C3.1 Neutrophils & macrophages

Question 1

name the type of receptors on macrophages

Answer 1

macrophage scavenger receptor
Fc receptors
PRR/ TLR e.g. CD14 for LPA
Interact w/ pathogen associated molecular patterns

Question 2

what can effect macrophage activity

Answer 2

Cytokine receptors - e.g activator/deactivator
Chemokine receptors
Activity can be primed so that they are more responsive to a 2nd signal – experimentally can use LPS and IFNy

Question 3

describe process of phagocytosis

Answer 3

must be next to target
chemotaxis impt (cells in right place)
form pseudopodia and enclose target in a phagosome
rate of phagocytosis enhanced (>4000x) if the target opsonised
(if target too large to be phagocytosed) macrophages can fuse - form ‘foreign-body giant cells’ or organised structures with other cells – granulomas – formation involves T cells to ‘direct’ cells

Question 4

describe Respiratory burst

Answer 4

occurs during phagocytosis
requires: NADPH oxidase (respiratory burst oxidase)
> When phagocyte activated the cytosolic components & cytochrome b558 assemble in the cytoplasmic membrane
involves P & cytoskeletal elements

Question 5

define role of NADPH oxidase

Answer 5

catalyzes reduction of oxygen –> superoxide O2-
Xanthine oxidase can also generate O2- (impt in NADH oxidase deficient mice)
froms phagolysosome: activates myeloperoxidase

Question 6

what causes formation of hypochlorite HOCl

Answer 6

myeloperoxidase (MPO)
myleoperoxidase deficiency occurs in 1:200-4000 individuals (asymptomatic unless another condition present, suggests: other mechanisms can compensate) – only in neutrophils
In all cases - waste form macrophages/neutrophils excreted by exocytosis

Question 7

Describe Phagolysosomal enzymes

Answer 7

Hydrolytic enzymes – pH 4.5-5.0 (optimal pH)
pH maintained by a membrane bound ATP-dependent pump - H+ ATPase
(Enzymes:) nucleases, proteases, glycosidases, lipases, phosphatases, sulfases, phospholipases
significant contribution to cell killing
Results in the generation of antigenic peptides for class II MHC loading

Question 8

what does oxidative mechanisms produce

Answer 8

reactive oxygen species (ROS)
Respiratory burst – using NAPDH-oxidase complex
hydrogen peroxide -> myeloperoxidase ->hypochlorous acid
= most bactericidal oxidant in neutrophils - chlorinate targets and effect their function
(Also get production hypobromous/hypoiodous acid)

Question 9

activation of what cell can cause iNOS (NOS2) expression

Answer 9

activated macrophages: IFNy, LPS or muramyl dipeptide
Th2 inhibit NO production via IL4 or IL23
Th1 enhance NO production via IFNy

Question 10

Describe Neutrophil non-oxidative mechanisms

Answer 10

• Granulocytes
• Enzymes e.g present in azurophil granules = specific small storage granules: Block respiratory burst with inhibitor & still get killing of bacteria
• Neutrophil elastases - produced as a preproenzyme – needs to b cleaved. Packaged with proteinase 3 and cathepsin G in primary granules
  Neutrophil elastase e.g. OmpA - if mice deficient 50% more likely to die
• TNF alpha – directly toxic

Question 11

Types of cell death via neutrophils (5)

Answer 11

1. Apoptosis - requires caspase/ no inflammation
2. Autophagy - programmed cell death via cellular starvation (no inflammation)
3. Pyroptopsis - programmed = cell lysis, Inflammation & phagocyte recruitment
4. Oncosis - cell death: lysis (pore formation) = inflammation
5. NETosis - involves oxidative stress = inflammation

Question 12

detail apoptosis via

• intrinsic signal
• extrinsic signal
• TNF
• how apoptosis is delayed

Answer 12

intrinsic signal e.g. high ROS (necrosis)
extrinsic signal e.g. TNFa, engagement Fas w/ FasL or TRAIL w/ TRAIL-R2/TRAIL-R3
TNF low conc (0.1ng/ml) delays apoptosis but high conc (10-100ng/ml) increases incidence
delayed if exposure to inflammatory mediators e.g. IL1, IL2, IL6, IL8, IL15, LPS from bacteria

Question 13

describe defenins

Answer 13

cysteine-rich cationic peptides
Effect gram -ve and +ve bacteria, fungi, some enveloped viruses
Inhibit protein kinase C

Question 14

why arent phagocytes damaged by ROS (6)

Answer 14

1. mitochondria decrease O2 via cytochrome oxidase
2. Mild-uncoupling e- pathway by FFA: keep proton potential below threshold to stimulate superoxide production
3. Cytochrome C - catalyses oxidation of superoxide ion to oxygen
4. superoxide dismutase - coverts superoxide –> hydrogen peroxide (escapes mitochondrion as more permeable to it)
5. Antioxidants: tocopherol, CQH2, ascorbate and low molecular weight anti-oxidants - quench ROS
6. Glutathione peroxidase and catalase - decompose hydrogen peroxide

Question 15

what does high [ROS] result in

Answer 15

Some anion acids become oxidised by ROS (permeability transition pores)
cytochrome C into cytosol = catalyses oxidation of superoxide ion to oxygen
Oxidation of cytochrome b5: disrupts ETC (no O2 –>superoxide ion)
Inhibition of Krebs —> prevents hydrogen peroxide to hydroxyl radical