Exam 1 (Lectures 3&4) - Complement

Question 1

Define complement, complement fixation, complement cascade, and complement activation.

Answer 1

1) Complement = distinct plasma proteins that react with one another to opsonize pathogens and induce a series of inflammatory responses to fight infection

2) Complement Fixation = the process of binding serum complement to the product formed by the union of an antibody and antigen

3) Complement Cascade = part of the immune system that enhances the ability of antibodies and phagocytic cells to clear microbes and damaged cells from an organism, promote inflammation, and attack the pathogen’s cell membrane

4) Complement Activation = generates active components with various effector functions
- MAC (lysis of bacteria)
- complement-mediated
opsonization
- enhancement of local
inflammation

Question 2

Where is complement made?

Answer 2

Liver

Question 3

List the ways the compliment cascade is initiated and the molecules responsible for the initiation.

Answer 3

1) Classical pathway = Complement-fixing antibody (IgM, IgG) binds antigen

2) Alternative = initiated when C3 (protein molecule) breaks down into C3a and C3b; C3b will bind to any membrane nearby to initiate complement cascade (innate)

3) Lectin = MBL (mannose-binding lectin) binds to mannose on microbes to initiate the cascade (innate)

Question 4

Under normal circumstances, complement fixation by the classical pathway is not occurring in the bloodstream even though antibody and complement components are circulating together. Please explain.

Answer 4

In order for complement cascade to be initiated, antibody has to be bound to antigen; (C1 will only bind to antibody that is connected to antigen, opening up the C1 binding site of the antibody) out in normal circulation, this is not happening.

Question 5

List the classes of antibody that can fix complement and explain why IgM is the most efficient.

Answer 5

IgG and IgM; IgM is more efficient because it is a pentamer. (One IBM molecule can initiate complement cascade by itself because it has multiple C1 binding sites).

Question 6

Define membrane attack complex (MAC) and explain its role in defense against microbes.

Answer 6

MAC is a cytolytic effector cell of innate and adaptive immunity; it forms pores in the plasma membranes of pathogens/target cells, leading to osmolysis

Question 7

List 4 functions of complement that contribute to eliminating bacteria, including molecules and cells involved for each.

Answer 7

1) Lysis of bacteria: molecule, MAC

2) Opsonization: molecule = C3b; cell = neutrophil and macrophage

3) Chemotaxis: molecule = C5a and other “a” molecules; cell = neutrophil

4) Increased vascular permeability: molecule = C5a; cell: neutrophil

Question 8

In general, explain how complement is regulated and the consequences if it is not regulated.

Answer 8

1) Short-acting components that are quickly hydrolyzed into inactive states so they act at the location of activation.

2) Soluble proteins regulate complement activation by inhibiting or slowing it down (host inhibitor molecules).

3) If not regulated, animal dies.

Question 9

List 2 ways the complement system can be inactivated in a blood sample.

Answer 9

1) Chelate calcium
-results in inactivation of complement and the clotting cascade (EDTA)

2) Heat to 56 degrees C for 30 minutes
- destroys complement but
does not harm antibodies

Question 10

Describe a neutrophil.

Answer 10

Granulocytes with the primary function of phagocytizing and destroying bacteria and fungi.

The predominant circulating WBC; circulate for 8-10 hours

Part of the innate immune system; first responders to arrive in high numbers to sites of inflammation

Question 11

List two or three important surface receptors on a neutrophil and explain their importance.

Answer 11

1) Inflammatory mediator receptors = leukotrienes, chemokines; cytokines bind to receptors and stimulate immune response

2) Selectin and Integrin receptors = Cell adhesion molecules

Question 12

Describe the margination pool and how that influences neutrophil counts in blood samples from animals that are stressed.

Answer 12

The margination pool is in post-capillary venules. Selectins are expressed here and neutrophils loosely bind to endothelium via selectin adhesion molecules and roll along the venule.

When an animal is stressed, the endothelium is signaled to up regulate selectins causing more neutrophils to bind so they can exit the bloodstream and get to the danger.

The neutrophil count in a blood sample (circulating pool) is lower than the number of neutrophils in the blood because of the margination pool.

Question 13

Define selectin, integrin, diapedesis, and chemotaxis and use these terms to describe the steps involved in a neutrophil going from the bloodstream to the site of infection.

Answer 13

1) Selectin = Adhesion molecule that will loosely bind neutrophils to the post-capillary venule endothelium

2) Integrin = Adhesion molecule that tightly binds neutrophils to endothelium
- during inflammatory stimulus,
endothelium up-regulated integrin
receptor molecules to which
neutrophilic surface integrin
molecules bind tightly to stop
rolling

3) Diapedesis = Process of cells squeezing between endothelial cells to leave the blood stream and enter the tissues

4) Chemotaxis = Cellular movement in response to extracellular chemical gradient
- neutrophils leave the bloodstream
to follow the chemotactic gradient

Question 14

Explain why animals with leukocyte adhesion deficiency (a deficit in integrin molecules) die from overwhelming bacterial infections despite having neutrophilia (increased numbers of neutrophils in the bloodstream).

Answer 14

These animals have a loss of integrin binding.

When this is lost, neutrophils are not bound to sites with inflammatory stimulus so there is no diapedesis or chemotaxis taking place. This results in the neutrophils being unable to reach the bacteria to phagocytize it.

Question 15

Define opsonize, phagocytosis, phagosome, and phagolysosome.

Answer 15

1) Opsonize = Prepare cell to be phagocytized (eaten); done by C3b

2) Phagocytosis = Ingestion of bacteria or other material; done by phagocytes

3) Phagosome = Vesicle formed around particle engulfed by phagocyte via phagocytosis

4) Phagolysosome = Cytoplasmic body formed by fusion of phagosome with lysosome; happens within phagocytes

Question 16

Explain the killing mechanisms of a neutrophil, including the important molecules involved (e.g. toxic products produced by the oxidative burst and bactericidal molecules in granules), how they contribute to killing, and their relative potency.

Answer 16

1) Lytic enzymes and anti microbial peptides from granules (lysosomal contents)
- Primary granules (hydrolases, lysozyme, defensins, and myeloperoxidase)
- Secondary granules (lysozyme, lactoferrin, collagenase, elastase, gelatinase)

2) Oxidative burst: most potent killing mechanism
- Occurs in phagolysosome
- Oxidative enzyme in membrane initiated chemical reaction that results in potent killing products (bleach, aldehydes,
hydrogen peroxide)

3) Neutrophil extracellular traps (NETs)
- Nuclear material and granular proteins that are released extracellularly by stimulated neutrophils
- Creates mesh that traps bacteria allowing antimicrobial proteins to kill bacteria
- Form of neutrophil death called NETosis

Question 17

Define defensins including function and location in the body.

Answer 17

Defensins are cationic antimicrobial peptides that form an essential element of innate immunity.

Located in areas of hydrophobicity so they can integrate into negatively charged microbial membranes (found in phagocytic cell granules)

Form a pore in membranes to lyse bacteria (similar to MAC)

Question 18

List the 5 cardinal signs of inflammation and the changes that occur to cause the sign.

Answer 18

1) Redness = Caused by vasodilation and increase blood flow to area

2) Heat = also results from vasodilation

3) Pain = Results from vasoactive substances acting on and damaging nerve endings

4) Swelling = Caused by the increase permeability and leakage of fluid and cells into tissues

5) Loss of function = due to pain and swelling

Question 19

Describe how neutrophil function can be decreased. Give an example of when you might want to decrease neutrophil function. Give an example of a common consequence of neutrophil function.

Answer 19

1) Neutrophil function can be decreased during:
- Distress
- Drug treatment with glucocorticoids
- Viral infections, bacterial virulence factors that allow bacteria to resist neutrophil killing
- Neonates and periparturient period have less efficient neutrophil function

2) We would want to decrease neutrophil function if they are causing a problem.

3) Common consequence = bacterial and fungal infection

Question 20

List the types of clinical conditions that stimulate an eosinophil response and explain how an eosinophil response can be enhanced by adaptive immunity.

Answer 20

1) Parasitic infections and allergies

2) Humoral adaptive immunity
- T Helper 2 cytokines, IL4, IL5, IL13
- IgE binds to parasite and attracts eosinophil via FcER

Question 21

Compare and contrast the macrophage and the neutrophil, including origin, life-span in tissues, location, functions, and killing mechanisms, type of inflammation, type of lesion produced with an accumulation in tissues.

Answer 21

1) Origin = both originate from hematopoietic stem cells in bone marrow

2) Life-span = Macrophages have a long life span; neutrophils have a short one

3) Killing mechanism = Macrophages utilize lysosomal enzymes, oxygen radicals, and nitric oxide; neutrophils utilize lysosomal enzymes, oxidative bursts, and NETosis.

4) Type of inflammation = Macrophages cause chronic inflammation; neutrophils cause acute inflammation

5) Type of lesion = Macrophages result in granulomas; neutrophils cause abscesses

Question 22

List three central roles the macrophage plays in the immune system.

Answer 22

1) Initial defense

2) Antigen presentation to T cells

3) Effector functions performed by activated macrophage

Question 23

List the names, locations, and functions of the resident macrophages in the body.

Answer 23

1) Osteoclast = Located in bone; functions to remodel bone

2) Kupffer cell = Located in liver; functions to remove bacteria from portal venous system

3) Serosal macrophage = Located in serosal surfaces of pleural or peritoneal cavity; functions to kill invading bacteria and remove dead and dying cells

4) Microglial cell = Located in the brain; functions to remove dead cells from the brain to maintain normal physiologic state

5) Mesangial cell = Located in the kidney; functions to trap and remove bacteria from glomerulus

6) Pulmonary alveolar macrophage (PAM) = Located in the alveolus of the lung; functions to phagocytize particles and digest/kill them

7) Pulmonary intravascular macrophage (PIM) = Located in the endothelium of lung capillaries; functions to remove microbes from blood