Phagocytosis

Question 1

what are the main roles of phagocytosis?

Answer 1

→to protect the body from pathogens

→ to dispose of damaged/dying (apoptotic) cells

→ to process and present antigens (Ag) This processing/presenting of antigens activates the adaptive immune system.

→links the innate and adaptive immune system
.• Destruction of the pathogen via respiratory burst

• Activation of genes leading to cytokine and chemokine release

Question 2

what are types of phagocytes and what is their origin?

Answer 2

→neutrophils
→macrophages (M)
→dendritic cells

origin: myeloid lineage; generated in bone marrow
function : identify, ingest and destroy pathogens have receptors for opsonins

Question 3

other cells (apart from phagocytes)

Answer 3

→mast cells
→ eosinophils
→ basophils (myeloid lineage)
→natural killer (NK) cells
→(lymphoid lineage; bone marrow)

Question 4

what are properties of neutrophils?

Answer 4

→(polymorphonuclear (PMN)
→leukocytes most abundant WBCs (circulating in blood) →early response (inflammation)
→ phagocytosis and killing of microbes enzymes: lysozyme, collagenase, elastase

• Perform NETosis- produce NETs that stops bacteria from colonising a tissue

Question 5

what is the lifespan of neutrophils?

Answer 5

→8-10 hours in blood

→4-5 days in tissues

Question 6

what are properties of macrophages? (include lifespan)

Answer 6

→monocytes (blood 20-40hrs)
→efficient phagocytosis
→killing of microbes
→secrete inflammatory factors (cytokines) => inflammation

Question 7

where are dendritic cells found and what is their function?

Answer 7

→skin
→ mucosa
→ tissues
→capture microbes
→phagocytosis
→not just to eliminate present Ag to T cells
→link innate and adaptive immune response

Question 8

how do dendritic cells signal for T cell activation?

Answer 8

Signal 1
→ antigen recognition by MHC:peptide (Major histocompatibility complex) onto TCR

Signal 2
→co-stimulation by CD80/CD86 (cluster of differentiation - protein on Dcells activated by B cells)
→onto CD28 (proteins on T cells that provide co-stimulatory signals for T cell activation/survival)

Signal 3
→ cytokines released by macrophages

Question 9

what are the steps for phagocytosis?

Answer 9

→Chemotaxis (mobilisation to site of infection/injury) →Recognition and attachment to microbe/dead cells →Engulfment
→Killing/digestion of ingested microbe/dead cells

Question 10

what is chemotaxis?

Answer 10

→ movement of cells towards site of infection

→guided by chemoattractant

Question 11

what are chemoattractants released by?

Answer 11

→bacteria - N-formyl-methionine-leucine-phenylalanine peptides (fMLP)
→inflammatory cells chemokines (IL-8)
→ damaged tissues

Question 12

what are the requirements of recognition of pathogens in phagocytosis?

Answer 12

→Requirements
→react to invading pathogens (foreign)
→no reaction to body’s own tissues (self)

Question 13

definition of PAMPs?

Answer 13

→PAMPs (pathogen associated molecular patterns) = structures shared by groups of related microbes

Question 14

what are PAMPs?

Answer 14

→present on pathogens and not on host cells
→invariant structures: shared by an entire class of pathogens
→essential for survival of pathogens
→prevents pathogen evasion of immune responses
→e.g. ds viral RNA=> replication
→e.g. lipopolysaccharide (LPS) => bacterial membrane

Question 15

what are Pattern recognition receptors (PRRs)?

Answer 15

→present on phagocytes (and other cells, e.g. epithelia) →recognize PAMPs
→detect foreign invaders or aged/damaged host cells

Question 16

where are toll like receptors found?

Answer 16

→ plasma membrane

→endosomal membrane

Question 17

C-type lectin receptors (CTLRs)

Answer 17

PRR e.g. mannose receptor

Question 18

NOD-like receptors (NLRs)

Answer 18

PRR reside as free proteins in cytoplasm

Question 19

RIG-like helicase receptors (RLRs)

Answer 19

PRR cytosolic receptors for viral dsRNA

Question 20

Scavenger receptors

Answer 20

PRR various bacterial wall components (CD14 scavenges LPS-LBP)

Question 21

what do Toll-like receptors (TLRs) do?

Answer 21

→ essential roles in innate immunity
→ conserved during evolution
→ human TLRs recognize PAMPs
→ stimulate production of inflammatory cytokines

Question 22

what PAMPs are recognized by TLRs?

Answer 22

→ lipolysaccharide (gram negative)
→ lipoteichoic acid (gram positive)
→ bacterial DNA sequences (unmethylated CpG)
→ single/double-stranded viral RNA
→ glucans (fungi)

Question 23

what is the role of opsonization?

Answer 23

facilitates phagocytosis (recognition of microbes)
→ Opsonized microbes can be phagocytosed easier (via receptors for opsonins on phagocytes)

→ Clinical note! => Encapsulated microorganisms require opsonization with antibodies to be effectively phagocytosed because the capsule deflects phagocytes

Question 24

how is opsonization done?

Answer 24

→coating of microbes with opsonins (such as proteins of the complement system, C3b, C4b, and antibodies, immunoglobulin).

Question 25

how is a phagosome formed and matured?

Answer 25

Microbe makes contact with the membrane
→actin cytoskeleton rearrangement
→membrane remodelling occurs to form pseudopods
→which wrap membrane around microbe = forming phagosome
→Lysosomes fuse forming phagolysosome
→Pathogen destruction

Question 26

what are the lysosomes involved in oxygen dependent killing of pathogens?

Answer 26

→proteolytic enzymes (cathepsins): degrade microbes
→lysozyme: breaks bacterial walls
→lactoferrin: binds iron => not enough left for bacteria
→defensins: destroy bacterial walls

Question 27

describe the oxygen-dependent killing of pathogens?

Answer 27

→resting phagocyte = NADPH + oxidase are not bound →activated phagocyte = assembly of NADPH oxidase generation of superoxide anion

Question 28

what are the oxygen-dependent killing equations?

Answer 28

O2 → O2 (superoxide)
(OXIDASE)

H2O + O2- → H2O2;OH (hydrogen peroxide; hydroxyl radicals)
Through DISMUTASE

arginine + O2 → NO + citrulline
iNOS (inducible NO synthase)

NO + H2O2 → peroxynitrite radicals oxidising radicals (ROS and NOS) kill phagocytosed microbes

Question 29

blocking phagocyte attachment

Answer 29

→Streptococcus pneumoniae – encapsulated bacteria

Question 30

blocking engulfment

Answer 30

Yersinia

Question 31

blocking destruction

Answer 31

Salmonella – resistant to ROS(reactive oxygen species)

Mycobacterium – blocks phagosome-lysosome fusion

Question 32

killing of phagocytes

Answer 32

Staphylococcus aureus – toxin => damages membranes

Question 33

what are other types of prey that phagocytes go after?

Answer 33

→Micro-organisms
→Damaged or dying cells
→normal turnover of cells = 100-200 billion cells/day apoptosis (programmed cell death)
fast, efficient removal by phagocytes
‘silent removal’: no inflammation
phagocytes: discriminate apoptotic vs. viable cells

Question 34

how does phagocytosis of apoptotic cells occur?

Answer 34

at-me signals apoptotic
→cell attraction as phosphatidylserine is expressed on apoptotic cell
→cell binds to phagocytic receptor
→Rac bound to GDP → Rac bound to GTP
→which then generates myosin-II and force generation for wrapping by pseudopods

→Repulsion on viable cell

Question 35

how does discrimination of apoptotic / viable cells happen?

Answer 35

Apoptotic cells ‘Eat-me’ signals
→Recognized by phagocytes => promote engulfment

→Viable cells ‘Don’t eat-me’ signals
→ Recognized by phagocytes => no engulfment

Question 36

what happens after phagocytes have taken up apoptotic cells?

Answer 36

→ secrete‘pro-healing’ cytokines
→ reduce inflammation (e.g. IL-10)
→ promote wound healing (e.g. TGB-β)
→ presentation of self antigens
→ role in maintenance of self tolerance

Question 37

How can efferocytocytes reduce inflammation?

Answer 37

Blocking TLR signalling
Blocking proinflammatory cytokine signalling (IFN-α)
Blocking (TNF-α expression)
Promote (IL-10 secretion)