Lecture 2 - First line and innate defence

Question 1

Commensal microbiota: what are they, why are they present, and how does the body use them as an innate defence?

Answer 1

Microbiota that cause little host damage that may work in a mutualistic fashion

They can be used to aid the body

• Compete with pathogenic bacteria for nutrients and adherent sites
• Strengthen barriers and stimulate epithelial cells to make antimicrobial peptides

Question 2

Extracellular pathogens: how does innate defence fight against it?

Answer 2

Usually susceptible to killing by phagocytes

Question 3

Intracellular pathogens: how does innate defence fight against it?

Answer 3

• Destroy pathogens before they enter cells e.g. release of soluble factors or phagocytosis before become intracellular
• Innate immune killing of of infected cells – NK cells

Question 4

Specialised innate immune defence mechanisms

Answer 4

• Chemical barriers - bile, pH, etc
• Antimicrobial proteins - lysozyme and phospholipase A2, etc
• Antimicrobial peptides –Defensins; Cathelicidines; Histatins. β-defensins packaged into lamellar bodies in lung and skin
• Lectins e.g. RegIIIγ – produced by paneth cells in gut.

Question 5

Soluble molecules: how have they been used as a defence?

Answer 5

Usually pattern recognition receptors and effector molecules at same time – most simple form of innate immunity

Question 6

Lysozyme: what is its antimicrobial function?

Answer 6

Digestion of cell walls of gram-positive/negative bacteria

Question 7

Defensins: what is its antimicrobial function?

Answer 7

Form pores to disrupt pathogen cell membrane

Question 8

What cells are tissue resident and which are circulatory?

Answer 8

Tissue resident:
Tissue-resident macrophages, dendritic cells, monocyte-derived inflammatory macrophages

Circulatory:
Lymphocytes, monocytes

Question 9

How do tissue resident cells fight against pathogens and what happens if they fail?

Answer 9

Tissue-resident macrophages can locally control pathogens and attempt to destroy invading pathogens

If they fail (or maybe even if they don’t fail) then they will call in circulating innate cells

Upon entering tissue monocytes will differentiate into inflammatory macrophages

Question 10

Phagocytosis: what is it and what is the process?

Answer 10

The cellular internalisation and degradation of substances

• Initiated when certain receptors on cell surface are engaged (e.g. MR, scavenger receptors)
• Bound material is internalised by enclosing a membrane around it, forming a phagosome
• Phagosome becomes acidified which kills most pathogens
• Phagosome fuses with lysosome(s) - phagolysosome
• Killing continues (augmented by ligation additional receptors e.g. fMLP)

Question 11

ROS: what does it stand for, what examples are there, how are they generated,

Answer 11

Reactive oxygen species

Include superoxide anion (O₂⁻) and hydrogen peroxide (H₂O₂).

• O₂⁻ - Generated by multicomponent, membrane-associated NADPH oxidase
• H₂O₂ - Generated by

Question 12

NADPH oxidase: what is it, what does it do, where is it found, and what is its use of oxygen called?

Answer 12

Multicomponent, membrane-associated protein

Generates superoxide anion in lumen of phagolysosome – further chemical and enzymatic reactions produce H2O2, hypochlorite and hypobromite.₂

One set of subunits in neutrophil granules/macrophage lysosome; other components in cytosol

Transient oxygen consumption by the cell - Respiratory Burst

Question 13

fMLP receptor: what is it, what does it do, what is its ligand, and what is the process of its activation?

Answer 13

Formalmethionine-leucyl-phenylalanine receptor

Induces ROS

N-formylmethionine – amino acid present prokaryotes but not eukaryotes.

• fMLP receptor recognises fMLP
• Rac2 is activated
• Phagosomes formed
• Rac2 induces assembly of a functional NADPH oxidase in the phagolisosome membrane, resulting in superoxide production
• Acidification produced by ion influx releases granule proteases from granule matrix

Question 14

CGD: what is it, what is it caused by, and what does it cause?

Answer 14

Chronic granulomatous disease

Genetic deficiency of NADPH oxidase

• Less able to kill phagocytosed microorganisms and clear infection
• Unusually susceptible to bacterial and fungal infections.

Question 15

NK cells: how do they know which cells to destroy?

Answer 15

Interferons - typically IFN-β/γ

Question 16

General process of the destruction of a virally infected cell

Answer 16

• IFN-β/γ produced as a response to virus
• Activate STAT1 and STAT2 which combine to form IRF9 to form ISGF3
• Resistance to viral replication induced by using Mx proteins, 2’-5’ linked adenosine oligomers, and the kinase PKR
• Induce IFIT proteins which supress viral RNA translation
• Increase MHC I expression and antigen presentation
• Activate dendritic cells and macrophages
• Activate NK cells to kill virally infected cells
• Induce chemokines to recruit lymphocytes

Question 17

Inflammation: what are the three roles?

Answer 17

• Supply additional effector cells and molecules to site of infection
• Induce local blood clotting, physical barrier stop spread of infection into blood stream
• Promote repair of injured tissue

Question 18

Inflammation: what is the early process?

Answer 18

• Neutrophils attracted then monocytes that differentiate into macrophages
• Endothelial cells become less tightly joined together – exit of fluid and proteins from the blood
• Endothelial cells become more adhesive - circulating leukocytes can bind
• Edema – plasma proteins (complement or MBL) become adherent in tissue
• Clotting – clots can be found around pathogens
• Macrophages and Neutrophils – secrete lipid mediators inflamm e.g. prostaglandins, leukotrienes.

Question 19

Perivascular Macrophages: what are they, why are they useful, and what pathogens target them?

Answer 19

Macrophages that are localised right next to the vasculature

Allows ‘hotspot’ recruitment of neutrophils into tissue

Staphyloccocus aureus targets these macrophages to prevent neutrophil recruitment

Question 20

Cytokines: in what signalling manners can they act and what are the main families of them that are active in innate immunity?

Answer 20

Autocrine - self-acting
Paracrine - adjacent cell signalling
Endocrine - distant cell signalling

• IL-1 family – most cytokines produced by inactive proprotein cleaved by inflammasome
• Hematopoietins – growth and differentiation factors e.g. GM-CSF stimulates monocyte/neutrophil production in bone marrow
• Interferons – type I IFN anti-viral responses
• TNF family – TNF-α multiple key roles inflammation e.g. activator of vascular endothelium and fever

Question 21

Chemokines: what are they, what do they do, what are examples of major chemokines, and what do they do?

Answer 21

Induce cell migration

Chemokines can also act as vasoactive mediators help cells get across endothelium.

• CCL2 – attracts monocytes from blood stream
• CXCL8 – migration of neutrophils

Question 22

Long range effect of macrophages: what overall effects do they cause on the body and what specific effects may they cause?

Answer 22

The typical “feeling ill” symptoms

• Increase body temperature using TNF-α, IL-6, IL-1β (endogenous pyrogens) or LPS (exogenous pyrogen)
• PGE₂ – hypothalamus – increased heat from brown fat – loss of excess heat from skin
• Mechanism to stop pathogen growth – controversial

Question 23

Endogenous pyrogens: what does it mean, what are some examples, what systems are affected by them, and what do they cause?

Answer 23

Cytokines that promote fever

TNF-α, IL-6, IL-1β, etc

• Liver - acute-phase proteins, c-reactive protein, mannose binding lectin - activation of complement (opsonisation)
• Bone marrow - neutrophil mobilisation - phagocytosis of pathogens
• Hypothalamus - increased body temperature - decreased pathogen replication and increased antigen processing/adaptive immune response
• Fat and muscle - increased metabolism, increasing temperature - decreased pathogen replication and increased antigen processing/adaptive immune response
• Dendritic cells - TNF-α stimulates dendritic migration to lymph nodes and maturation (initiation of adaptive immune response)

Question 24

C-reactive protein: what is it, what does it do, and how is it used clinically?

Answer 24

Member of pentraxin family – five identical subunits

• Bind phosphocholine on bacteria and fungal cell walls
• Acts as opsonin and can also activate complement cascade

Commonly used a measure of inflammation in clinic