Immuno 4 - Allergy

Question 1

What is the difference in response to a bacterium/virus/ fungi vs a response to a worm/ venom/ allergen?

Answer 1

bacterium/virus/ fungi –> Th1 response against intracellular parasites –> driven by pathogen associated molecular receptors

worm/ venom/ allergen –> Th2 response against extracellular parasites –> due to loss of tissue function (functional change recognised by immune system)

In contrast to immune responses to bacteria, virus and fungi, immune responses to worms, venoms and allergens tend to react to tissue damage caused by these agents rather then relying on direct recognition of the pathogen

Worms, venoms and allergens are far more diverse than bacteria

Question 2

What triggers mast cell degranulation,
what is being released
what is being targeted by drugs
what is the function of these mediators?

Answer 2

What triggers mast cell degranulation –> IgE cross linking
what is being released –> PG, Leukotrienes, histamine, proteases

what is being targeted by drugs –> histamine, leukotrienes

what is the function of these mediators –> act on endothelium –> worm + allergen expulsion, enhanced epithelial barrier function

more detail in flashcard 13

Question 3

Where are innate lymphoid cells found?

Answer 3

Mucosal barriers (skin, respiratory , GIT)

Question 4

What is the function of ILC2 (innate lymphoid cell 2)?

Answer 4

secretes predominantly IL5, IL13, AREG (amphiregulin)

Also secretes IL-4, IL-9

Implicated in allergic asthma, allergic rhinitis, food allergy, eosinophilic oesophagitis

AREG - important in epithelial barrier repair in skin + respiratory tract

 In allergic disorders, upregulation of the activity of the innate lymphoid cells tends to overcome steady state inhibition exerted by tissue CD-4 regulatory T cells

Question 5

Which markers do CD4 Th2 cells express and which cytokines do they release?

Answer 5

CD4 Th2 express
Transcription factor GATA-3
Signal transduction protein STAT-6

Release
IL4 - helps B cells produce IgE, involved in Th2 development, particularly in the context of antigen presentation to a naïve/memory CD4 T cell
IL5 - expands and activates eosinophils
IL13 - stimulates mucus production

Question 6

key cytokine in the development and expansion of eosinophils

Answer 6

IL-5

Question 7

IgE

Which receptors does it bind to and where

Answer 7

High affinity receptor (FcεR1) on mast cells, basophils, eosinophils and DC

Low affinity’ (FcεR2) receptor on above cells +
B cells, respiratory and GI epithelial cells

Question 8

Factors that promote IgE prodution

Answer 8

Antigen dose
Physical properties of antigen

Route of exposure
Oral exposure - immune tolerance
Skin + respiratory exposure - IgE sensitisation
Length of exposure
Low dose, accumulative exposure over time –> will promote IgE rather than IgG

Question 9

How does oral exposure to antigen promote tolerance?

Answer 9

 When an allergen is ingested through the oral route, T-regs (from GI mucosa) inhibit IgE synthesis

• Oral tolerance requires induction of CD4+ T-reg cells
• T-regs inhibit multiple pro-allergic functions such as inhibiting DC APC function, secretion of IL-10, etc.

Question 10

Summary of Th2 immune responses

Trigger
What does the trigger release 
Where do these factors act
What do they induce
Where do these factors act

Answer 10

Trigger
Stressed or damaged endothelial cells secrete
IL25
IL33
GMCSF - granulocyte macrophage colony stimulating factor
TSLP - thymic stromal lymphopoietin

Factors act on
Tissue immune cells (DC, basophils, T2 innate lymphoid cells)
Neurones

Induce Th2 cell immune responses –> release of IL4, IL5, IL9, IL13

Th2 cytokines secreted by tissue lymphocytes act on effector cells -( basophils, oeosinophils, epithelial cells, B cells, sensory neurones, endothelium, smooth muscle cells) –> expel pathogen / allergens + repair tissue damage

IL4 - helps B cells produce IgE, involved in Th2 development, particularly in the context of antigen presentation to a naïve/memory CD4 T cell
IL5 - expands and activates eosinophils
IL13 - stimulates mucus production

Question 11

Where are the 2 types of mast cells found and what do they release?

Answer 11

MC skin –> Tryptase

MC airways –> Chymotryptase

Question 12

Mast cell degranulation is triggered by

Answer 12

o IgE/IgG receptors which respond to antibody-antigen cross linking

IgE receptor FcεR1
IgG receptor FcγR1, FcγRIA

o G-protein-coupled receptors which are ligands for soluble mediators (complement and drugs)

Question 13

Mast cell degranulation leads to

Answer 13

o Vasodilatation
o Recruitment of soluble proteins and inflammatory cells to site of infection
o Increase rate of lymphatic flow back to regional lymph nodes to enhance adaptive immune responses
o Smooth muscle contraction in lungs and gut – may help expel pathogens
o Activation of sensory neurones in respiratory cutaneous tissues– itch, sneeze

Question 14

What needs to happen for L4 to be induced and why is it important?

Answer 14

o Plays a crucial role in the development of Th2 immune responses
o Is only induced following peptide-MHC presentation to naïve/memory Th2 cell receptor

Question 15

Which mechanism is responsible for the

immediate (2-3h, max 4h)
delayed (after 12h)

symptoms of allergy

Answer 15

• Rapid onset of symptoms (within 2-3h, max. 4 hours)  caused by release of inflammatory mediators following allergen cross linking of IgE on surface of mast cells and basophils
• Delayed symptoms (after 12 hours)  caused by CD4 T2 cell (IL-4, IL-5, IL-13) immune responses and eosinophil related tissue damage

Question 16

Describe the hygiene hypothesis

Answer 16

Hygiene Hypothesis  lack of childhood exposure to infectious agents increases susceptibility to allergic diseases by suppressing natural development of the immune system

 In essence, this hypothesis states that the default system for the development of the immune system in neonates and children is the Th2 immune response and that exposure to infection deviates this to a Th1 immune response

Question 17

What biological mechanism can protect against development of asthma?

Answer 17

D. Increased secretion of pro-inflammatory innate cytokines by PBMC following exposure to environmental microbial product may protect against development of asthma

Question 18

Allergic illnesses in

infants
children

Answer 18

infants
atopic dermatitis
food allergy

children
asthma
allergic rhinitis

Question 19

Clinical features of IgE allergic responses

Symptom onset
systems involved

Answer 19

within m or up to 3h after exposure
Symptoms reproducible
Symptoms may be triggered by co-factors (i.e. NSAIDs in asthma, virus in children to VIF)

 Symptoms – at least 2 organ systems usually involved
• Skin symptoms  urticaria, angioedema
• GI symptoms  D&V
• Respiratory tract symptoms  SoB, cough, wheeze
• Vasculature symptoms  hypotension, impending doom

Question 20

Positive and negative control in skin prick

Positive skin prick test

what needs to happen before the test

Answer 20

positive control - histamine
negative control - diluent

• Positive outcome = wheal ≥3mm than negative control
• Antihistamines discontinued for 48 hours beforehand (checked with positive control)

Question 21

positive and negative predictive value of skin prick test

Answer 21

positive = poor positive predictive value - high false positive rate
Positive predictive value is the probability that subjects with a positive screening test truly have the disease.

negative = excellent negative predictive value usually >95%
This characteristic can predict how likely it is for someone to truly be healthy, in case of a negative test result

Question 22

How do IgE RAST (radioallergosorbent) sensitisation blood tests work?

Answer 22

o (1) Allergen bound to sponge in a plastic cap and patient’s serum is added
o (2) Specific IgE (if present) binds to allergen
o (3) Anti-IgE antibody tagged with a fluorescent label is added
o (4) Amount of IgE/Anti-IgE is measured by fluorescent light signal
o Very reliable, expensive

Question 23

Indications for an IgE blood sensitisation test vs skin prick test

Answer 23

No access to SPT and/or IDT

Patients who can’t stop anti-histamines

Patients with a history of dermatographism, extensive eczema

Patient with a history of anaphylaxis

Decision on who needs food challenge

Prediction for resolution of egg, milk, wheat allergy

Monitor response to anti-IgE therapy

Question 24

How do we diagnose allergic disease

Answer 24

• History – key to the diagnosis

• Examination
o Dry skin
o Wheeze

• Allergen-specific IgE (sensitisation) tests - these tests do not predict the severity of the reaction, but larger skin wheals + higher specific blood IgE values are more likely to be associated with an allergic disorder
o Skin prick and intradermal test
o IgE blood test
o Clinical history is used to select what allergens should be tested by skin prick and/or blood tests

•	Functional allergen tests
o	In vitro tests 
	Basophil activation
	Serial mast cell trypase – useful for the dx of anaphylaxis, particularly that occurring under GA
o	Ex vivo tests
	Open or blinded allergen challenge

Question 25

Next investigation if skin prick test is negative but hx is convincing

Answer 25

Intradermal tests  More sensitive but less specific than SPT  can be used if SPT to allergen is negative but convincing history

Question 26

What does component resolved/molecular allergen testing test for and what are the indications for it?

Answer 26

 Blood test to detect IgE sensitisation against individual protein within whole allergen extract  Indications • Detect primary sensitisation • Confirm cross-reactivity • Define risk of serious reaction for stable allergens • Improve diagnostic sensitivity for components poorly represented in whole food extracts • Improve diagnostic sensitivity for unstable molecules in whole food extracts  Second line – reflex test for positive whole allergen blood tests

Question 27

How long does serial mast cell tryptase need in order to reach peak concentration in blood and when does it return to the baseline?

Answer 27

o Peak concentration = 1-2 hours; baseline = 6-12 hours

Question 28

• Serial mast cell tryptase test use

Answer 28

o In vitro test o Measures tryptase o Useful if diagnosis of anaphylaxis uncertain (e.g. hypotension/rash in anaesthesia, particularly serial analysis of mast cells and serum tryptase) o Reduced sensitivity for food-induced anaphylaxis

Question 29

What does the basophil activation test measure?

Answer 29

o Measurement of basophil response to allergen IgE cross-linking o Upregulation of some basophil surface proteins following exposure to the allergen o Activated basophils increase expression of CD63, CD203 and CD300 protein on cell surface o Increasingly used in the diagnosis of food and drug allergy – surrogate marker for challenge tests

Question 30

• Gold standard for food and drug allergy diagnosis

Answer 30

 In vivo test: dou ble blinded allergen challenge (challenge tests – food and drug allergy)

Question 31

What should children with moderate/severe atopic dermatitis be tested for?

Answer 31

o Moderate/severe atopic dermatitis = important risk factor for food allergy – indication for allergy testing even in absence of clinical history

Question 32

• IgE-mediated food allergy syndromes:

Answer 32

o Anaphylaxis  Peanut, tree nut, shellfish, fish, milk and eggs are most common  Natural history dependent on food o Food associated exercise induced anaphylaxis  Food induces anaphylaxis if individual exercises within 4-6 hours of ingestion  Common food triggers are wheat, shellfish, celery o Delayed food-induced anaphylaxis to beef, pork, lamb  Symptoms occur 3-6 hours after eating red meat and gelatin  IgE antibody to oligosaccharide alpha-gal (α1, 3-galactose) found in gut bacteria  Induced by tick bites which should be avoided o Oral allergy syndrome  Limited to oral cavity, swelling and itch  Only 1-2% cases progress to anaphylaxis  Onset after pollen allergy established  Affects adults > young children  Respiratory exposure to pollen (birch)  results in IgE directed to homologous proteins in stone fruits (apple, pear) vegetables (carrot) and nuts (peanut, hazelnut)  Cooked fruits, vegetables and nut cause no symptoms  heat labile allergens detected by component allergen tests

Question 33

Sensitivity test + indications Skin prick (intradermal) RAST IgE blood test Component Resolved Diagnostics (CRD)

Answer 33

``` Skin prick (intradermal) To exclude allergy (high NPV) ``` RAST IgE blood test To exclude allergy (high NPV) When skin prick is not possible (i.e. cannot stop antihistamine) Component Resolved Diagnostics (CRD) Detect IgE to peanut and hazelnut components in specific situations (i.e. poorly represented components in whole food) Second line – reflex test for positive whole allergen blood tests

Question 34

Diagnostic test + indications Serial mast cell tryptase Basophil activation Allergen challenge

Answer 34

Serial mast cell tryptase Diagnosis of anaphylaxis (uncertain cause) Basophil activation Diagnosis of food and drug allergy (surrogate marker for challenge tests) Allergen challenge Diagnosis of food and drug allergy Gold standard