4 - innate immunity

Question 1

3 “main” parts

Answer 1

physical and chemical barriers

cellular responses

activation of adaptive immune system

Question 2

Anatomical barriers

Answer 2

Skin (antimicrobial peptides, fatty acids in sebum)

Mouth and upper alimentary canal (enzymes, antimicrobial peptides, sweeping of surface by directional flow of fluid towards stomach)

Stomach (low pH, digestive enxymes, bile salts, antimicrobial peptides, fluid flow toward intestine)

Small intestine (digestive enzymes, antimicrobial peptides, fluid flow to large intestine)

Large intestine (normal intestinal floea complete with invading microbes, fluid/feces exprelled from rectum)

Airway and lungs (cilia sweep mucus outward, coughing, sneezing expel mucus, macrophages in alveoli of lungs)

Urogenital tract (flushing by urine and mucus, low pH, antimicrobial peptides, and proteins

Salivary, lacrimal, and mammary glands (flushing by secretions and mucus, antimicrobial peptides and proteins)

Question 3

Lysozyme

Answer 3

enzyme

saliva, tears, fluids of respiratory tract

cleaves peptidyglycan in bacterial cell walls

Question 4

surfactants

Answer 4

lubricating lipids and proteins on epithelium of respiratory tract

Question 5

antimicrobial peptides

Answer 5

less than 100 AA long (othervise protein)

main types = alpha- and beta-defensins, cathelicidin, and histatins.

Question 6

PRR

Answer 6

pattern recognition receptors

recognize PAMPs and DAMPs, can be inside or on cell surface

All white myeloid blood cells express them, subsets of the lymphocytes also. Cells especially exposed to infectious agents also

Question 7

PAMPs and DAMPs

Answer 7

pathogen-associated molecular patterns

damaga-associated molecular patterns

Question 8

TLR

Answer 8

Toll-like receptor (13)

extracellular region = leucine-rich repeats (LRRs).

upon binding of ligand they dimerize to either homo- or heterodimer

Question 9

TLR signaling pathways

Answer 9

Activate NF-kB (k=kappa), important for activating expression of many innate and inflammatory genes.

can induce subsets of proteins that are effective in combating the specific antigen

Type I interferons !!!! (IFN-alpha and -beta). Activation of INTERFERON REGULATORY FACTORS (IRFs) is essential for inducing transcription of these.

Two key adaptors: MyD88 (most TLRs) and TRIF (TLR 3 + 4 (endosomes, not plasma membrane))

Question 10

MyD88

Answer 10

The MyD88 pathway activates the transcription factors (TFs) AP-1 and NF-κB, but
can also induce the TF IRF7, leading to increased production of IFN α and β.

Question 11

TRIF

Answer 11

The TRIF-pathway results

in the activation of TFs IRF3 and IRF7, and may also activate AP-1 and NF-κB

Question 12

Which TLRs bind what?

Answer 12

Bacteria: 1, 2, 4, 5, 6, 9, 11, 13

Virus: 3!, 4, 7!, 8!, 9, 13

the others can bind fungi ++

Question 13

CLR

Answer 13

c-type lectin receptor

not TLR-family

expressed on monocytes, macrophages, DCs, neutrophils, B-cells and T cell subsets.

generally recognize carb components of fungi, mycobacteria, ciruses, parasites, some allergens.

CLR signaling activates TFs which induce effector gene expression (IRF5, NF-kB, AP-1, NFAT). Induce the expression of cytokines.

Question 14

NLR

Answer 14

Nod-like receptor and nucleotide oligomerization domain(leucine-rich repeat-containing receptor

bind intracellular DAMPs and PAMPs and other harmful substances

Some NLRs can assemble into inflammasones, large protein complezes that cleave and activate the large precursors of the proinflammatory proteins IL-1beta and IL-18

Question 15

NOD1 and NOD2

Answer 15

bind breakdown products of bacterial cell wall peptidoglycans

Initiate signaling pathways that activate the NF-kB, MAPK, and IRF pathways

Question 16

ALRs

Answer 16

AIM2-like receptors

bind long dsDNA from bacteria and viruses (cytosol)

form inflammasomes that promote inflammation, some ALRs may also induce IFN production

Question 17

RLRs

Answer 17

RIG-I-like receptors

RNA helicases that function as cytosolic PRR

recognize viral dsRNA

activate IRFs and NF-kB, inducing expression of interferons and cytokines

Question 18

cGAS and STING

Answer 18

cytosolic DNA from viruses/bacteria activates DNA sensor cGAS (cyclic GMP-AMP synthase), which synthetizes cGAMP, leasing to activation of STING (ER associated) (stimulator of interferon genes), triggering signaling pathways activating IRF3, NF-kB, leading to synthesis of type I IFNs and cytokines

Question 19

Type I interferons

Answer 19

IFN alpha and beta

Bind to IFNAR (IFN alpha receptor), expressed in most cells

Initiates a cascade that eventually allows IFN entry into the nucleus,
where they can stimulate the transcription of ISGs (interferon-stimulated genes). In particular, four ISGs are
important for inhibiting viral replication;

1) Protein kinase R inhibits the translation of eIFα, an initiation factor
whose blocking will inhibit all protein synthesis in a cell, both cellular and viral

2) 2’,5’-Oligoadenylate A
stynthethase (OAS) induces degradation of viral RNA by RNase L

3) Mx group proteins inhibit both the
transcription and assembly of viral components

4) IFIT (IFN-induced proteins with tetratricopeptide repeats)
also block viral RNA translation, as well as inactivate the eIF3 translation initiation factor

Question 20

cytokines

Answer 20

The immune response is also reliant on cytokines, small signaling molecules that can activate and/or regulate
many different cells. Three of the most common ones are IL-1, TNF-α, and IL-6, which all act to increase
vascular permeability during an infection, as well as recruiting and activating immune cells to the site of
infection. IL-1 also recruits MyD88 to its receptors, thus activating the MyD88 pathway. TNF-α activates
macrophages and TAK1, the latter is also an important part of the MyD88 pathway. This will lead to increased
activation of the TFs, and thereby an increased production of ISGs

Question 21

Chemokines

Answer 21

small proteins

chemoattractants for immune cells

som are responsible for migration of white blood cells through the body, others have key roles in early immune and inflammatory responses (attract cells that contribute to clearing the infection/damage and to amplifying the response.

Question 22

Enzymes: iNOS and COX2

Answer 22

key roles in the generation of antimicrobial and proinflammatory mediators

iNOS for formation of NO, which kills phagocytosed microbes.

COX2 (induced by PRR activation in monocytes, macrophages, neutrophils and mast cells) is key to forming prostaflandins, potent proinflammatory proteins

Question 23

Phagocytosis

Answer 23

1) bacterium binds to PRRs on membrane evaginations (pseudopodia)
2) bacterium is ingested, forming phagosome
3) phagosome fses with lysosome
4) bacteria is killed and then digested by low pH-activated lysosomal enzymes
5) digestion products are released from cell

Blood-monocytes, macrophages, neutrophils, DS in tissues are the main phagocytosers.

Question 24

Phagocytic receptors

Answer 24

Most PRRs induce phagocytosis (but nor TLRs)

Enhanced by opsonization: phagocyte recognition of soluble proteins (opsonins) that have bound to microbial surfaces.

MBL and CRP are opsonins, along with many components of the complement system

Question 25

processes that kill phagocytosed microbes

Answer 25

in most phagocytes the phagosome fuses with lysosomes, in neutrophils htey fuse with primary and secondary granules.

the contents of the lysosomes include:

• antimicrobial proteins and peptides (incl defensins ans cathelicidins)
• low pH (due to the activity of a vacuolar ATPase proton pump)
• hydrolytic enxymes (incl lysozyme and proteases)
• and specialized molecules that mediate oxidative attack

Question 26

Oxidative attack

Answer 26

occurs in neutrophils, macrophages, DCs

employs highly toxic reactive oxygen species (ROS) and reactive nitrogen species (RNS). Damage mibrobial membranes and intracellular components.

Question 27

elimination of intracellular bacteria

Answer 27

Autophagy

the bacteria is surrounded by a membrane to form an autophagosome that then fuses with lysosomes.

Activated by NOD1 and NOD2

Question 28

Cell turnover and clearance of dead cells

Answer 28

Dead and dying cells express DAMPs on the surface, signaling for phagocytes to eat them. receptors of phagocytes recognize these and do

Example of DAMP: the lipid mediator lysophosphatidic acid (chemoattractant function)

Tumor cells may escape phagocytosis by expressing CD47 protein (“don’t eat me” signal), inhibiting phagocytosis by macrophages.

Question 29

regulated cell death

Answer 29

contributes to pathogen elimination

regulated cell death = cell death induced by receptor-activated signaling pathways

One form of RCD = apoptosis, induced by TNF binding + NK cells and Tc cells

two other forms: NETosis + pyroptosis

NETosis
NET = neutrophil extracellular traps
made of chromatin with associated antimicrobial proteins
traps and kills pathogen and the neutophil

Pyroptosis:
typically of macrophages. induced by inflammasome activation. Several benefits;
1) death of the infected macrophage = prevents further spread
2) important fot the release of mature IL-1beta and IL-18, and DAMPs that may enhance local inflammatory responses

Question 30

Local inflammation

Answer 30

triggered by innate immune responses

1) tissue damage and bacteria cause resident sentinel cells to release chemoattractants and vasoactive factors that trigger local increase in blood flow and capillary permeability.
2) Permeable capillaries allow an influx of fluid (exudate) and cells
3) neutrophils and other phagocytes migrate to the site of inflammation (chemotaxis)
4) phagocytes and antibacterial substances destroy bacteria

redness, swelling, heat, pain (and loss of function).

proinflammatory cytokines: IL-1beta, TNF, IL-6

Question 31

Innate lymphoid cells

Answer 31

ILCs
lack the BCR and TCR, instead respond to other signals including infection+ damage+ stress, to enchance and regulate innate and inflammatory repsonses

Question 32

NK cells

Answer 32

activated by infection, cancer, DAMPs. immeatiate response.

enhanced by IFN-alpha (produced early in viral infections, example of positive feedback).

secrete cytotoxic mediators (perforin and granzymes, kills cells by inducing apoptosis), cytokines (some do) most commonly TNF-alpha and IFN-gamma (TYpe II INF).

Nk receptors:
1) activating receptors - have specificity for various cell surface ligands that serve as indicators of infection/cancer/stress. Specificity for proteins upregulated on infected/malignant/stressed cells, rather than pathogen-stuff.

2) inhiboitory receptors - rec membrane proteins (usually MCH) on normal healthy cells, thus inhibitng the NK from killing the cell. Vuris infected/tumor cells usually lose expression of MCH proteins - NK attack!!

Question 33

ILC populations produce distinct cytokines and have different roles

Answer 33

6 populations have been identified, divided into three groups based on type of proteins, and functions.

Nk cells in the blood. Other ILCs are found mainly in epithelial barrier tissues, inclusing mucosal tissues in intestine + lungs and glandular tissues (salivary glands).

Except for NK cells and ILC3, most ILCs dont have PRRs and are not directly activated by pathogens. Instead, they are activated by cytokines and other mediators produced by local epithelial cells, macrophages and DCs after PRR stim by PAMPs

Question 34

ILC group 1

Answer 34

includes NK and ILC1 cells

share the production of IFN-gamma and TNF-alpha

NK cells have cytotoxic activity

Question 35

ILC group 2

Answer 35

produce IL-4, -5, -9 and -13

innate protection against parasitic worms

IL-5 activates eaosinophils

IL-13 promotes smooth mucle contraction, mucus production, recruitment of activated macrophages

ILC2s also secrete amphiregulin (mediates tissue repair during infection-resolution)

Question 36

ILC group 3

Answer 36

vary a bit in the mediators they produce and their funcitons

LTi = lymphoid tissue inducer. Essential for the development of secondary lymphoid organs

produces LT-alpha and beta (members of TNF family, important for sec lymph organ development), IL-22 (induces intestinal epithelial cells to produce antimibrobial peptides + other molecules that help them resist bacterial infection), IL-17 promotes local inflammatory responses important against yeast and fungi.

Question 37

Similarities between ILCs and Th

Answer 37

ILC1 and Th1 produce IFN-gamma + TNF

ILC2 and Th2 produce IL-4, IL-5 and IL-13

ILC3 and Th17 produce IL-17 and IL-22

Question 38

Regulation of innate and inflammatory responses

Answer 38

septic schock - circulatory and respiratory collapse

major cause of sepsis = gram neg bact cell wall component LPS (aka endotoxin). This is a logand for TLR4. LPS = highly potent inducer of TNF-alpha, IL-1beta and IL-6, chemokines, antimicrobial components.

danger: fluis loss into tissue (as part of inflammatory response) -> lower blood pressure. Together with the clotting factors stimulated by TNF, could result in capillary clotting.

Question 39

Innate and inflammatory responses are regulated

Answer 39

Inflammation is controlled by both positive and negative feedback mechanisms. One example of positive
feedback is the production of the cytokines IL-1β and TNF-α, which will induce further production of
themselves. Negative feedback mechanisms also regulate the immune response and inflammation to prevent
damage to the host. Regarding the same example with IL-1β and TNF-α; these cytokines promote production of
receptors they can bind to in a soluble form. These receptors will bind and prevent the cytokines from further
stimulating an immune response.

LPS tolerance can be developed if the macrophage is continually exposed to it (no response)

Question 40

Interactions between the innate and adaptive immune system

Answer 40

DCs carry microbes to lymphoid tissues to present them to T and B cells. The pathogen can be attached to the cell surface or in a phagosome. DC travels through lymphatic vessels. Ususally present on MCH class II

The binding of a PAMP to a DC causes it to mature to perform this function.

Matuer DCs have the ability to activate naïve T cells that eventually wil become mature cytotoxic and helper T cells.

extracellular bacteria and endosomal nucleic acids from internalized bacteria stimulate the DC to produce IL-12 - ACTIVATES TH1 (secrete IFN-y + activates macrohpages and NK to eliminate the pathogens + infected cells).

PAMPs from helminths and bacteria + fungi that bind to the plasma membrane and and cytosolic PRR -> DC not able to secrete IL-12, produces IL-10 instead (IL-4 and -13 from nearby cells (basophils and ILC2s)). ACTIVATES TH2 (produces cytokines to activate other leykocytes to release mediators that help clear these pathogens)

fungal PAMPs bind CLR dectin-1 in DC, ACTIVATES TH17 (secretes IL-17, activates production of mediators tha recruit inflammatory cells to the site).

Activation of TLRs in precence of vitamin A induces the DC to secrete IL-10 + TGF-beta, which together convert the naïve T cell to T regulatory cell (inhibit other immune reponses).

Question 41

Some PAMPs can activate B cells independent of helper T cells

Answer 41

naïve B cells typically recuires multiple signals (Ag binding to BCR + sigs from TH cells) to mature into Ab-secreting plasma cells.

However, B cells have TLRs, and binding of PAMPs will activate signaling pathwats that can add to or substitute for the normally-required signals.

Example: LPS at high concentrations can activate B cells in mice. I humans, no TLR4 (reg LPS) in B cells, but they can be activated by CpG DNA (microbial).

Question 42

Adjuvants

Answer 42

adjuvants that have been shown to enhance immune responses contain ligands for TLRs or other PRRs.

killed or inactivated pathogen vaccines have adjuvants naturally

Question 43

Some pathogen clearance mechanisms are common to innate and adaptive immune response

Answer 43

the adaptive immune system has co-opted several pathogen clearance mechanisms, such as opsonization and complement-activation, so that they contribute to Ab-mediated pathogen elimination

opsonization = binding of soluble proteins to microbial surfaces to help pahgocytes recognize and phagocytose them