The Immune System in Action (12-17)

Question 1

After the immature B cell has formed, what happens next in B cell development?

Answer 1

B cells are tested for reactivity to self antigens, called B cell tolerance

occurs in the spleen in two stages:
1. T1 - B cells are checked for autoreactivity, if they receive a strong signal if they encounter self antigens they are deleted, B cell peripheral tolerance
2. T2 - ?

B cells move through white pulp between T1 and T2

Question 2

In what stages does the T cell develop?

Answer 2

occurs in thymus, called ‘thymocytes’
leave bone marrow and travels through blood by following chemokine signals

stages of development are defined by expression of CD molecules:
- 4 stages of double negative expression (DN1-4)
- 1 double positive stage (DP)
- then express either CD4 or CD8

after DN3 is where gamma delta T cells can arise
alpha chain recombination occurs after DN4
positive selection takes place after DP so only T cells w good receptors remain
after SP, negative selection occurs to filter out autoreactivity

Question 3

Describe beta selection in T cell development.

Answer 3

Occurs between DN3 and DN4

• pTalpha (pre alpha receptor) joins rearranged beta chain, forming pre T cell receptor
• pTα is required for expression of the rearranged β chain on the cell surface and complex with CD3 (needed to signal)
• Expression of the pre-TCR is enough to induce signalling. Ligand binding is not required.

Question 4

Describe positive selection in T cell development?

Answer 4

occurs during DP stage

it is the selection of cells w functional TCRs able to intercts w self MHC-peptide
only T cells able to interact w MHC and signal through their TCR are useful

a lot of the alpha chains generated by gene rearrangement will not be able to make functional TCRs, determines MHC restriction

this is done by cortical thymus epithelial cells (cTEC) present peptides to DP cells both in MHC I and II

Question 5

Reminder: What cells do CD4+ and CD8+ T cells develop into?

Answer 5

CD4+: Th or Treg cells (if intermediate affinity for MHC)
CD8+: CTL

Question 6

Describe negative selection in T cell development.

Answer 6

occurs during SP stage

deletion or modification of autoreactive CD8 and CD4 cells in thymus
recognition of cell would lead to autoimmunity
done by SP T cells interacting w:
- DCs: present self antigens from blood/lymph
- mTECs (medullary thymic epithelial cells): present self antigens from tissues areound the body, tissue restricted antigens (TRAs)

high affinity binding = apoptosis
intermediate affinity binding = Treg
low affinity binding = survival

negative selection is central mechanism for central tolerance of T cells

Question 7

How are mTECs able to present antigens from elsewhere in the body?

Answer 7

AIRE (autoimmune regulator) protein drives expression of tissuerestricted antigens (TRAs) in mTECs

Question 8

What are the two arms of tolerance?

Answer 8

central tolerance: occurs in generative lymphoid organs during development of the immune system (embryonic/neonatal)
immature lymphocytes specific for self antigens undergo:
- deletion (apoptosis): T and B cells
- receptor editing: B cells
- Differentiation into Treg cells: CD4 T cells

peripheral tolerance: occurs in peripheral tissues
mature self-reactive lymphocytes that escape central tolerance are
- rendered nonresponsive (anergy)
- deleted
- suppressed by Tregs

peripheral tolerance prevents reactivity to antigens which will not be present in thymus such as food and commensal microbes
is a failsafe for mature and self reactive T cells that escape central tolerance

Question 9

Peripheral tolerance: How is anergy induced in T cells?

Answer 9

• absence of costimulation from APC eg reactive to self peptide
• engagement of inhibitory receptors

Question 10

Peripheral tolerance: How are T cells deleted?

Answer 10

mitochondrial pathway
- repeated signalling downstream of TCR without costimulation induces BIM/BID, leading to cytochrome c release from mitochondria -> activation of caspases and apoptosis

Fas/FasL
- Fas expression induced in T cells. binding of FasL to Fas activates caspases

Question 11

Peripheral tolerance: How are T cells suppressed by Tregs?

Answer 11

contact-dependent mechanisms: CTLA-4
1. blocks B7 (CD80/86) APC so not available for effector T cells to bind to CD28
2. removes B7 from APC
3. CTLA-4 binding to B7 sends inhibitory signals to APC -> less antigen presentation -> less production of cytokines by APC

cytokine-mediated mechanisms
1. production of suppressive cytokines (IL-10, TGFbeta)
2. consumption of available IL-2 by CD25 on surface so other T cells cant use it

Question 12

What is FOXP3?

Answer 12

TF needed for Treg differentiation and function

Question 12

What are two types of Tregs?

Answer 12

Tregs are generated
- in the thymus ((n)atural Tregs): generated during negative selection
- in the periphery ((i)nduced Tregs): generated in the presence of TGFbeta

Question 12

What CD receptors do Tregs express on their surface?

Answer 12

CD25 - high affinity IL-2 receptor
CTLA-4 - inhibitory receptor

Question 13

What is sex bias in the context of autoimmunity?

Answer 13

higher incidence of autoimmunity in women
- hormonal differences
- x-inactivation
- impact of pregnancy on immune system, perhaps

Question 14

How can infection trigger autoimmunity?

Answer 14

through activation of APCs during somatic hypermutation

APC presenting self antigen at time of infection can cause self reactive T cell to become activated

through molecular mimicry
a microbe that has the same antigen as a self antigen
eg production of antibodies against streptococcal cell wall cross react w heart valve tissue (carditis) or skin tissue (erythema marginatum)

Question 15

How can physical trauma cause autoimmunity?

Answer 15

there are immunologically privileged sites eg eyes, brain, testis, and developing fetus

trauma to blood-site barriers can cause autoimmunity by allowing lymphocytes to enter

Question 16

What is Rituximab?

Answer 16

an anti CD20 molecule that depletes B cells developed for B cell-lymphomas
approved for treatment against:
- rheumatoid arthritis
- ANCA-associated vasculitis
- pemphigus vulgaris

not specfic for autoreactive B cells
more vulnerable to infections/cancer

Question 17

List some properties of gamma delta T cells.

Answer 17

adaptive lymphocytes, but innate like properties
limited RAG-dependent TCR diversity
confined to skin and epithelial barriers, without recirculation
not MHC restricted, can directly recognise Ag
functions still being clarified

Question 18

Reminder: What signals are involved in T cell activation?

Answer 18

1. TCR-peptide MHC engagement
2. co-stimulation eg CD28:CD80/86 (B7.1/B7.2)
3. cytokines eg IL-12, IL-4

all from APC
type of cytokine produced decides what T cell the t cell differentiates into

type of cytokine produced depends on type of challenge encountered

Question 19

What is the major co-stimulatory signal provided by in T cells

Answer 19

CD28
effective at mediating tolerance

IL-2 produced by activated APCs and is part of positive feedback loop which keeps cell alive and allows clonal expansion

Question 20

How do CD4+ T cells initiate responses to diverse pathogens?

Answer 20

Th1 produces inflammatory cytokines which activate macrophages for intracellular pathogens which cant be reached by complement or ab - engulf entire infected cell

Th17 produces IL-17 which activates neutrophils which is good at killing extracellular pathogens such as bacteria and fungi

Th2 activates eosinophils for removal of helminths and ectoparasites and viruses (Abs)

Question 21

What TFs specify CD4 T cell fate?

Answer 21

Tbet ->
Gata3 ->
Rorγt -> Th17
Foxp3 -> Treg

Question 22

How do CD4 T cells help B cells?

Answer 22

• recognising peptide in MHC class II
• deliver cytokines such as IL-4 and IFN-γ
• surface ligation of B cell CD40 by T cell CD40L

occurs in lymph node

Question 23

How does activation of macrophages by Th1 eliminate pathogens?

Answer 23

- iNOS in macrophages kills intracellular pathogens - causes tissues inflammation - raise of temperature reduces viral replication

Question 24

What is interesting about the interaction between Th1 and Th2?

Answer 24

mutually antagonistic inhibit each other, if one is present more than the other, that ones response will become dominant IL-10 indirectly inhibits Th1 IFN-gamma inhibits Th2

Question 25

What are some properties of Tregs?

Answer 25

defined as CD4+CD25+ which express Foxp3 mice deficient in CD25, IL-2, CTLA-4, Foxp3, IL-10 or TGF-beta develop severe immunopathology Tregs suppress all functional CD4 subsets - IL-10 and TGF-beta

Question 26

What are CTLs?

Answer 26

cytotoxic T cells CD8+, only one type of CD8 T cell require CD8+ cells require IL-2, need CD4+ T cell help essential for defence against viruses immune surveillance against tumours graft rejection

Question 27

How do T cells kill?

Answer 27

perforins create membrane pores and CTLs release granule contents into target cell granules specifically target so not released into general environment as they are v powerful, ensure they wont kill healthy cells forms junction, actin moves back so granules pushed forward into cell Fas/FasL-mediated killing FasL on CTL interacts w Fas on target cell causes apoptosis

Question 28

What are proto-oncogenes?

Answer 28

genes, if mutated or incorrectly regulated, contribute to cancer progression protein products stimulate cell growth and division eg growth factors and their receptors positive regulation mutated versions are known as oncogenes

Question 29

What are tumour suppressor genes?

Answer 29

protein products stop cell growth and promotes apoptosis prevent unwanted proliferation of damaged cells eg p53 - induces death in cells w DNA damage negative regulation

Question 30

What is tumour vascularisation?

Answer 30

tumours induce angiogenesis, the formation of new blood vessels directly into the tumour to be supplied w oxygen and nutrients allows benign tumour to become malginant

Question 31

Is susceptibility to cancer purely genetic?

Answer 31

no, incidence of cancers is higher in immunosuppressed patients

Question 32

If cancers present self antigens, how does immune system respond?

Answer 32

tumour specific antigens - present on tumour cells but not normal cells - viral proteins of cause by oncogenic virus - mutated versions of normal proteins eg tumour suppressors - chromosomal rearrangements creating proteins which are fusions of two genes tumour associated antigens - antigens found on normal cells but at higher levels on tumours - antigens found on normal cells at different stages of development eg onco-fetal or testis antigens

Question 33

What is a common method used by tumours to escape immune responses?

Answer 33

loss of MHC I no MHC I prevents CD8 T cells from initiating a cytotoxic response can be overcome by NK cells, which detect downregulated MHC on cells upregulation of stress proteins such as MIC-A and MIC-B push NK cells towards activation detection of MHC I reduces expression of stress proteins

Question 34

What role do macrophages play in cancer suppression?

Answer 34

- activate other immune cells via cytokine release eg IL-12 - induce apoptosis - phagocytose cancerous cells however macrophages can become tumour-associated (M2-like TAMs) - suppress immune response via other cytokines such as TGF and IL-10 - promote invasion and metastasis - provide growth factors - promote angiogenesis

Question 35

How can antibodies be used as a therapeutic?

Answer 35

Neutralisation – prevent receptor ligation Activation – trigger receptor ligation Antibody dependant cellular cytotoxicity - depletion Antibody-drug conjugates -targeted delivery of treatment

Question 36

How can monoclonal antibody (Mab) treatments be used to target tumours?

Answer 36

direct targeting - block growth factor receptors eg Trastuzumab - deplete tumour cells expressing tumour associated antigens eg rituximab target the tumour immune response - block checkpoint inhibitors - enhance BiTEs to enhance T cell function

Question 37

What are BiTEs?

Answer 37

fusion proteins consisting of two single chain variable fragments (scFv) derived from different antibodies one binds to CD3 and the other a tumour cell which draws T cell and tumour closer together and induces T cell signalling will result in directed tumour lysis and help to drive anti-tumour response

Question 38

What are CAR T cells?

Answer 38

have the TcR replaced w chimeric fusion protein consisting of various domains from BcR, TcR, and costimulatory molecules to create a super receptor to enhance T cell activation can also have CAR NK and CAR M cells

Question 39

What are the levels of IgM and IgG during primary and secondary response?

Answer 39

IgM peaks earlier than IgG, but IgG levels are much higher than IgM in secondary and tertiary responses

Question 40

Is memory a property of the system or is it carried by cells that have an enhanced functions?

Answer 40

both cells are same but over period of time, increase of affinity: property of system somatic hypermutation inserts point mutation in B regions: enhanced function

Question 41

What are some characteristics of memory B cells?

Answer 41

express switched isotypes (A, G, E), some remain IgM which have special function, "stem cells of immune system" express somatically mutated Ig V genes express higher affinity Ig lower activation thresholds increased levels of adhesion molecules (CD44, LFA-1)

Question 42

Where are memory B cells generated and activated?

Answer 42

generated in secondary lymphoid tissues activated in germinal centre Bcl6 TF crucial for GC development Blimp1 necessary for plasma cell development activated B cells create short lived B cells or proliferate to GCs, where they can differentiate into long-lived plasma cells or memory B cells

Question 43

What is the structure of a GC?

Answer 43

formation is at border between B cell and T cell zones in follicles , both are necessary for proliferation two zones - proliferating cells - cells which present antigen so necessary for maturation initiation of GC reaction - antigen: localisation of immune complexes onto follicular dendritic cells (FDC) - signals from T cells: memory is characteristic of T dependent responses

Question 44

What occurs in germinal centres?

Answer 44

- clonal expansion of antigen-specific B cells - affinity maturation -- somatic hypermutation -- antigenic selection both are crucial components of memory B cell formation

Question 45

What cells are present in GCs?

Answer 45

centroblasts - B cells activated and rapidly divided centrocytes - B cells with receptors with higher affinity helper T cells FDCs - explained in another card

Question 46

What are FDCs?

Answer 46

follicular dendritic cells have chains of beads along dendritic process - iccosomes, which contain electron dense antigens presented to B cells which internalise them do not present to T cells intermediary APC required for presentation to T cells b cell that are not selected by antigen on FDC soon die and are taken up by tangible body macrophages

Question 47

What are some characteristics of memory T cells?

Answer 47

- change in the isoform of CD45 expressed - changes in expression of adhesion olecules eg CD44, ICAM-1, as well as chemokine receptors - altered migration pathways - lower activation thresholds - clonal selection may lead to affinity maturation - no isotype switching - no somatic mutation - dont require germinal centres or any morphological distinct site - dont mutate receptors

Question 48

How do T cells migrate and enter lymph nodes?

Answer 48

- naive T cells enter lymph nodes from the blood by crossing high endothelial venules (HEV) - changes in adhesion molecules and chemokine receptors allow entry in both inflamed and non-inflamed tissues - hence many memory cells enter lymph nodes from the tissues via afferent lymphatics - some memory cells still enter lymph nodes across HEV

Question 49

What are homing potentials of T cells?

Answer 49

expression of different adhesion molecules and chemokine receptors changes what T cells can enter each tissue has a different combination code to enter naive and memory T express CCR7 so can enter lymph nodes effector memory T cells do not express CCR7, instead enter skin or mucosa

Question 50

What are two theories of memory T cell differentiation?

Answer 50

1. divergent two different conditions = two different cells (effector or memory) 2. linear eg naive differentiates into effector, then if conditions are right, differentiates into memory entry into memory pool may simply be a result of resistance to or escape from activation induced cell death (AICD)

Question 51

is a memory cell inherently long lived or does a memory cell require external stimuli for long term survival?

Answer 51

B and T memory cells require external signals

Question 52

What are the properties of cytokine effects?

Answer 52

pleiotropy - multiple biological effects on different cell types redundancy - different cytokines have similar effects on same cells synergy - combined effects greater than additive effect of each cytokine antagonism - effect of one cytokine inhibits effect of another

Question 53

What are some properties of cytokine receptors?

Answer 53

- extracellular and intracellular domains - 2 or 3 receptor subunits (alpha, beta, gamma) - one is cytokine-specific, the other is shared - shared subunits result in redundancy - cytoplasmic tail of receptor subunits transduces signals

Question 54

How are cytokine signals transduced?

Answer 54

binding of a cytokine to the receptor induces phosphorylation by a kinase leads to recruitment of specific STATS type of STAT contributes to type of response

Question 55

How can cytokine receptors be negatively controlled?

Answer 55

- degradation of signal transduction elements - receptor antagonists block receptors - soluble receptor or antibodies bind away from cell - altered receptors cannot signal

Question 56

What is the function of IL-3 and IL-5?

Answer 56

produced by Th2 IL-3: stimulates growth and differentiation of mast cells and eosinophils IL-5: 1. stimulates growth and differentiation of eosinophils 2. pleiotropic - also switches B cells to IgA

Question 57

What is the function of GM-CSF?

Answer 57

granulocyte macrophage colony stimulating factor produced by Th17 essential for differentiation of granulocytes, macrophages and dendritic cells

Question 58

What is the function of IL-1?

Answer 58

produced by macrophages pro-inflammatory vasodilation blood vessels become leaky, allowing more immune cells into site of infection systemic effect: act on hypothalamus to raise temperature - fever IL-1 primarily made by macrophages but can be made by sentinel cells (DCs and fibroblasts)

Question 59

What is Anakinra?

Answer 59

IL-1Ralpha IL-1 has destructive potential, can lead to joint pain and degradation so is closely regulated

Question 60

What is the function of IL-6?

Answer 60

predominantly made h macrophages main trigger of acute phase response in the liver acts w IL-23 to promote differentiation of Th17 increased levels in rheumatoid arthritis patients, treated w Tocilizumab antibody therapy which blocks IL-6 receptor

Question 61

What is the function of TNFalpha?

Answer 61

predominantly made by macrophages, but also Th1 cells tumour necrosis factor acute effects - septic shock response to bacterial endotoxins. if every cell produces TNFalpha, every blood vessel become leaky and blood leaks into tissues → clots → organ failure - anti-TNFalpha mAbs abolish spetic shock eg infliximab chronic effects - causes consumption of fat reserves - wasting in malignancy and parasite infection - involved in rheumatoid arthritis

Question 62

What is the function of IL-12 and IL-23?

Answer 62

produced by Th1 and Th17 respectively both cytokines share the p40 subunit, but drive alternative inflammatory T cell subsets

Question 63

What is the function of IL-17?

Answer 63

produced by Th17 important in defense against extracellular bacteria and fungi acts to recruit and expand neutrophils by secreting GM-CSF; phagocytes which clear tissues of extracellular pathogens

Question 64

What is the function of IL-4 and IL-13?

Answer 64

produced by Th2 used to remove parasites - induce hyper-proliferation of epithelial cells - airway mucus, fibrosis

Question 65

What is the function of interferons?

Answer 65

- block viral replication - alpha and beta (type 1 IFNs) are important anti-viral molecules - gamma (type 2) is v powerful and pleiotropic cytokine. produced by Th1 --on macrophages, enhance NO killing of intracellular pathogens, also raises IL-1 production -- on T cells, recruits more Th1 (inflammatory) -- On b cells, increases IgG_2a and IgG_3 class switch -

Question 66

What is the difference between type 1 and type 2 interferons?

Answer 66

only difference is that type 1 are stable in acidic conditions and type 2 are not

Question 67

What is the function of IL-10?

Answer 67

- produced by Tregs - indirectly inhibits Th1 cells - antagonises IFN-gamma - anti-inflammatory - inhibits proliferation and function of T cells - reduces macrophage activation eg IL-1 - prevents runaway inflammatory reactions blocking of IL-1- causes earlier death but parasiteamia is reduced this is due to higher necrosis rate of organs

Question 68

What is the function of TGF-beta?

Answer 68

- produced by Th0 - blocks cell proliferation - inhibits inflammatory cytokine production - induces more Tregs released in an inactive form, latency associated peptide (LAP) needs to be cleaved to activate it integrins expressed on surface of cells cleave LAP

Question 69

What is the function of IL-2?

Answer 69

major T cell growth factor, promotes T cell proliferation in any cells bearing an IL-2R can act in an autocrine manner needed by effector T cells and by Tregs knockout mice die due to lack of Tregs