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A10HA > 1 - Neutrophils pt. 1 > Flashcards

1 - Neutrophils pt. 1 Flashcards

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1
Q

Examples of phagocytes

A
  • neutrophil * (blood)
  • monocyte * (blood)
  • eosinophils & basophils (blood)
  • mast cells (present in tissues)
  • MACROPHAGE - present in tissues, ingest micro-organisms, dead/senescent cells - stimulate adaptive system by antigen presentation
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2
Q

Elements involved in phagocytosis

A
  • membrane bound granules containing AMPs enzymes and other factors
  • opsonin
  • pattern recognition receptors on cell
  • pathogen associated molecular patterns
  • RAB GTPases, lysosomal enzymes or anitoxidants
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3
Q

How are particles opsonised?

A

Particles can be opsonised by complement, AB, integrins

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4
Q

What induces internalisation?

A

Phagocytic receptors induce internalisation

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5
Q

Chemotaxis

A

Cells directed by peptides, complement components, cytokines

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6
Q

Part 1 of phagocytosis steps

A
  1. phagocytic receptors induce internalisation
  2. chemotaxis
  3. opsonisation
  4. clustering of receptors on cell signals engulfment
  5. binding - binding between PRR and PAMP, binding between PRR and opsonin, initiation of cytoskeletal reorganisation in the phagocyte
  6. activated receptor complex engages adaptor proteins, resulting in lipid remodelling
  7. cytoskeletal altercations mediated by GTPases directing actin assembly
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7
Q

Part 2 of phagocytosis steps

A
  1. internalisation - Rab GTPases direct conformational changes to membrane and components of the cytoskeletal mesh
  2. destruction of internalised particles
    • phagosome, containing internalised non-self material
    • RabGTPases direct the fusion of membranes and formation of the phagosome and phagolysosome
    • granule contents gain access to particles inside the phagolysosome
  3. killing mechanisms
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8
Q

What are the killing mechanisms

A
  • enzymes
  • anti-microbial peptides (AMPs)
  • reactive oxygen species (ROS)
  • reactive nitrogen intermediates (RNI)
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9
Q

Multi-phase process of phagocytosis

A
  1. attraction/ movement via chemotaxis
  2. recognition at the membrane surface
  3. membrane and cytoskeletal alterations essential for invagination
  4. attachment of lysosomes
  5. generation of reactive O and N species
  6. killing of microbe
  7. externalisation/degranulation
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10
Q

How can phagocytes be seen?

A

Using an electron scanning micrograph

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11
Q

How many cells are produced each day and how long do they live?

A
  • continuously generated - up to 2 x 10^11 cells/day
  • short life span - estimates vary from 8 hours to 4-5 days
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12
Q

Where are neutrophils localised?

A
  • neutrophil localisation to inflammation sites vital for infection clearance
  • BUT, excess infiltration/activation can cause chronic inflammation, limit injury repair and lead to loss of organ function
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13
Q

What are the three main types of granules

A
  1. Primary - peroxidase-positive
  2. Secondary - specific (peroxidase negative)
  3. Tertiary - gelatinase (peroxidase negative)
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14
Q

Peroxidase positive

A

Peroxidase-positive Azurophilic (aka primary), containing bactericidal permeability increasing protein [BPI], elastase, cathepsin G, myeloperoxidase, defensins, azurocidin

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15
Q

Peroxidase negative

A

Peroxidase negative:
a) Secondary (aka specific) containing NADPH oxidase, lactoferrin
& b) tertiary (gelatinase) granules, MMP9: matrix metalloproteinase 9

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16
Q

How many main phases of movement of neutrophils?

A

Three main phases of movement:
1. Early recruitment (forward migration)
2. Amplification
3. Resolution (death/reverse migration)

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17
Q

Why is neutrophil movement relevant?

A

Relevance:
* Not enough neutrophils – infection risk
* Inappropriate resolution – chronic inflammation
* Drug discovery

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18
Q

Neutrophil tracking

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A

Neutrophil trafficking to LN in homeostasis and following inflammation. 1. Neutrophils are produced in BM and released into blood
2. Local inflammation results in recruitment of neutrophils to inflamed tissue and to draining LN via blood and lymphatic vessels
3. Neutrophils also traffick to LN via blood and lymphatic vessels in homeostatic conditions and can egress via efferent lymphatic vessels

19
Q

Efferocytosis

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A

Ingestion of apoptotic cells by phagocytosis

20
Q

Key molecules involved in neutrophil trafficking

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A

TISSUE RECRUITMENT
- PSGL-1: P-selectin glycoprotein ligand 1
- LFA-1: lymphocyte associated antigen 1
LYMPHATIC MIGRATION
- Mac-1: macrophage 1 antigen
- CCR7: C-C chemokine receptor 7
LYMPH NODE
- ICAM-1: intercellular adhesion molecule
- PNAd: peripheral node addressin
BLOOD MIGRATION
- CXCR4: CXC chemokine receptor
- High endothelial venules

21
Q

Phases of neutrophil recruitment

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A

a. earliest signals
b. further recruitment of early neutrophils
c. amplification of neutrophil response
d. extra levels of signalling in infection

22
Q

Phases of neutrophil recruitment: Earliest signals

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A
  • The recruitment of neutrophils to a site of damage occurs in several phases
  • Damage-associated molecular patterns (DAMPs) are released at a tissue injury site and promote the release of hydrogen peroxide (H2O2) and direct the recruitment of early-arriving neutrophils through the SRC family kinase LYN
23
Q

What are DAMPs? (a)

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A
  • Damage Associated Molecular Patterns
  • e.g., DNA, proteins (incl. heat shock proteins), uric acid
24
Q

What is detection by neutrophil mediated by? (a)

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A

Detection by neutrophil mediated by >30 pattern recognition receptors (PRRs)

25
What is recruitment directed by? (a)
1. enzyme phosphorylation 2. signalling pathway
26
Phases of neutrophil recruitment: Further recruitment of early neutrophils
- DAMPs also induce the production of CXC-chemokine ligand 8 (CXCL8) family chemokines and leukotrienes from surrounding tissue cells to further recruit neutrophils
27
Phases of neutrophil recruitment: Amplification of neutrophil response
- Early-arriving neutrophils are then themselves activated to both directly and indirectly promote further secretion of CXCL8 family chemokines and leukotriene B4 (LTB4) to induce neutrophil recruitment from the circulation and amplification of the response
28
Phases of neutrophil recruitment: Extra levels of signalling in infection
In an infection, extra layers of signalling exist to prolong and amplify neutrophil infiltration, including the release of pathogen-associated molecular patterns (PAMPs) and the involvement of other recruited immune cells, such as macrophages, dendritic cells (DCs) and T cells
29
How does infection give rise to additional signals? (d)
- Additional signals from PAMPs prolong and increase neutrophil infiltration - Other immune cells are recruited, e.g. macrophages - Cellular interactions vary with different pathogens, e.g. DC involvement well-known with Leishmania infection - T-cells likely to be involved with persistent inflammation
30
Neutrophil recruitment cascade
1. Recruitment initiated by endothelial changes 2. Recruitment promoted by inflammatory mediators, e.g. histamine, cytokines 3. Endothelium stimulated directly by PRR binding 4. Binding leads to increased expression of adhesion molecules, namely selectin, integrin 5. Upregulation of selectin maximizes recruitment
31
Rolling vs adhesion, crawling and transmigration
Rolling is mostly selectin-dependent, whereas adhesion, crawling and transmigration depend on integrin interactions
32
Neutrophil movement
ROLLING: - tethering - slow rolling ADHESION: - full arrest - firm adhesion CRAWLING - intraluminal crawling TRANSMIGRATION
33
How many methods of transmigration?
Two possible methods of transmigration are acknowledged: paracellular (between endothelial cells; a) and transcellular (through endothelial cells
34
Rolling neutrophils and crawling neutrophils
- Chemokines lining the luminal part of endothelium activate rolling neutrophils, thus inducing conformational changes of neutrophil surface integrins and allowing for subsequent events - Crawling neutrophils follow the chemokine gradient along endothelium, which guides them to the preferential sites of transmigration
35
Extravasation
- crossing the endothelium - requires adhesion proteins
36
Paracellular
- thought to occur at areas of fewer junctional proteins and of less organised alignment
37
Transcellular
- endothelial cell forms projections which attach to the neutrophil
38
What do domes rely on?
Domes rel on adhesion protein enrichment, actin-binding proteins, cytoskeletal modulation
39
What is phagocytosis?
Phagocytosis is a multi-phase, complex process, performed by specialized immune cells, principally neutrophils and macrophages
40
What do neutrophils form?
Neutrophils form part of the initial response to pathogens, containing diverse selection of antimicrobial proteins distributed between 3 different granule types
41
What does neutrophil movement involve?
Neutrophil movement/migration varies with wounding/infection signals and is dependent on adhesion molecules
42
Why is the neutrophil life span/migration important?
Neutrophil life span/migration important in balance between acute/chronic inflammation, and involves trafficking between blood, sites of inflammation and lymph nodes
43
How does a neutrophil pass across the endothelium?
Neutrophil passage across the endothelium can be via paracellular or transcellular routes

A10HA (18 decks)

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