Hypersensitivity Láng Orsolya MD Ph D Dept Genetics

Hypersensitivity Láng, Orsolya MD, Ph. D Dept. Genetics, Cell & Immunobiology, Semmelweis University Lecture ED 2015 www. dgci. sote. hu

Hypersensitivity - Tolerance Hypersensitivity: Tolerance: Immune reaction leads to pathology upon recognition of either harmless environmental antigens or self-antigens immunological unresponsiveness to (self)-antigens (tolerogen) that have the capacity to elicit an immune response Autoimmunity: Autoreactivity is inevitable Dysregulation or failure of self-tolerance => autoimmune disorder Common: adaptive immune system

Hypersensitivity (HS) – Allergy § The most common immunological abnormality. § Increasing number of affected people (25 -40%) § Potential reasons: Environmental pollution (? ) Lack of selection (? )

General mechanism First exposure Sensitization Repeat exposure Tissue injury or disease Robert Royston Amos ("Robin") Coombs: § British immunologist, § co-discoverer of the Coombs test (Arthur Mourant and Rob Race) in 1945 Gell - Coombs classification of hypersensitivity in 1963

Four classifications § § Type I (Immediate) hypersensitivity Type II (cytotoxic) hypersensitivity Type III (immune complex mediated) hypersensitivity Type IV (delayed) hypersensitivity

Types of hypersensitivities (Gell - Coombs classification) Type Mediator I Ig. E mediated Mast cells and basophils Mechanism Time Disorders Immediate 1 -2 mins Allergy Anaphylaxy (local and systemic) Histamine II Cell or matrix associated antigens connecting Ig. G 4 -6 hrs Transfusion reaction Erythroblastosis fetalis Myasthaenia gravis Basedow disease III Soluble antigen -Ig. G immunecomplex 2 -8 hrs Arthus reaction RA, SLE Delayed 2 -3 days Mantoux test Chronic allergy Contact dermatitis Complement IV T cell mediated T-cell response to antigens

Type I. hypersensitivity (HS) Immediate HS or Allergy , Atopy – inherited tendency to respond immunologically to inhaled or ingested allergens with increased Ig. E production

Common types of immediate HS Skin contact Inhaled allergens Ingestion Hives Urticaria Hay fever Food allergies or injection Bronchial asthma Anaphylaxis

Main characteristics of the allergenes Small size proteins or glycoproteins: - Carried on desiccated particles (pollen grains or mite feces), where they are very stable Have enzyme activity that facilitate the transmucosal penetrance Small, molecular wheight is 5 to 70 k. Da (dustmite: der p 1 15 k. D), High solubility Small dose (ragweed: 1µg/year) MHCII binding

What do they have in common? ? Hevein domain

Cross-reactive allergenes http: //ainotes. wikispaces. com/Pollen+Food+Allergy+Syndrome

Type I. hypersensitivity Afferent phase Ig. E production Hypersensitivity IL 4, IL 13 Degranulation First exposure Class switch APC and Th 2 activation Ig. E production Ig. E+ memory B-cells cross -linked. Ig. E Mast cells Basophils FcᵋRI receptor Repeat exposure ACTIVATION and DEGRANULATION Nature Review, Drug Discoverys alapján

Class switch of BCR genes INF IL 5 IL-4 IL-13 13

Fce receptors on mast cells and basophils FcƐRI (high affinity) MC are always coated with Ig. E bound receptors Kd= 10 -11 M [Ig. E] = 10 -9 M FcƐRII (low affinity) (CD 23) FcƐRIIa FcƐRIIb on B cells, T cells, Mφ-s, DC-s and basophils

Signaling induced by cross-linking of Ig. E 15

Effects of Ig. E cross-linking (phospholipids)

Degranulation of MC § Other degranulators § Immune § Anaphylatoxins: C 5 a, C 3 a § Modulator § IL 3 § Non Immune § § § Bacterial products, gastrine, physical (cold), stress, chemical (gases, smoke), etc. Intracellular signalling: Ca++ Inhibitors of degranulation § Pharmacotherpy: cromoglicinum 17

Cells Molecules Effect Preformed granules Histamine Increases permeabilty, SM contraction Tryptase Carboxypeptidase Chymase Acidic hydrolase Cathepsin G Mast cell, basophils Pro-inflammatory mediators released by mast cells, basophils and eosinophils Mechanism of production Heparin anticoagulant Chondroitinsulphate E Released from the membrane PGD 2 Vasodilatation, brochoconstriction, chemotaxis LTB 4 Bronchoconstriction, mucus secretioon, increased vasopermeability LTC 4 LTE 4 PAF Scell proliferation imflammation CCL 2, CCL 3, CCL 5 Chemotaxis MBP( major basic protein) Toxic compound ECP( Eosinophil catioinic protein) Eosinophils Primary mediators are in preformed granules Mast cell proliferation, inflammation, Ig. E synthesis, activation of eosinophyls TNF Preformed granules IL 3, IL 4, IL 5, IL 6, IL 13 GM-CSF Nucleus, de novo synthesis Leukocyte activation and chemoattraction Tissue destruction peroxidase Lysosomal hydrolase Lysophospholipase Nucleus, de novo synthesis RANTES, IL 8, eotaxin Chemotaxis IL 10 B cell proliferation Gilfillan et al. Nature Reviews Immunology 6, 218 -230 (March 2006) | doi: 10. 1038/nri 1782

Physiological effect of mast cell degranulation

Histamine receptors

Histamine is the key mediator and their receptors H 1: e. g GIT, bronchoconstriction ↑; vessel permeabilty↑ H 2: e. g. vasodilataion ↑; secretion of exocrine gland(e. g. gastric acid)↑; secretion (H 3: neural system) H 4: eosinophil chemotaxis

Role of the Mast cells (MC) MC (Connective tissue) Intravenous , high dose capillaries Systemic anaphylaxis Subcutanous, low dose capillaries urticaria Mucosal MCs Inhalation, low dose SM in brochus Ingestion SM in intestin Hay fever, Food allergy: Bronchial asthma diarrhea, vomiting utricaria, anaphylaxis

Immediate reaction in respiratory tract Nature Medicine 18, 693– 704 (2012) doi: 10. 1038/nm. 2755

Immediate and late phase symptoms PEFR = peak expiratory flow rate After 2 hrs After 1 day

Chronic inflammation and complications (tissue remodelling) Nature Medicine 18, 693– 704 (2012) doi: 10. 1038/nm. 2755

Atopy § Atopy is the term for the genetic trait to have a predisposition for localized anaphylaxis. § Atopic individuals have higher levels of Ig. E and eosinophils.

Allergy – Multifactorial disorder Genetics FcƐRI beta chain - 11 q 13 IL-3, IL-4, IL 5, IL-9, IL-13 and GMCSF coding genes - 5 q 31 MHCII allels - 6 p Allergy Dysregulaion of the Immune system Activation of Th 1 and Th 2 subpopulation Ig. E production Immunodeficiency Increased eosinophil count Environmental factors Failure of tolerance during childhood

DER p 1 in the faeces of the house dust mite penetrates the airway epithelium 28

Bronchial asthma normal bronchiole severe asthma 29

Anaphylaxis • Anaphylactic shock is the most serious • Symptoms are directly related to the massive release of vasoactive substances leading to fall in blood pressure, shock, difficulty in breathing and even death. • It can be due to the following: – Horse gamma globulin given to patients who are sensitized to horse protein. – Injection of a drug that is capable of acting as a hapten into a patient who is sensitive, ie, penicillin. – Following a wasp or bee sting in highly sensitive individuals. – Foods – peanuts, shellfish, etc.

Staphylococcus exotoxins may serve either as superantigens or allergens Non specific T-cell activation ! 31

Therapy – Avoidance of known allergens – Localized reactions use OTC antihistamines and decongestants. – Asthma uses combination – antihistamines, bronchodilators and corticosteroids. – Systemic use epinephrine – Hyposensitization – inject antigen to cause production of Ig. G which binds to antigen (allergen) before it reaches Ig. E coated cells. – Monocolonal anti-Ig. E – inject, binds to receptors on mast cells blocking them from the Ig. E.

Administration of increasing doses of antigen desensitization

DESENSITIZATION – Allergen-specific immunotherapy Repeated administration of the sensitizing allergen usually by subcutaneous injection or, more recently, by sublingual application. Both Ig. E- and Ig. G-specific antibodies increase during postdesensitization therapy. Ig. G acts as blocking Ab http: //www. fpnotebook. com 34 http: //www. voedselallergie. nl/allergic-and-non-allergic-hypersensitivity-to-food

Anti-Ig. E therapy Monoclonal antibody Nature Reviews Immunology 8, 218 -230 (March 2008) 35

In vivo test - Intradermal allergy test (Prick test) § Small amount of allergen injected into skin § Look for wheal formation of 3 mm or greater in diameter § Simple, inexpensive, can screen for multiple allergens. § Stop anti-histamines 24 -72 hours before test. § Danger of systemic reaction § Not for children under 3

In vitro test § Measure total Ig. E or antigen-specific Ig. E in serum § Less sensitive than skin tests. §R IST, RAST, Allergen specific and Microarray will be covered later. RAST Radio. Allergo. Sorbent Test Protein array

Anaphylaxis vs. Anaphylactoid reaction (pseudoallergy) Anaphylaxy/ allergy Anaphylactoid reaction/ pseudoallergy Ig. E mediated allergic reaction in sentizized patient triggering material is capable of direct mast cell or basophil degranulation to induce, and thus cause histamine release • Foods (particularly nuts and seafood) • Drugs (particularly beta-lactam antibiotics) • Insect stings/bites (bees, wasps and fire ants) • Latex • …. • Drugs (particularly NSAIDs, aspirin, opioids) • Radiographic contrast media (CT/MR) • High histamine containing food (red wine), bacterial toxins generated from histidine (scombroid fish - fish that were inadequately refrigerated or preserved after being caught) 4 type of pseudoallergy based on COX 1 inhibition Ibuprofen v. COX 1 inhibitor No test! http: //allergycases. blogspot. hu/2010/07/allergic-and-pseudoallergic-reactions. html

Type II. hypersensitivity (HS)

Type II. hypersensitivity Cell surface antigen (Ig. G v. Ig. M) Antigens: Intrinsic - autoantigen, Membrane component(receptor) Extrinsic antigen RBC – tarnsfusion, Rh incompatibility Absorbed drugs or metabolits

Pathomechanisms of type II. hypersensitivity Opsonization => phagocytozis Fc -receptor mediated phagocytosis/ cell lysis (macrophage, NK cell, neutrophil & eozinophil) Complement activation=> cell lysis ADCC (antibody dependent cellular cytotoxicity) Abnormal physiologic response: Ab inhibits binding of neurotransmitter anti Ach R: myasthenia gravis Ab stimulate receptor anti-TSH R: autoimmune thyroiditis

Hemolysis Transfusion reaction Incompatible transfusion - Ig. M Polytransfused - Ig. G Multipara - Ig. G Rh incompatibility Complication: Erythroblastosis fetalis Not only RBC, but Platelets or leukocytes

Erythroblastosis fetalis Haemolytical disease of the newborn

Erythroblastosis fetalis Passive immunization with anti-Rh

Haemolytic anaemia and thrombocytopeny Drugs/ metabolits can act as a hapten – e. g. penicillin

Drug allergy (penicillin)

Altering signal transduction - Myastenia Gravis + Autoimmune thyroiditis Graves- Basedow ptosis 3 hrs after methyl prednisolon treatment exophtalmus

Type III. HS Soluble antigen-antibody complex - IMMUNE COMPLEX disease

Pathomechanism Solubile Ag - Ab (Ig. G or Ig. M) => IC => Complement activation=> inflammation and tissue injury Mechanism of the tissue destruction similar in all tissue Severity depends upon: size of the IC, ratio of Ag/Ab, affinity of Ab, isotype of Ab

The consequence of the tissue damage depends on the site of deposition 1. Local immune complex disease Arthus reaction – skin necrotic vasculitis – vascular wall pneumonitis – farmer’s lung 2. Acute-systemic immune complex disease acute serum disease (7 -10 days) 3. Chronic immune complex disease SLE Rheumatoid arthritis

Pathomechanism – Arthus reaction Ag exposure or vaccination Ag –Ab complex Complement activation Mast cell degranulation Fcgamma. RIII Inflammation 4 -12 hrs severe pain, swelling, induration, edema, hemorrhage, and occasionally by necrosis

Key mediators: C 3 a C 5 a Arthus reaction (experimental): - local response induced by intradermal injection of the Ag in sentitized patient

Vasculitis Farmer’s lung Actinomyces: Saccharapolyspora rectivirgula

Type IV. HS T cell mediated disease Delayed HS

Ags inducing type IV. HS DTH – delayed Type Hypressensitivity It takes aprx 12 hrs as Th 1 cells are involved Contact Ag Nickel salts, chromate Poision ivry, oak picricchlorine Hair stain Microbial induced IC bacteria Mycobacterium tuberculosis, leprae Lysteria monocytogenes Brucella abortus Fungi Candida albicans Pneumocystis carinii Cryptococcus neoformans histoplasmosis Viruses Herpes simplex Small pox Measles

Type IV. HS Syndrome Antigen Symptomes Late type hypersensitivity Proteins Insect protein Mycobacterial protein (tuberculin, lepromin) Local skin swelling: Erythema Induration (hardening) Cellular infiltration Dermatitis Contact hypersensitivity Haptens Pentadeca-chatechol (poison ivy) Paraphenylene diamine (hair stain) Metallic ions: Nickel, Chromium Local skin reaction: Erythema Cellular infiltration Blisters Intra-epidermal foci Gluten sensitive enteropathy Gliadin (grain protein) Celiac disease – flour sensitivity Atrophy of microvilli in the small intestine Undernutrition Damaged exocrine secretion of pancreas

Th 17 Neutrophil recruitment

Contact reaction Langerhans scells uptake the Ag Ag presentation to TH 1 cells INFgamma Macrophage activation Activation of the keratinocytes Macrophage activation inflammation TNF-alpha, beta Tissue destruction

A schematic view of the sensitization phase Tetsuya Honda (2013) 133, 303 -315. doi: 10. 1038/jid. 2012. 284

Activation of the innate immune system by contact allergens (haptens, Ni 2+)

Upon reexposure to haptens

Tuberculin Hypersensitivity Mantoux test (it detects infection) • Maximum at 48 -72 hours • Inflitration of lesion with mononuclear cells • Responsible for lesions associated with bacterial allergy – cavitation, caseation, general toxemia seen in TB • May progress to granulomatous reaction in unresolved infection

Granuloma Formation

Diagnostic test: Patch test – Epicutan test Allergen containig adhesive tape is fixed to the back, evaluation after 48, 96 hrs and later (4. -6. days). Steroids, antihistamins may influence the test result

Diagnosis based on the reaction

Latex allergy Type I. HS Type IV. HS Not immunological Latex-allergen Chemicals, soapa Urticaria Anafilaxy DTH Dermatitis Irritative contact dermatitis

It is important to distinguishi: § side effect § Toxicity § Intolerance § Idiosyncrasy § immun mediated reaction - hypersensitivity

Penicillin allergy Type I. Drug allergy Type II. drug induced anaemia Penicillin-protein Penicillin-RBC Ig. M, Ig. G Ig. E target Systemic anaphylaxis urticaria target Hemolytic anaemia Penicillin-protein Ig. G Complement Ig. E-Mast cell Type IV. HS Penicillin Type III. HS TDTH Ig. G immuncomplex Macrophage activation target Serum sickness glomerulonephritis target Contact dermatitis

Allegies in dentristry Chlorhexidine: including allergy (Type I HS ) and allergic contact dermatitis/stomatitis (Type IV HS) Flouride - Type I and VI. HS skin rashes, mouth lesions Metals- e. g. Ti : Type I and VI. HS stomatitis, facial erythema

MELISA test (Memory Lymphocyte Immunostimulation Assay ) §clinically useful in identifying metal hypersensitivities 1. Blood sample 2. Lymphocytes are isolated and then incubated for 5 days with individual metals. 3. in HS patient if T cells re-encounter that metal in the culture, they proliferate, 4. Assessment is based measurement of T cell proliferation has occurred in response to that metal.