1. HYPERSENSITIVITY REACTIONS

2. HYPERSENSITIVITY REACTIONS
Innocous materials can cause hypersensitivity in certain individuals
unwanted inflammation
damaged cells and tissues
Non-proper reaction of the immune system to foreign substances
Mainly harmless substances – after second or multiple times

3. TYPES OF HYPERSENSITIVITY REACTIONS
Type I
„immediate”
Type II
Type III
Type IV
„delayed”
Antibody mediated
T cell mediated
specific IgE
cell surface antigen specifically reacting with antibody
aspecifically
deposited soluble immuncomplex
MHC restricted T cell activation
mediators produced by mast cells
FcR mediated inflammation, inhibition of cell functions
FcR mediated complement activation, inflammation
cytokines, cytotoxicity
„classical allergy”
newborn haemolytic anaemia, penicillin sensitivity, M. gravis
Serum sickness, SLE
Contact dermatitis
M. Gravis – acethilcolin receptorokhoz kötődő IgG ellenanyagok blokkolják a jelátvitelt
mostly appear together with autoimmune diseases

4. ANTIBODY MEDIATED HYPERSENSITIVITY REACTIONS Certain drug allergies
Hay fever
Asthma
Systemic anaphylaxis
Certain drug allergies
(penicillin)
Serum sickness
Arthus reaction

5. TYPE I HYPERSENSITIVITY REACTION
ALLERGY

6. DIFFERENCES OF IMMUNE RESPONSES INDUCED BY ALLERGENS AND PATHOGENS
Deviving
Escape mechanism
DANGER SIGNAL
Microbial DNS, CpG-ODN
dsRNS  IFN
LPS (Gram-), PG (Gram+)
HSP
Inflammatory cytokin
TNF-, IL-1
DC/TISSUE DEMAGE
Activation of innate immunity
Inflammation, neurotransmitters (VIP)
ALLERGEN
Non deviding
Act together with other environmental effects
DANGER SIGNAL
Ni, Mn, Co, Sn chlorides
Ion channel inhibitors
Energy homeostasis
DC/TISSUE DEMAGE
TNCB, DNCB
Der p1 – cisztein proteáz
papain – meat processing
CD25, CD23 cleavage
Structure of epithelial „tight junction” is demaged
Bee bite (toxin)
Substilyzin – washing powder

7. T cell priming and polarization
ALLERGENS USUALLY ENTER THE BODY VIA MUCOSAL SURFACES AND THEY ARE PRESENT AT A LOW DOSE DC
B cell
antigen presentation
Th2
Th2
T cell priming and polarization
allergy response
soluble proteins on te surface of small particles (pollen, dust mite „drops”)
small molecular weight, soluble
trans-mucosal entry, enzymatic activity
low dose (ragweed: 1µg/year)

8. Mechanism of the initiation of Th2 response
IL-4
CD4+ T
IL-4
IL-10
Allergen
Mucosa

9. allergy GENETIC/ENVIRONMENTAL PREDISPOSITION TO ALLERGY
Genetic factors
chromosome 11q
FcεRβ chain gene
IL-3-5 IL-9, IL-13 GMCSF
HLAII DRB1*015
Inproper immunregulation
Th1/Th2 inbalance
regulation of IgE synthesis
immunodeficiency
high eosinophil counts
allergy
Environmental factors
lack of tolerance

10. Mast cell degranulation, wheel and flare reaction
Ragweed
Saline
Histamine

11. MAST CELL RESPONSE TO SURFACE FcRεI CROSSLINKING LATE MEDIATORS
EARLY MEDIATORS
Biogenic amins – histamin
Enzymes – triptase, chymase, carboxypeptidase
LATE MEDIATORS

12. The effect of mast cell degranulation varies with
the tissue exposed to allergen

13. Systemic anaphylaxis is caused by allergens
that reach the blood stream

14. Types of IgE-derived allergic response
SYNDROME
ALLERGENS
ROUTE OF ENTRY
RESPONSE
systemic anaphylaxis
drugs
anti-serum
peanuts
intravenous
peroral
edema, increased vascular permeability
tracheal occlusion
circulatory collapse, death
acute
urticaria
bug bite
allergy test
subcutan
local increase in blood flow and vascular permeability
allergic
rhynitis
pollen
dust mite drops
inhaled
irritation and edema of nasal mucosa
airway inflammation
asthma
animal fur
bronchial constriction, increased mucus production
food allergy
nut, peanuts,
fish, shellfish
milk, eggs
vomiting, diarrhea
pruritis (itching)
urticaria (hives)
anaphylaxia (rare)

15. Short/Common ragweed (Ambrosia artemisiifolia)

16. Short/Common ragweed (Ambrosia artemisiifolia)
Mugwort (Artemisia vulgaris)
Green leaf back
White leaf back

17. Mugwort (Artemisia vulgaris)

18. Mugwort (Artemisia vulgaris) – ?
Wormwood (Arthemisia absinthium) – Absinthe (thujone: max 35 mg/l)
Fekete üröm kivonat van az unikumban
Az Abszint eredeti fogyasztásmódja: kockacukron keresztül csepegtetett víz opálossá teszi az eredetileg áttetsző zöld folyadékot.
A tujon tartalma a fogyasztók lassú elbutulásához vezetett – lásd az arckifejezést az „Abszintivók” képen (Edgar Degas) [dögá]
Most 35mg/l-ben maximalizálták.

19. HYPERSENSITIVITY REACTIONS INDUCED BY IMMUNE COMPLEXES
TYPES II and III
Type II hypersensitivity
IgG tpye antibodies bound to the cell surface or to tissue antigens
cells expressing the antigen become sensitive to complement mediated lysis or to opsonized phagocytosis
frustrated phagocytosiss  tissue demage
the antibody inhibits or stimulates target cell function
no tissue damage (e.g. M. gravis – receptor blocker antibodies)

20. MECHANISMS OF TYPE II HYPERSENSITIVITY REACTIONS
Hemolytic anemia of newborns
Erythroblastosis fetalis
Drug induced
Hemolytic anemia
Trombocytopenia
Agranulocytosis
Penicillin-based antibiotics Anti-arythmic quinidin
Goodpasture syndrome (type IV collagen)
Pemphigus vulgaris
(desmosomal antigens)
Damage of epidermal and mucosal junctions, acantholysis

21. DEVELOPMENT OF DRUG SENSITIVITY Drug-modified cell surface proteinTh B
Drug-modified cell surface protein
Healthy cell
IgG type antibodies

22. FRUSTRATED PHAGOCYTOSIS MEDIATED BY IgG TYPE ANTIBODIES
Binding Opsonization Internalization Enzyme release
The tissue, which can not be phagocytosed, is damaged
Internal or absorbed antigen (drug)
Opsonized surface Binding Frustrated Enzyme release
phagocytosis

23. Examples - Type II hypersensitivity
Newborn haemolytic anaemia
Transfusion reaction
Hyperacut allograft rejection
Drug-derived
Haemolitic anaemia
Thrombocytopenia
Agranulocitosis
Penicillin-based antibiotics
Anti-arithmic quinidin
Goodpasture syndrome (kidney, membrane basalis, collagen type IV)
Myasthaenia gravis (anti-acetyl-choline receptor antibodies)
Basedow-disease (anti-TSH-receptor antibodies)
Pemphigus vulgaris (mucosal bubbles)  against desmosomal antigens, interruption of epidermal and mucosal connections, acantolysis (desintegration into single cells)

24. TYPE III HYPERSENSITIVITY Antibodies binding to soluble antigens
Small circulating immune complexes
Depends on:
Size of immune complexes
Antigen-antibody ratio
Affinity of antibody
Isotype of antibody

25. PATHOGENICITY OF IMMUNE COMPLEXES
Pathogenic immune complexes
Formed in the blood and than are deposited in tissues
Formed in situ at the site of antigen localization
Mechanisms of tissue demage is independent on the site of deposition
Steps of tissue demage
Formation immune complexes in the blood
Deposition depends on the size, composition and cytophylic properties of the antibody (IgM, IgG, IgA)
FcγRIII has a pivotal role – expressed by basophylic granulocytes, NK cells
Permeability of endothelium
Tissue demage
Increased permeability of blood vessels
Reqruitment of neutrophils – enzymes, chemoattractans, dilatators, prostaglandins
fibrosis
Consequences of tissue demage depends on the site of deposition
Arthus reaction – local reaction in skin
Infectious diseases – morbilli – erythema, vasculitis
Acute serum disease – 7 – 10 days
Polyclonal antibodies against snake venom produced in horses (anti-streptococcal)
Immune suppresszive anti-lymphocyte globulin
Bacterial trombolytic streptokinase – treatment of miocardial infarction
Subacute bacterial endocarditis – pathogens are not eliminated
Chronic viral hepatitis
SLE – small vessels, kidney, joints, skin, heart, serosal surfaces

26. THE PROCESS OF TISSUE DAMAGE CAUSED BY IMMUNE COMPLEXES
Blood vessel wall permeability
Frustrated phagocytosis
Immune complexes activate the complement system, neutrophils, bazophil granulocytes and thrombocytes

27. Arthus-reaction Localized Type III hypersensitivity
Local vasculitis develops as a result of immune complex deposition
Inhaled antigens (fungi, animal feces) may induce similar reaction in the lung
IgG type antibody
‘Farmer lung’ and ‘piegeon breeder lung’
The Arthus reaction involves the in situ formation of antigen/antibody complexes after the intradermal injection of an antigen. If the animal/patient was previously sensitized (has circulating antibody), an Arthus reaction occurs. This manifests as local vasculitis due to deposition of immune complexes in dermal blood vessels. Activation of complement and recruitment of PMNs ensue resulting in an inflammatory response.

28. MANIFESTATION OF TYPE III HYPERSENSITIVITY IN SLE
Facial, malar "butterfly" rash with characteristic shape across the cheeks. Discoid lupus erythematosus (DLE) involves mainly the skin, it is relatively benign compared to systemic lupus erythematosus (SLE). In either case, sunlight exposure accentuates this erythematous rash. A small number (5 to 10%) of DLE patients go on to develop SLE (usually the DLE patients with a positive ANA).
Here is a more severe inflammatory skin infiltrate in the upper dermis of a patient with SLE in which the basal layer is undergoing vacuolization and dissolution, and there is purpura with RBC's in the upper dermis (which are the reason for the rash).
                   
             

29. DEPOSITION OF IMMUNE COMPLEXES IN THE SKIN OF SLE PATIENTS
When immunofluorescence staining with an antibody to complement or immunoglobulin is performed, a brightly fluorescent signal staining the dermal epidermal junction is visable indicating immune complex deposition.
Immunofluorescence staining pattern with antibody to IgG staining immune complexes at the dermal-epidermal junction. If such a pattern is seen only in skin involved by a rash, then the diagnosis is probably DLE, but if this pattern appears even in skin uninvolved by a rash, then the diagnosis is SLE.

30. RENAL FAILURE IN IMMUNECOMPLEX DISEASES
One of the feared complications of the rheumatic diseases is renal failure. This is most likely to occur in SLE. Here is a glomerulus in which the capillary loops are markedly pink and thickened such that capillary lumens are hard to see. This is lupus nephritis.
Here is a glomerulus with thickened pink capillary loops, the so-called "wire loops", in a patient with lupus nephritis. The surrounding renal tubules are unremarkable.

31. Anti -nuclear antibody
ANA
Anti -nuclear antibody
This is the so-called "nucleolar pattern" of staining in which the bright fluorescence is seen within the nucleoli of the Hep2 cells. This pattern is more suggestive of progressive systemic sclerosis.
This is the so-called "speckled" pattern of staining which is more characteristic of the presence of autoantibodies to extractable nuclear antigens, particularly ribonucleoprotein. This pattern is not very specific, but may be seen with an entity called "mixed connective tissue disease" which is a mix between SLE, scleroderma, and polymyositis, but without serious renal or pulmonary disease. The autoimmune diseases are very hard to classify, even for the experts.
This is the so-called "rim" pattern that is more characteristic of SLE.

32. TYPE IV HYPERSENSITIVITY REACTION
T CELL MEDIATED PROCESS
MACROPHAGES ARE INVOLVED

33. Type IV hypersensitivity reaction
Chemokines, cytokines,
cytotoxins

34. Delayed-type (TYPE IV) Hypersensitivity

35. Delayed-type (Type IV) Hypersensitivity

36. Delayed-type hypersensitivity (DTH) (e.g., tuberculin skin test)
TH1 from a previous immunization (memory)

37. Mycobacterium protein (PPD)
Tuberculin skin test
Ag = antigen
Mycobacterium protein (PPD)
Introduction of Ag

38. Chemical Mediators of DTH

39. DTH as a result of a contact-sensitizing agent*
Contact Dermatitis
*a contact-sensitizing agent is usually a small molecule that penetrates the skin then binds to self-proteins, making them “look” foreign

42. Poison ivy
Anacardiaceae (family), Toxicodendron (genus)
Toxicodendron radicans or Rhus toxicodendron

44. Delayed-type hypersensitivity is mediated by T cells

45. Delayed-type Hypersensitivity
A positive tuberculin skin test is a DTH reaction

46. PRACTICAL ASPECTS OF TRANSPLANTATION IMMUNOLOGY

47. Transplantation reactions
REJECTION
HLA-A, B, C, DR, DQ, DP, minor histocompatibility antigens
foreign MHC-antigens recognized by T cells
Direct: self T cells - donor APCs CD8+ T cells
Indirect: self APC presents donor MHC-derived peptides
CD4+ T cells
inflammatory cytokine release
Hyperacut rejection
Causes: previous immunization against alloantigens, preformed anti-HLA-antibodies, blood group incompatibility, xenotransplantation
antibodies bound to endothel
activation of the complement system
thrombosis in venules
vascularis necrosis
Therapy resistent

48. HLA typing used for transplantations diagnostical value
MHC I: HLA-A, HLA-B, HLA-C
MHC II: HLA-DP, HLA-DQ, HLA-DR
used for transplantations
Most important is the most polymorphic HLA-B and HLA-DR, HLA-C does not matter)
diagnostical value
Connections between the HLA alleles and diseases
Serotyping - microcitotoxicity tests (1)
Based on the serological reaction between the TARGET cells and the typing serum.
Complement-mediated lysis induced by antibodies recognizing MHC I and/or MHC II cell surface molecules as antigens
There is NO reaction in the case of serotype identity
(dead cells can be visualized by specific dye)
Typing sera containing antibodies to Class I and II proteins are collected from multiparous women, or individuals who had received multiple blood transfusions (immunized against multiple HLA alleles). The specificity of thes polyclonal test sera has been validated by international workshops.
The procedure is carried out in Terasaki microtiter plates (with 10µl working aliquots) due to the limited quantity of typing sera. Polyclonal typing sera have been replaced by monoclonal antibodies with unlimited availability.

49. Serotyping - technology
HLA-D (MHC II) antigens are also studied on nylon column separated B cells
The polymorphism of HLA-D antigens could be studied by mixed lymphocyte reactions (MLR)
Bidirectional MLR – lymphocytes of both donors are responding
Unidirectional MLR – lymphocyte proliferation of one donor is blocked (irradiation, Mitomycin treatment)
THE RATIO OF ALLOREACTIVE LYMPHOCYTES IS AN INDIVIDUAL IS 1 – 10 %
Microtiter plates

50. Genomic DNA based examinations
Serotyping have some limits:
– crossreactions (HLA-B27 – HLA-B7)
– there are no sera available against HLA-C because of its low immunogenicity
– some subtypes cannot be discriminated
Genomic DNA based examinations
PCR-SSP is using the PCR amplification reaction directly to detect HLA polymorphisms. Primers can be constructed specifically to complement HLA polymorphisms; if the primers bind the complementary polymorphism and amplify the gene segment, then the PCR product can be detected by standard techniques. It has been constructed an array of PCR primers complementary to the range of HLA polymorphisms
HLA-A …

51. PCR-SSOP (sequence specific oligonucleotid probe)
The examined HLA genes are amplified by PCR with non allele specific primer pairs. The amplicons are immobilized to nitrocellulose membranes or microtiter plates and are hybridized by HLA allele specific labeled oligonucleotides.
The label can be enzimatic, fluorescent or radioactive
SBT (sequence based typing)
allotypes can be evaluated by sequencing the MHC region of the genomic DNA
(minor mutations can be examined)

52. HLA allotypes and diseases
(autoimmune)
HLA
frequency
concerned
control
SLE
DR3 55 20 B8 50
Hydralazine(?) induced lupus erythematosus
DR4 73 32
Basedow-disease 56 25 43
Active chronic hepatitis 21
Sclerosis multiplex
DR2 60 30 B7 37 24
Myasthenia gravis 44
Autoimmune IgA glomerulonephritis 53 19
Type I diabetes 72 49 22 40
Addison-disease (idiotopic, autoimmune) 70 46 23
Sjörgen-syndrome
Coeliaca 79
DR7 15 68
Goodpasture-syndrome 88 29
IgA loss 81
Dermatitis herpetiformis 82 75
De Qervain-thyreoiditis
B35
Reiter-syndrome
B27 9
Felty-syndrome 95
Diseases
HLA
frequency
concerned
control
Narcolepsia
DR2
100 22
Bechterew-disease
B27 89 9
Adrenogenitális syndrome
salt lost
late
virilizing
Bw47 36 1
B14 57 4 B5 48 10
Psoriasis vulgaris
Cw6 56 15
B13 24 8
B17 27
DR7 23
Idiophatic haemochromatosis A3 76 28
Bechet-disease
B51 50 11
Gold induced thrombocytopenia
DR3 13
Gold induced leucopenia 47
HLA allotypes and diseases
A kontroll csoport a vizsgált betegcsoporttal azonos helyről származik. Ez az oka a kisebb eltéréseknek ugyanazon HLA típusok között.
ref: Klinikai immunológia (II. klinikum) (OHVI 1990 szerk.: Szegedi, Gergely, Sipka, Szemere)
Stenszky Valéria: Autoimmun betegségek genetikai vonatkozásai

53. HLA-A alleles described until october 2006
498