L2- Innate Immunity

Question 1

Innate immunity serves three essential functions that protect us against microbes and tissue injury, what are they?

Answer 1

• Innate immunity is the initial response to microbes that prevents, controls, or eliminates infection of the host by many pathogens
• Innate immune mechanisms eliminate damaged cells and initiate the process of tissue repair
• Innate immunity stimulates adaptive immune responses and can influence the nature of the adaptive responses to make them optimally effective against different types of microbes

Question 2

What are the two major types of responses of the innate immune system that protect against microbes?

Answer 2

Inflammation and antiviral defense

Question 3

Describe innate immunity (immediate 0-4 hours)

Answer 3

Infection

Recognition by performed, nonspecific and broadly specific effectors

Removal of infectious agent

Question 4

Describe early induced innate response (early 4-96 hours)

Answer 4

Infection

Recognition of microbial-associated molecular patterns

Inflammation recruitment and activation of effector cells

Removal of infectious agent

Question 5

Describe phases of adaptive immune response ( later >96 hours)

Answer 5

Infection

Transport of antigen to lymphoid organs

Recognition by naive B and T lymphocytes

Clonal expansion and differentiation to effector cells

Removal of infectious agent

Question 6

Is there a change in the response or magnitude of the innate immune response to a microbe upon repeat exposure?

Answer 6

No there isn’t. In contrast, repeated exposure to a microbe enhances the rapidity, magnitude, and effectiveness of adaptive immune responses

Question 7

How many molecular structures can the innate immunity recognise as opposed to the adaptive immune system?

Answer 7

(~1000) that are either products of microbes or are expressed by injured or dead host cells.

By contrast, the adaptive immune system potentially can recognise millions of different molecular structures of microbes, and can also recognise non-microbial environmental antigens as well as self antigens that are normally present in healthy tissues

Question 8

List some mechanical barriers that prevent pathogens from crossing epithelia and colonizing tissues

Answer 8

Epithelial cells joined by tight junctions

Longitudinal flow of air or fluid

Movement of mucus by cilia

Tears

Nasal cilia

Question 9

List some chemical barriers that prevent pathogens from crossing epithelia and colonizing tissues

Answer 9

Fatty acids

Low pH

Enzymes (pepsin)

Enzymes in tears (lysozyme)

Question 10

List an example of a microbiological barrier that prevent pathogens from crossing epithelia and colonizing tissues

Answer 10

Normal flora

Question 11

What are 3 ways the epithelial barrier protects the body from microbes

Answer 11

Physical intact barrier

Killing of microbes by locally produced antibiotics (defensins and calthelicidins)

Killing of microbes and infected cells by intraepithelial lymphocytes

Question 12

What are defensins?

Answer 12

Defensins are small (3–4 kDa) positively-charged antimicrobial peptides, which are able to disrupt bacteria, fungi, parasites, and some enveloped viruses by forming multimeric pores in the cell membranes of these pathogens.

Electrostatic attraction and the transmembrane electric field bring the defensins into the lipid bilayer.

They also activate cells involved in the inflammatory response to microbes.

The defensins belong to one of two families, the α- and β-defensins.

Question 13

What cells produce defensins?

Answer 13

Defensins are produced by cells including epithelial cells of mucosal surfaces and by granule-containing leukocytes, including neutrophils, natural killer cells, and cytotoxic T lymphocytes.

Question 14

What is the role of intraepithelial T lymphocytes?

Answer 14

recognize and respond to commonly encountered microbes

Question 15

Where are intraepithelial T lymphocytes found?

Answer 15

IELs are distributed in the epithelium of the intestine and found both along the basement membrane and
between epithelial cells in the lateral intercellular space

Question 16

What is the common characteristic of intraepithelial T cells compared with T cells

Answer 16

Limited diversity of their antigen receptors.

Question 17

What antigen receptors do intraepithelial T cells have?

Answer 17

Some intraepithelial T lymphocytes express the conventional αβ form of TCR, which is present on most Tcells in lymphoid tissues and the circulation.

Other T cells in epithelia express a form of antigen receptor called the γδ TCR that may recognize peptide and nonpeptide antigens

Question 18

The pattern recognition of the immune system are nonclonally distributed. What is meant by this?

Answer 18

Identical receptors are expressed on all the cells of a particular type, such as macrophages. Therefore, many cells of innate immunity may recognize and respond to the same microbe

Question 19

How do B and T cells each express a unique receptor?

Answer 19

Rearrangement of gene segments during lymphocyte development

Question 20

How many innate immune receptors are capable of recognizing microbes?

Answer 20

It is estimated that there are about 100 types of innate immune receptors that are capable of recognizing about 1000 PAMPs and DAMPs

Question 21

What receptors are responsible for recognising microbes in adaptive immunity

Answer 21

Immunoglobulin [Ig] and T cell receptors [TCRs])

Because of their diversity they are able to recognize millions of different antigens

Question 22

List 4 examples of pattern recognition receptors of innate immunity

Answer 22

Toll-like receptor

N-Formyl methionyl receptor

Mannose receptor

Scavenger receptor

Question 23

Signalling from cell surface involves cytosolic and nuclear phases. Explain the process.

Answer 23

Ligation of a receptor activates cell and induces signalling, which typically involves an initial cytosolic phase when the cytoplasmic portion of the receptor or of proteins that interact with the receptor may be enzymatically modified.

This often leads to the activation or nuclear translocation of transcription factors that are silent in resting cells, followed by a nuclear phase when the transcription factors orchestrate changes in gene expression.

Question 24

The innate immune system recognizes microbial elements that are often essential for survival of the microbes. This ensures that the targets of innate immunity cannot be discarded by microbes in an effort to evade recognition by the host. Provide 2 examples.

Answer 24

Double-stranded viral RNA, which is an essential intermediate in the life cycle of many viruses

LPS and lipoteichoic acid are structural components of bacterial cell walls that are recognized by innate immune receptors; both are required for bacterial survival.

Question 25

What are PAMPS?

Answer 25

Pathogen associated molecular patterns

Question 26

What are PRR's and where can they be found?

Answer 26

pattern recognition receptors. They can be found extracellular, cytosolic and endosomal.

Question 27

What are Toll-like receptors?

Answer 27

Toll-like receptors are a class of proteins that play a key role in the innate immune system. They are single-pass membrane-spanning receptors usually expressed on sentinel cells such as macrophages and dendritic cells, that recognize structurally conserved molecules derived from microbes

Question 28

How many Toll-like receptors are there?

Answer 28

There are nine different functional TLRs in humans, named TLR1 through TLR9

Question 29

Examples of cell wall bacterial products that bind to TLRs

Answer 29

LPS and lipoteichoic acid, which are constituents of the cell walls of gram-negative bacteria and gram-positive bacteria, respectively TLR4 binds to LPS TLR2 binds to Lipoteichoic acid

Question 30

Example of nucleic acid TLR ligands

Answer 30

Double-stranded RNAs, which make up the genomes of some viruses and are generated during the life cycle of most RNA viruses but are not produced by eukaryotic cells; Single-stranded RNAs, which are distinguished from cellular cytoplasmic single-stranded RNA transcripts by their location within endosomes and by their high guanosine and uridine content; and unmethylated CpG dinucleotides, which are common in prokaryotes but rare in vertebrate genomes

Question 31

How does the lysozyme destroy Gram-positive bacterial cell wall?

Answer 31

Lysozyme catalyzes hydrolysis of 1,4-beta linkages between N-acetylmuramic acid and N-acetyl-D-glucosamine residues in peptidoglycan, the major component of Gram-positive bacteria cell walls

Question 32

What happens once there is TLR recognition of microbial ligands?

Answer 32

Results in gene expression. Activation of several signaling pathways and ultimately transcription factors, which induce the expression of genes whose products are important for inflammatory and antiviral responses

Question 33

Signalling pathways are initiated by ligand binding to the TLR at the cell surface or in the endoplasmic reticulum or endosomes leading to.....which causes......

Answer 33

Leading to dimerization of the TLR proteins. Ligand-induced TLR dimerization is predicted to bring the TIR domains of the cytoplasmic tails of each protein close to one another. This is followed by recruitment of TIR domain–containing adaptor proteins, which facilitate the recruitment and activation of various protein kinases, leading to the activation of different transcription factors.

Question 34

The major transcription factors that are activated by TLR signaling pathways are...

Answer 34

nuclear factor κB (NF-κB), activation protein 1 (AP-1), | interferon response factor 3 (IRF3), and IRF7

Question 35

What are CDS's?

Answer 35

Cytosolic DNA sensors (CDSs) are molecules that detect microbial double-stranded (ds) DNA in the cytosol and activate signalling pathways that initiate antimicrobial responses, including type 1 interferon production

Question 36

What is STING?

Answer 36

STING is an endoplasmic reticulum–localized transmembrane adaptor protein that binds to cGAMP and activates the TBK1 kinase, (stimulator of IFN genes) pathway is an important mechanism of dsDNA-induced activation of type 1 interferon responses

Question 37

When a cytosolic DNA receptor sensors foreign dsDNA it activates a series of events leading to type 1 IFN gene expression. Explain.

Answer 37

Cytosolic dsDNA, usually of microbial origin, activates the enzyme cGAS (cyclic GMP-AMP synthase), which generates a signalling molecule called cGAMP (cyclic GMP-AMP). STING is an endoplasmic reticulum–localized transmembrane adaptor protein that binds to cGAMP Once STING adaptor protein has bound to cGAMP it activates the TBK1 kinase, which phosphorylates and activates the IRF3 transcription factor, leading to type 1 IFN gene expression.

Question 38

Explain the importance of C-type Lectin receptors for microbial carbohydrate detection.

Answer 38

Recognition of carbohydrates on the surface of microbes facilitate the phagocytosis of the microbes and the secretion of cytokines that promote inflammation and subsequent adaptive immune responses (C-type lectin = protein that binds carbohydrates (hence lectins) in a Ca++-dependent manner (hence C-type)

Question 39

Where can you find C-type lectin receptors?

Answer 39

Receptors are integral membrane proteins found on the surfaces of macrophages, DCs, and some tissue cells. Other lectins are soluble proteins in the blood and extracellular fluids

Question 40

There are several types of plasma membrane C-type | lectins with specificities for different carbohydrates. List 4 of these carbohydrates.

Answer 40

Mannose, glucose, N-acetylglucosamine, and β-glucans

Question 41

What are NOD-like receptors?

Answer 41

NOD-like receptors (NLRs) are a family of more than 20 different intracellular pattern recognition receptors that activate downstream signaling cascades leading to activation of components of the inflammasome.

Question 42

What is the inflammasomes made of?

Answer 42

1. NLRP3 (sensor) 2. Adaptor (ASC) 3. Capase-1 (inactive)

Question 43

Briefly explain what happens when NLRP (subfamily of NOD-like receptors) are activated?

Answer 43

The NLRP subfamily of NOD-like receptors respond to cytosolic PAMPs and DAMPs by forming signalling complexes called inflammasomes, which generate active forms of the inflammatory cytokines IL-1 and IL-18

Question 44

Describe in detail how the inflammasome is created and the role of caspase 1.

Answer 44

* After binding of a ligand, multiple identical NLRP3 proteins interact to form an oligomer, and each NLRP3 protein in the oligomer binds an adaptor protein called ASC. The adaptors then bind an inactive precursor form of the enzyme caspase-1 through interactions of caspase-recruitment domains on both proteins. Caspase-1 becomes active only after recruitment to the inflammasome complex * Caspase-1 cleaves the inactive cytoplasmic precursor forms IL-1β and IL-18 to generates active forms which then leave the cell and perform various proinflammatory functions (Pro-IL-1β is cleaved to become IL-1β)

Question 45

NLRP-inflammasome responses are induced by a wide variety of cytoplasmic stimuli that are often associated with infections and cell stress. Provide some examples.

Answer 45

• Microbial products: Bacterial molecules, such as flagellin, muramyl dipeptide, LPS, and pore-forming toxins, as well as bacterial and viral RNA • Environmentally derived crystalline substances: Asbestos and silica • Endogenously derived crystalline substances: monosodium urate, cholesterol * Reduction in cytosolic potassium ion (K+ ) concentrations induced by some bacterial pore-forming toxins * Reactive oxygen species, toxic free radicals of oxygen that are often produced during cell injury

Question 46

Which is best-characterized inflammasome?

Answer 46

The NLRP3 (nucleotide-binding domain, leucine-richcontaining family, pyrin domain–containing-3)

Question 47

NLRP3 inflammasome has demonstrated to contribute to what disorder?

Answer 47

NLRP3 inflammasome is expressed in various cells of the cardiovascular system, including in cardiomyocytes, endothelial cells, and immune cells. The activation of the NLRP3 inflammasome contributes to cardiovascular disorders, including atherosclerosis, myocardial ischemia, hypertension, infectious cardiac disease, cardiomyopathy, and heart failure. Targeting NLRP3 inflammasome via specific pharmacotherapy, nonspecific pharmacotherapy, and nonpharmacotherapy should be beneficial for the control of cardiovascular disorders

Question 48

What are RIG-1 like receptors?

Answer 48

RIG-like receptors (RLRs) are cytosolic sensors of viral RNA that respond to viral nucleic acids by inducing the production of the antiviral type I interferons

Question 49

RIG-1 like receptors recognise what type of RNA/DNA?

Answer 49

RLRs can recognize double-stranded RNA and RNA-DNA heteroduplexes, which include the genomes of RNA viruses and RNA transcripts of RNA and DNA viruses.

Question 50

The two best characterized RIG-1 like receptors are..?

Answer 50

RIG-I (retinoic acid–inducible gene I) and MDA5 (melanoma differentiation-associated gene 5)

Question 51

What makes RIG-I and MDA5 the best RIG-1 like receptors to talk about?

Answer 51

RIG-I and MDA5 recognize different sets of viral RNAs that are characteristic of distinct viruses, partly based on the length of the RNA ligands. RLRs also can discriminate viral single-stranded RNA from normal cellular single-stranded RNA transcripts

Question 52

Where are RIG-1 like receptors expressed?

Answer 52

RLRs are expressed in a wide variety of cell types, including bone marrow–derived leukocytes and various tissue cells. Therefore, these receptors enable the many cell types susceptible to infection by RNA viruses to participate in innate immune responses to these viruses.

Question 53

How do RIG-1 like receptors initiate signalling?

Answer 53

On binding viral RNA, the RLRs initiate signalling events that lead to phosphorylation and activation of IRF3 and IRF7, as well as NF-κB, and these transcription factors induce production of type I interferons

Question 54

What is a major way that the innate immune system blocks viral infections?

Answer 54

Inducing the expression of type I interferons whose most important action is to inhibit viral replication.

Question 55

Define Type I interferon.

Answer 55

Type I interferons are a large family of structurally related cytokines that mediate the early innate immune response to viral infections and most important role is to inhibit viral replication.

Question 56

What are the most important type 1 interferons?

Answer 56

IFN-α (~13 different closely related proteins) and IFN-β (a single protein).

Question 57

What immune cell is a major source of IFN-α?

Answer 57

Plasmacytoid DCs. May also be produced by mononuclear phagocytes

Question 58

What cells produce IFN-β?

Answer 58

IFN-β is produced by many cell types in response to viral infection.

Question 59

The most potent stimuli for type I interferon synthesis are...?

Answer 59

Viral nucleic acids

Question 60

Recall what RIG-like receptors, DNA sensors and TLR have in common?

Answer 60

RIG-like receptors and DNA sensors in the cytosol, and TLRs 3, 7, 8, and 9 in endosomal vesicles, recognize viral nucleic acids and initiate signaling pathways that activate the IRF family of transcription factors, which induce type I interferon gene expression. See past notes for each one if unsure.

Question 61

IFN allow cells to reach an antiviral state. How?

Answer 61

Type I interferons, signaling through the type I interferon receptor, activate transcription of several genes that confer on the cells a resistance to viral infection called an antiviral state 1. Inhibition of viral protein synthesis 2. Degradation of viral RNA 3. Inhibition of viral gene expression and virion assembly.

Question 62

Provide two examples of Type I interferon induced genes and what they do.

Answer 62

* double-stranded RNA–activated serine/threonine protein kinase (PKR), which blocks viral transcriptional and translational events, and * 2ʹ,5ʹ-oligoadenylate synthetase and RNase L, which promote viral RNA degradation

Question 63

What is meant by paracrine action of type I interferon?

Answer 63

virally infected cell secretes interferon to act on and protect neighbouring cells that are not yet infected

Question 64

What effects do type 1 interferons have on viral gene expression?

Answer 64

* The antiviral action of type I interferon is primarily a paracrine action in that a virally infected cell secretes interferon to act on and protect neighbouring cells that are not yet infected. * The effects of type 1 interferons are not specific to viral gene expression, and part of the ability of these cytokines to block the spread of infection is due to their toxicity to host cells that are near infected cells * Interferon secreted by an infected cell may also act in an autocrine fashion to inhibit viral replication in that cell

Question 65

Describe Natural killer cells main function

Answer 65

Major function is killing infected cells, similar to cytotoxic T lymphocytes (CTLs); ready to do so once they develop, without further differentiation (hence natural) Produce IFN-γ, which activates macrophages to destroy phagocytosed microbes

Question 66

Where are natural killer cells derived?

Answer 66

NK cells are innate lymphoid cells (ILCs), bone marrow derived cells with lymphocyte morphology

Question 67

If natural killer cells are lymphoid cells, bone marrow derived with lymphocyte morphology then why are they not can’t lymphocytes

Answer 67

Produce cytokines similar to those made by T cells but lack T cell receptors (TCR). They are referred to as “lymphoid cells,” not “lymphocytes,” because they do not express clonally distributed diverse antigen receptors like the T lymphocytes they otherwise resemble

Question 68

NK cells do not express CD3, the pan-T cell receptor. T or F?

Answer 68

True. This is a characteristic of T cells

Question 69

What is perforin?

Answer 69

perforin, a protein that facilitates the entry of other granule proteins called granzymes into the cytosol of target cells, to initiate a sequence of signaling events that induce apoptosis and cell death

Question 70

What is one type of cell that makes perforin?

Answer 70

NK cells

Question 71

NK cells respond to IL-12 produced by macrophages and in return they...

Answer 71

NK cells respond to IL-12 produced by macrophages and secrete IFN-γ which increases the capacity of macrophages to kill phagocytosed bacteria, similar to IFN-γ produced by T cells. This IFN-γ–dependent NK cell–macrophage interaction can control an infection with intracellular bacteria such as Listeria monocytogenes for several days or weeks and thus allow time for T cell–mediated immunity to develop and eradicate the infection

Question 72

NK cells distinguish infected and stressed cells from healthy cells, and NK cell function is regulated by a balance between signals that are generated from....

Answer 72

activating receptors and inhibitory receptors

Question 73

What do the NK receptors do?

Answer 73

These receptors recognize molecules on the surface of other cells and generate activating or inhibitory signals that promote or inhibit NK responses In general, the activating receptors recognize ligands on infected and injured cells, and the inhibitory receptors recognize ligands on healthy normal cells

Question 74

What does the activating receptor on NK cells respond to?

Answer 74

stimulates the killing activity of the NK cells, resulting in destruction of stressed or infected cells.

Question 75

What does the inhibitory receptor on NK cells respond to?

Answer 75

Engagement of inhibitory receptors shuts off NK cell activity and prevents destruction of healthy cells

Question 76

Explain the anti-viral function of NK cells: antibody-dependent cellular cytotoxicity (ADCC)

Answer 76

A mechanism of cell-mediated immune defense whereby an effector cell of the immune system actively lyses a target cell, whose membrane surface antigens have been bound by specific antibodies. > Antibodies bind to viral infected cells > NK cells bind to antibodies with CD16 receptor > NK cell releases cytotoxic granules and antiviral cytokines to kill the infected cells

Question 77

Can an NK cell recognise a stressed cell in the absence of a antibody?

Answer 77

Can recognise a stressed cell in the absence and presence of an antibody.

Question 78

In the case of chronic or systemic inflammation, how do NK cells respond?

Answer 78

NK cells are biased to produce IFN-γ in many conditions, there are situations of chronic or systemic inflammation that promote IL-10 secretion. NK can also cross-talk with DC in many different ways, including the NK cell killing of immature DC and the promotion of DC maturation by NK cell–derived IFN-γ and TNF-α, which leads to enhanced antigen presentation to T cells. Through these biological activities, NK cells participate in the shaping of the subsequent immune response; NK cells boost or dampen macrophage and T cell responses through IFN-γ or IL-10 secretion, respectively.

Question 79

What is the two-signal hypothesis of immunity?

Answer 79

The activation of lymphocytes requires two distinct signals, the first being antigen and the second being molecules that are produced during innate immune responses to microbes or injured cells. This idea is called the two-signal hypothesis for lymphocyte activation The requirement for additional stimuli triggered by innate immune reactions to microbes (signal 2) ensures that adaptive immune responses are induced when there is a dangerous infection and not when lymphocytes recognize harmless antigens, including self antigens Examples: * IL-12 stimulates the differentiation of naive CD4 + T cells to the Th1 subset of effector cells * IL-6 promotes the production of antibodies by activated B cells

Question 80

Bacterial LPS (lipopolysaccharide), a major PAMP, is recognised by which PRR?

Answer 80

TLR-4 on Langerhans cells in the periphery, stimulating them to migrate to lymph nodes and activate CD4+ T cells and thereby initiate adaptive immune responses

Question 81

What are Langerhans cells?

Answer 81

immature dendritic cells (DC) resident in skin that ingest microbes and their products and break them down

Question 82

The activation of DC cells induces what two changes in the cells?

Answer 82

* Activated Langerhans cells become migratory cells that enter the lymphatic system and move to lymphoid tissues, eg. Lymph nodes, where they become mature DC * These mature DC have enhanced ability to stimulate T cells because of an increase in the number of MHC molecules on their surface and expression of co-stimulatory molecules CD80, CD86