Relationship between Th1/Th2 cytokine patterns and the arthritogenic response in collagen-induced arthritis

It is hypothesized that the balance of cytokines produced by Th1/Th2 subsets of T helper cells plays an important role in the development of autoimmune diseases. Murine collagen-induced arthritis (CIA) is an example of an autoimmune disease in which immunization with cartilage-derived type II collagen induces, firstly, a T cell response to type II collagen and, secondly, the manifestation of a destructive inflammatory response in affected joints. We have investigated the role of Th1/Th2 responses in the development of CIA by monitoring levels of interferon (IFN)-γ (a Th1 cytokine) and interleukin (IL)-4 and IL-10 (Th2 cytokines), and IL-1β and tumor necrosis factor (TNF) (pro-inflammatory cytokines) produced by cultured draining lymph node cells (LNC) from collagen-immunized DBA/1 mice during the induction phase of arthritis and throughout the time of clinical manifestation and subsequent remission of the disease. Although a transient increase in IL-10 was detected 3 days after immunization, Th2 cytokine production was found to be almost completely suppressed 6 days after immunization. In contrast, IFN-γ was detected in LNC cultures as early as 6 days after immunization and the addition of type II collagen to the culture medium resulted in an approximately 10-fold increase in IFN-γ production, indicating that a predominantly Th1 response had become established by this time. IFN-γ production by LNC was found to be further increased at the time of clinical manifestation of arthritis and could be up-regulated by co-culture with type II collagen. IL-10 was not detected in LNC cultures at the onset of arthritis and IL-4, although present, was found to be markedly suppressed in LNC cultures containing type II collagen. These findings indicate that Th1 responses are predominant at the time of onset of arthritis and that the activation of collagen-specific Th1 cells may result in suppression of Th2 activity. IFN-γ production declined progressively during the progression and subsequent remission of arthritis whereas levels of IL-10 increased and low, though persistent, levels of IL-4 were detected throughout this period. High levels of IL-1β and TNF-α production were detected at the onset of the disease. The role of Th1 responses in the development of CIA was further emphasized by the observation that immunization of mice with type II collagen in incomplete Freund's adjuvant, which normally fails to induce arthritis, resulted in a predominantly Th2 cytokine profile.     

The role of the B cells in the adoptive transfer of collagen-induced arthritis from DBA/1 (H-2q) to SCID (H−2d) mice

Collagen-induced arthritis (CIA) can be transferred from DBA/1 to SCID mice when native type II collagen (CII) is administered together with spleen cells, arthritis appearing some 14 days after cell transfer. In the present study, we demonstrate that both donor T- and B-lymphocyte populations play a role in this model, and that arthritis arises in SCID recipients of either murine or bovine native CII. Furthermore, the requirement for administration of soluble native CII can be replaced by subarthritogenic doses of serum from Wistar rats with CIA. In this case a fully developed arthritis appears as early as 2 days after cell transfer. However, protein G-purified IgG from CIA rat serum together with splenocytes from arthritic DBA/1 mice does not transfer arthritis. A key role of B cells in this model appears to be the production of a humoral arthritogenic factor since arthritis can be successfully transferred to SCID mice by CIA rat serum administered together with a B cell-depleted splenocyte population consisting of T cells and donor-histocompatible antigen-presenting cells. By contrast, transfer of disease cannot be achieved by co-administration of CIA rat serum and purified donor T cells, indicating that the presence of donor antigen-presenting cells is a requirement for adoptive transfer of arthritis. We propose that joint damage initiated by arthritogenic product(s) of the B cell lineage releases soluble antigens that are presented to T cells which perpetuate the disease. The finding that arthritis can be generated in SCID recipients of CIA rat serum together with splenocytes from non-arthritic DBA/1 mice immunized with denatured CII supports the hypothesis that T cells with specificity for denatured joint components perpetuate disease initiated by humoral factors.     

P-selectin glycoprotein ligand 1 therapy ameliorates established collagen-induced arthritis in DBA/1 mice partly through the suppression of tumour necrosis factor

We investigated the therapeutic potential of P-selectin glycoprotein ligand (PSGL)-1 in established collagen-induced arthritis (CIA) in DBA/1 mice. PSGL-1 is the high-affinity specific ligand for P-selectin and is thus important in cell recruitment to inflammatory sites. I-316 PSGL-1 or rPSGL-1Ig fusion protein were administered to mice after the onset of clinical arthritis for 10 days, and the effect of treatment on both clinical and histopathological progression of disease was studied. It was found that both PSGL-1 biologicals effectively suppressed progression of clinical arthritis, and this was accompanied by protection against damage of joint tissues. We sought to investigate a mechanism underlying the effect of rPSGL-1Ig on the reduction of clinical arthritis. Blockade of PSGL-1/P-selectin interaction blocks recruitment of leucocytes, thus we observed a notable reduction in viable cell numbers of synoviocytes from rPSGL-1Ig treated mice. In view of this finding we suspected an effect of treatment on the production of pro-inflammatory mediators such as bioactive tumour necrosis factor-alpha (TNF) in synovial membrane ex vivo cell cultures. Production of TNF was reduced in arthritic mice that had been treated with rPSGL-1Ig. To further investigate the mechanism of rPSGL-1Ig, we explored the possibility that PSGL-1 might also have a direct signalling effect on TNF release from inflammatory cells. Thus synoviocyte cultures from arthritic mice were incubated with rPSGL-1Ig. A significant reduction in the spontaneous bioactive TNF release from these cultures was noted. We therefore confirmed these surprising findings using cultures of a mouse macrophage like cell line RAW 264.7, stimulated by LPS. Our results indicate that both forms of PSGL-1 have significant therapeutic effects in CIA murine model of RA. The mechanism of action involves reduced cellularity of synovium as anticipated, along with a reduction in TNF production from inflammatory cells in the synovium. The latter mechanism needs further mechanistic analysis.     

Suppression of inflammatory responses after onset of collagen-induced arthritis in mice by oral administration of the Citrus flavanone naringin

Rheumatoid arthritis (RA) is closely related to the pathogenesis of tumor necrosis factor α in lesions. We investigated the suppressive effects of a Citrus flavanone naringin on inflammatory responses in mice with collagen-induced arthritis (CIA), a mouse model for RA. To investigate potential preventive and therapeutic effects of naringin, mice were given naringin orally three times a week from the second immunization with collagen (day 21) and from day 31, when symptoms of CIA had reached a plateau, respectively. In both cases, inflammation-related clinical scores for knee joints were significantly reduced by administration of naringin. Histological analyses demonstrated that representative phenomena, such as damage to interchondral joints, infiltration of inflammatory cells and pannus formation, were significantly depressed by treatment with naringin. In addition, increases in the expression of high-mobility group box-1 protein in the joints of mice with CIA were suppressed by naringin. These results suggest that oral administration of naringin might be effective for treating human patients with RA.     

PGE2受体EP2和EP4调节CIA小鼠脾B细胞表面分子和细胞因子表达

目的: 探讨前列腺素E₂(PGE₂)受体EP2和EP4在胶原诱导性关节炎(CIA)小鼠脾B细胞免疫调节中的作用。方法: 建立CIA小鼠模型,用CD19⁺ 免疫磁珠分选脾B细胞,流式细胞术检测MHCⅡ、CD80和CD86的表达,实时荧光定量PCR技术检测EPs 和细胞因子IFN-γ、TNF-α、IL-6、IL-4、IL-10和TGF-β的表达。结果: 小鼠脾B细胞表达EP的4个亚型,CIA模型小鼠EP2和EP4表达增加;EP2阻断剂可以降低MHCⅡ、CD80和CD86的表达,而EP4阻断剂对CD80没有明显影响;EP2和EP4阻断剂均可以降低IFN-γ、TNF-α 和IL-6 的表达(P<0.05或P<0.01),促进IL-10的表达(P<0.01或P<0.05),并可以分别促进IL-4和TGF-β的表达(P<0.01)。结论: PGE₂可通过EP2/EP4调节B细胞表面分子和细胞因子参与CIA发病,EP2/EP4有可能成为类风湿关节炎治疗的新靶点。     

IL-35 is a novel cytokine with therapeutic effects against collagen-induced arthritis through the expansion of regulatory T cells and suppression of Th17 cells

Epstein-Barr virus-induced gene 3 (EBI3) and the p35 subunit of IL-12 have been reported to form a heterodimeric hematopoietin in human and mouse. We have constructed a heterodimeric protein covalently linking EBI3 and p35, to form a novel cytokine which we now call IL-35. The Fc fusion protein of IL-35 induced proliferation of murine CD4(+)CD25(+) and CD4(+)CD25(-) T cells when stimulated with immobilized anti-CD3 and anti-CD28 antibodies in vitro. The IL-35-expanded CD4(+)CD25(+) T cell population expressed Foxp3 and produced elevated levels of IL-10, whereas the IL-35-induced CD4(+)CD25(-) T cells produced IFN-gamma but not IL-4. The in vitro expanded CD4(+)CD25(+) T cells retained their suppressive functions against CD4(+)CD25(-) effector cells. Furthermore, when cultured with soluble anti-CD3 antibody and antigen-presenting cells, IL-35 suppressed the proliferation of CD4(+)CD25(-) effector cells. Moreover, IL-35 inhibited the differentiation of Th17 cells in vitro. In vivo, IL-35 effectively attenuated established collagen-induced arthritis in mice, with concomitant suppression of IL-17 production but enhanced IFN-gamma synthesis. Thus, IL-35 is a novel anti-inflammatory cytokine suppressing the immune response through the expansion of regulatory T cells and suppression of Th17 cell development.     

青藤碱对胶原诱导性关节炎小Toll样受体4/髓样分化因子88通路的影响

目的 探讨青藤碱(sinomenine,SN)对胶原诱导性关节炎(collagen-induced arthritis,CIA)小Toll样受体4(toll like receptor 4,TLR4)/髓样分化因子88(myeloid differentiation factor 88,MyD88)信号通路诱导炎症的影响.方法 6周龄DBA/1小30只,随机分为正常对照组、CIA组及SN组,每组10只.其中CIA组及SN组小鼠尾根部初次免疫及加强免疫Ⅱ型胶原乳剂诱导CIA模型.小鼠初次免疫d28开始灌胃给药,SN组予SN治疗,正常对照组和CIA组予生理盐水作为对照.连续治疗20 d后处死小鼠,观察药物对小鼠关节肿胀的影响,收集膝关节滑膜及血清.Western blot法检测滑膜TLR4及MyD88蛋白表达,酶联免疫吸附试验(enzyme-linked immunosorbent assays,ELISA)检测血清炎性因子肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)的水平.结果 经治疗后,SN组小鼠关节肿胀情况较CIA组改善明显,差异有统计学意义[d41(5.50±1.38)比(9.67±2.34),P<0.05;d48(1.67±0.52)比(3.33±1.21),均P<0.05];TLR4、MyD88蛋白表达及血清炎性因子TNF-α均较CIA组下降,差异有统计学意义[TLR4/β-actin:(0.33±0.05)]比(0.62±0.05)];MyD88/β-actin:(0.10±0.02)比(0.33±0.03);TNF-α:(169.16±10.68)比(347.97±13.45),均P<0.05].结论 SN可改善CIA小鼠的关节炎症,该作用可能与其抑制TLR4/MyD88信号通路有关.     

Prevention and amelioration of collagen-induced arthritis by blockade of the CD28 co-stimulatory pathway: requirement for both B7-1 and B7-2

Collagen type II-induced arthritis (CIA) is an experimental model of arthritis that has been successfully used to dissect the pathogenesis of human rheumatoid arthritis and to identify potential therapeutic targets. We have used this model to evaluate the role of T cell co-stimulation in both disease development and progression. T cell co-stimulation is provided by ligation of CD28 with either B7-1 or B7-2 present on antigen-presenting cells and can be prevented by a soluble form of CTLA-4 (CTLA-4Ig) which binds with high affinity to both B7-1 and B7-2. We found that administration of CTLA-4Ig at the time of immunization prevented the development of CIA and was associated with lack of lymphocyte expansion within the draining lymph node and failure to produce anti-collagen IgG1 or IgG2a antibodies. To determine which CD28 ligand plays a more dominant role in CIA, we treated mice with monoclonal antibodies (mAb) against either B7-1 or B7-2. Neither anti-B7-1 nor anti-B7-2 had any effect on the course of CIA when given alone, but resulted in reduced incidence and clinical scores when given together. Interestingly, when treatment was delayed until after the onset of clinical disease, both CTLA-4Ig or anti-B7-1 plus anti-B7-2 mAb still ameliorated disease. Effective treatment was associated with a reduction in interferon-γ production by lymph node cells following stimulation in vitro, suggesting that Th1 responses were diminished. This study points to a critical role of CD28 co-stimulation in the development and perpetuation of CIA in DBA/1 mice. Interestingly, it demonstrates an active role for T cells in the later stages of this disease and implicates both B7-1 and B7-2-mediated co-stimulation in the pathogenesis of CIA.     

T cell regulation of collagen-induced arthritis in mice. II. Immunomodulation of arthritis by cytotoxic T cell hybridomas specific for type II collagen

Injection of native type II collagen (CII) to susceptible strains of mice (H-2^q) induces a rheumatoid arthritis-like disease. To study the role of CD8⁺ T cells in the collagen-induced arthritis (CIA),we generated CII-specific T cell hybridomas by fusion of cells from arthritic C3H.Q mice and an AKR thymoma. Two hybrid clones (P3G8 and P2D9) were selected for their ability to lyse syngeneic CH-pulsed macrophages and recognize different antigenic epitopes in association with K^q molecules. When these T cell clones were irradiated and inoculated into (C3H.Q × AKR)F₁ mice 21 days prior to priming with native CII/ complete Freund's adjuvant, the incidence and the duration of CIA were significantly reduced in comparison to groups receiving saline or control T cell hybridoma. Furthermore, both anti-CII T cell hybridomas were able to attenuate CIA in highly susceptible inbred strains of mice and this suppression was antigen and disease specific. The protective activity seems to require intact cells as neither membrane fractions nor cytosolic preparations of the hybridoma T cells retained the vaccinating activity. Most importantly, one of the hybrid clones (P3G8) had a therapeutic effect on CIA since its administration to arthritic DBA/1 mice on day 30 after priming down-regulated the ongoing disease. Taken together, these findings suggest that anti-CII cytotoxic T cell clones can vaccinate against CIA and even reverse the disease.     

Anti-IL-12 antibody prevents the development and progression of collagen-induced arthritis in IFN-γ receptor-deficient mice

In several models of inflammation, including collagen-induced arthritis (CIA), the disease-promoting effect of IL-12 has been attributed to its well-known ability to produce IFN-γ. However, IFN-γ receptor knockout (IFN-γ R KO) mice of the DBA/1 strain have been reported to be more susceptible to CIA than corresponding wild-type mice, indicating the existence of an IFN-γ-mediated protective pathway in this model. In the present study the development of CIA was found to be completely prevented by pretreatment with a neutralizing anti-IL-12 antibody, not only in wild-type, but significantly also in IFN-γ R KO mice. In both strains of mice, the protective effect of anti-IL-12 was associated with lower production of anti-collagen type II antibodies. In vivo stimulation with anti-CD3 antibody in arthritic IFN-γ R KO mice resulted in production of higher levels of circulating IFN-γ, TNF and IL-2 than in corresponding control mice that had not received the arthritis-inducing immunization. This was not the case in arthritis-developing wild-type mice. Furthermore, the protective effect of anti-IL-12 antibody in mutant, but not in wild-type mice, was associated with lower circulating IFN-γ, TNF and IL-2 and higher IL-4 and IL-5 cytokine levels following an anti-CD3 challenge. The data indicate that IL-12 promotes the development of arthritis independently of its ability to induce or favor production of IFN-γ. In fact, any IFN-γ produced in the course of the disease process rather exerts a protective effect. Furthermore, our study suggests that, in the absence of a functional IFN-γ system, endogenous IL-12 exerts its disease-promoting effect by favoring production of other Th1-associated cytokines (IL-2 and TNF), by inhibiting development of IL-4- and IL-5-producing T cells and by stimulating production of anti-collagen autoantibodies.     

Exposure to mercuric chloride during the induction phase and after the onset of collagen-induced arthritis enhances immune/autoimmune responses and exacerbates the disease in DBA/1 mice

In susceptible mice, mercuric chloride induces a systemic autoimmune response that is characterized by elevated serum levels of immunoglobulin G1 (IgG1) and immunoglobulin E (IgE), production of anti-nucleolar antibodies (ANolAs) and the formation of renal IgG deposits. We have previously shown that mercury can also enhance immune/autoimmune responses in mouse strains genetically prone to develop spontaneous autoimmune disease. Here, we investigated whether mercury can enhance the severity of murine collagen-induced arthritis (CIA), an inducible (acquired) autoimmune disease that cannot be induced by mercury itself. While mercury administered prior to the induction phase of CIA exerted little, if any, influence, administration of mercury during the induction phase and following onset aggravated the symptoms of this disease and increased the serum levels of IgE and IgG2a antibodies directed against collagen type II (CII). Furthermore, while animals injected with mercury alone exhibited a significant decrease in the ratio of the levels of interferon-gamma (IFN-gamma) to interleukin-4 (IL-4) mRNA in their spleens, this ratio was increased in mice with CIA, with or without administration of mercury. Finally, the production of anti-nuclear antibodies, a hallmark of autoimmunity in response to mercury, was observed in all mice with CIA treated with this heavy metal. Our findings suggest that exposure to mercury during the development of CIA may influence immunological factors in such a way as to synergistically promote disease development.     

Dendritic cells modulated by innate immunity improve collagen-induced arthritis and induce regulatory T cells in vivo

Dendritic cells (DCs) mediate interactions between innate and specific immunity and may induce regulatory mechanisms. We investigated the effects of modulated DCs in mice with collagen-induced arthritis (CIA) and tested the responses of cells to induced naturally occurring regulatory T cells. DCs were stimulated or not with DNA or lipopolysaccharide (LPS) for 24 hr. DC maturation was assayed, and then modulated DCs were intraperitoneally injected on day 14 into DBA/1 mice to treat CIA. In addition to arthritis scores and type 2 collagen (CII) response, the induction of CD4(+) CD25(+) T cells was analysed by flow cytometry in peripheral blood and the expression of Foxp3, transforming growth factor (TGF)-beta, interleukin (IL)-10 and cytotoxic T-lymphocyte antigen (CTLA)-4 was quantified. Finally, the expression of indoleamine-2,3-dioxygenase (IDO) was assayed in DCs. In comparison with LPS-stimulated DCs, plasmid-stimulated DCs expressed lower levels of major histocompatibility complex (MHC) class II, CD40, CD80 and CD86 molecules and secreted less IL-12p70, interferon (IFN)-gamma, IL-10 and TNF-alpha, displaying a semi-mature phenotype. Compared with non-stimulated DCs, stimulated DCs improved arthritis scores when injected after immunization, without modifying the T helper type 1 (Th1)/Th2 balance of the immune response against collagen. Stimulated DCs induced markers for regulatory T cells (Foxp3, TGF-beta1 and CTLA-4) in vivo. Only LPS-stimulated DCs expressed IDO, which may explain their better therapeutic efficacy. Regulatory mechanisms were induced using DCs modulated by innate immunity stimulators. Innate immunity mechanisms do not require the presence of the disease-causing antigen, even in T- and B-cell specific diseases. Our results have implications for the treatment of rheumatoid arthritis, an autoimmune disease whose triggering antigen has not been identified, and substantially clarify the role of regulatory T cells in CIA.     

Exacerbation of autoimmune arthritis by copolymer-I through promoting type 1 immune response and autoantibody production

Copolymer-I (COP-I) is an unique immune regulatory polymer that has been shown to suppress experimental autoimmune encephalomyelitis (EAE) and is a treatment option for multiple sclerosis (MS). To investigate whether its immune suppressive effects can be extended to other autoimmune diseases, we treated mice with COP-I during the induction of collagen-induced arthritis (CIA). Our results show that COP-I treatment exacerbated CIA, leading to faster onset, more severe and longer-lasting disease. The mechanisms underlying the exacerbation of CIA by COP-I treatment include enhanced activation and inflammatory cytokine production by autoreactive T cells and elevated production of autoreactive antibodies. In addition, germinal center response was significantly enhanced by COP-I treatment. Thus, great caution should be taken when COP-I is to be used in MS patients with other autoimmune complications or its potential therapeutic effects are to be extended beyond autoimmune demyelinating diseases.     

Conversion in vivo from an early dominant Th0/Th1 response to a Th2 phenotype during the development of collagen-induced arthritis

Over the past decade, the central role of T cells in the process of collagen-induced arthritis (CIA) has been extensively documented. The inflammatory features of CIA and its successful modulation after treatment in vivo with Th2 lymphokines, known to down-regulate proinflammatory cytokines, classify CIA as a Th1-mediated disease. However, no direct evidence for the presence of the different T helper subsets has been obtained. To identify the collagen-specific CD4⁺ T cell subset(s) developing during the course of CIA, lymph nodes from susceptible DBA/1 mice (H-2q) were harvested at different times after injection of bovine type II collagen in Freund's complete adjuvant and checked by enzyme-linked immunospot assay for the production of interferon (IFN)-γ and interleukin (IL)-4. The results clearly showed that type II collagen-specific T cells secreting either IFN-γ, IL-4, or both, develop early in vivo, before the onset of arthritis: the number of IFN-γ-secreting cells was already maximal 15 days after immunization, whereas more IL-4-secreting cells were found at day 30, just before the onset of clinical arthritis. Another strategy was to establish collagen-specific CD4⁺ T cell lines and sublines in vitro and to analyze their lymphokine secretion pattern. Lines generated 8 days after immunization displayed a mixed lymphokine secretion pattern characteristic of Th0 cells or of a mixture of Th1 and Th2 cells. After limiting dilution of a day 8 line, 60% of the growing sublines were Th0-like (secreting IFN-γ, IL-4, and IL-5), and 25% were Th1 (secreting IFN-γ). By day 25 post-immunization, 33% of the generated sublines were Th0-like, 11% Th1, and 56% Th2 (secreting IL-4 and IL-5). Moreover, all the sublines raised from the lymph nodes of arthritic mice harvested at day 55 secreted high amounts of Th2 lymphokines, and only 3 out of 14 also produced some IFN-γ. This study demonstrates that during the course of CIA the collagen-specific CD4⁺ T cell response shifts in vivo from a dominant Th0/Th1 response to a clear Th2 phenotype. These results contribute to our understanding of the collagen-specific CD4⁺ T helper subsets which develop during the induction and clinical phases of CIA.     

Blocking of YY1 reduce neutrophil infiltration by inhibiting IL-8 production via the PI3K-Akt-mTOR signaling pathway in rheumatoid arthritis

Systemic lupus erythematosus (SLE) is a complex autoimmune disorder whose pathology involves multiple immune cell types, including B and T lymphocytes as well as myeloid cells. While it is clear that autoantibody-producing B cells, as well as CD4⁺ T cell help, are key contributors to disease, little is known regarding the role of innate lymphoid cells such as natural killer (NK) cells in the pathogenesis of SLE. We have characterized the phenotype of NK cells by multi-color flow cytometry in a large cohort of SLE patients. While the overall percentage of NK cells was similar or slightly decreased compared to healthy controls, a subset of patients displayed a high frequency of NK cells expressing the proliferation marker, Ki67, which was not found in healthy donors. Although expression of Ki67 on NK cells correlated with Ki67 on other immune cell subsets, the frequency of Ki67 on NK cells was considerably higher. Increased frequencies of Ki67⁺ NK cells correlated strongly with clinical severity and active nephritis and was also related to low NK cell numbers, but not overall leukopenia. Proteomic and functional data indicate that the cytokine interleukin-15 promotes the induction of Ki67 on NK cells. These results suggest a role for NK cells in regulating the immune-mediated pathology of SLE as well as reveal a possible target for therapeutic intervention.     

T helper subset involvement in two high antibody responder lines of mice (Biozzi mice): HI (susceptible) and HII (resistant) to collagen-induced arthritis

CD4⁺ T helper cells play a critical role in the chronicity of collagen-induced arthritis (CIA) in mice. The present results focus on the involvement of Th1 and Th2 subsets at the initial stage of the experimental disease in two lines of mice selected for high antibody production: HI that is susceptible, and HII that is resistant to CIA. Both lines are known to be H-2^q, display an identical full set of V-β genes, and mount similar antibody responses to both heterologous and autologous CII. The kinetic analysis of local T cell and anti-bovine CII antibody responses was followed by Elispot assays, the production of interferon-γ (IFN-γ) and IgG2a being considered indicative of a Th1 profile, and interleukin-5 (IL-5) as well as IgG1-IgE, of a Th2 profile. The number of IL-5 Elispots is constantly higher in susceptible than in resistant mice. The IFN-γ production is rather low in HI compared to HII, and besides, preferential help is observed for the Th2-dependent IgG1-IgE isotype-producing B cells in HI, while a switch-over toward IgG2a anti-CII isotype is found in HII. These results suggest that a Th1 preeminence at the onset of the anti-CII response is decisive in the resistance to CIA.     

Modulation of type II collagen-induced arthritis in DBA/1 mice by intravenous application of a peptide from the C1q-A chain.

In this report we are able to show that intravenous (i.v.) application (day 0) of a nonapeptide (residues 26-34) from the human C1q A-chain (designated peptide A-C1q) prior to intradermal (i.d.) administration of chicken type II collagen (CII) in arthritis-susceptible DBA/1 mice (H2q), leads to abrogation of polymorphonuclear neutrophil (PMN) invasion into the joints. This nonapeptide exhibits epitope characteristics and high homology to residues 137-147 of CB11 (a cyanogen bromide fragment of chicken CII, known to contain both arthritis inducing and suppressing determinants). Arthritis index was lowest in animals pretreated i.v. with CII (as internal control), though animals pretreated i.v. with peptide K (residues 137-147 with an additional glycine residue from CB11) or peptide A-C1q exhibited comparative arthritic indices. Only in the arthritis-positive control group (day 0: PBS i.v.) did i.d. application of CII lead to invasion of PMN into the synovial layer and the joint space. Analysis of antibody (Ab) responses at day 48 after i.v. immunization (day 0) and CII challenge (day 7) revealed IgE-Abs to native CII and also to native C1q. IgG titers to CII were highest in animals pretreated with peptide A-C1q. Abs from this group, exhibiting activity to peptide A-C1q (immunizing antigen), were of mainly IgG1 and IgG3 isotypes. Evaluation of the immune response following i.v. application of peptide A-C1q or CII, prior to i.d. CII administration, in DBA/1 mice, revealed IgM responses to peptide A-C1q and peptide K, but not to CII. Intravenous application of peptide A-C1q led to generation of IgG3-Abs reacting only with peptide A-C1q and peptide K, but not with native CII. Additionally, i.v. application of peptide A-C1q elicited IgG responses to a pentapeptide, resembling amino acid residues 26-30 (K-G-E-Q-G) of the C1q A-chain. This five residue antigenic determinant is present in peptide K, in chicken and human CII as well as in human C1q. No specific IgE response to any of the antigens tested could be detected. Since a peptide from the C1q A-chain is both capable of eliciting immune responses and modulating CII-induced arthritis in mice, we postulate that the collagen-like complement component C1q is involved in the development of CII-induced inflammatory arthritic lesions, and may represent, in vivo, the early antigen responsible for inducing anticollagen antibodies prior to CII in hyaline cartilage becoming available as antigen.     

Therapeutic effects of estradiol benzoate on development of collagen-induced arthritis (CIA) in the Lewis rat are mediated via suppression of the humoral response against denatured collagen type II (CII)

The effects of estradiol benzoate (EB) on the development of anti-CII antibodies and their pathogenic potential were studied during the progress of established CIA in the rat. CIA was induced in mature female Lewis rats by two subcutaneous inoculations containing bovine native CII (BCIIn), emulsified in Freund's incomplete adjuvant. Clinical arthritis fully developed by day 18 and then EB (1 mg/kg body wt per day, diluted in corn oil (CO)) was administered intramuscularly every second day thereafter. Antibodies binding four different CIIs (bovine or rat, either native or heat-denatured) were detected in sera and joint tissue extracts by means of solid-phase ELISA. Pharmacological doses of EB (>0·2 mg/kg body wt per day) caused significant remission of established CIA 5-7 days after treatment, and selectively suppressed the production of antibodies specific for denatured CII. To evaluate the arthritogenic potential of circulating anti-CIId IgG, transfer experiments were performed. IgG anti-CIIn, purified from EB-treated CIA rats, was not arthritogenic, whereas IgG anti-denatured (CIId), purified from CO-treated CIA rats, caused severe passive arthritis. Furthermore, pretreatment with rat CIId protected against subsequent induction of CIA, and this protection was associated with suppressed antibody production against CIId. Collectively, our results indicate that antibodies specific for CIId are involved in the pathogenesis of CIA, and that oestrogen-related remission of clinical arthritis may be caused by a selective suppression of antibodies produced against degraded/denatured CII.