Phagocyte function and Macrophages Flashcards
Why is phagocytosis important?
*Protects against invading organisms
*Processes foreign antigens
*Presents parts of foreign antigens activating adaptive immune system
(links innate and adaptive immunity)
*Purges debris and damaged/dying host cells
Which cells are phagocytic?
-Macrophage
-Neutrophil (Polymorphonuclear leukocyte, PMNs)
-Dendritic cell
-eosinophil
-basophil
Where do phagocytes act ?
- Sites of infection (attracted by inflammatory response)
- Mucosal lymphoid tissue (patrolling of Respiratory & GI tracts, Liver)
- Lymph nodes (filtering from Lymph fluid)
- Spleen (filtering from Blood)
List the process of phagocytosis in simple terms
- Activation of resting phagocytes
- Chemotaxis
- Attachment and signalling
- Phagosome maturation
- Killing and antigen presentation
Describe the Activation of resting phagocytes
Describe the Chemotaxis of phagocytes (Step 2)
Movement towards an increasing concentration of attractant bacterial protein, capsule, cell wall, complement (C5a), chemokine (CXCL-8)
Compared the enhanced and unenhanced attachment of phagocytes (step 3)
Unenhanced
Pattern Recognition Receptors (PPRs) for
Pathogen-Associated Molecular Patterns (PAMPs)
(eg. Microbe cell wall components, viral DNA)
Enhanced
Opsonin Recognition receptors for Antibody (Fc) or Complement (C5a) fractions previously fixed to microbe components
Describe unehanced attachment of phagocytes, giving examples (step 3)
pattern recognition receptors (PRRs) are designed to recognise a few highly conserved structures Pathogen-Associated Molecular Patterns (PAMPs) (examples in image)
PAMPs can be hidden/absent by pathogen diversity in cell wall structure
Unenhanced - Pattern recognition receptors (PRRs) can be one of two:
Endocytic pattern recognition receptors
Signalling pattern recognition receptors
Describe Unenhanced - Endocytic pattern recognition receptors
Mannose receptors
bind to terminal mannose and fucose on microbial glycoproteins/lipids
Scavenger receptors
bind to other bacterial cell wall components
e.g. Lipopolysaccharide (LPS), peptidoglycan
Describe Unenhanced - Signalling pattern recognition receptors
Toll-like receptors
different TLRs bind different microbial molecules but TLR’s do NOT induce phagocytosis
Binding to bacterial and fungal components:
transmits a signal to the nucleus inducing cytokine production e.g. IL-12, TNF-a, IL-6
leads to innate immune defences such as inflammation, activation of other phagocytes
Binding to viral components:
triggers anti-viral interferons (IFN)
List the types of TLRs, their ligands and cytokines
TLR4 detects Gram-negative bacteria because of its ability to recognize endotoxin. It can also activate the macrophage by a second pathway that is initiated by Trif, which leads to a secondary production of IFNbeta and
autocrine activation of additional macrophage genes.
CD14 is a GPI-linked membrane protein that facilitates the recognition of LPS by TLR4 so that it increases LPS sensitivity. Recently CD14 has also been shown to facilitate recognition of ligands by TLR2 and TLR3.
TLR4 also recognizes degraded extracellular matrix and the nuclear protein high mobility group 1 protein (HMGB1) which can be released by necrotic cells, an example
of damage-associated molecules.
Describe enhanced attachment
More specific and efficient
Mediated through opsonins
C3b + C5a promote enhanced attachment
Fc fragments of IgA and IgG
Macrophages use both enhanced and unenhanced attachment
Neutrophils use mainly enhanced attachment
Opsonic receptors require antibody or complement to recognize the target
Bacteria opsonized by C3 fragments or antibody engage complement receptors (CR) or Fc receptors (FcR).
Monocytes and macrophages express a range of receptors (CR1, CR3, CR4) for C3 cleavage products that may become
bound to pathogens, immune complexes or other
complement activators.
Describe phagosome formation
- The microbe attaches to macrophage receptors on its cell surface
- This causes actin cytoskeleton rearrangement, whereby the Actin filaments align in pseudopodiaaround the microbe
- The microbe is slowly engulfed by the cell membrane which invaginates to form a vesicle on its inner side
- the vesicle then separates from the cell membrane forming a phagosome.
Describe phagosome maturation
- electron pump uses ATP to pump protons into the phagosome, lowering the pH inside - has a direct effect on microbial growth
Destruction
-Phagosome fusion with lysosome form phagolysosome
-Oxygen-dependent – ROS, NO : Oxygen-independent – enzymes, Proteases, Nucleases
Processing
-MHC-II presentation of antigens, TLR signalling inducing pro-inflammatory cytokine release
How do lysosomes contribute to Oxygen-independent killing?
Lysosomes also contain
Lysozyme – breaks down proteoglycans in bacterial cell walls
Cathepsins – proteolytic enzymes that degrade bacteria
Lactoferrin – deprives bacteria of iron
Defensins – small peptides that form channels in lipid bilayers
Describe Oxygen-dependent killing
the generation of reactive oxygen and nitrogen species.
Oxidation of proteins, carbohydrates, DNA destroys function
Oxidation of lipids can destabilise cytoplasmic membrane structure
How does Streptococcus pneumoniae evade phagocytosis?
Streptococcus pneumoniae – has a capsule
Resists unenhanced attachment by preventing binding ofpattern recognition receptors
Resists C3b opsonisation
How does Yersinia evade phagocytosis?
Yersinia – depolymerise actin preventing engulfment
How do Salmonella and Mycobacterium Block phagolysosome killing
Salmonella – more resistant to toxic ROS and defensins
Mycobacterium – blocks phagosome fusing with lysosome
How does Staphylococcus aureus Kill phagocytes
Staphylococcus aureus – produces leukocidin,
a toxin which damages membranes
Why is Phagocytosis of apoptotic cells important?
Q. Give an example in the immune system where macrophages phagocytose apoptotic cells.
Important for regulation of immune responses
Prevents leakage of cytotoxic or antigenic contents
Rapid, efficient and ‘silent’ (no inflammation)
A. Thymocytes that fail the processes of positive and negative selection die by apoptosis and are phagocytosed by thymic macrophages. Also B cells that die within lymphoid follicles are taken up by tingible body macrophages
Which phagocytes eat apoptotic cells?
Professional phagocytes – macrophages etc.
mobile and can infiltrate tissues
‘Amateur’ phagocytes – essentially any cell can take up a neighbouring dying cell
slower kinetics than the professional phagocytes
What are the signals from apoptotic cells?
‘find-me’ signals attracting phagocytes
soluble and secreted by apoptotic cells
‘eat-me’ signals activating phagocytes
expressed on the apoptotic cell surface
‘don’t eat-me’ signals preventing phagocytosis
NOT expressed by dying cells
Describe the Host “Find me” signals
Chemoattractants
-Lysophosphatidylcholine (LPC)
-ATP
-Chemokines e.g. CXCL3 and CXCL5 (neutrophil
chemoattractants)
Apoptotic neutrophils produce ‘find me’ signals (e.g. lysophosphatidylcholine) that attract macrophages
Describe the Host “Eat me” signals
Normally expressed on the inner leaflet of the plasma membrane. In apoptosis PtdSer is translocated to the outer part of the lipid
bilayer. Involves a flipase and scramblase
enzyme
The appearance of phosphatidylserine (PS) in the outer leaflet of the plasma membrane is characteristic of apoptotic cells. PS recognition by PS-binding proteins stimulates the uptake of apoptotic cells and the production of antiinflammatory mediators, especially TGFb, which inhibit production of proinflammatory chemokines and cytokines.
Describe scenarios where there is no full phagocytosis but just host cell housekeeping
a) RPE (retinal epithelial cells) in the eye remove
fragments of photoreceptors that die by apoptosis in response to light – essential for
maintaining vision
b) Microglia in the brain help trim and drive neuronal structuring by “nibbling” them (trogocytosis)
Describe don’t eat me signals
Host cells control the phagocytic activity of macrophages by displaying the ‘don’t eat me’ signal CD47. CD47 engages a receptor in macrophages called SIRPalpha that through
its immunoreceptor tyrosine-based inhibitory motif (ITIM) motif inhibits the uptake process. CD47 expression by tumor cells has been proposed as an immunosurveillance escape mechanism
CD31 expressed on endothelial cells, platelets, macrophages and Kupffer cells, granulocytes,
lymphocytes (T cells, B cells, and NK cells), megakaryocytes, and osteoclasts binding normally leads to repulsion signals. During apoptosis CD31 signalling is disabled so that the phagocyte is not actively rejected
What are the consequences of apoptotic cell removal?
Secretion of ‘pro-healing’ anti-inflammatory cytokines (e.g. IL-10, TGF-β)
-reduce inflammation
-promote wound healing
Presentation of antigens
-maintenance of self tolerance
-release of unprocessed pathogens/antigens
Describe the Defects in apoptotic cell clearance - links with disease
Leakage of content from non-cleared apoptotic cells act asninflammatory factors
Lack of suppression of inflammatory and immune responses
* no release of anti-inflammatory cytokines e.g. TGF-b, IL-1
Onset of autoimmune disorders linked to inefficient removal of apoptotic cells
* knock-out mice studies (systemic lupus erythematosis)
Describe the Defects in phagocytosis- links with disease
Chronic granulomatous disease Missing NADPH oxidase components
Chediak-Higashi Syndrome Hyperpigmentation/albinism delayed phagosome-lysosome fusion low chemotaxis
Both lead to recurrent infections
List the Macrophage subsets and their role
Describe the M1 phenotype
Tissue damage/microbial entry triggers TH1 driven inflammation
Monocytes recruited from circulation/bone-marrow
Monocytes differentiate into macrophages which can help activate DC’s to stimulate Th1/NK cells which then release IFN-γ
IFN-γ promotes Macrophages to become M1-phenotype
M1 inflammatory phenotype produce NO, ROS, IL-1, TNF & MMPs
Inflammatory microbial response
Describe the M2 phenotype
Alarmins (IL-25, IL-33 TSLP) stimulate Th2 and
other cell subsets to release IL-4 & IL13
Macrophages develop M2 and later regulatory
phenotypes which begin to accumulate
M2 Macrophages antagonise M1
Macrophages and promote wound healing
Regulatory Macrophages stop wound healing when
complete
Describe how Helminth infection can also promote Macrophage
proliferation in situ
Nematode worm infection drives high levels of IL-4
through Th2 and other cell types
High IL-4 promotes M2 macrophage proliferation in
situ
M2 macrophages kill the pathogen
High IL4 and M2 macrophage presence can also produce a M1/M2 combined phenotype that is proliferative
