Autoimmunity

AUTOIMMUNITY A major cause of morbidity and mortality in the world

Autoimmune diseases Systemic Lupus Sclerodermia Vasculitis Organ-specific Type 1 diabetes endocrine pancreas Asthma lung Rhumatoid arthritis joints Myositis muscle Thyroiditis (Graves, Hashimoto) thyroid APECED endocrine glands Multiple sclerosis nervous tissue Anemia, thrombopenia, neutropenia blood cells Psoriasis, eczema skin Gastritis stomach Colitis (Crohn, UC) colon Myasthenia neuromuscular junction Uveitis eye Rhinitis nose … ..

Many autoimmune diseases are difficult or impossible to cure for the obvious reason that the focus of the immune response – self antigens- cannot be eliminated The ultimate mechanism involves a failure of tolerance and the accumulation of irreversible damage in the target tissues

Proof of autoimmunity Damage in tissues due to immune cell infiltration Disease transferable by lymphocytes and / or serum Presence of cells/antibodies directed against self components

Scheme of immune cell function Central Repertoire Selection Homeostatic expansion Half-life Self agonist selection Antigen-driven expansion Innate cell priming Secondary lymph organ Effector functions DEATH SELF TISSUE SELF TISSUE SURVEY/REPAIR Peripheral pool Regulatory activity Memory

Epidemiology Diseases affecting young adults Female > male Triggering and / or evolution often influenced by infectious diseases, pregnancy, traumatisms Familial trends North-South gradient Higher prevalence in nordic countries Inverse distribution with infectious diseases

Genetics Evaluate the degree of familial clustering Concordance rates between monozygotic versus dizygotic twins Increased risk of developing the disease in affected families Genetics of single gene disorders Risk confered to one individual is high but impact on the population is minimal due to the presence of rare variants Deterministic relationship between the variant and the disease state Genetics of common autoimmune diseases Common gene variants Complex inheritance mode Complex phenotypes Likely overtime selection of gene variants by environment

AIRE and central tolerance Autoimmune polyendocrine syndrome (APS-1): a multiple attack against endocrine organs, skin and other tissues The AIRE gene controls the expression level of several promiscuously expressed antigens in the thymus Lack of AIRE leads to impaired tolerance by failure of central deletion

FOXP3 and regulatory T cells FOXP3, transcription factor of the forkhead family Lack of FoxP3 in mouse (scurfy mouse) leads to the absence of regulatory T cells Mouse equivalent of the rare human IPEX disease (immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome)

FAS and lymphocyte apoptosis Fas (CD95), prototypic death receptor of the TNF family binds Fas-L Lpr/lpr and gld/gld strains of mouse share a common phenotype of lymphoproliferation and autoimmunity Lpr codes for Fas and gld for Fas ligand The APLS (autoimmune lymphoproliferative syndrome) disease mimics the mouse phenotype Fas contributes to the deletion of activated lymphocytes

Le modèle murin lpr / gld Mutations de Fas (lpr) ou Fas-L Lymphadénopathie et autoimmunité de type lupique Homme: APLS IL-2 Expansion Mort Tolérance périphérique par délétion Fas-médiée

CPA Fas TNF-R Fas-L TNF CD40-L, TNF CD40 Activation Survie Mort Costimulation CD28 -> NF-kB

Répertoire périphérique Répertoire naïf tolérisé mémoire Temps

Voie Fas Déficit Fas / Fas-L et caspases -> ALPS: Lymphoprolifération et Autoimmunité

Impact of single gene disorders on AI Central Repertoire Selection Homeostatic expansion Half-life Self agonist selection Antigen-driven expansion Innate cell priming Secondary lymph organ Effector functions Death SELF TISSUE SELF TISSUE SURVEY/REPAIR Peripheral pool Regulatory activity Memory AIRE FOXP3 FAS

Genetics of complex autoimmune diseases

Complex autoimmune diseases Chronic conditions elicited by a loss of immunological tolerance to self antigens Clinical manifestation: organ(s) failure Physiopathology: immune-mediated inflammatory disorder (IMID concept) 4-5% of the population, females > males Most common AIDs: Type 1 diabetes, Rhumatoid arthritis, lupus, Graves’ disease, multiple sclerosis, pernicious anemia 1 in 30 individuals affected: major health problem (2 x cancer ?)

Impact of complex gene disorders on AI Central Repertoire Selection Homeostatic expansion Half-life Self agonist selection Antigen-driven expansion Innate cell priming Secondary lymph organ Effector functions DEATH SELF TISSUE SELF TISSUE SURVEY/REPAIR Peripheral pool Regulatory activity Memory

Strategy for analysis 1. Defining immune phenotypes 2. Phenotype / Genotype linking 3. Defining gene polymorphism 4. Identifiying pathogenic mechanisms 5. Defining epistatic interactions (additive / synergistic / suppressive) 6. Reconstituting the pathology

Immune features in AID Autoreactivity Autoantibodies RA, MG, SLE T lymphocytes T1D, Thyroiditis Altered regulation IPEX, APECED Cell activation Innate immune cell activation Lymphocyte hyper/hypo activation SLE Inflammation (C’, …) Pemphigus Homeostatic phenotypes Cell death T1D, SLE Microenvironmental alterations SLE Various AID display at various levels these phenotypes Some are responsible for the pathogenesis, others are associated to it, few are specific for a given pathology

Directly pathogenic (myasthenia (AChR), FVIII deficiencies, Graves’ disease (TSH), autoimmune hemolytic anemia, neutropenia, thrombocytopenia) FcR dependent effects and innate cell degranulation Immune complexes (mixed cryoglobulinemia, SLE, RA, vasculitis) The case of Autoantibodies

Immune complexes Polyclonal Ab Multi-epitope Ag binding Optimal concentrations Deposition on basal membranes and inflammation/destruction

Complement J. Bordet (1870-1961) C’ is a system of plasma proteins that interacts with pathogens to mark them for destruction by phagocytes

Figure 2-19 - Direct binding on bacteria C-reactive protein (Pneumococcal C polysaccharide) - Ag-Ab complexes

C3 convertase Hydrolysis of C3 causes initiation of the C’ alternative pathway

C5 convertase Surface-bound C3 convertase deposits large numbers of C3b fragments on pathogen surfaces and generates C5 convertase activity

Small fragments of some Ct proteins can initiate a local inflammatory response C5a

Direct pathogenic role of autoantibodies Myasthenia gravis Anti-acetylcholine receptor on the postsynaptic neuronal plate Blocks acetylcholine neuromuscular transmission Leads to muscle paralysis Improvement by plasma exchange and IVIG Presence of C’ fixing on myelinated nerves Goodpasture’s syndrome: Anti-glomerular basement membrane Ab Kidney failure

Immune complexes, C’ and FcR IC deposition, C’ activation Kidney, lung, blood vessels -> kidney failure, lung hemorrhages, vasculitis Hepatitis C associated cryoglobulinemia (purpura, glomerulonephritis, motoneuritis) Role of C’: anti-C5 inhibitor antibody (eculizumab) in RA IC-mediated FcR activation Linked polymorphism of low affinity IgG FcR with AID (CD32, CD16) -> impact on phagocytic property of APC IVIG useful in idiopathic thombopenic purpura (engagement of inhibitory Fc  RIIB)

Pathogenic role for B cells Efficacy of B cell depletion in some AID Rituximab : anti-CD20 Ab CD20 marker of B cells, absent from plasma cells Modes of action B cell -> autoAb depletion Delayed reconstitution Abrogation of B-cell dependent downstream effectors including T-B cooperation (i.e. improvement of lupus by anti-CD40L mAb) and neolymphogenesis ITP: no direct correlation between auto-Ab levels and B cell depletion using Rituximab RA: impact on rhumatoid factor levels

Modulation of B cell survival The BLys/BAFF pathway, members of the TNF/TNF-R family Required for B cell survival Increased serum BLys/BAFF levels in Sjogren, SLE patients Transgenic BLys mice develop Sjogren’s and lupus-like diseases Blocking BLys signals improves SLE in mouse

Other players Lymphocytes: T, NK, regulatory, … Innate cells Acute phase proteins Tissues (mode of response to stress, fragility, organisation)

Effector mechanism in RA GPI / Ig C3b deposition C3 convertase C5 convertase ALTERNATIVE C’ PATHWAY C5a C5aR properdin Fc  R GPI / Ig Factor B Neutrophils Mast cells Fc  R ARTHUS REACTION AMPLIFICATION LOOP

RF in RA Anti-IgM anti-Fc IgG Ab Heavily mutated VH regions witnessing memory B cells

Mouse models of RA Collagen-induced arthritis (CIA) ZAP70 mutation RF transfer does not cause disease Role of B cells in pathology : altered APC ? KRN model TCR Tg mouse: anti-RNase / I-Ak crossed on NOD -> joint inflammation, synovitis, infiltration (neutrophils ++, few T/B cells), cartilage and bone resorption; no RF, vast IgG deposits in many organs on basal membranes Essential contribution of H-2 g7 (disease in B6 g7 ), alloreactivity of the Tg TCR, partial central and peripheral deletion T and B cell dependent, requires T-B MHC restricted interaction, antigen presentation by B or APC, transferable by serum (IgG1 dominant) -> transient disease Glucose-6-phosphate isomerase, an ubiquitous self antigen seen by Tg T cells and immune Ig Requirement for FcR  , C5/C5aR, C3/B factor for disease induction (C5a is strongly attractive for neutrophils) -> Fc  RIII expressed by numerous innate cells, activated by Ig (Arthus reaction), importance of the alternative C’ pathway C3b2-IgG-properdin-factor B complex (neutrophils secrete properdin), amplification via C5 activation GPI in RA ? Evidence for affinity matured and somatically mutated IgG in the sera of RA patients, high concentration of GPI on synovial lining and arterioles, possible binding to a membrane receptor (growth factor function ?)

Non-immune and environmental parameters Infections Trigger or protector Difficult to identify (delay, variations, non contingent with AID) Food Mode of life (stress, neural factors) Sex: hormones, pregnancy, puberty, Age

Linkage studies Inheritance of AID susceptibility is complex MHC exerts a predominant influence Many genomic segments show weak statistical association (lod scores from 2-5 versus 30 for a fully penetrant Mendelian disease locus) Multifactorial diseases result from the combined impact of multiple susceptibility genes, further enriched by poorly defined environmental factors Complex but chronic so in principle accessible to therapeutic interventions if predictions are reliable

Family studies in human AIDs Genetic influence Multigenic  s 10-40 0.2-1 2-4 3-4 12-15 Rhumatoid arthritis 0.2 2-5 2-5 24-57 Lupus 0.1 3-5 0-5 25 Multiple sclerosis 0.4 6 0-13 30-50 Diabetes % Non-twin siblings Dizygotic twins Monozygotic twins Population prevalence Concordance rate (%) Disease

MHC: a central player 15% MHC identical siblings MHC weight Other genes 0.4 6 0-13 30-50 Diabetes % Non-twin siblings Dizygotic twins Monozygotic twins Population prevalence Concordance rate (%) Disease

Complex diseases Genetic contribution MHC locus Environmental / individual contribution Extrapolated from global familial risk of disease

Linkage to MHC alleles Suggestive of T cell mediated dys-immunity Suggestive of a restricted set of primary autoantigens (still ill-defined) Existence of MHC molecules with peculiar antigen-binding properties Peptide register shifting in MHC class II groove renders the predictive task more difficult

Difficulty of linkage analysis Most MHC-linked diseases are multigenic Strong linkage desequilibrium in the MHC locus

MHC susceptibility locus for T1D HLA-DQ  1, HLA-DR  1 : still candidate alleles , not definitively proven Mouse equivalent: I-A g7 (Tg allo-MHC protective) Frequency of MHC susceptibility alleles in Caucasian population: 53% => More frequent than protective alleles !! Inheritance Recessive mode for susceptibility Dominant mode for protection Incomplete penetrance of susceptibility alleles

T1D And MHC Conserved Extended Haplotypes 0.14 B57, SC61, DR7 0.21 B44, FC31, DR7 0.22 B7, SC31, DR2 Protective 0.45 B35, SC31, DR5 0.77 B44, SC30, DR4 1.0 B35, FC(3,2)0, DR1 1.0 B60, SC02, DR6 Neutral 2.1 B8, SC01, DR3 4.4 B62, SC33, DR4 5.8 B62, SB42, DR4 14.6 B18, F1C30, DR3 17.0 B62, SC31, DR4 Susceptibility Ratio (T1D:normal) CEHs [ HLA-B, complotype, DR ] CEH type

Genetic complexity and heterogeneity Association studies differ between ethnic groups Genetic localisations of AIDs vary between disease models T1D red MS blue RA green

Single Nucleotide Polymorphism SNP Most abundant form of DNA variation Around 7 million SNPs with MAF around 5% (minor allele frequency) and 4 million SNPs with MAF between 1 and 5%, plus numerous rare SNPs Relation between common variants and human phenotypes (height, eye color, disease susceptibility) not known Global approach for a dense SNP map : oligonucleotide arrays Hinds et al (Science 2005): 1.6 million SNPs mapped covering 95% of the genome (inter-SNP intervals < 50kB) -> informative of population ancestry, but in agreement with the notion that most common DNA variations are shared across human populations

Causal variants Table 2 papier Rioux 10 millions SNPs with a minor allele frequency greater than 1% Common haplotypes

Penetrance of susceptibility genes Basic observation: discordance rate between monozygotic twins => incomplete penetrance of susceptibility genes Environmental triggers (difficult to define !) Intrinsic parameters: parental imprinting, allelic exclusion of BCR and TCR, monoallelic expression of cytokines, NKR Importance of other non-MHC genetic loci

Features of SLE Disease heterogeneity in humans (glomerulonephritis, hemolytic anemia, vasculitis, arthritis, cutaneous lesions, …), drug inducibility Complex interactions between hormonal, genetic and environmental factors 9:1 female gender bias Lethality linked to glomerulonephritis (IC) Numerous antoantibodies to nuclear antigens Existence of various mouse models displaying some features of the human disease

Autoantibodies in SLE Antinuclear Ab Sensitive test for SLE Not SLE specific Not associated with severity of SLE (flares without ANA) High ANA levels in the absence of disease Anti-Sm Ab specific for SLE Anti-RNP Ab associated with subset of patients with mixed connective tissue disease Anti-phospholipid Ab predisposition to thrombotic events Anti-ribosomal P Ab increased risk of CNS disease Anti-Ro (SSA) Ab increased risk of heart block in newborns of SLE patients

Immunological diagnosis Autoantibodies Antinuclear Ab Anti ds DNA Ab Anti Sm Ab Complement: decrease C3, C4 (disease activity) Inflammatory markers (sedimentation rate, acute phase proteins) Anemia, …

Spontaneous murine models of SLE -> Analysis of phenotypes

Genetic factors Large number of loci Epistatic interactions Linked gene clusters Suppressor genes Threshold effects

Mode of analysis Immunological tools: defining cell phenotypes Genetic tools: linkage analysis, backcrossing studies, transgenic approaches Biochemical tools: probing candidate pathways

Candidate susceptibility genes HLA: DR-B1 (DR2, DR3) Genes of the HLA region in linkage desequilibrium: MHC class III (TNF, TAP, HSP) Fc  R: low affinity receptor for IgG and IgG IC Complement: classical pathway

Genetic association studies of systemic lupus erythematosus. Multiple alleles 18q21 151430 Bcl-2 297C/416G 10q24 134638 FAS – 844C 1q23 134637 FASL 230A 10q11.2-q21 154545 MBL AQ0 6p21 120790 C4 TNFB*2 6p21 153440 TNFβ TNF2 6p21 191160 TNFα DR2/DR3 6p21 152700 142860 HLA-DR3, -DR2 PD-1.3A 2q37 600244 PDCD-1 +49G 2q33 123890 CTLA-4 Multiple alleles 1q31-32 124092 IL10 F176 1q22-23 146740 FcGR3A R131 1q22-23 146790 FcGR2A Associated allele(s) Cytogenetic location OMIM a Gene

Examples of genetic manipulations that lead to lupus-like disease in mice. No single gene defect is sufficient for lupus development Defective hormone signaling ERα Cytokine production abnormalities TGFβ, IL-10, IFN-γ (Tg) IL-4 (Tg) Dysregulated lymphocyte activation caused by intracellular signaling molecule mutations SHP-1, Lyn, PKC-d, P21, E2F2, Stra13, Gadd45a, Rasgrp1, Fli-1 (Tg), SOCS-1, LIGHT (Tg) TSAd Dysregulated lymphocyte activation caused by receptors and their ligands FcγRIIB, TACI, CD22 TGFβRII, CD45*, PD-1 BAFF (Tg) Defective clearance of apoptotic cells, cell debris, and/or ICs SAP, DNase, Mer, MFG-E8, IgM (secreted), C1q, C4 Defective apoptosis Fas, Fas ligand, Bcl-2 (Tg), IEX-1(Tg) Mechanism Targeted gene product a

Lupus nephritis: pathogenic Ab

Physiopathology of Lupus Apoptosis defect Defective clearance Of apoptotic cells Loss of tolerance To apoptotic self Complement deficiency Immune complex deposition Hyperactivation of self reactive B cells The tolerance hypothesis The clearance hypothesis

SLE pathogenesis as a progression of successive stages In the lupus prone NZM mouse, Sle1+2+3 are the major susceptibility loci Triple congenic mice on a B6 background reconstitute the pathology (Koch’s postulate) Sle1 C’, Sap Loss of tolerance to chromatin leading to ANA production, B cell defect Sle2, lpr, yaa, Blys Progressive expansion of autoimmune response with epitope spreading Sle3 Dysregulation of CD4+ T cells, non T-cell autonomous Sle6 FcR End organ damage culminating in fatal lupus

Physiopathologie du lupus D’après Morel et al, Immunity 1999 Sle1 Sle2,3 Lpr, gld, yaa Sle6 Rupture de tolérance / chromatine ANA Expansion, ANA ++ Lésion d’organe Lupus fatal NZM loci

Sle1, a cluster of functionally related genes Human and mouse studies converge on mouse telomeric chr1 and human 1q21-44 Mutations of genes encoded in this region FcgRIIA, FcgRIIIA, FcRg and SAP modulate lupus susceptibility Backcross studies B6.Sle1 selective loss of tolerance to chromatin (H2A/H2B/DNA subnucleosomes) and spontaneously activated T and B lymphocytes But normal immune responses and normal lymphocyte apoptosis

Contribution of B cells in Sle1 effect Bone marrow reconstitution studies: B6.Sle1 bm -> B6: autoantibodies, activ CD4+ B6 bm -> B6.Sle1: normal Cotransfer of bone marrows: B6’, B6.Sle1 Ig Allotype marker: « a » for B6’, « b » for B6.Sle1 Autoantibodies of the « b » allotype Activ B cells of the « b » allotype B cells carry the Sle1 phenotype in a cell-autonomous fashion but autoantibodies are non nephrogenic

Dissecting further Sle1 Genetic recombinants in Sle1 -> 1a, 1b, 1c using new microsatellite markers polymorphic between B6 and B6.Sle1 Congenic lines on B6 background (> 800 mice analysed !!) Autoantibodies and activ CD4+ cells: 1b > 1a and 1c 1a + 1b = 1a + 1b + 1c = WT Sle1 Nephrogenic Ab ? : cross Sle1 congenic mice on the NZW background and analyse proliferative GN: GN correlates with Sle1b in (NZWxB6.Sle1)F1 and activ CD4+ T cells but not with higher autoAb titers Additional locus in Sle1b controls the extent of GN (Sle1d => Suppressive effect ?) Candidate gene for Sle1c: CR2 ?

Regulating the Sle1 effect Sle1 Sle2,3 Lpr, gld, yaa Sle6 Rupture de tolérance / chromatine ANA Expansion, ANA ++ Lésion d’organe Lupus fatal Sles1 Sles1 is present in the lupus-prone NZM strain and attenuates the pathology in this strain via epistatic interactions

Apoptotic cells must be cleared At least two steps: - clearing - digesting and masking

Opsonisation via C1q Flexibility and versatility of ligand recognition Heterotrimer Modular assembly Charge Spatial orientation Symmetrical trimer of  -sandwich subunits (TNF superfamily) Important role in Clearance Altered self Infectious nonself Inflammation

C1q-based recognition C1q engagement leading to MAC activation immune complexes (IgG) -> MAC non immune molecules (polyanions such as LPS, DNA, RNA,polysaccharides, viral membranes, Prp, and unidentified microbial components) opsonisation CRP, SAP, pentraxin : regulators of opsonin activity towards altered self debris Fibronectin, fibrin, …: involved in wound healing natural antibodies In conclusion: Higly diversified recognition modules with combinatorial potential Recognition of ALTERED SELF versus NON-SELF recognition leads to differential effector functions (opsonisation versus lysis ?)

Le rôle du complément D’après Janeway-Travers, Immunobiology

Le facteur H régule l’activation du C’

CR1, récepteur à C3b, C4b, iC3b (CD35) Expression par B, neutro, mono, GR, FDC, ep glomérulaire Cofacteur de I (serine estérase) et H : conversion C3b -> C3bi Fonction d’opsonisation des structures fixant C3b, C4b , iC3b Rôle régulateur des seuils d’activation du lymphocyte B

Rôle des pentraxines Proteine C Réactive (CRP) : - fixation sur les corps apoptotiques (phosphorylcholine) - recrutement de C1q, activation du C’ et donc opsonisation - recrutement du facteur H qui bloque les C 3 et C5 convertases 2. Serum amyloid protein (SAP) Fixation et solubilisation de la chromatine libre TGF  CR1 C3b C3bi Corps apoptotiques CRP CRP CRP C1q C1q C1q H C3b C3b Phagocyte

Pentraxins Protomers Oligomers CRP SAP

A famous pentraxin: C-reactive protein Highly conserved plasma protein (vertebrates and invertebrates) Marker of Acute Phase Response (associated with clotting proteins, C’, anti-proteases, transport protein) Plasma concentration increased by inflammation (x > 1000), induced by IL-6, IL-1  (via C/EBP) Produced mainly by hepatocytes (  g/ml range) Recognition molecule for patterns exposed during cell death and on pathogen surfaces

C-reactive protein complexed with phosphocholine 5 non covalently associated protomers, symmetrically arranged around a central pore

Plasma membrane asymmetry phosphatidylserine and phosphatidyletholamine phosphatidylcholine and sphingomyelin glycosphingolipids

Interaction of CRP with C1q Recognition face binds ligand : « masking effect » ? Effector face binds C1q or Fc  R

Effector function: opsonin ? Activator of C’–mediated opsonisation (up to C3 convertase) due to recruitment or induction of inhibitory factors (H, CD59, DAF, …) FcR-mediated phagocytosis

CRP role Hiding SELF ? rather anti-inflammatory towards self components (ex: lupus) -> proinflammatory towards bacteria, provides protection towards PC and PE+ bacteria

C’ bridges innate and adaptive immunity Carbohydrate-rich particles lacking sialic acid Immune complexes Non immune molecules (CRP, SAP, polyanions, Microbial ligands)

Apoptose et lupus ? Cellules apoptotiques, sources d’autoantigènes ? Clairance des cellules apoptotiques Réponse immunitaire anti-corps apoptotiques Rôle du complément dans la clairance Rôle des anticorps anti-C1q

Déficit en complément et prédisposition au lupus % LUPUS Homme Concordance Souris entre jumeaux C1q 93 90 ++ C1r/s 57 67 C4A/B 75 80 ++ C2 10 +/- C3 rare - MAC rare -

Phenotypes of SLE patients Low CRP levels in serum Reduced DNAse 1 activity C’ defects Reduction in the degradation capacity of necrotic-derived chromatin

Cellules apoptotiques, source d’autoantigènes ? Auto-anticorps du lupus Corps apoptotiques a) Ag intracellulaires -> chromatine intracellulaire + -> spliceosome + -> snRNP + b) Antigènes membranaires : Phospholipides (PS) + c) Protéines plasmatiques :  2 glycoprot I, C1q + Ag ubiquitaires, abondants => rupture de tolérance improbable NéoAg ? Externalisation Ag CA in vivo rares ?

Présentation antigénique par les CPA infection capture réplication Virus MHC TCR T CPA Signal 1 MHC I MHC II Voie exogène Voie endogène

Expérience: 1. Immunisation de souris avec des tumeurs syngéniques apoptotiques 2. Injection de tumeurs normales -> mort ? A) Immunisation avec des DC « pulsées » -> protection anti-tumorale => immunisation B) Immunisation avec des Macrophages -> croissance tumorale => tolérance Conclusions 1. Les cellules apoptotiques peuvent être immunogènes 2. La CPA impliquée (ou son état) décide de l ’issue D ’après Ronchetti et al, 1999

Cross-Présentation antigénique Corps apoptotiques MHC TCR T CPA Signal 1 MHC I MHC II Voie « croisée » Régulation ?

Facteurs régulateurs Complément Nature de la CPA Auto-anticorps Inflammation

C’ et inflammation C’ consommé en phase active de lupus - C1q, C2, C4, plus rarement C3 (formes sévères) Dépôt de C’ -> associé aux IC -> pas strictement corrélé aux lésions tissulaires Clairance des IC : CR versus FcR ? Rôle des anticorps anti-C1q ? -> Vascularite urticariante (HUVS) -> Lupus (1/3 patients +)

Expérience: 1. Péritonite stérile (thioglycolate IP) -> macrophages péritonéaux 2. Injection de thymocytes apoptotiques -> clairance in vivo ? Souris Clairance Lupus Contrôle +++ - C1q KO - +++ C4 KO + ++ C3 KO +++ - Conclusion: La déplétion en C1q entrave la phagocytose des corps apoptotiques et prédispose au lupus

CPA Virus CPA primée Le deuxième signal est induit par l ’inflammation et permet l ’expression des molécules B7 par les CPA Inflammation poly Antigène Signal 1 Signal 2 B7 TCR CD28 Virus BCR T B = CPA HELP Anticorps Effecteurs Le rôle de l ’inflammation

Apoptose et inflammation macrophage CPA primée B7 Elimination discrète ! Agitateurs Toll, IFN, TNF, IL-1 Convocation du lymphocyte et identification du cadavre Cross-présentation Co-stimulation Recrutement Toll

Corps apoptotique Phospholipides annexin V,  GPI C1q Ag nucléaires snRNP 1. Expression des autoAg à la membrane des corps apoptotiques 2. Clairance médiée par les auto Ac -> capture Fc  R-dépendante 3. Consommation de C1q -> perte du pouvoir « solubilisant » du C ’ pour les complexes immuns 4. Conséquence systémique -> dépôts d ’IC dans les tissus 5. Début des lésions tissulaires Le rôle des auto-anticorps ?

Corps apoptotique Macrophage Cellule dendritique C ’ PCR Corps apoptotique C ’ PCR Auto Ac CR1, CD36, …. CR1, CD36, …., FcR IL-1, TNF  Présentation croisée TGF  , IL-10 Rupture de tolérance Tolérance FcR ?

Corps apoptotique Macrophage « Eat me » signals « Cool down » or « Turn me on » signals CHO iC3b PS C1q b2GPI TSP ox LDL CD14 lectin b2 intégrine PS-R C1q-R CR1 CD36, intégrines CD68 ... Distribution tissulaire des récepteurs - différentielle - régulée Fonctions différentes des récepteurs Tether -> Trigger

Apoptose Clairance Prédisposition Déficit en C ’, CRP, … Défaut de maturation des CPA... Excès de mort Lésion Lupus Infection Gènes etc Gènes

Autoantibody-mediated pathogenesis of SLE. Lymphocyte activation in GC IC formation Tissue inflammation and lesions Apoptosis defect But not all autoAb and IC are pathogenic

D’après Immunological Rev, 2001 Clairance des IC: rôle du C’ et des GR

Lymphocyte activation defect Multiple alleles 18q21 151430 Bcl-2 297C/416G 10q24 134638 FAS – 844C 1q23 134637 FASL TNFB*2 6p21 153440 TNFβ TNF2 6p21 191160 TNFα PD-1.3A 2q37 600244 PDCD-1 +49G 2q33 123890 CTLA-4 Multiple alleles 1q31-32 124092 IL10 Associated allele(s) Cytogenetic location OMIM a Gene

Gadd45a deficiency Growth-arrest and DNA damage-inducible gene Downstream effector of p53 Participates to growth arrest, genomic stability and apoptosis KO mouse: lupus-like disease

Immune phenotypes Glomerulonephritis Autoantibodies Unabated T cell proliferation upon stimulation

Excessive inflammation Chronic IFN pathway activation

Impact on B cell function ? Excessive autoantigen production Engagement of immunogenic rather than tolerogenic Ag presentation Hyperactivation of B cells Abnormal regulation of Ig switch and production of high affinity autoantibodies Abnormal capture / clearance of IC complexes Abnormal immune deposits, innate cell activation and tissue response to inflammation

Conclusions Genetics: identify susceptibility and protective alleles using TagSNPs Function: gather markers of disease evolution during the infraclinic development in human, then modelie in animals Intervention: skew/reset immune responses, immunotolerize, induce lymphoid regeneration

The classical pathway is initiated by activation of the C1 complex

The mannose-binding lectin pathway is homologous to the classical pathway

Ct activation is largely confined to the surface on which it is initiated

The terminal Ct proteins polymerize to form pores in membranes that can kill pathogens

Ct control proteins regulate all three pathways of Ct activation and protect the hosts from its destructive effects

Ingestion of complement-tagged pathogens by phagocytes is mediated by receptors for the bound complement proteins

Autoimmunity

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Autoimmunity