Animal Models of Rheumatoid Arthritis

Animal Models of Rheumatoid Arthritis Fei F. Shih, MD PhD Markey Program Sept 14, 2007 Rheumatoid arthritis •RA is a major inflammatory disease affecting upwards of 1% of US population. •Cardinal feature: symmetrical polyarthritis: distal >proximal •Extra-articular manifestations: malaise, vasculitis, pericarditis, pneumonitis, hematologic abnormalities, autoantibodies (RF, CCP) •Genetic correlation: twin concordance 15-30% Goals of Animal Models Predisposition •Genetic: N.Am Indians •HLA DR4 assoc •Shared epitope •Female Inciting Event ???? Infection? EBV Parvovirus Injury Effectors: Cellular Adaptive Innate Soluble factor Antibodies Complement cytokines Therapy: NSAIDS RA meds Anti-TNFa IL-1Ra Gene therapy ImmunoRx Requirements for Animal Models Reproducible Fast Identifiable effectors Available reagents for experimental manipulations 5. Correlation with human disease 1. 2. 3. 4. Effectors in RA: QuickTime™ an d a TIFF (Uncompressed) decompressor are need ed to see this picture . RA as an immune reaction: role of adaptive immunity (T cells) • MHC and T cells: – association with particular HLA DR4 locus (especially shared epitope LLEQKRAA or LLEQRRAA among several HLA DRb alleles) – Particular HLA-DRB1 alleles associated with more severe disease: suggest presentation of disease-associated peptides to particular CD4+ T cells – large numbers of CD4+ T cells in RA lesions, – skewed TCR usage (expansion of particular, – transient amelioration of RA with CD4 depletion – Therapy with anti-T cell medication (cyclosporin) can help RA as an immune reaction: role of adaptive immunity (B cells) • Important antigen presenting cells to T cells • Production of autoantibodies: – Rheumatoid factor(Ab that bind t o the Fc portion of IgG): only established marker for RA by ACR criteria, found in 6080% of RA pts, but low specificity – Antibodies to cartilage components: collagen II (more reactivity to denatured than native, some to xenogeneic collagen) Seen in 30-70% of RA pts – Anti-keratin antibodies in 40-50% of RA pts. Antigen is a deiminated form of filaggrin. These are generally called anticitrullinated peptides from filaggrin. 67% RA pts but less than 5% of normal controls. – Antibodies to nuclear proteins: ANA, hnRNP – Antibodies to glucose-6-phosphate isomerase RA as an immune complex disease: role of complements Figure 2-19 RA as an immune complex disease: role of FcR (I) RA as an immune reaction: innate immunity • Dendritic cells – Presentation of antigens to T cells – Prominent producer of inflammatory cytokines such as TNFa, IL1, IL6 • Macrophages – Two types of synoviocytes: macrophage-like vs fibroblastlike – Macrophage are prominent feature of pannus – Prominent producer of inflammatory cytokines • Neutrophils – Prominent cell type in inflammed synovium and fluid – Mediators of tissue damage by toxic oxygen radicals and secretion of pro-inflammatory mediators Putting it together Animal Models (induced) • Xeno-reactivity – Collagen-induced arthritis – Antigen-induced arthritis – Proteoglycan induced arthritis • Microbial products(reactive arthritis) – Streptococcal cell wall arthritis – Zymosan(glycan derived from yeast)-induced arthritis – Adjuvant arthritis: only in rats by immunizing with killed mycobacterium tuberculosis, associated with anti-hsp response • Others – Pristane-induced Collagen induced arthritis (CIA) • Requires both CD4+ T and B cells for full spectrum of disease • Role of Antibodies – Passive transfer of disease with CII abs and LPS in susceptible strains in 2-3 days, transient, independent of MHC – Complements and FcR are critical for disease • Role of cytokines – IL-1 and TNFa are required for inflammation – IL-6 not required • Correlation with human disease – Antibodies to CII can be seen in RA patients and normal controls. – Most antibodies are directed toward denatured CII and not native CII, especially in normals. – CII antibodies does not correlate with disease incidence or severity, not assessed routinely for RA •Xeno-reactivity: sequestered antigen and breakdown of tolerance –Earliest animal model of RA, initially established in rats, but can be seen in mice and primates as well –Produced by injecting mice with bovine CII in CFA, initial reactivity to heterologous collagen that subsequently reacts with autologous CII –Particular MHC susceptibility (H-2q, H-2r), restricted TCR Vb genes –Disease in 3-5 weeks after immunization –Affects front and rear paws, sometimes spine, tail, and ear Streptococcal cell wall antigen: role of infection • Infectious agent – Single IP injection of SCW induces chronic erosive arthritis in female rats – Active component is peptidoglycan polysaccharide (PGPS) • Initial stage: (Days 3-10) – T cell independent, complement dependent – Synovial infiltration by PMN and macrophages (days 3-10) • Chronic phase: (2-4 weeks) – Reactivation of arthritis requires T cells, complement independent – Extensive synovial hypertrophy – Cycles of remission and reactivation that can be triggered by microbial productds and inflammatory cytokines • Proposed mechanisms: – SCW activates PMN, macrophages in the joints via TLR – Transient joint inflammation with release and exposure of autoantigen – Inflammation + sequestered antigen, altered antigen-->T cell activation Adjuvant arthritis (AA) • Injection of rats with adjuvant (Complete Freud’s adjuvant) – Produced by injecting rats with CFA (mineral oil with heat killed mycobacteria) – Can also be produced by IFA, suggest that oil is critical – Heat shock protein appear to be the auto-antigen – Similar mechanism for pristane induced arthritis • Proposed mechanism – Macrophages phagocytosed non-degradable oil--> activated and produce large quantities of pro-inflammatory cytokines (IL-1, TNFa, and IFN-g) – Inflammation results in joint damage, exposure of autoantigens, and abnormal expression of MHC II to adaptive immunity – Activation of T and B cells (anti-hsp T cells, hyper Ig) • Microbial product as adjuvant – AA (16-19 days) faster and more severe than pristane-induced arthritis (3-6 months) – More erosive disease with AA than pristane Spontaneous arthritis • Defects in tolerance – MRL lpr: defect in anti-apoptotic molecule Fas,SLE – NZBxNZW: murine SLE – K/BxN: KRN TCR transgenic mouse – SKG: defect in zap 70 – HTLV tax tfransgenic • Cytokine Disregulation – – – – Hu TNF transgenic TNF gene mutation in AUUUA motif IL-1RA knockout IL-6 receptor mutation KRN TCR Transgenic Model of Rheumatoid Arthritis Spontaneous arthritis(K/BxN) Articular • spontaneous • chronic, progressive • symmetrical • distal > proximal • erosive--> joint destruction Systemic • immune hyperreactivity • hypergammaglobulinemia • auto-antibodies (not RF) • splenomegaly Arthritis in K/BxN Mice Normal K/BxN Arthritis in K/BxN Mice Normal K/BxN Kinetics of Arthritis and AntiGPI Ig Seroconversion Pre 5.5 Acute Chronic 10000000 7 Pre 10 10 10 10 1000000 6 Acute Chronic mean ankle thickness (mm) 5 mean ankle thickness (mm) 4.5 serumanti-GPI titer serum a-GPI titer 3 4 5 6 7 age (wk ) 8 9 10 11 5 100000 4 10000 4 1000 103 100 102 3.5 3 101 1 age (wk) 10 2.5 1 3 4 5 6 7 age (wk ) 8 9 10 11 age (wk) K/BxN K/BxN Tg(-) Tg(-) Initiation of an autoimmune response Thymus Lymph node Joint KRN KRN aGPI aGPI Passive Ab transfer arthritis Accum of aGPI in joint Spontaneous arthritis Defects in immune tolerance: – HTLV-tax • Tg for the pX region of the human T cell leukemia virus 1 (Tax transcription factor) • Dysfunction in immune tolerance and cytokine production • BALB/c most susceptible (72%), C57Bl/6 (2%) • AutoAbs: RF, anti-CII, anti-hsp • High levels of inflammatory cytokine production • Highly susceptible to CIA – SKG • Natural mutation in Zap70, key component in TCR signal transduction machinery • Defective signaling “amplitude”-->decreased sensitivity to antigens and negative selection • Escape of autoreactive T cells into the periphery: T cells can transfer disease (does not need B cells) • AutoAbs: RF, anti-CII, anti-Hsp, increased IgG and IC in serum Spontaneous arthritis • Cytokine transgenics and knockouts – Human TNF transgenic mice • Constitutive expression of TNF • 100% mice affected, beginning at 3-4 weeks of age, joint destruction by 9-10 weeks • More severe on DBA/1 but does not need T or B cells • Blocked by abs to TNFa and by ab to IL-1R1. – IL-1ra knockout mice on BALB/c • Needs lymphocytes • Autoabs: RF, anti-CII-abs, anti-dsDNA – Mutant IL-6 Receptor mouse: • point mutation in a phosphatase-binding site of the gp130 subunit of IL-6R • Require lymphocytes • Thymic tolerance impaired • autoAbs are produced Mechanisms of inflammatory arthritis SKG Goals of Animal Models Predisposition •Genes Inciting Event ???? Infection? EBV Parvovirus Injury Effectors: Cellular Adaptive Innate Soluble factor Antibodies Complement cytokines Therapy: NSAIDS RA meds Anti-TNFa IL-1Ra Gene therapy ImmunoRx