Amelioration of established murine collagen-induced arthritis with anti-IL-1 treatment.

Inflammatory cytokines have been implicated in the pathogenesis of rheumatoid arthritis. To validate a key role for IL-1 in arthritic processes we have studied the protective effect of neutralizing antimurine IL-1 antibodies in the murine collagen-induced arthritis (CIA) model. Combination of anti-IL-1 alpha and anti-IL-1 beta given before onset of arthritis was shown to prevent disease completely. Remarkably, a single treatment was also highly effective in the established phase of arthritis, reducing both inflammation as well as cartilage destruction. Suppression was most pronounced with the combination, but anti-IL-1 beta alone also induced significant relief. Finally, we studied the protective effect of IL-1 neutralization on cartilage metabolism in a unilateral expression model of collagen arthritis. To this end zymosan was injected in one knee joint before onset of disease, resulting in accelerated expression in that particular joint and the draining paw. Anti-IL-1 treatment started after accelerated expression of arthritis was able to fully normalize chondrocyte synthetic function, which was highly suppressed in the control group. It is concluded that IL-1 is an important determinant in both inflammation and cartilage destruction in collagen arthritis, and this may have implications for therapy in human arthritis.     

Anti-CD2 (OX34) MoAb treatment of adjuvant arthritic rats: attenuation of established arthritis, selective depletion of CD4+ T cells, and CD2 down-modulation

Anti-CD2 MoAbs have previously been shown to induce tolerance and to block B cell differentiation, T cell and monocyte activation. Since these immune functions are important in joint inflammation, we asked whether administration of the anti-CD2 MoAb OX34 has a beneficial effect on established rat adjuvant arthritis, a model of human rheumatoid arthritis, and how it affects CD2-bearing leucocyte subsets. Female Lewis rats with established adjuvant arthritis received a total of 5 mg OX34 or isotype-matched control MoAb starting on day 15 after adjuvant injection. Weight and arthritis score (AS) were measured in a blinded fashion. Peripheral blood cells were analysed for numbers of leucocyte subsets at various time points. Animals were killed on day 30 and lymphatic organs were processed for immunohistology. Clinically, OX34 treatment led to increased body weight and reduced AS. Although OX34 binds to CD4⁺ and CD8⁺ T cells in a comparable fashion, OX34 treatment reduced CD4⁺ T cells, but not CD8⁺ T cells. Among CD4⁺ T cells CD45RC⁺ (‘naive’) T cells virtually disappeared; CD45RC⁻ (‘recently activated’) T cells were slightly reduced. A reduction of CD4⁺ T cells was also found in the lung, liver, bone marrow, spleen and lymph nodes. Down-modulation of the CD2 molecule by OX34, again, affected CD4⁺ T cells, suggesting a specific signal for CD4⁺ but not CD8⁺ T cells. In conclusion, the anti-CD2 MoAb OX34 attenuates established rat adjuvant arthritis. In spite of similar binding to CD4⁺ and CD8⁺ T cells, OX34 depletes only CD4⁺ T cells and down-modulates the CD2 molecule on these cells. These results suggest a therapeutic benefit from CD2-directed therapy for chronic types of arthritis.     

伴和不伴类风湿性关节炎患者腰椎管狭窄后路减压融合过程中隐性失血的对比

背景:腰椎管狭窄症后路手术除术中出血和术后引流外,还存在大量的"隐性失血"。合并类风湿性关节炎患者可能会影响围术期出血尤其是隐性失血,此前并无报道。目的:针对合并类风湿性关节炎的腰椎管狭窄患者与非类风湿性关节炎行腰椎后路手术时术中出血量、术后引流量以及隐性失血情况进行对比,并探讨类风湿性关节炎患者隐性失血的危险因素。方法:回顾性纳入了65例合并类风湿性关节炎的腰椎管狭窄患者(类风湿性关节炎组),筛选87例未合并类风湿性关节炎的腰椎管狭窄患者(非类风湿性关节炎组),所有患者均采取椎弓根螺钉+钛棒+椎间融合器内固定系统进行腰椎后路减压融合和后外侧融合治疗,术中行自体骨后外侧植骨。提取信息包括人口统计学信息、类风湿性关节炎信息(如类风湿性关节炎病史、Steinbrocker分级、抗类风湿性关节炎药物)、手术信息以及出血量相关指标。以术中出血量、术后引流量和隐性失血作为主要指标;以手术时间、术前术后红细胞压积和血红蛋白及其变化值、手术前后贫血数量、术后新发贫血数量、自体血和异体血输注量等作为次要指标。结果与结论:①类风湿性关节炎组腰椎管狭窄患者平均年龄为(65.97±8.02)岁,平均体质量指数为(25.76±3.68)kg/m^2,非类风湿性关节炎组中患者在性别比例、年龄和手术节段数上均与之匹配;②类风湿性关节炎组中患者平均病程为(16.78±12.73)年,其中单药或联合口服改变病情抗风湿药者最常见,2组在椎弓根螺钉数和椎间融合器置入数量上差异均无显著性意义,围术期并发症发生率2组差异亦无显著性意义;③主要结果对比显示2组在总失血量、术中出血量和术后引流量方面差异无显著性意义,而隐性失血以及隐性失血所占总失血量比例在非类风湿性关节炎组中更低(P<0.001,0.012);根据手术节段数进行分层分析,长节段(≥3节段)手术中非类风湿性关节炎组中隐性失血和隐性失血所占总失血量比例均优于类风湿性关节炎组;④次要指标对比红细胞压积改变值(P=0.021)在非类风湿性关节炎组小于类风湿性关节炎组但血红蛋白减小值2组差异无显著性意义;术后2组新发贫血以及贫血加重情况相比差异无显著性意义,异体血输注和手术时间相比差异也无显著性意义;⑤对类风湿性关节炎组患者隐性失血进行多元线性回归分析显示,类风湿性关节炎的Steinbrocker级别高、未服用改变病情抗风湿药、血红蛋白变化和输注异体血为隐性失血的独立危险因素;⑥提示类风湿性关节炎组和非类风湿性关节炎组在总失血量、术中出血、术后引流和手术时间上无差异,而隐性失血以及隐性失血所占总失血量比例类风湿性关节炎组高于非类风湿性关节炎组,尤其是长节段手术;类风湿性关节炎组的Steinbrocker分级高、未服用改变病情抗风湿药、血红蛋白改变较多以及输注异体血为隐性失血的独立危险因素。     

Human IgG anti-F(ab'')2 antibodies possess rheumatoid factor activity.

Individual preparations of affinity purified anti-F(ab')2 antibodies and anti-Fc antibodies isolated from the sera of patients with rheumatoid arthritis (RA), were examined for reactivity with the Fab and Fc fragments of human IgG. Western blot assays demonstrated specific interaction of affinity-purified anti-Fab antibodies with both Fab and Fc molecules. Approximately one-half of the anti-Fab antibody preparations studied contained IgG antibodies reactive with Fab and Fc fragments in ELISA, suggesting the existence of naturally occurring epibody-like autoantibodies in these patients. Thirteen of 14 affinity-purified anti-Fc antibody preparations contained IgG cross-reactive with Fab molecules in ELISA. Double-adsorption assays on affinity columns demonstrated that a minimum of 14%, and possibly as much as 50%, of the IgG anti-Fab antibodies reacted with the Fc of IgG. Conversely, a minimum of 12%, and possibly as much as 70%, of the IgG anti-Fc antibodies reacted with IgG Fab molecules. Anti-Fab antibodies isolated from non-RA individuals also exhibited anti-Fc reactivity in ELISA, demonstrating the presence of these dual-reactive antibodies in other autoimmune and normal individuals. These studies establish the presence of naturally occurring IgG autoantibodies reactive with both the Fab and Fc fragments of human IgG. Their existence emphasizes the potential of anti-immunoglobulin antibodies to recognize a multiplicity of antigens, possibly including other members of the immunoglobulin supergene family.     

Efficient generation of respiratory syncytial virus (RSV)-neutralizing human MoAbs via human peripheral blood lymphocyte (hu-PBL)-SCID mice and scFv phage display libraries

RSV is one of the major causes of pneumonia and bronchiolitis in infants and young children and is associated with high mortality. RSV neutralizing human antibody (hu-Ab) is known to mediate resistance to viral infection as well as to be an effective treatment for severe lower respiratory tract RSV infection. We have previously demonstrated that human primary and secondary immune responses can be established in severe combined immunodeficient mice engrafted with human peripheral blood lymphocytes (hu-PBL-SCID). By combining this animal model with the single-chain Fv antibody (scFv) phage display library technique, we were able to investigate further its clinical potential by generating a panel of human scFvs that exhibit both high F glycoprotein (RSV-F) binding affinities ( approximately 108 M(-1)) and strong neutralizing activities against RSV infection in vitro. Sequencing analysis of the randomly isolated anti-RSV-F scFv clones revealed that they were derived from different VH families with mutations in the complementarity-determining region 1 (CDR1). The results suggest that: (i) RSV-F-specific human immune responses and affinity maturation can be induced in hu-PBL-SCID mice; and (ii) this approach can be applied to generate large numbers of human scFvs with therapeutic potential. Despite the fact that hu-PBL-SCID mouse and human scFv phage display library have individually been established, our approach contributes a simple and significant step toward the generalization of antigen-specific human monoclonal antibody (hu-MoAb) production and their clinical applications.     

Maintained pregnancy levels of oestrogen afford complete protection from post-partum exacerbation of collagen-induced arthritis.

Pregnancy is known to influence the course of rheumatoid arthritis (RA) in women, as well as type II collagen-induced arthritis (CIA) in DBA/1 mice. A characteristic feature is the remission during gestation and the exacerbation of the diseases during the post-partum period. In the case of CIA in DBA 1 mice, two hormonal changes have been assumed to be critical for the induction of the post-partum flare: (i) the fall in steroid hormone levels from those present during pregnancy; and (ii) surges of prolactin (PRL) release at and after delivery. Our results show that treatment with oestradiol during a short period immediately after parturition protects the mouse from a post-partum flare of the disease, and that treatment with bromocriptine, a drug known to inhibit the endogenous PRL release, has a significant though less marked effect. Studies of lactating (i.e. animals with physiological stimulation of endogenous PRL release) and non-lactating arthritic mice revealed no clear-cut differences, indicating that PRL is of minor importance for the induction of the post-partum flare. Some steroids other than oestradiol, which may be implicated in the exacerbation of arthritis, namely progesterone and hydrocortisone, had no clinical effect. Analyses of agalactosyl IgG levels in mice with CIA, and anti-collagen II antibodies in sera collected at the end of the experiments revealed no significant differences between the oestradiol and the control groups. The successful oestradiol treatment of the mice indicates that the drop in endogenous oestradiol levels prior to delivery ends the oestrogen-mediated protection against arthritis during pregnancy.     

Correlative studies of rheumatoid factors and anti-viral antibodies in patients with rheumatoid arthritis.

An analysis of the relationship between the immune response to ubiquitous herpes family viruses, namely Epstein-Barr virus (EBV), cytomegalovirus (CMV), and varicella-zoster virus (VZV) and the presence of rheumatoid factors (RF), which are autoantibodies characteristic of patients with rheumatoid arthritis (RA), was conducted. Antibody profiles (RF, anti-viral antibodies) were monitored in the serum of the RA patients, and in normal individuals. No patient was found to have circulating RF in the absence of anti-viral antibodies. When the patients and normal controls were subdivided according to the presence of serum RF, it was found that when RF were present, the frequency of anti-CMV antibodies, but not anti-EBV or anti-VZV antibodies, was significantly higher (P = 0.02) when compared with RF-negative individuals. The titres of anti-CMV but not anti-VZV antibodies were found to increase in the RA patients with disease duration. To see if these viruses could stimulate RF production in vitro, peripheral blood mononuclear cells (PBMC) isolated from the patients and normal controls were stimulated with viral antigens. PBMC from normal controls, but not from RA patients, appeared to be responsive to viral antigen stimulation and produced RF. These data suggest that the immune response to CMV, to a greater extent than to EBV or VZV, correlates with the presence of RF.     

Clonality of T lymphocytes expanded with IL-2 from rheumatoid arthritis peripheral blood,synovial fluid and synovial membrane

The association of rheumatoid arthritis (RA) with particular MHC class II genes suggests that autoantigen-spccific T cell clones present in joints could be central to the pathogenesis of the disease. Previous investigations on the clonal diversity of T cells infiltrating the rheumatoid synovial membrane have yielded conflicting results. With the use of Southern blot analysis, we investigated the clonalily of rheumatoid T cell lines expanded from peripheral blood, synovial fluid and synovial tissue. From peripheral blood lymphocyte (PBL) of RA patients and healthy normal controls, we also checked the consequences of two different culture conditions on the clonality of these cell lines. From control PBL, we found that in vitro non-specific expansion of non-clonal T cell populations does not create artefactual clonal selection. However, growing T cells in ritro with IL-2 seems to be able to lead to preferential expansion of cells bearing IL-2 receptor (IL-2R). We identified such in vivo activated IL-2-sensitive T cell clones frequently in RA synovial tissue (8/13) and more rarely in synovial fluid and peripheral blood (3/12). One patient presents the same T cell receptor gene rearrangements in synovial membrane of two affected joints. In RA synovial tissue, the frequency of these IL-2-responsive T cells is most prevalent among actively inflamed membranes removed early in the disease process. The role and the relevance to the disease of these I L-2-responsivc T cells remain to be elucidated.     

可溶性TRAIL重组蛋白治疗CIA实验研究

用sTRAIL重组蛋白治疗CIA模型 ,探讨sTRAIL用于自身免疫病治疗的可能性。将鸡肋骨II型胶原蛋白注射大鼠 ,建立了RA的动物模型———胶原诱导性关节炎 ,自制的可溶性TRAIL蛋白治疗CIA ,观察治疗后关节炎指数与血清中炎性细胞因子的变化。结果是sTRAIL 30 0ng/只 (0 1ml/次 )局部应用于关节炎部位 ,皮下注射 1周以后 ,sTRAIL应用的大鼠症状较对照组指数评分稍有下降 ;患病大鼠较正常大鼠IFN γ和TNF α表达量增加 ,而经过sTRAIL治疗后两者表达量都有显著下降 (P <0 0 5 )。提示sTRAIL局部应用后可缓解关节炎的局部炎症 ,可能是通过抑制激活的淋巴细胞 ,减少炎症细胞因子的分泌量和对炎症细胞的趋化而起作用     

Adhesion of peripheral blood lymphocytes of children with arthritis to human umbilical vein endothelial cells.

To determine whether adhesion of peripheral blood lymphocytes (PBL) of patients with juvenile rheumatoid arthritis (JRA) may be enhanced, adhesion of PBL of children with JRA, children with seronegative spondyloarthropathies (SSA), age-appropriate and adult controls, to human umbilical vein endothelial cells (HUVEC) was assessed in vitro. B and CD4 T lymphocytes in initial, adherent, and non-adherent cell fraction were identified by flow cytometry. B lymphocytes of all the younger subjects combined had a higher adherence to activated HUVEC compared with B lymphocytes of the adult donors. Except for greater adherence of HLA-DR+ CD4 T cells, lymphocytes of children with JRA showed no enhanced adhesion to either unactivated or activated HUVEC. The percentage of B cells adherent to activated HUVEC in each of the subject groups was 1.5-3.6-fold higher than adherent CD4 T lymphocytes. Surface analyses indicated higher percentages of CD49d (alpha 4)+ and CD29 (beta 1)+ CD4 T lymphocytes in adherent cells, but less of a differential in CD49 (alpha 4)+ and no difference in CD29 (beta 1)+ B lymphocytes. There were fewer Leu-8 (L-selectin)+ B and Leu-8+ CD4 T cells among adherent cells. The data suggest a greater adhesive capacity of B lymphocytes compared with CD4 T lymphocytes which is unrelated to disease, and the possibility that B lymphocytes may utilize adhesion molecules distinct from those of CD4 T lymphocytes. Only a small subset of T cells of patients with JRA may have an enhanced capacity for adhesion to endothelium.