Module 3 - Innate Immunity, Induced Response

What is the difference between immediate and induced immune response?

Immediate: mediated by preformed effector proteins and resident effector cells, includes alt complement pathway, other protein systems, and resident macrophages

Induced: mediated by inflammation and nonspecific leukocytes, includes resident and circulating macrophages, granulocytes, monocytes, dendritic cells, NK cells, lectin+classical pathway, and interferons

What bridges the gap between immediate and induced innate responses?

Resident macrophages; pathogens coated in complement are recognized by CR on macrophages triggering phagocytosis and release of cytokines to recruit more leukocytes which helps this transition

What are PRRs?

Pattern recognition receptors; these recognize pathogen’s surface proteins and abnormal self cells

What are PAMPs?

Pathogen-associated molecular patterns, distinct pathogenic structures that PRRs bind to

What are DAMPs?

Damage-associated molecular patterns, abnormal cell antigens that PRRs bind to

What are CLRs?

C-type lectin receptors, transmembrane glycoproteins with variable number of C-type lectin domains (CTLDs) that binds to carbs

What are TLRs?

Toll-like receptors, made up of a horseshoe-shaped pattern recognition domain that can dimerize upon PAMP binding, recognize a variety of different molecules, can be found in plasma membrane or endosomes

What are NLRs?

(Nucleotide oligomerization domain)-like receptors, recognize debris from phagocytized bacteria that escapes the phagolysosome and enters the cytoplasm

What are ALRs?

(absent-in-melanoma)-like receptors, form an inflammasome (activated by pro-caspase 1 to caspase 1) in the cytoplasm that recognizes double stranded DNA from phagocytized pathogens or damaged host cells in PAMPs

What are RLRs?

(retinoic acid-inducible gene-1)-like receptors, target double stranded RNA (viruses) and located in cytoplasm, viral RNA binding → dimerization of RIG-1-like helicases with mitochondrial antiviral signaling protein (MAVS) on the outside of the mitochondrial membrane, this leads to activate interferons

What are scavenger receptors?

Broad classes of PRRs with a larger range of molecule recognition (only includes CLRs and TLRs; expressed by macrophages, dendritic cells, neutrophils, B cells, and other cells to trigger phagocytosis, cell adhesion, and changes in intracellular signaling; if no infection, imp for clearing cell debris from stress and death

What are dectins?

A type of CLR that can identify carbs in fungal and bacterial cell walls.

Describe the process of CD206.

aka Mannose receptors

1) The mannose receptor has 10 extracellular domains of 4 types, this recognizes mannose residues of a PAMP

2) Surface sugars of a bacterium are bound by 2 CTLD domains

3) Macrophage ingests bacterium by receptor-mediated endocytosis

4) Bacterial degradation begins in the endosome

5) Endosome fuses with lysosome to gorm a phagolysosome in which bacterium is further degraded

6) Mannose receptor is returned to cell surface and repeat

If a CLR doesn’t have lectin domain, what is the other type?

Collagen-like, good for recognizing bacterial cell wall components and has 2 types:

SR-A1: recognizes LPS, LTA, hepatitis C virus, B-amyloid, and heat shock proteins

MACRO: recognizes LPS, intact Gram (+) bacteria, and proteins

Describe the activation of the NF-kB transcription factor.

When TLRs bind to PAMPs, these triggers an intracellular signaling cascade leading to the activation, this leads to production of pro-inflammatory cytokines to mediate inflammation and recruit leukocytes

What is the significance of genentic polymorphism in TLR genes?

Different alleles of the same TLR gene can produce different TLF variants (allotypes), this mostly impacts membrane TLRs and their structures, which can give greater disease susceptibility, ie TLR4 change = inc risk of septic shock

Ligand binding involving NLRs will lead to what?

NOD dimerization which signals to NF-kB activation

What are the two proteins that can undergo NOD dimerization?

NOD1: recognizes γ-glutamyl diaminopimelic acid from peptidoglycan of Gram(-) cell walls, expressed on many cells

NOD2: recognizes muramyl dipeptide found in most peptidoglycan, expressed only on myeloid and lymphoid leukocytes

What are IFN?

Interferons, small proteins produced by viral-infected cells, functions to interfere with viral replication, alert neighboring cells of virus, and make viral-infected cells more susceptible to NK cells

List the IFN classes

I: a, b, d, k, e, w

II: y

III: l

Describe the anti-viral response of interferons

1) Virus infects an epithelial cell that responds by secreting the cytokine IFN-B

2) IFN-B is bound by the cell’s type I interferon receptors, stimulating an autocrine IFN-a response

3) IFN-B binds to type I IFN receptors on adjacent uninfected cell, giving paracrine IFN-B response

What are pDCs?

Plasmacytoid dendritic cells, secrete up to 1000x more IFN, located in blood and lymphoid tissues, detect viruses through endosomal TLRs: 7 - ssRNAs, 9 - unmethylated CpG motifs in dsDNA

What are 2 cytokine classes crucial to the immune system?

IFNs and ILs

Which IL is the master regulator of inflammation?

IL-1B (IL-1 family is highly conserved)

Where is IL-1B stored?

Macrophages form large amounts of IL-1B stored as an inactive (pro) form in the cytoplasm, ready to release anytime a pathogen is near

Describe the process of IL-1B activation

1) dsDNA is bound to PRR

2) AIM2 receptors become activated and starts forming inflammasome

3) During this process Caspase 1 cleaves pro form of IL-1B + others

4) Pyroptosis occurs, inflammasomes form a pore to release large amounts

What is pyroptosis?

Cell death as a result of pore formation for the secretion of IL-1B

Why is IL-1B the master regulator?

1) Inc speed of recognizing PAMPS, efficiency of phagocytizing and secreting molecules in response

2) Pro-inflammatory cytokines secreted will recruit other effector immune cells

What is TNF-a?

Once bound to its receptor on endothelial cells and neutrophils, it triggers the expression of cell surface adhesion molecules

What are the adhesion molecules of endothelial cells?

Intercellular adhesion molecules (ICAMs)

What are adhesion molecules of neutrophils?

Leukocyte function-associated antigen-1 (LFA-1, integrin)

What is margination?

Occurs when neutrophil adhesion molecules transiently associate with endothelial cell adhesion molecules

1) L-selectins bind to carbohydrates on E-selectins (CD34)

2) LFA-1 binds to ICAM-1

aka rolling of neutrophils along the endothelium of blood vessel cell wall

What is CXCL8?

Attract other effector cells to an infected spot with chemotaxis (the closer you are, the higher [C] of them)

Describe the binding process of CXCL8.

1) CXCL8 binds to its receptor on neutrophil forming complex with G protein

2) Placement of GDP by GTP activates G protein and it dissociates from the receptor

3) G protein dissociates to a and By subunits to associate with other proteins

Describe the process of extravasation.

1) Rolling adhesion, adhesion molecules to bind to selectins on endothelial cells

2) Tight binding, CXCL8 binds to neutrophil receptor

3) Diapedesis, neutrophil squeezes inbetween endothelial cells at junctions

4) Migration, neutrophil follows gradient of chemoattractants towards infection

Contrast the differences in phagocytosis between neutrophils and macrophages

1) Neutrophils are much better at recognizing a wider range of PAMPs due to their PRRs

2) Also have complement receptors to help enhance neutrophil phagocytosis + fuses cytoplasmic granules with phagolysosome

What are primary (azurophilic) granules?

Contains proteins and peptides that distrupt and digest pathogens (i.e. defensins, lysozymes)

What are secondary (specific) granules?

Contains…

1) Lactoferrin that restricts bacterial growht by competing for metal ions

2) NADPH oxidase for respiratory burst (creates reactive O2 species to kill bacteria but also kills neutrophils)

What are tertiary (gelatinase) granules?

Gelatinase restricts bacterial growth by competing for metal ions

What are secretory vesicles?

Adhesion molecules and glycoproteins needed for margination and extravasation

What are 2 ways that neutrophils die?

1) Apoptosis: programmed cell death

2) NETosis: neutrophil cell death that results in decondensed chromatin and proteins that trap and kill pathogens

What are the proinflammatory cytokines systemic effects on the following body parts?

1) Liver

2) Bone Marrow

3) Hypothalamus

4) Fat, Muscle

1) Acute-phase proteins CRP and MBL → activates complement opsonization

2) Mobilize neutrophils → phagocytosis

3/4) Increase body temperature (pyrogen) → decreased viral and bacterial replication

What is CRP?

C-reactive protein, pentraxin that binds to bacterial (or yeasts, fungi, and some protozoa) to act as an opsonin

What is Serum amyloid A?

A small protein that polymerizes to form a pentagon, to which it binds to and stimulates macrophages to secrete proinflammatory cytokines

What is MBL?

Mannose-binding lectin, C-type lectin that binds to mannose containing carbs if pathogens and acts as an oponin, also triggers lectin pathway

Describe the lectin pathway form MBL to C4b

1) MBL associates with MASP zymogens to form MBL complex

2) When MBL binds to pathogen mannose-containing carb, one MASP-2 is activated, then it cleaves itself then other MASP-2

3) MASP-2 cleaves C4 → C4a + C4b, where C4b attaches to microbial surface

Describe the lectin pathway from C4b to C3b

4) MASP-2 cleaves C2 → C2a + C2b

5) C2a + C4b → C4bC2a (classical C3 convertase

6) C4bC2a binds to C3 → C3a + C3b, C3b attaches to the microbial surface

Describe the initiation of CRP in the classical pathway

1) CRP attaches to pathogen surface carbs (i.e. phosphorylcholine

2) CRP interacts with C1

3) Pathogen-bound CRP binds to C1q zymogen stalks → one C1r to cleave itself, the other C1r + two C1s proteases (now active to cleave C4

4) Cleaves C2+C4 to leave C2a+C4b to bind together then to C3

5) C4bC2a binds to C3 → C3a + C3b, C3b attaches to the microbial surface

What are innate lymphoid cells?

Nonspecific immune cells of the lymphoid lineage

What are NK cells?

Cytotoxic ILCs that circulate in the blood and enter infected tissues

What are ILCs (1-3)? and their specific functions

“Helper” cells that reside in tissues

1: Inflammatory activation of macrophages

2: Noninflammatory activation of macrophages

3: Promoting phagocytosis and secretion of antimicrobial peptides

What are CD56bright NK cells?

Weakly cytotoxic, more committed to making cytokines and resides in tissue, give rise to CD56dim NK cells

What are CD56dim NK cells?

Differentiated cytotoxic lymphocytes, comprised more than 90% of NK cells in blood

Describe the pathway leading up to inducing apoptosis.

1) Virus infects cells and triggers the interferon response

2) Type 1 interferon drives proliferation of NK cells

3) Type 1 interferon drives differentiation of NK cells into cytotoxic effector cells

4) Effector NK cells kill virus-infected cells by inducing apoptosis

How is NK cytotoxicity regulated?

1) NK cells can only degranulate after close contact with target cell (this forms an immunological response)

2) NK cell activation is dependent on several NK receptors interacting with target cell ligands (i.e. activating signals > inhibitory signals, then NK will activate + ADCC needs to recognize tumor antigens present)

3) Inhibitory receptors that recognized healthy target cell ligands prevent NK cells from targeting healthy cells (i.e. recognizing MHC class I receptor = no lysis)

NK expresses _ of the 10 TLRs

3, TLR378

Describe the process of macrophage activation of NK cells

IL-12 secreted by macrophages controls the activation and recruitment of NK cells

1) Macrophages secrete proinflammatory cytokines, including IL-12, to recruit naive NK cells

2) NK cells form an immunological synapse with macrophages, strengthening NK activation via proinflammatory cytokines

3) IL-12 stimulates naive NK cells to differentiate into CD56bright NK cells that secrete IFN-y

4) IFN-y from NK further activates macrophages increasing their efficiency

Dendritic cells function?

Scan the environment for infection, phagocytize pathogens, and stimulate lymphocytes to respond to infection

When DCs phagocytize and process pathogens, their cell surface proteins change and NK cells recognize to form an immunological synapse with them

If NK cells outnumber DCs..

NK cells divide and differentiate into cytotoxic effector cells thus killing dendritic cells

If NK cells are less than DCs..

NK cells use cytokines to stimulate DCs to differentiate into a mobile form, that leaves the infected tissue into the lymphatic system to induce an adaptive response