadministered into the ankle joints of the animals. As indicators of severity of arthritis, swelling of the ankle and articular index (AI) scores were evaluated, while histopathological observation of articular tissue was also performed. RESULTS: In the cultured human RA synoviocytes, overexpression of the ASK1DeltaN significantly reduced cell viability and induced apoptosis. In the CIA rats transduced with the ASK1DeltaN gene, arthritis was significantly promoted in terms of the swelling of the ankle joints and elevation of the AI scores. Histopathological observation also revealed that the constitutively active ASK1 induced massive infiltration of inflammatory cells into the synovial membrane as well as proliferation of synovial fibroblasts. Degeneration of the synovial membrane was not evident. CONCLUSION: Adenoviral transduction of ASK1DeltaN induced apoptosis in RA synoviocytes in vitro, but not in CIA synovium in vivo.     

Effects of combination M40403 and dexamethasone therapy on joint disease in a rat model of collagen-induced arthritis

OBJECTIVE: To investigate the effects of combination therapy with M40403, a superoxide dismutase mimetic (SODm), and dexamethasone (DEX) on collagen-induced arthritis (CIA) in rats. METHODS: CIA was elicited in Lewis rats by an intradermal injection of 100 mul of an emulsion of bovine type II collagen (CII) in Freund's incomplete adjuvant (IFA) at the base of the tail. On day 21, a second injection of CII in IFA was administered at the base of the tail. RESULTS: Lewis rats developed erosive arthritis of the hind paw when immunized with an emulsion of CII in IFA. The histopathology of CIA included erosion of the articular cartilage at the joint margins and subchondral bone resorption. Immunohistochemical analysis for nitrotyrosine, poly(ADP-ribose) polymerase (PARP), inducible nitric oxide synthase (iNOS), and cyclooxygenase 2 (COX-2) revealed positive staining in inflamed joints of collagen-treated rats. The combination therapy with M40403 2 mg/kg and DEX 0.01 mg/kg significantly reduced the development of the inflammatory process and reduced the degree of staining for iNOS, COX-2, nitrotyrosine, and PARP. No significant difference in the degree of staining between the combination therapy and the higher dose of DEX (0.1 mg/kg) was found. Furthermore, radiographic evidence of protection from bone resorption was apparent in the tibiotarsal joints of rats that received the combination therapy. CONCLUSION: This study shows that combination therapy with M40403 and DEX reduced the degree of chronic inflammation and tissue and bone damage associated with CIA in the rat. It supports the possible use of SODm in combination with steroids to reduce the dose necessary and the side effects related to the use of steroids in the management of chronic diseases such as rheumatoid arthritis.