L5-6: Phagocytosis + Complement System

Opsonization

Coating of + charged opsonins (ex. IgG, IgM, C3b etc.)

Allows neutrophil to bind bacteria (normally both - so repel)

Fc Receptors

Neutrophil receptor that binds antibodies that are found to antigens (esp. IgG antibody)

C3b receptors

Neutrophil receptors that bind to C3b when it coats bacteria etc.

Phagosome

Membrane bound vesicle that has ingested microbe/material inside (ie. neutrophil engulfing opsonized bacteria)

Phagolysosome

Phagosome (neutrophil + bacteria) and lysosome fused, becomes digestive vacuole with enzymes that can kill microbes

Hydrolases

Primary Granule

Breaks covalent bodys through water

Degrades dead bacteria + dead tissues

Lysozymes

Primary + Secondary granule

Breaks down peptidoglycan in gram-positive bacteria

Found in body secretions

Defensins (“Antimicrobial Peptides”)

Primary Granule

Cationic proteins that kill bacteria (especially gram +)

Hydrophobic outside and hydrophillic inside → allows it to form a pore in membranes

Myeloperoxidase

Primary Granule

Oxygen mediated killing mechanism

Lactoferrin

Secondary Granule

Binds with iron → prevents bacteria from aquiring it (results in death)

Collagenase

Secondary Granule

Degrades CT → allows it to move to site of inflammation

What is the active ingredient in bleach?

Hypochloride

NETs / NETosis

Neutrophil Extracellular Traps

Neutrophil recognizes pathogen through receptors → releases its contents → traps bacteria and antimicrobial proteins kill it (instead of phagocytosis)

Is NETosis a result of neutrophil death?

No, it’s an active response to inflammatory stimuli

What is pus?

Neutrophils bursting after extended phagocytosis of bacteria

Macrophage Phagocytosis

As macrophages kill pathogen, they take “souveniors” (antigens) of their targets to present to lymphocytes → important APCs for adaptive immune system

What roles do complement and antibody receptors play in phagocytosis?

??

M1 Macrophages

Focus on killing pathogens

More phagocytic activity

More lysosomal enzymes

M2 Macrophages

Focus on Repair

Repair tissues

Reduced microbial killing

Is myeloperoxidase present in neutrophil or macrophage killing mechanisms?

Neutrophil

It’s a very HARSH enzyme → neutrophils are harsh killers versus macrophages are more gentle

What cytokines induce M2 macrophages?

IL-4

IL-13

IL-10

What cytokines induce M1 macrophages?

IFN-y

What is arginine converted to destroy bacteria?

Arginine → Nitrous Oxide

M1 cells

What is arginine converted to repair tissues?

Arginine → Citrulline

What causes a macrophages to remove dying neutrophils? Why is this removal important?

Absense of CD31 ligands on the receptors of neutrophils

• Prevents damaged caused by escaping neutrophil enzymes

What are the first cells to arrive at inflammation sites? The second?

1) Neutrophils

2) Macrophages

What are the most effective opsonins?

Antibodies

Complement

If a macrophage doesn’t eat a dead/dying neutrophil, what could happen?

The neutrophil could release enzymes that cause damage

What is the complement system?

Proteins activated when antigen + antibodies combine → causes enzyme cascade → cell lysis + opsonization

What is the complement protein that causes opsonization, flagging the bacteria for destruction?

C3b

What are the 3 pathways of the complement system?

1. Classical

2. Alternative

3. MB-Lectin

Have certain triggers to prevent the system from damaging beneficial cells

How do the complement pathways differ? How are they similiar?

Initiation of each cascade is different

Once C3b is made → cascade is same

What complement pathways are a part of the innate vs. adaptive immune response?

Innate: Alternative + MBL pathways

Adaptive: Classical

What immunoglobulins are needed to intiate the classical complement pathway?

1x IgM OR

2x IgG

Classical Pathway

1. Antigen + Antibody binds → conformational change

2. C1 binds to Ag-Ab complex

3. C1 cleaves C4 into C4a and C4b

• C4a → goes away

• C4b → binds to membrane + C2

4. C1 cleaves C2 → becomes C4bC2b (C3 convertase)

5. C3 convertase cleaves C3 → C3a + C3b

• C3b → opsonin

6. C3b binds C5

7. C4bC2b cleaves C5 → intitates terminal pathway

Alternative Pathway

1. C3 breaks into C3a + C3b

2. C3b carbonyl group binds to cell surfaces

3. Factor B → binds to C3b

4. Factor D → cleaves Factor B-C3b complex

5. C3bBb (C3 convertase) → C3 breaks down and continues on pathway

What is the C3 convertase of the alternative pathway?

C3bBb (Factor B + C3b)

What is the C3 convertase of the classical pathway?

C4bC2b

Are antibodies required for the MB-Lectin Pathway?

No

MBL Pathway

1. MBL binds to mannose

2. Activates MASP-2

3. MASP-2 claves C4 → cascade initiated

What is MBL?

Mannose Binding Lectin

Soluable PRR

What is mannose?

PAMP located on microbes

What is MASP-2?

MBL-associated serine protease

Has similiar activity to C1

What is the sequence for phagocytosis?

1. Chemotaxis: phagocytes arrive to infection site

2. Adherence: phagocyte adheres to target

3. Ingestion: englufment of target

4. Destruction: killing and digestion

“CAID”

Egestion only occurs in…

macrophages

What fuses with phagocytes to help kill microbes?

Lysosomes → has enzymes that kill pathogens

How do neutrophils bind to opsonized microbes?

Through specific receptors

1. Antibody receptor

2. Complement receptor

What are the professional phagocytes?

Neutrophils

Macrophages

What are 3 killing processes of the engulfed bacteria/pathogen?

1. Lytic enzymes + Antimicrobial peptides from granules

2. Oxidative Metabolism (respiratory burst)

3. Neutrophil Extracellular Traps

Oxygen mediated killing mechanism / “respiratory burst”

• Most potent killing mechanism of a neutrophil

• Occurs in phagolysosome

• Killing products:

  • Hypochlorite

  • Hydrogen peroxide

  • Aldehydes

  • Oxygen radicals

Chronic Granulomatous Disease

Defective oxidative enzyme → can’t kill bacteria → frequent infections/inflammation