L13- Mucosal Immunity

Question 1

The mucosal immunity system comprises innate and adaptive immunity and comprises of multiple sensing and defence mechanisms. what are some of these cellular entities?

Answer 1

• Fluid Phase components-
Antibody, peptides, enzymes

• Stationary components-Epithelium, connective tissue cells, mast cells, fibroblasts
• Mobile Components- Neutrophils, monocytes, T-cells, B-cells, NK cells, Dendritic cells

Possesses sensing functions due to receptors on cell surface or in cytoplasm

Question 2

Describe the layers of the mucosal surface.

Answer 2

The mucosal surface comprises of;

The epithelial layer – cells lining the tissue and at the interface with the lumen;

• The lamina propria – a collection of loose connective tissue with blood vessles, lymph vessels, nerves
• The muscularis mucosa smooth muscle and connective tissue below lamina propria

Question 3

Purpose of cell turnover

Answer 3

Renewal and elimination of dead cells

Question 4

Purpose of cell shedding

Answer 4

Elimination of infected cells

Question 5

Purpose of cell death

Answer 5

Elimination of infected cells

Question 6

Define autophagy

Answer 6

Destruction of cytoplasmic pathogens

Question 7

What do goblet cells do?

Answer 7

Produce secreted gel-forming mucin glycoproteins, trefoil peptides and RELM- beta

Question 8

What do paneth cells do?

Answer 8

Produce antimicrobial peptides, lectins and cytokines

Question 9

What is the role of the mucosal leukocytes and stroma?

Answer 9

• Regulates secretory cell differentiation
• Modulate epithelial secretory function
• Produce secretory IgA

Question 10

Describe the environment for the outer mucus layer

Answer 10

• Non-sterile, viscous due for mucin glycoproteins
• Degrading mucus
• Microbes utilise mucin carbohydrates for energy (including mucosal microbiota)

Question 11

Describe inner mucus layer

Answer 11

• Relatively sterile

- Rich in antimicrobial peptides and antibodies to protect epithelial layer

Question 12

The major tight junction proteins include…

Answer 12

Occludin 
Zona occludens 
Claudin 
Cateninin 
Cingulin 
E-Cadherin

Question 13

Describe how bacteria can get past the tight junction and the role of DC’s

Answer 13

Some invasive bacteria can enter epithelial cells directly

Whereas others utilize M cells, which are situated over lymphocytic aggregates, to breach the barrier.

A newly discovered route across the epithelium uses uptake by the projections that dendritic cells extend into the intestinal lumen
This pathway can be used by noninvasive bacteria and may allow uptake of soluble antigens and presentation of microbes to intraepithelial lymphocytes or those in aggregates below M cells.

Under resting conditions, infiltrating DCs establish loose contacts with preexisting epithelial TJs. Upon bacterial infection, DCs are recruited from the blood and activated, probably via epithelial cell signals. They up-regulate the expression of occludin, which in turm allows DCs to compete for epithelial occludin and open up the TJs, like a zip.

Infiltrating DCs then face the gut lumen and can directly sample bacteria Bacterial components such as LPS trigger the reorganization of TJ proteins via up-regulation of ZO-1 and the disappearance of occludin, thus allowing the DCs to detach from junctions with epithelial cells and to migrate into the draining lymph nodes

Question 14

What molecules are involved with resolution of inflammation?

Answer 14

Acute phase proteins
Antioxidants 
Clara cell protein 
Proteases
Protease inhibitors 
ECM production

Question 15

What molecules are involved in inflammation

Answer 15

Adhesion molecules
MHC class I and II
Cytokines
Growth factors
Endothelin 
Prostaglandins 
Leukotrienes
Nitric oxide species

Question 16

Some viruses can enter the epithelium by which receptor?

Answer 16

ICAM-1 also known as CD54

Question 17

Some bacteria enter epithelium via…

Answer 17

TLR and NOD2, M cell entry, DC route…

Question 18

Provide an example of a receptor where an allergen can enter epithelium.

Answer 18

PAR receptor

Question 19

What are some PAMP examples that can be picked up by epithelium sensing.

Answer 19

Gram positive-
Peptidoglycan (TLR2)
Polysaccharides 
Lipoteichoic acid (TLR2)

Gram negative-
LPS (lipopolysaccharides)
(TLR4)

Both +ve and -ve bacteria- 
Fimbriae
Lipopeptides
Lipoproteins 
Muramyl dipeptide (mycobacterium)
RNA (TLR3)
CpG (TLR9)
Flagellin (TRL5)

Viruses-
RNA

Fungi-
Mannans
Glycans

Question 20

Some extracellular components that are picked up by epithelium sensing include..

Answer 20

Toxins
Superantigens
Proteases
Other enzymes

Question 21

What do PRR do once they recognise PAMPS?

Answer 21

Involved in phagocytosis and cell activation

Pro-inflammatory mediators and cytokine release

Induce inflammation and activation of cells

Activates adaptive immune responses

Question 22

What innate functioning molecules of epithelium have a low molecule weight?

Answer 22

Prostanoids
Leukotrienes
Nitric oxide

Question 23

What innate functioning molecules of epithelium have a high molecule weight?

Answer 23

Cytokines
Chemokines
Antimicrobial peptides
Enzyme inhibitors
Enzymes

Question 24

What are lipid mediators and is their function?

Answer 24

Lipid mediators derive their name from the fact that ther are derived from cell membrane phospholipid which is converted to arachidonic acid by phospholipase. AA is the converted to Either prostanoids or leukotrienes by COX or lipoxygenase.

These mediators play an important role in both homeostasis and inflammation because they can induce vasodilation, permeability and cell recruitment.

Question 25

What are cytokines?

Answer 25

Cytokines play a major role in acute and chronic inflammation due to their capacity to recruit inflammatory cells and cytokines such as IL-1, IL-5, IL-6, IL-11, TNF- and GM-CSF, chemokines such as IL-8, eotaxin, MIP-1- and RANTES, and LIF are very important. These cytokines stimulate the proliferation of cells such as T cells Activate and chemoattract inflammatory cells such as neutrophils and eosinophils Induce the expression of adhesion molecules Alter tissue permeability Induce the synthesis of cytokines and mediators such as prostaglandins and nitric oxide.

Question 26

What cells produce cytokines?

Answer 26

They are produced by a variety of cells including T cells, macrophages, mast cells and monocytes as well as epithelium. In this regard, epithelium is a significant source of cytokines

Question 27

There are two major classes of antimicrobial peptides (AMP). What are they?

Answer 27

Cathelicidin LL-37 and a -and b-defensins

Question 28

How are antimicrobial peptides able to kill pathogens?

Answer 28

The peptides are cationic and have affinity to the negatively charged bacterial membrane and induce bacterial killing by lysis.

Question 29

The skin is 1.8sqm. What is the mucosa?

Answer 29

400sqm

Question 30

What are some special stats to remember about the mucosa that involve adaptive immunity?

Answer 30

* Comprises 85% of all lymphoid tissue * Involves 67% of all antibody production * Involves fluid, stationary & mobile phases * Involves specialised lymphoid tissue

Question 31

IgA in mucosal surfaces properties?

Answer 31

Found in secretions, transferred to epithelium via polyIgR Both serum and secretory forms may comprise IgA subclasses. IgA1 predominates in serum whereas both predominate in secretions. IgA2 lacks a 13 AA residue section in its hinge region which helps confer resistance to bacterial IgA-specific proteases.

Question 32

IgG in mucosal surfaces properties?

Answer 32

Found in secretions, predominates in male and female genital tract, cervical, vaginal fluid, semen, present also in nasal secretions, transferred to epithelium via neonatal FcR

Question 33

IgM in mucosal surfaces properties?

Answer 33

Can be secreted if IgA deficient

Question 34

How does IgA confer resistance against IgA-specific proteases?

Answer 34

IgA2 lacks a 13 AA residue section in its hinge region which helps confer resistance to bacterial IgA-specific proteases.

Question 35

IgA exists as a secretory form (monomer) and as a serum form (dimer)? T or F?

Answer 35

False, it’s the opposite. IgA exists as a serum form (monomer) and as a secretory form (dimer)

Question 36

What is the joining of the J chain like for secretory IgA?

Answer 36

The secretory form has two IgA molecules joined together by a J chain - a small protein (15kDa) that forms disulphide bonds with the heavy chains of the 2 molecules. It is also found with polymeric IgM and is also important for binding of secretory IgA to the poly-Ig receptor for transcytosis to the lumen. The polyIg receptor gets cleaved to leave behind a remnant called secretory piece (SC).

Question 37

Where is serum IgA produced?

Answer 37

Serum IgA produced by bone marrow

Question 38

Where is secretory IgA produced?

Answer 38

Secretory IgA Is produced by mucosal tissue

Question 39

What are some special features of secretory component of IgA to remember?

Answer 39

- Synthesised by epithelial cells - Can be found free or attached to Ig - 80kDa protein derived from PolyIg receptor - Free form plays role in innate immunity - Helps to anchor sIgA to mucus

Question 40

Describe features of the J chain.

Answer 40

- 15.6 kDA protein - Associated with polymeric Ig - Synthesised by plasma cells - One J chain per molecule - Initiates polymerisation of Ig and optimises binding to Sc

Question 41

Does IgA1 have a hinge region?

Answer 41

Yes

Question 42

Does IgA2 have a hinge region?

Answer 42

Lacks 13aa, renders it resistant to protease digestion

Question 43

What is the concentration of serum IgA1 in solution?

Answer 43

90%

Question 44

What is the concentration of serum IgA2 in solution?

Answer 44

Only 10%

Question 45

What is the concentration of secretory of IgA1 in solution?

Answer 45

50% just like IgA2.

Question 46

Describe the mucosal Ab function.

Answer 46

Mucosal B cells terminally differentiate to become mucosal plasma cells, most of which produce dimeric IgA that is exported into secretions as secretory IgA (sIgA) to intercept antigens and pathogens, and to prevent mucosal invasion. Neutralizing IgG is also present within mucosal tissues; mucosal IgG might be derived from local plasma cells or from blood, by diffusion from local fenestrated capillaries.

Question 47

__________ bind to (pathogen-derived) antigens and prevent or inhibit their attachment to and/or invasion of epithelial cells.

Answer 47

``` Secretory immunoglobulins (SIgs) bind to (pathogen-derived) antigens and prevent or inhibit their attachment to and/or invasion of epithelial cells. ```

Question 48

The ________ complex mediates intracellular neutralization of pathogens that have invaded the epithelial cells.

Answer 48

The IgA–'joining' (J) chain–polymeric-immunoglobulin receptor (pIgR) complex mediates intracellular neutralization of pathogens that have invaded the epithelial cells.

Question 49

In addition, antigen excretion through the secretion of immunoglobulins maintains ________ at mucosal surfaces. Secretory antibodies mediate removal of antigens from the mucosal lamina propria, in which antigens are bound to _____ IgA and are subsequently transported to the luminal surface of the epithelial cells following endocytosis of the __________ and its release into the _______.

Answer 49

In addition, antigen excretion through the secretion of immunoglobulins maintains (homeostasis) at mucosal surfaces. Secretory antibodies mediate removal of antigens from the mucosal lamina propria, in which antigens are bound to (dimeric) IgA and are subsequently transported to the luminal surface of the epithelial cells following endocytosis of the (antigen–dimeric IgA–J chain–pIgR complex) and its release into the (mucosal lumen).

Question 50

What bacteria use proteases to degrade IgA1?

Answer 50

Particularly ones that colonise mucous membranes Neisseria meningitidis Streptococcus pneumoniae Haemophilus influenzae (which cause bacterial meningitis) Neisseria gonorrheae.

Question 51

How are proteases able to break down IgA1?

Answer 51

Enzymes are secreted and cleave IgA after proline residues (After PT or PS) in the hinge region of IgA1 to produce Fab and Fc fragments. Whilst the Fab fragments would still bind to bacteria, they would be less efficient at removing bacteria - non-agglutinating and less inhibitory.

Question 52

How is IgA2 able to resist protease cleavage?

Answer 52

It is missing 13aa that miss the proline residues (PT and PS) and therefore cannot be cut.

Question 53

What is pIgR?

Answer 53

pIgR is a 5 domain cell surface protein involved in transport (transcytosis) of secretory IgA from basolateral side to apical side of epithelium.

Question 54

What is nFcR?

Answer 54

nFcR is a MHC related protein note the three domains which are similar to MHC Class 1 and contain b 2 microglobulin. This is involved in transcytosis of IgG across epithelium, catabolism and placental transfer

Question 55

How does secreted Ig exocytose across the epithelium?

Answer 55

The polymeric-immunoglobulin receptor (pIgR) expressed on the basolateral side of columnar epithelial cells and specifically binds the 'joining' (J) chain as well as regions of the immunoglobulin. Following internalization of the pIgR–pIg–J chain complex, the vesicles containing the complex are directed to the apical membrane for exocytosis. Following exocytosis at the apical membrane, the extracellular portion of the pIgR is proteolytically cleaved by an endopeptidase and released. The 80 kDa extracellular portion of the pIgR, known as the secretory component, remains associated with the pIgA and pIgM (not shown), forming secretory IgA and IgM.

Question 56

Where can the secretory component of Ig also be found?

Answer 56

The secretory component is also found in a free (unoccupied) form in the mucosal lumen as a result of transcytosis and cleavage of unbound pIgR – function unknown.

Question 57

The secretory part of the Ig is a seperate component. What is it’s function?

Answer 57

The secretory component may protect secretory immunoglobulin from stomach acid and intestinal proteases and may also target secretory immunoglobulin to mucus by selectively binding to mucins.

Question 58

How is IgG transferred across to the fetus and what else is this receptor used for?

Answer 58

IgG transferred across epithelial tissue by the neonatal Fc receptor (FcnR) which is related to MHC Class I molecules. FcnR plays a major role in transferring IgG from mother to the fetus but is also involved in transferring IgG from the circulation to the gut and lung. IgG is pinocytosed and then binds FcnR at acidic pH in endosomes. It is then released into the lumen when it comes into contact with neutral pH. This system may also be exploited to transfer lung delivered material into the blood stream. This process is also involved in regulation of IgG catabolism.

Question 59

List some mucosa-associated lymphoid tissue.

Answer 59

Mucosa-associated lymphoid tissue (MALT): Lymphoid structures and aggregates associated with mucosa. Subdivided based on anatomy: N(asal)ALT, G(ut)ALT, S(kin)ALT and B(ronchus)ALT. Comprises the tonsils, nose, Peyer's patches, isolated lymphoid follicles, appendix or caecal patch and mesenteric lymph nodes. They are enriched in conventional and unconventional lymphocytes and specialized dendritic cell and macrophage subsets. They provide the first line of defence against entry of pathogens across the mucosal barrier.

Question 60

What is Peyers patch?

Answer 60

Groups of lymphoid nodules (5-200) that are present in the small intestine (usually the ileum). They occur massed together on the intestinal wall, opposite the line of attachment of the mesentery. Peyer's patches consist of a dome area, B cell follicles and interfollicular T cell areas. High endothelial venules are present mainly in the interfollicular areas. They posses no afferent lymphatics, thus they collect antigen from the lumen.

Question 61

What specialised cells are found in follicle-associated epithelium (FAE)?

Answer 61

Microfold (M) cells. The majority of FAE cells are enterocytes with apical microvilli coated by a thick brush border glycocalyx. FAE enterocytes do not transport antigens, but they might contribute to antigen sampling by sensing luminal pathogens and their products and releasing cytokine and chemokine signals that attract and activate DCs. CCL, CC-chemokine ligand; CCR, CC-chemokine receptor; TLR, Toll-like receptor.

Question 62

What are M cells?

Answer 62

M cells are called microfold cells. They are found in follicle-associated epithelium specialized for endocytosis and rapid transepithelial transport of intact antigens and microorganisms into intraepithelial pockets that contain B and T cells and occasional dendritic cells (DCs).

Question 63

Do M cells secrete mucus of enzymes?

Answer 63

No. The do not have a thick glycocalix and play a role in allowing antigen to gain entry into Peyers patches to activating immune responses. M cells sample and uptake antigens at their apical membrane, encase them in vesicles to transport them to the basolateral membrane of M cells, and from there deliver antigens to the nearby lymphocytes.

Question 64

Which intestinal pathogens exploit M cells as their portal of entry to invade the host and cause infections?

Answer 64

Yersinia, Shigella, Vibrio cholerae, the pathogenic strain of Escherichia (O157:H7), Salmonella, Campylobacter spp, poliovirus, Human immunodeficiency virus 1 (HIV-1) and prion disease all exploit M cells for ease of entry.

Question 65

How does Salmonella enter the gut?

Answer 65

Via M cells or enter dendrites of dendritic cells that are sampling the gut luminal contents.