Esa 3 - I & I Session 2

Front 1

How does the innate immune system detect pathogens?

Back 1

The cells of the innate immune system have germline encoded receptors ( ie they don’t change and adapt ) which recognise PAMPS ( pathogen associated molecular patterns).
PAMPS are highly conserved, essential for microbial viability, not present in higher eukaryotes. Examples of PAMPS are lipopolysaccharides, flagellin ( component of flagella).

Front 2

What is the basic structure of an antibody molecule?

Back 2

An antibody is comprised of 2 heavy chains in the middle and 2 light chains, one on either side of the H pair. The Fab portion of the Ig binds to antigens and the Fc (fragment crystalisation) is responsible for effector functions and binds to Fc receptors on phagocytes.
Each chain is bound to the adjacent chain via a di-sulphide bond. The bonds divide the chains into domains: constant and variable.
The constant domains are the same in all antibodies of the same isotope, class. The variable region, the FAB portion, changes between different clones and confers antibody specificity. Heavy chains have a single variable domain and 3 constant domains (CH1,CH2,CH3) or 4 as in Ig E and a hinge region which allows flexibility of the Fab arms.
The fab arm binds to epitopes on antigens. Epitopes are small molecules on antigens which are recognised by antibodies. An antigen may have many different epitopes and so different antibodies can bind to it.
There are 2 classes of light chains – lambda and kappa. An individual Ig molecule will only include one class of L chains. The L chain has one constant domain and one variable domain, which forms the FAB region

Front 3

Describe the effector functions of igM.

Back 3

IgM – is a large pentamer or monomer and is involved in agglutination of pathogens, Complement activation (classical pathway), opsonisation triggering phagocytosis, and is the principal antibody class in primary response of the adaptive immune system. It is found in the blood.

Front 4

Describe the functions of IgG.

Back 4

IgG – is a monomer that is involved in secondary adaptive immune response, it crosses through the placenta during gestation conferring immunity to the foetus which lasts for 3 months after birth , it performs opsonisation and complement activation (classical pathway), it is involved in neutrilisation, and antibody dependent cellular cytotoxicity ( ADCC –NK cells). IgG is found in the blood and tissues.

Front 5

Describe the functions of IgA.

Back 5

IgA – Is a monomer or dimer that is secreted at mucosal surfaces and neutralises microbes and toxins. It is attached to a secretory component at the mucosal surface. It is also found in blood and colostrums ( breast milk)

Front 6

Describe the functions of IgE.

Back 6

IgE – is a monomer which attaches to mast cells and eosinophils causing degranulation, release of granules containing heparin and histamine and ROS and enzymes to kill parasites. In anaphylactic shock, many IgE bind to mast cells, cross link and cause mass degranulation resulting in anaphylactic shock.

Front 7

Describe the functions of IgD.

Back 7

IgD – is a monomer which is the b cell antigen receptor in the b cell membrane. It is co-expressed with IgA on naive B lymphocytes.

Front 8

How are the ig monomers of the pentameric IgM linked? What happens when IgM binds to an antigen?

Back 8

Igm monomers are linked by a J chain. On binding to an antigen the IgM undergoes a conformational change that allows it to activate complement.

Front 9

How does IgA travel from the blood across the cell and onto the mucosal surface.

Back 9

As IgA passes from the blood stream into an epithelial cell, it receives a secretory component that helps to transport the antibody across the cell and stay attached at the mucosal surface.

Front 10

What do the letters of the different classes of immunoglobulins represent?

Back 10

IgA – alpha heavy chains, IgG – gamma heavy chains, IgM – mu heavy chain, IgD –delta heavy chain, IgE-epsilon heavy chain
These constant heavy chain regions are changed during class switching

Front 11

Describe somatic gene rearrangement.

Back 11

Somatic genetic rearrangement of the germline DNA coding for the heavy and light chains takes place within each b cell in primary lymphoid tissue. Chromosome 2 encodes the kappa chain, chromosome 14 encodes the heavy chains and chromosome 22 encodes the lambda chain.
In light chains the V region genes recombine with a J ( joining) gene which together combine with C (constant) segment. Transcription occurs to produce mRNA, followed by splicing to remove introns producing a mature Mrna. Translation then occurs to produce a light chain polypeptide.
In the heavy chain there is an additional variable segment called D (Diversity) which joins to V and J segments. Transcription and translation occurs to produce a heavy chain polypeptide. The heavy chains and light chains then join to form an immunoglobulin molecule.
There is also a gene segment determining whether the ig is membrane bound (surface receptor) or secreted.

Front 12

Describe the sequence of events of gene rearrangement in b cells in primary lymphoid tissue.

Back 12

The heavy chains coded for on chromosomes 14 are the first to be generated.
1- Joining and Diversity genes combine on both chromosomes
2- The J-D pair combines with a Variable gene on ONE of the chromosomes. If that is a success then the light chains will then be coded for. If it is not successful, the variable gene on the SECOND chromosome will be used. If that is not successful then apoptosis occurs
3- V-J combine to form kappa chain ( chromosome 2) which combine with heavy chain (V-J-D). If it fails then the heavy chain joins with V-J of SECOND kappa chromosome 2. If that fails go to first chromosome coding for lambda light chain ( chromosome 22). If that fails go to second chromosome coding for lambda light chain.

Front 13

Describe the processes which cause the great number of antibody specificities.

Back 13

1. Combinatorial diversity – there are many different V,D and J segments resulting in many different possible combinations which form the variable region. Takes place in primary lymphoid tissue (somatic gene rearrangement)
2. Junctional diversity – between V-J on light chains and D-J on heavy chains, nucleotides may be inserted, catalysed by an enzyme.
3. Somatic hypermutation – mutation of the already formed variable region that takes place in secondary lymphoid tissue. Antigenically activated b cells are more likely to undergo point mutations in the variable regions of heavy and light chains resulting in mutant surface immunoglobulins. The antibody specificity may therefore be altered to have a lower or higher affinity. If it has a higher affinity for the antigen then it is more likely to be stimulated to proliferate therefore increasing the number of b cells with high affinity surface immunoglobulins. This is known as affinity maturation of antibody.

Front 14

What is a pre b cell?

Back 14

A pre b cell is a b cell where only the heavy chain has undergone gene rearrangement and been transcribed. It becomes a mature b cell once the light chain has been transcribed and joined with the heavy chain.

Front 15

Describe the sequence of events involving antibodies, FDCs, iccosomes and tingible body macrophages.

Back 15

Secreted antibodies interact with antigens to form an immune complex. The complex binds to follicular dendritic cells, due to its receptors for Fc and C3b, in the germinal centres of secondary follicles of lymph nodes. The surface membrane along with the immune complex breaks off to form iccosomes. Iccosomes present the original antigen to b cells which have high affinity surface immunoglobulins for this antigen. These b cells survive to become memory cells or plasma cells. B cells with low affinity die by apoptosis and are phagocytosed by macrophages in the germinal centres. These macrophages are then called tingible body macrophages. B cells with high affinity are saved from apoptosis by t helper cells interacting with B cell presented peptide derived from the original antigen. The CD40 ligand interacts with CD40 on B cell triggering the formation of Bcl-xl protein which prevents apoptosis.

Front 16

Describe class switching.

Back 16

Class switching involves the heavy chain constant region of antibodies only. It occurs during affinity maturation stimulated by T cells. In the primary adaptive response, where the antigen is first exposed, IgM is produced in large numbers but in the secondary adaptive response, higher affinity antibodies are produced which have switched classes to IgG.

Front 17

Describe the b cell receptor complex on b cells.

Back 17

The b cell receptor complex consists of the membrane bound antibody ( IgD, ( along with IgA in naive b cells) and several protein chains – Ig alpha and Ig beta chains. All together they constitute a disulphide linked hetero-dimeric complex. They are involved in expression of membrane bound immunoglobulin and in signal transduction.

Front 18

Describe opsonisation.

Back 18

Opsonisation is coating of micro-organisms with plasma proteins to increase their adherence to phagocytic cells. The main opsonins are IgG, and activated C3 (c3b). IgG binds to the bacterial epitopes recognised by FAB portion. Activation of c3 exposes a thiol bond which forms ester or amide bonds with the bacterial surface structures. Phagocytic cells have fc receptors for the fc region of IgG and c3b receptors.

Front 19

What is complement?

Back 19

Complement is a serum constituent which complements the action of specific antibodies in killing pathogens. The complement system is a cascade of proteases which ends in the formation of MAC –membrane attack complex. The membrane attack complex damages the pathogen by pore formation and osmotic pressure. Complement components are labelled 1-9 and MAC uses C5-C9. C9 is the transmembrane pore which resembles the perforin found in granules of cytotoxic t cells which kills viruses. Thus bacteria and eukaryotic cells are killed by transmembrane pore c9 and viruses are killed by transmembrane pore of lytic granules of cytotoxic t cells.
Complements kill mico-organisms directly by formation of c5b-9 MAC and indirectly by opsonisation followed by phagocytosis and intracellular killing.

Front 20

Describe the 3 different pathways of complement activation.

Back 20

The object of all pathways is to create c3b – activated c3 which performs opsonisation.
The classical pathway is activated by binding to antibodies
- when c1q, the first subcomponent of c1 complex, binds to 2 adjacent IgG molecules or a single IgM molecule. This activates C1s subcomponent to become an active protease which cleaves C4 and C2 to form the classical pathway c3 convertase. The classical pathway c3 convertase is C4b2a ( a complex formed from activated c4 (c4b) and activated c2 (c2a) which activates c3b. c3a - chemotaxis.
The alternative pathway is activated by directly binding to pathogen
- The initial turnover is driven by spontaneous hydrolysis of thioester bond c3 to c3b (h2O). C3b (H2O) can bind to factor B and D and is stabilised by protein properdin to form a low efficiency c3 convertase ( c3b(h20)Bb). When c3b binds to micro-organisms via exposed thiol group allowing an ester or amide bond to form, activation of alternative pathway is amplified so that a large amount of c3b(h20)Bb is formed.
The lectin pathway is activated by manose binding lectins binding to manose in the absence of antibodies
- it involves an analogue of C1q, MBL( manose binding lectin) which binds directly to the mannose residues on micro-organisms in the absence of an antibody. This binding activates MASP-2 (MBL associated serine protease) which is an analoque of C1s and so cleaves c4 and c2 like in the classical pathway to form activated c4 and c2.
The alternative and lectin pathway are involved in the innate immune system and the classical pathway is involved in the adaptive immune system
Once c3b has been formed some binds to c3 convertase to form c5 convertase which activates c5 to form c5b. C6,C7,C8 and C9 bind together along with c5b to form the MAC which is inserted into cell membranes to produce lysis.

Front 21

What in the alternative complement cascade is relatively resistant to factors H and L?

Back 21

C3b like molecule (H20) combine with factor B, D and properdin to form a low efficiency C3 convertase which is resistance to factors H and L.

Front 22

What is attached to antibody molecules at mucosal surfaces?

Back 22

A secretory component helps transport IgA across the epithelium and onto the mucosal surface. IgA prevents the attachment of pathogens to mucosa.

Front 23

Describe TCR gene rearrangement. How is it different to B cell receptors?

Back 23

Expression of the TcR-t cell antigen receptor confers antigen specificity to the t cell. The great diversity of TcR is achieved by TcR gene rearrangement of V,D,J and C gene segments, catalysed by RAG enzyme ( recombination activating gene). The combinatorial diversity increases the number of antigen specificities in the t cell repertoire. Individual antigens are recognised by clones of T cells which recognise specific epitopes within the antigen. Unlike immunoglobulins, TcRs don’t undergo class switching or somatic hypermutation.
There are 2 forms of TCR
- 90% are alpha/beta TCR
- 10% are gamma/delta TCR – main roles are cytokine secretion and cytotoxicity
a/b tcr positive cells express either CD4+ or CD8+ surface markers. The cd4 t cells are helper cells involved in cytokine secretion and co-ordination of the immune response. They help b cells make antibodies and class switching. The cd8 t cells represent cytotoxic cells which kill cells invaded by intracellular parasites. All t cells also express CD3.

Front 24

DESCRIBE THE MECHANISMS BY WHICH T LYMPHOCYTES RECOGNISE ANTIGENS.

Back 24

For t cells to recognise antigens they must be processed and presented by an antigen presenting cell eg dendritic cell. T lymphocytes recognise antigen in the form of short peptides associated with self major histocompatibility complex, MHC class 1 or class 2, at the surface of the APC.
MHC 1 – present antigens to CD8 cells
- HLA-A,B and C
MHC 2 – present antigen to CD4 cells
- HLA-DR,HLA-DP and HLA-DQ
There is great polymorphism in HLA meaning that no-one has an identical HLA type. Different HLA molecules bind to different peptides. HLA molecules are the main antigenic targets in organ graft rejection.

Front 25

Which MHC is associated with increased risk of anklosing spondylitis?

Back 25

HLA- B27

Front 26

Describe the class 1 MHC pathway of processing of protein antigens

Back 26

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The class 1 MHC pathway is the endogenous pathway involving antigens found in cells (endogenous).
Viral proteins enter a host cell or APC and are cleaved by the proteasome( antigen processing) to produce peptides. These peptides are transported into the ER via TAP (transport associated with antigen presentation). The peptides are assembled into the MHC and an exocytic vesicle transports the MHC-peptide to the surface of the cell to be presented. Specific T killer cells bind to the MHC complex and trigger destruction of the invaded cells by perforin released from granules that forms pores, through which proteases are released and apoptosis is triggered. MHC class I proteins are consitutively expressed by most somatic cells.
TAP deficiency can cause chronic respiratory infections.



What types of infections are asplenic patients particularly susceptible to?
Asplenic patients following a spleenectomy are particularly susceptible to encapsulated bacterial infections eg streptococcus pneumoniae. This is because the polysaccharide capsule that encircles the virus is fairly difficult to phagocytose. The spleen is the largest organ of secondary lymphoid tissue and so by removing the spleen much fewer b cells are activated, and so few opsonins are produced and phagocytosis is not as effective.

Front 27

Describe the class 2 MHC pathway of processing of protein antigens.

Back 27

The class 2 MHC pathway is the exogenous pathway.
The antigen is taken up into the APCs by endocytosis or phagocytosis. The lysosomes fuse to the phagosome and release its contents which breakdown the antigen into small peptides. The class 2 MHC is synthesised and released from the ER via a vesicle which binds to the peptide vesicle to form a MHC-peptide complex which is then presented on the surface of the cell. Specific T helper cells ( cd4) bind to the MHC-peptide and are activated to enhance the adaptive immune system by secreting cytokines which stimulate the further proliferation of specific clones of high affinity b cells. The peptides derive from exogenous antigens such as soluble proteins or extracellular organisms eg bacteria. MHC class II proteins are constitutively secreted by proffesional APCs – macrophages, dendritic cells, b cells.

Front 28

Describe the role of costimulatory proteins?

Back 28

Costimulatory proteins are required for the production and release of IL-2 which triggers T cell proliferation and differentiation. B7 protein on APC binds to CD28 on t cells. If there is no costimulation then the T cell on binding to the MHC-peptide complex is not activated and is unresponsive – anergy. The cell will die. On activated T cells CTLA-4 is expressed which has a higher affinity for B7 on APC than CD28. This causes the inhibition of costimulation and so the t cell is inhibited.

Front 29

Describe how exotoxins can cause septic shock.

Back 29

Exotoxins are released from certain bacteria eg staphlococcal aureus food poisoning, streptococcal pyogenes (scarlet fever), clostridium perfringens (gas necrosis) and act as superantigens. They bind to the MHC class II and cd4 T cell receptor and act as the peptide, binding to the beta chain. This causes widespread activation, stimulating many cytokines and causing toxic shock syndrome, hypotension, fever and multi-organ failure.

Front 30

Describe the difference between immature and mature dendritic cells.

Back 30

Immature dendritic cells are found in the periphery where they take up antigen and migrate to regional lymph nodes. They have absent or low costimulation molecules, adhesion molecules, cytokine receptors and reduced levels of MHC class II. When they reach the lymph node, they process the antigens and mature.
Mature dendritic cells have high levels of costimulation molecules, adhesion molecules and cytokine receptors and very high levels of MHC class II and thus can activate naive antigen specific t cells.
Hence this is a method of self tolerance as antigen presentation outside of lymphoid tissue usually lacks costimulation so the t cells are unresponsive ( ANERGISED).
Mature dendritic cells also influence to balance of Th1 (cell mediated response) or Th2 (humoral response).

Front 31

What types of infections are asplenic patients particularly susceptible to?

Back 31

Asplenic patients following a spleenectomy are particularly susceptible to encapsulated bacterial infections eg streptococcus pneumoniae. This is because the polysaccharide capsule that encircles the virus is fairly difficult to phagocytose. The spleen is the largest organ of secondary lymphoid tissue and so by removing the spleen much fewer b cells are activated, and so few opsonins are produced and phagocytosis is not as effective.

Front 32

Describe the 4 phases of the adaptive immune response.

Back 32

1. Induction phase: APC migrates to lymphoid tissue with antigen, matures and presents it to naive antigen specific t cells, creating effector T cells. effector T cells can then recognise the MHC-antigen complex presented by ANY cell and not just APC.
2. Activation phase: once activated the effector cells undergo clonal expansion and differentiation to become fully activated effector cells ( helper t cells or cytotoxic t cells) or remain in partially activated state where they re-circulate as memory cells. Memory cells don’t require costimulation like naive t cells and thus respond quickly to subsequent exposure to antigen.
3. Effector phase: effector cells migrate to site of infection and perform their actions whether this is cytolytic killing of viral infected cells, induction of delayed type hypersensitivity reactsions and macrophage activation or promotion of antibody production by T helper cells. t helper cells co-ordinate the antigen specific immune response by providing help through cytokine release for cells in class switiching and affinity maturation of antibody responses, for CD8 t cells in the induction of effector CTL and for enhancement of NK cell function.
4. Resolution: non specific phagocytic cells remove debris and macrophages and fibroblasts resolve any damage caused by infection and promote healing.

Front 33

What are regulatory T cells?

Back 33

Regulatory T cells are a subset of t cells which inhibit immune responses once an appropriate immune attack has been made. They are very important in preventing autoimmune diseases.