Faculty of Medicine of Sousse

University of Sousse
2022/2023

1st year of Medical studies

Subject matter 14

Mechanisms of Hypersensitivity
Part1: HS Type I
Part2: HS Type II
Part3: HS Type III
Part4: HS Type IV

Pr. Ag. Nadia Idriss

Department of Immunology

1
I. General Introduction

➢ Hypersensitivity reactions may result from uncontrolled or excessive responses

against foreign antigens or from a failure of self-tolerance.

➢ During these abnormal reactions, the first exposure to the antigen “sensitizes”
lymphocytes; any secondary exposure will induce a damaging reaction with clinical
manifestations; and the response is specific to a particular antigen.

➢ We distinguish 4 types of hypersensitivity reactions and they are classified based of

the effector mechanism (cells and mediators) responsible for tissue injury:

➢ Classification of hypersensitivities: Gell and Coombs classification

- Hypersensitivity type I (immediate (few minutes to install), dependent on IgE)

- Hypersensitivity type II (antibody-mediated HS, dependent on IgG and IgM against
cell surface)
- Hypersensitivity type III (immune complexes-mediated HS)
- Hypersensitivity type IV (delayed HS, dependent on T cells).

2
 Part I: Type I hypersensitivity (HS): Allergy
Immediate Type HS

Educational objectives:

1. Define type I hypersensitivity

2. Describe the immunological mechanisms of immediate hypersensitivity
3. Enumerate the different forms of type I HS
4. List the tests to biologically diagnose type I HS
5. Recognize the non-specific treatment of immediate HS
6. Explain the desensitization therapy and its effect on the immune system

Course plan
1. Introduction
2. Immunological mechanisms of the allergy
1.1. Sensitizing phase
1.2. Effector phase of the allergy
a. Immediate release of the granules content
b. Delayed release of newly formed mediators
3. Types of allergens
4. Biological diagnosis of allergy:

a. Diagnosis of atopy

b. Identification of the allergen

5. Treatment of immediate HS
5.1. Non-specific treatments
5.2. Specific treatments

1. Introduction

✓ In genetically predisposed patients, we observe an abnormal and excessive immune

response (excessive production of IgE specific for an allergen) to an environmental
antigen called allergen, usually harmless.

✓ Atopy: the genetic tendency to be sensitized and produce IgE as a reaction to

external substances.

✓ Allergies are very common and are increasing in prevalence.

3
✓ This condition results in multiple, non-specific, and recurrent symptoms after each
new exposure.
✓ The allergen are more often ingested or inhaled.

✓ 2 phases: sensitizing phase (clinically mute) and effector phase which is symptomatic.

✓ The clinical manifestations could be respiratory (allergic rhinitis, allergic asthma),

ocular (conjunctivitis), digestive (vomits, diarrhea, abdominal pain), or skin
manifestations (urticarial, atopic dermatitis).

✓ Other cells and mediators involved:

o Mast cells are mainly responsible of clinical manifestations of the effector
phase. Also, basophils play such role.
o Other cells are involved in tissue lesions: neutrophils that aggravate the
lesions, and eosinophils which play an important role in asthma.

2. Immunological mechanisms of the allergy

1.1. Sensitizing phase

✓ This phase can last few weeks to few months after the first exposure to the
allergen.

✓ The antigen is captured by the dendritic cells. They induce a cellular immune
response by interacting with T lymphocytes which differentiate into Th2 in a
cytokine particular environment rich in IL-4, IL-5, and IL-13. A specific B response
is set up producing IgE antibodies.

✓ The cytokines also induce the increase of the number of mast cells, basophils,
eosinophils.

✓ The IgE produced against the antigen bind to mast cells and basophils, by their Fc
constant fragment to the high affinity receptor for IgEs (Fcε RI) expressed by
those cells. On the cell surface, IgE persist for several months.

✓ A low affinity receptor for IgE, FcεRII or CD23 is present in particular on

eosinophils.

✓ IgEs leave their variable region, Fab, free for the corresponding allergen.

4
 1.2. Effector phase of the allergy
✓ Characterized by an immediate degranulation (release of the granules content) of
mast cells essentially. Basophils can also play this role (depending on the site and
mode of introduction of the antigen).

✓ After a new contact with the allergen. The latter will be recognized by the IgE linked
with high affinity to FcεRI on the surface of mast cells.
✓ If two adjacent IgEs are linked to the allergen (multivalent or at least divalent
allergen) the mast cell will be activated.

a. Immediate release of the granules content

✓ The activation leads to an immediate secretion of the granule content in the
extracellular medium.
✓ Those granules contain preformed mediators, mainly vasoactive amines (histamine),
lysosomal enzymes and chemokines.

✓ Those mediators are responsible of rapid and brutal response (in a few minutes)

➔Immediate phase of type I hypersensitivity.

5
➢ Clinical consequences:
- The histamine produced diffuses through the tissues to bind to its receptors,
Causing: vasodilation, an increase in capillary permeability, a bronchoconstriction.
- Released proteases, such as tryptase, initiate local inflammation and tissue
lesions.
- Chemokines attract and activate leukocytes.
- The symptoms depend on the type of allergen, the portal of entry and the patient.
o Urticaria (papules, pruritus)
o Edema of deep tissues (angioedema)
o Bronchoconstriction and hypersecretion of bronchial mucus (asthma
attack).
o Circulatory insufficiency by vasodilation (a shock can occur)

Urticaria Angiodema

b. Delayed release of newly formed mediators

➢ It is a late phase of the re-exposure (after a few hours of the immediate phase)
characterized by delayed production of newly formed factors.
- Lipid mediators deriving from membrane phospholipids (activation of
phospholipase A2, which releases arachidonic acid and lysophosphatidylcholine)
o Prostaglandins (PG D2), Leukotrienes (LT-C4): contraction of bronchial
smooth muscle, mucus secretion, mucous edema.
o LT B4: chemoattractant of neutrophils and activation of their oxidative
functions.
o Platelet Activator Factor (PAF): activation of platelets (micro thrombosis)
and contraction of bronchial smooth muscle.
- Cytokines and growth factors
o Pro-inflammatory agents (TNF-α…)
o IL-4, IL-13: Th2 response
o IL-5 which promotes the recruitment and activation of eosinophils.
➢ The late phase may contribute to the installation of a chronic inflammatory reaction
with tissue remodeling.

6
 3. Types of allergens
The allergens are harmless antigens that induce an allergic response: proteins from pollen,
milk, fruit, dust mites, food or animal dander…

➢ Classification of the allergens:

- Pneumallergens (inhaled)
- Trophallergens (ingested)
- Transcutaneous allergens
- Drug allergens
- Occupational allergens
- Venoms.
➢ Food allergy: prevalent
- In adults : rosacea (flowering plants), latex group, umbellifers (aromatic flowering
plants), nuts, cereals, peanuts, sesame, egg white, crustacean, fish, additives, ….
- In infants: white egg, peanut, cow milk, legume, nut, fish, additive, cereal, latex
group…
➢ Pneumallergens: the most frequently found are:
- Seasonal pneumallergen:
o Grass: chaff, cat grass or dactyl (summer to autumn)
o Trees: birch, oak…
o Herbaceous: ambrosia (spring)
- Perennial pneumallergens:
o Mites
o Pet allergens: cat
o Commensal pneumallergens: Aspergillus (saprophytic fungus) ,
Cladosporium (fungus).

4. Biological diagnosis of allergy:

Biological examinations must be prescribed in addition to a meticulous examination which
remains a fundamental step in the diagnosis of an atopic state.

4.1. Diagnosis of atopy

- Personal and family history of allergy
- Total blood count: Eosinophil > 4% (or 0.4g/l)
- Total IgE assay: indications are limited

Physiological increase until adolescence: <10-20UI/ml

Normal rate from adolescence: <200-250UI/ml

- Non-quantitative multi-allergen screening tests (Phadiatop®, Alatop®)

7
 4.2. Identification of the allergen

a. Examination

To look for circumstances triggering or increasing clinical signs, seasonality,…

b. Immediate reading skin test

- Prick test: drop of allergen + puncture through the drop with an IDR (intra-dermo
reaction) needle: direct intradermal injection of the allergen.

Reading at the 15-20th minute: measurement of the papule and erythema

The predictive value of skin tests varies according to the allergen in question, age, ethnic
origin and allergic pathology of the patient.

c. Orientation tests by groups of allergens

d. Monospecific tests: Specific IgE

Recombinant antigens are increasingly used: latex, peanut....

e. Multi-allergen tests with qualitative or semi-quantitative response per allergen

➢ Good diagnostic value for pneumallergens but limited for trophallergens

➢ Beware of cross-allergenicity between pollen and food.

8
Attention, the positivity of specific IgE of an allergen corresponds to a sensitization with
respect to this allergen but does not in any case allow making the diagnosis of a clinical
allergy on its own.

5. Treatment of immediate HS
5.1. Non-specific treatments

➢ A healthy lifestyle
➢ Steroidal anti-inflammatories
➢ Antihistamines (anti H1): during conjunctivitis, rhinitis or urticaria
➢ Degranulation inhibitors (sodium cromoglycate)
➢ Sympathomimetic bronchodilators (salbutamol…): used in asthma
➢ Anti-IgE antibodies: complex with IgE and prevent them from binding to mast cells
and basophilic PNs.

5.2. specific treatments

➢ Avoid the allergen (once identified)

➢ Allergen-specific immunotherapy (desensitization)

- Based on the administration of increasing doses of the allergen

- Administered subcutaneously or sublingually
- Induces production of allergen-specific regulatory T cells (produce IL-10)
- Induces inhibition of Th2 pathway
- Leads to class switch in favor of IgG (especially IgG4) instead of IgE
- Takes a period of 3 to 5 years to be effective.

9
 Type II hypersensitivity (antibody-mediated
hypersensitivity)

Educational objectives:

1. Defining type 2 hypersensitivity

2. Explain the mechanisms of lesions induced by antibodies
3. Identify the different clinical forms of type 2 hypersensitivity
4. Describe the processes and clinical consequences of the main pathologies due to
cytotoxic antibodies

Course plan:

1. Introduction
2. Mechanisms of action of produced antibodies
3. Transfusion Reactions: rare
a. Antigens and antibodies
b. In case of incompatible transfusion
c. Blood group systems implicated in transfusion reactions
d. Post-transfusion allo-immunizations
4. Hemolytic disease of the new born (HDNB)
4.1. Mechanism of HDNB
4.2. Consequences for the fetus
4.3. Treatment of the newborn
4.4. Prevention of HDNB
5. Hemolytic anemia induced by drugs
6. Autoimmune hemolytic anemia (AIHA)
6.1. Diagnosis of AIHA
6.2. Characteristics of warm AHAIs
6.3. Characteristics of cold AHAIs
7. Other autoimmune diseases with immune-mediated HS reaction
7.1. Autoimmune cytopenias
7.2. Myasthenia
7.3. Graves' disease
7.4. Pemphigus

10
 1. Introduction
Antibodies against cell surface or extracellular antigens cause diseases that are specific to
the tissues harboring those antigens. In most cases, these antibodies are autoantibodies, but
they may be produced against a foreign antigen that is cross-reactive with self-components
of tissues.

Those antibodies can be cytotoxic inducing destruction of cells or non-cytotoxic causing

functional changes without cell lysis.

The antigens are constituent of the cell membrane (autoimmune cytopenias) or adsorbed on
this membrane (hemolytic anemia induced by penicillin)

Type II hypersensitivity reactions are involved in: Transfusion reactions, Hemolytic disease of
the newborn, drug cytopenias, certain autoimmune diseases (autoimmune cytopenias,
pemphigus…)

2. Mechanisms of action of produced antibodies

Antibodies can cause tissue damage by 3 main mechanisms:

➢ Opsonization of cells: antibodies bound to a cell allow phagocytic cells with Fc

receptors to bind and phagocytize them
➢ Activation of the complement system which recruits neutrophils and macrophages
that cause tissue damage (Autoimmune hemolytic anemia)
➢ Antibody-dependent cellular cytotoxicity (ADCC): cytotoxic cells expressing receptors
binding the Fc region of antibodies present on the target cells (hemolytic disease of the
newborn HDNB).
➢ Possible binding to normal cellular receptors and interference with their function
(Myasthenia gravis and Graves disease).

11
 3. Transfusion Reactions: rare
a. Antigens and antibodies
The target antigens are ABO blood group system, antigens on red blood cells. The antibodies
produced are pre-existing IgM antibodies targeting the antigens that are absent on red
blood cells.

Blood type and natural antibodies

b. In case of incompatible transfusion

A “B blood type” patient receiving a transfusion with an incompatible blood (“A blood type”
for example) has natural anti-A antibodies. These antibodies will bind to the antigen A on red
blood cells forming immune complexes that activate the complement. The activation of
complement system induces the lysis of the red blood cells.

Transfusion accidents are rare but remain possible.

Clinical manifestations result from massive intravascular hemolysis (minutes or hours

following the transfusion) and the major risk is shock with collapse.

12
 c. Blood group systems implicated in transfusion reactions

Transfusion reactions can result from antibodies against other blood group antigens.
Especially Rhesus, Kell, Duffy, Mn systems.

d. Post-transfusion allo-immunizations

Patients with certain hematologic diseases or certain cancers need repeated blood
transfusions. They develop post-transfusion allo-immunizations with IgG class antibodies.

Incompatibilities result in delayed hemolytic reactions (2-6 days post-transfusion and the
destruction of red blood cells occurs at extravascular sites by phagocytosis.

4. Hemolytic disease of the new born (HDNB)

4.1. Mechanism of HDNB

➢ In the fetus, this disease is due to transport of maternal IgG antibodies specific for one
of the Rhesus (Rh) protein antigens (RhD) across the placenta. In fact, the D allele of
the Rh antigen creates the strongest immune responses.

➢ About 85% of people are Rh+. When a pregnant woman is Rh-(negative) and the
father is Rh+, the fetus could be Rh+.

➢ During a first pregnancy, before any contact with the fetal blood, the mother will not
develop an immune response against Rhesus D, because the trophoblast prevents the
passage of RBC from the fetus into the maternal circulation.

13
➢ However after placental separation at the time of birth, a large amount of blood from
the cord passes into the maternal circulation. These Rh+ fetal RBCs stimulate a T-
dependent immune response, resulting in the generation of memory B cells capable
of producing IgG antibody against RhD.

➢ In a subsequent pregnancy with a Rh+ fetus, maternal IgG can be transported across
the placenta, react with fetal Rh+ red cells, and activate complement, producing
hemolytic disease.

4.2. Consequences for the fetus

- Moderate to severe anemia.
- Jaundice
- Brain damage can occur because of bilirubin (which comes from hemoglobin)
which is fat-soluble and can accumulate in the brain (because the blood-brain
barrier not yet complete).
- Bilirubin being toxic to neurons, the outcome may be fatal in the absence of
treatment.

Mechanism of hemolytic disease of newborn

14
 4.3. Treatment of the newborn
- Blood transfusion in case of anemia.
- Skin exposure to ultraviolet radiation (UV) to rapidly degrade bilirubin.

4.4. Prevention of HDNB

➢ It can be prevented by administering to the mother antibodies directed against
Rhesus antigen, at about 28 weeks of gestation and again within 72 hours after the
first delivery.
➢ These antibodies bind to all fetal RBCs that enter the maternal circulation, facilitate
their elimination before the B cells have been activated

5. Hemolytic anemia induced by drugs

✓ Some antibiotics (penicillin, cephalosporins, streptomycin) as well as other drugs

(ibuprofen, naproxen) can bind non-specifically to proteins on the surface of the red
blood cell (adsorbed).
✓ Drug-protein complexes can induce the production of antibodies, in some cases.
✓ These antibodies bind to the adsorbed drug and lead to the lysis of the RBCs by the
complement (progressive anemia).
✓ Stopping the drug leads to the disappearance of hemolytic anemia.

15
 6. Autoimmune hemolytic anemia (AIHA)
They appear due to the destruction of red blood cells of auto-immune etiology
(autoantibodies). There are 2 types: warm AHAI and cold AHAI.

6.1. Diagnosis of AIHA

- Symptoms: Dyspnea, Weakness.
- Signs: Pallor, Jaundice, Splenomegaly.
- Total blood count: normochromic anemia, often normocytic and regenerative
(reticulocytosis with reticulocytes >120,000/mm3).

➢ To confirm hemolysis:
o Increase in bilirubin (unconjugated/indirect)
o Increase in LDH enzyme
o Decrease in haptoglobin.

➢ To confirm autoimmunity: by 'Direct Coombs test'

o Patient's red blood cells + anti-human Ig antibodies.
o If presence of anti-GR antibodies the test is positive and gives an
agglutination.
o It allow to detect an autoantibody on the red blood cells, to determine
the isotype (IgG, IgM..), to search for the presence of complement on the
RBCs.
➢ If negative, it does not eliminate the diagnosis of AIHA because the concentration of
the autoantibody may be low.

16
➢ Positive versus negative test:

6.2. Characteristics of warm AHAIs

- Occurs at 37 ºC (autoantibody binds best to RBCs at 37 degrees)

- It is usually caused by IgG coating red blood cells
- Hemolysis is extravascular (by phagocytosis in the spleen)
- Test for Coombs positive anti-IgG or anti-IgG and anti-C3 of the complement
- Etiologies: chronic lymphocytic leukemia, lupus, viral infections, idiopathic form
(50%)
- Treatment: corticosteroids, possible splenectomy.

17
 6.3. Characteristics of cold AHAIs

- IgM class antibodies react at temperatures below 37 ºC (bind to RBCs at low

temperature)
- IgM can activate the complement cascade
- Hemolysis is intravascular
- The Coombs test is positive with the complement anti-C3 antibody
- Etiologies: infections, lymphomas, idiopathic
- Treatment: Treat the underlying cause, keep the patient warm, possible
plasmapheresis (eliminates autoantibodies)

18
 7. Other autoimmune diseases with immune-mediated HS reaction

7.1. Autoimmune cytopenias

➢ Anti-platelet antibodies ➔ autoimmune thrombocytopenia (purpura on the skin
(hemorrhagic lesion due to extravasation of blood in the dermis)).

Purpura

➢ Antibodies directed against neutrophils➔ autoimmune neutropenia (risk of

occurrence of infections (bacteria and fungi)).

7.2. Myasthenia
Anti-acetyl choline receptor antibodies induce muscle weakness by blocking neuromuscular
transmission.

7.3. Graves' disease

Anti-TSH receptor antibodies bind to the TSH receptor in a similar way to TSH and deliver a
stimulus to the thyrocyte thus inducing hyperthyroidism.

7.4. Pemphigus
Pemphigus is a bullous cutaneous disease with the presence of antibodies directed against
the junction of epidermal cells: anti-desmosome antibodies.

19
 Type III hypersensitivity: Immune complexes-
mediated HS
Semi-delayed HS

Educational objectives:

1. Defining type III hypersensitivity

2. List the categories of immune complexes diseases
3. Describe the mechanisms favoring the persistence and deposition of immune complexes in the
tissues
4. Explain the effector mechanisms of tissue damage
5. Identify the particularities of the main diseases.

Course plan:

I. Introduction
II. Antigens involved in the formation of immune complexes
III. Factors favoring the deposition of ICs in tissues
IV. Effector mechanisms of lesions during hypersensitivity reaction type III
V. Manifestations due to the deposit of immune complexes during
hypersensitivity reaction type III
1. Circulating immune complexes
2. Immune complexes formed in situ: precipitating antibodies
2.1. Arthrus reaction
2.2. Extrinsic allergic alveolitis (EAA)
a. Farmer's lung disease :
b. The bird breeder lung:
VI. Detection of immune complexes
1. Immune complexes deposited in the tissues
2. Circulating immune complexes

20
 I. Introduction
The immune complexes that cause disease may involve either self or foreign antigens bound
to antibodies. These immune complexes are filtered out of the circulation in the small
vasculature, so their sites of ultimate damage do not reflect their sites of origin. These
diseases tend to be systemic with little tissue or organ specificity.

Under certain conditions, immune complexes can deposit in the tissues and be pathogenic
by triggering inflammation.

The diseases characterized by the deposit of immune complexes could be:

➢ Persistent infections: with persistent production of a small quantity of antibodies

inducing chronic formation of IC (leprosy, malaria, hepatitis viral, endocarditis).
➢ Autoimmune diseases: some of them are characterized by the production of high
quantity of autoantibodies. The high rates of autoantibodies may induce the
saturation of means of transport (on red cells) and elimination (by macrophages) of IC
(SLE, RA).
➢ Repeated introduction of external antigens: the antigen coming from animals, plants
or molds (example: in case of extrinsic allergic alveolitis (EAA) the antigen is inhaled
and induces the local formation in the alveoli of IC. An example of EAA: farmer's
disease).

II. Antigens involved in the formation of immune complexes

➢ Extrinsic antigens
- Heterologous serum proteins: during serotherapy
- Bacterial, viral, parasitic and fungal antigens: microorganisms
- Drugs (penicillin, sulfonamides, etc) : haptens

➢ Intrinsic antigens
- DNA (lupus)
- IgG (rheumatoid arthritis)
- Tumor antigens (cancers)

21
 III. Factors favoring the deposition of ICs in tissues
The deposit of immune complexes in tissues triggers inflammatory response. We distinguish
factors that favor the deposit of IC.

➢ Complement deficiency

Complement plays an important role in the solubilization and the transport of circulating IC.
In case of complement deficiency, ICs are no longer well transported by red blood cells and
will persist in the circulation (with risk of deposit in the tissues). In fact, subjects with C2 or
C4 deficiency often present with immune complexes.

➢ The size of the ICs

- The large complexes are quickly eliminated

- While the intermediate sized complexes are deposited more easily

- Immune complexes are filtered out of the circulation into the renal glomeruli. As much as
the ICs are smaller, the deeper they will deposit in the glomeruli. The damage will be more
serious.

➢ Deficit of phagocyte functions

-
A deficit of phagocyte function will result in the persistence of immune complexes
which will then be deposited
➢ Hemodynamic factors
- Some hemodynamic factors favor deposition of circulating immune complexes in
the endothelium and the basal vascular membrane:
o High pressure: in the glomerular capillaries the pressure is higher than in
the other capillaries, which favors the deposits in the kidney;
o Physiological zones of filtration as renal glomeruli and synovial capillaries,
facilitating deposits in the kidney and the joints;
o Areas of turbulence (arterial bifurcations): Vasculitis (inflammation in the
blood vessels).

➢ Antigen Affinity for certain tissues

- During lupus, exaggerated apoptosis induce the release into the circulation of DNA
associated with histones ➔ the histones because they are positively charged are
deposited in the negatively charged glomerular basement membrane.
- Subsequently, anti-DNA antibodies bind to the DNA in the kidney (ICs).

22
IV. Effector mechanisms of injuries/lesions during hypersensitivity
reaction type III

➢ These diseases are due to the formation and deposit of ICs. The ICs can settle near the
location where they are formed, or can be formed in the blood circulation and deposit
in the tissues.

➢ The complement system is then activated. The neutrophils and monocytes will be
attracted to the sites of deposition of ICs.

➢ The neutrophils, unable to phagocytize ICs, will release proteolytic enzymes and
lysozymes which will induce tissue-damages.

➢ Macrophages secrete pro-inflammatory cytokines (TNFα, IL1).

➢ Platelets are also activated by ICs, by binding to Fc receptors, inducing the release of
vasoactive amines and platelets derived growth factor.

23
V. Manifestations due to the deposit of immune complexes during
hypersensitivity reaction type III

➢ The extent of the reaction depends on the level of immune complexes and their
distribution in the body.

➢ When the Ag-Ab complexes are deposited in a tissue very close to the site of entry of
the antigen, the immune complexes are then formed in Situ and a local reaction
develops.

➢ When immune complexes are formed in the blood (circulating ICs), a reaction can
develop wherever these complexes are deposited.
o Serum sickness represents the typical model of acute illness with circulating
ICs containing external antigen
o ICs formed by internal antigens (as DNA) and autoantibodies are implicated in
the pathogenesis of systemic lupus erythematosus (autoimmune disease).

1. Circulating immune complexes

Serum sickness (as an example)

- Represents the reactions that occur after administration of xenogeneic proteins:

use of serotherapy.
 Production of antibodies directed against foreign proteins (from animals).

- The clinical manifestations are secondary to the deposition of immune complexes,

formed in the blood circulation, in the tissues.

- A single injection causes, between the 8th and 10th day, the following clinical
signs:
o Fever
o Arthralgia
o Vasculitis
o Glomerulonephritis
- The biological signs combine:
o Hypocomplementemia: complement proteins are consumed
o Proteinuria due to renal damage
- Evolution towards regression without sequelae.

24
 2. Immune complexes formed in situ: precipitating antibodies
We distinguish arthrus reaction and extrinsic allergic alveolitis that will be described.

2.1. Arthrus reaction

➢ An Arthus phenomenon can be observed during medication

➢ This phenomenon is due to the reaction of an antigen injected repeatedly under

the skin producing precipitating antibodies

➢ It is a necrotic reaction of semi-delayed hypersensitivity.

➢ The manifestations appear 8 to 10 hours after the injection of the antigen.
➢ The lesions are caused by polymorphonuclear cells attracted and activated by
locally formed immune complexes.

2.2. Extrinsic allergic alveolitis (EAA)

- Some inhaled particles can penetrate deep into the lungs and stimulate the
production of precipitating antibodies, causing local hypersensitivity reactions,
similar to the Arthus reaction.

- Repetitive introduction of antigens into the pulmonary alveoli induce the

production of antibodies against these antigens.

- These reactions affect the alveoli and bronchioles without airway obstruction.

25
 - They induce dyspnea and can lead to respiratory failure if the inhalation becomes
chronic, hence the need to avoid the allergen.

- Farmer's lung disease and bird breeder lung are the best-known examples of EAA
that occur in individuals exposed on a regular basis to inhalation of antigens.

a. Farmer's lung disease :

o It is caused by micro-organisms (actinomycetes) present in damp and
moldy hay.
o Pulmonary signs (cough, dyspnea) start six to ten hours after inhaling
contaminated hay dust.
o Gel immunoprecipitation assay reveals precipitation lines between the
patient's serum and the actinomycete extracts.

b. The bird breeder lung:

o Is seen in breeders of pigeons, parakeets, parrots…
o Avian antigens can cause pneumopathy of immunological origin similar to
that of farmers' lung.
o Precipitating antibodies are observed when the patient's serum is placed
in the presence of extracts from the defecations of these birds.

VI. Detection of immune complexes

1. Immune complexes deposited in the tissues

- By direct immunofluorescence on biopsies using fluorescein-labeled anti-
complement or anti-immunoglobulin antibodies.
- The search and identification of the causative agent (responsible antigen) is often
difficult.
2. Circulating immune complexes
- Circulating ICs are sought in the serum.
- The assay is performed by ELISA using the C1q test
- This test detects antibodies that activate complement by the classical pathway
- However the presence of circulating ICs does not mean a pathological
phenomenon

26
Type IV hypersensitivity: delayed type hypersensitivity
(DTH)

Educational objectives:

- Recognize the effector cells during delayed type hypersensitivity reactions

- Determine the three categories of delayed type hypersensitivity reactions
- Explain the immunological mechanism of DTH reactions
- Describe the tuberculin hypersensitivity
- Explain the formation of granulomas during Granulomatous hypersensitivity
- Determine the mechanism of Contact hypersensitivity reaction
- Cite tests to explore each type of the three forms of delayed type hypersensitivity

Course plan:

I. Introduction
II. Immunological mechanisms of the delayed type hypersensitivity
1. Sensitization phase
2. Onset of the DTH
III. Tuberculin type hypersensitivity: DTH reaction
1. Detection of tuberculin-type hypersensitivity
2. In vitro detection tests of tuberculin-type hypersensitivity:
IV. Granulomatous hypersensitivity
1. Circumstances of appearance
2. Immunological mechanisms
3. Diseases associated with granulomatous HS
V. Contact hypersensitivity (or allergic contact dermatitis)
1. Clinical presentation
2. Mechanism
3. Diagnosis of allergic contact dermatitis
4. Treatment of allergic contact dermatitis

27
I. Introduction

➢ The delayed type hypersensitivity (DTH) is the pathologic manifestation of the

cellular immunity. Then, DTH depends on antigen-specific Th1 type T cells. The
effector cells are T lymphocytes, monocytes/macrophages…).

➢ DTH is the set of local inflammatory manifestations, induced by a new penetration of

a sensitizing antigen. Then, we distinguish a sensitizing phase and an effector phase.

➢ The manifestations of the effector phase appear at the 12th hour, reaching a
maximum between 48-72 hours.

➢ There are 3 forms of delayed type hypersensitivity (DTH):

- Tuberculin hypersensitivity which is a defense reaction against intracellular
multiplying microorganisms.
- Granulomatous hypersensitivity which develops in the presence of resistant
intracellular multiplying microorganisms.
- Contact hypersensitivity reaction due to skin contact with harmless chemical
agents.

28
II. Immunological mechanisms of the delayed type hypersensitivity

➢ A physiological cellular immune response is a normal component of adaptive

immunity (cytotoxic response and Th1 response) and is essential for the control of
intracellular pathogens and tumor cells.

➢ In certain circumstances, if this response is exacerbated, poorly controlled and

unregulated, it leads to tissue damage.

1. Sensitization phase
- Once the antigen is introduced under the skin, it will be captured by immature
dendritic cells which will undergo a maturation process and migrate into the
secondary lymphoid organ.

- Dendritic cells (DCs) will present the antigens to T cells that will be activated and
differentiated in antigen-specific Th1 lymphocytes under the effect of IL-12
produced by DCs. Th1 cells will proliferate (clonal expansion) and give rise to
effector specific Th1 lymphocytes and to memory Th1 cells. These latter will
migrate to the periphery.

2. Onset of the DTH

➢ In vitro studies have made it possible to understand the immunological mechanisms

involved in DTH reactions, and to demonstrate that these reactions take place in two
distinct phases: an initial specific inducing/amplifying phase, then a non-specific
effector phase.

➢ Initial specific inducing/amplifying phase:

o Once the antigen is injected another time, it will be captured by the dendritic
cells of the dermis which present it to memory T lymphocytes of type Th1.
o More T cells are recruited and activated leading to their proliferation, and the
formation of new effector and memory Th1 lymphocytes.
➢ During the effector phase:
o The effector T lymphocytes release cytokines in particular TNF and IFNɤ,
which will recruit and activate leukocytes.
o These factors are also active on endothelial cells inducing the expression of
adhesion molecules and the release of chemotactic factors (IL-8, MCP-1).
o IFNɤ increases the expression of MHC molecules.
o Lymphokines are synthesized as LMIF (leukocytes migration inhibition factor)
and NAF (neutrophil activating factor), which explains the initial influx of
neutrophils, during the first hours of the reaction of HSR.

29
 o T lymphocytes produce factors that will recruit and activate monocytes-
macrophages and Langerhans cells: MCP (macrophage-chemotactic protein-
1), and MIF/MAF (macrophage migration-inhibiting factor/macrophage-
activating factor).
o Macrophages secrete Factors with microbicidal activity (lysosomal enzymes,
oxygen derivatives) and pro-inflammatory cytokines (IL-1, IL-6 and TNF).

o If the germ is resistant to lysis, the activation of macrophages is intensified

which can lead to tissue damage following the secreted factors.
o As an example, during tuberculosis, tissue damage is partly due to the
intensified activation of the immune system.

III. Tuberculin type hypersensitivity: DTH reaction

- It is a physiological defense reaction against microorganisms with intracellular
multiplication: Intracellular bacteria (mycobacteria, brucella, salmonella…),
parasites with intracellular multiplication (leishmania, toxoplasma, candida…).
- The stimulation of microbial antigen-specific T cells leads to macrophage
activation and localized inflammation.
- This response is essential for the control of intracellular pathogens.
- The presence of these microorganisms is detected by skin tests: intradermal test
(IDT).

1. Detection of tuberculin-type hypersensitivity

- Sensitized subjects: subjects contaminated with the tubercle bacillus (Koch's

bacillus- KB) or vaccinated with BCG. This phase takes a few weeks to happen.
30
- DTH reaction:
o An extract of cultures of tubercle bacilli (called tuberculin) is injected into
the derm (intradermal injection): a Kock Bacillus extract (0.1 ml of
tuberculin equivalent to 5 units) is injected into the anterior face of the
forearm.

o 24 hours later, DTH reaction to tuberculin appears: erythema with

induration, ± blisters (a fluid-filled elevation of the epidermis) and
necrosis.
o The reading after 48 to 72 hours, maximum: the diameter of the
induration is measured, not the erythematous area (red) around the
induration.
o The reaction is considered negative when the induration diameter is less
than 5 mm

o In fact, the skin becomes indurated following the infiltration of the dermis
by mononuclear cells (lymphocytes and macrophages).

- IDR it serves as a diagnostic test in the identification of agents involved in several

infections.

- The IDR with tuberculin is carried out to:

o Search for prior contact with BK (suspicion of infection with BK)
o Verify if the subject has taking the BCG vaccine
o Explore a deficit of cellular immunity (reduction of DTH reactions).

31
 2. In vitro detection tests of tuberculin-type hypersensitivity
- The patient's lymphocytes are cultured with the relevant antigen.
- If he has memory lymphocytes, they will proliferate and secrete cytokines that can
be measured (especially IFNɤ).
- This is not done routinely.

IV. Granulomatous hypersensitivity

1. Circumstances of appearance

➢ Granulomatous inflammation is an example of chronic inflammatory response.

➢ It follows the persistence of the infectious antigen with intracellular multiplication.

➢ The infectious agent, poorly eliminated, maintains a persistent inflammatory reaction

which can be a source of tissue damage

➢ It appears after 21 to 28 days of the contact.

2. Immunological mechanisms

- The persistence of the infectious antigen induces chronic activation of Th1

lymphocytes which secrete INFγ and TNF.

- Macrophages are massively recruited and are locally activated.

32
 Immunological mechanisms

- They differentiate into epithelioid cells and some merge to transform into giant cells
and form granulomas.

- The granulomas that form around the infectious focus limit the dissemination of
germs (diffusion in the rest of the body).

- Granulomas are made up of lymphocytes, macrophages, epithelioid cells, giant cells

and fibroblasts.

o Epithelioid cells: classically macrophages that have lost their function of

migration and phagocytosis but possess an important secretory function.
o Giant cells (or multinucleated cells) come from the fusion of epithelioid cells
present in granulomas.

- Activated macrophage cells secrete pro-inflammatory cytokines (IL-1, TNFα) with

exaggerated inflammation and tissue damage, and growth factors with fibrosis
formation.

33
 3. Diseases associated with granulomatous HS
Granulomatous HS develops in different diseases:

- Infectious diseases where the antigen is known: tuberculosis (pulmonary

granulomas), leprosy (cutaneous granulomas)
- Chronic inflammatory diseases where the antigen is unknown: sarcoidosis (lung),
Crohn's disease (bowel).

V. Contact hypersensitivity (or allergic contact dermatitis)

1. Clinical presentation

- Contact HS develops at the point of contact with an allergen which is a harmless

agent.
- It is frequent and often of professional origin (7% of dermatology consultations in the
USA).
- It is clinically characterized by eczema: red, dry and itchy patches (oval or circular-
shaped areas). It is an inflammation of the skin.

34
2. Mechanism
- This reaction is observed following a contact with agents of small size (known as
haptens), not immunogenic by themselves such as nickel (costume jewelry..),
chromium salts (construction trades..) , rubber derivatives (gloves, etc.)

- Two phases for the allergic reaction: sensitization and elicitation phases

- Sensitization phase:

o The agents are lipophilic and easily penetrate the epidermis where they
combine with self-proteins to form neo-antigens.
o Langerhans cells ingest the antigen and migrate through lymphatic vessels to
regional lymph nodes.
o In the paracortex, they present the antigen to CD4 T cells.
o CD4 T cells differentiate into memory cells and migrate to the skin

- Elicitation phase
o Upon subsequent introduction of the same hapten, Langerhans cells take up
the hapten-protein conjugate and move from the epidermis to the dermis
where they present the antigen to memory T cells.
o Memory T cells release interferon gamma which activates keratinocytes and
endothelial cells of the dermis.
o Activated keratinocytes secrete inflammatory cytokines (IL1, IL6…) and
chemokines.
o CD4 T cells and macrophages are attracted and infiltrate the epidermis where
they increase inflammation.

35
3. Diagnosis of allergic contact dermatitis

- Its diagnosis requires skin tests (Patch tests) (especially in a professional environment
where several chemicals are used).
- These tests make it possible to find the product responsible for allergic contact
dermatitis:
o A battery of known antigens is used: different dilutions of the haptens are
applied.
o The antigen is applied to the skin (percutaneous application).

- The reading is done after 48 hours.

- The test is positive in sensitized subjects.

Skin-patch test preparation

36
 Skin-patch test application

Skin-patch test reading

4. Treatment of allergic contact dermatitis

- It is necessary to eradicate the responsible agent if it has been determined (possibly

change profession).
- Apply corticosteroids locally.

37