Greg - Introduction and Complement

Question 1

Define the complement system

Answer 1

The complement system is a biological cascade made up of multiple serum proteins that is part of the innate immune system

Question 2

What is serum?

Answer 2

The liquid from clotted blood

Question 3

What is plasma?

Answer 3

The liquid part of nonclotted blood

Question 4

How was complement discovered?

Answer 4

Serum was taken from a patient and mixed with bacteria
It was observed that the bacteria were destroyed
However if you heat the serum and then carry out the experiment the bacteria are not destroyed, therefore it can be said that a heat-labile component in serum has the ability to lyse bacteria (complement)

Question 5

On a electrophoresis graph, where is complement found?

Answer 5

Complement is found in the beta globulin region

Question 6

What percentage of the globulin fraction of serum is made of complemet?

Answer 6

5%

Question 7

What causes the levels of complement to increase?

Answer 7

An immune response i.e. infection

Question 8

In an electrophoresis graph where are the antibodies found?

Answer 8

Found in gamma globulin region

Question 9

What are the four main functions of complement

Answer 9

Formation of MAC

Opsonisation

Disposal of immune complexes and apoptotic cells

Activation of immune responses e.g. vascular permeability, chemotaxis for phagocytes and degranulation

Question 10

What is the main detrimental role of complement

Answer 10

Inappropriate inflammation

Question 11

Give an example of a disease caused by inappropriate inflammation

Answer 11

Systemic lupus is caused by an over active complement system and it causes tissue damage

Question 12

How does complement cause lysis of pathogens?
(4)

Answer 12

Complement is deposited on the surface of pathogen

Complement forms the membrane attack complex (MAC)

This punches holes in the surface of the cell

The cell swells and bursts by osmosis

Question 13

How do complement proteins act as opsins
(3)

Answer 13

Complement proteins bind to pathogens and mark them for phagocytosis

The phagocytes express receptors such as CR1 which allows the cell to recognise complement attached to a pathogen

When the receptor meets the complement there is a conformational change and the pathogen is phagocytosed

Question 14

How does complement activate the inflammatory response?

Answer 14

If complement binds to the receptors found on granulocytes it causes these cells to degranulate e.g. mast cells/gatekeeper cells releasing histamine causing inflammation and vasodilation

Question 15

How do complement proteins clear immune complexes

Answer 15

Complement marks antibody-antigen complexes for phagocytosis in the liver and spleen

Question 16

What does CR1 stand for?

Answer 16

Complement Receptor 1

Question 17

How many proteins make up complement?

Answer 17

30+

Question 18

What cells make complement?

Answer 18

Hepatocytes (mostly produced here)
Epithelial cells of the gastrointestinal and genitourinary tracts
Monocytes/macrophages

Question 19

What type of proteins are complement proteins?

Answer 19

Acute phase proteins
Zymogens

Question 20

Complement proteins are zymogens, what does this mean?

Answer 20

Zymogens are inactive forms of enzymes
It means complement needs to be activated before working

Question 21

How are complement proteins activated?

Answer 21

Proteolytic cleavage (the chopping off of part of the protein)

Question 22

How are complement proteins numbered?

Answer 22

C1, C2, C3 …. C9

Question 23

After cleavage how are the components of complement named?

Answer 23

Usually the larger component gets named B and the smaller A

e.g. C3a and C3b

However for C2, C2a is the larger piece and C2b is the smaller piece

Question 24

What does the b part of complement proteins usually do?

Answer 24

b is usually the membrane-binding fragment

Question 25

How do you write an activated component?

Answer 25

You overline the components

Question 26

What are convertases?

Answer 26

Complexes with proteolytic activity e.g. C3 convertase cleaves C3 into C3a and C3b

Question 27

How many complement pathways are there?

Answer 27

Three: - classical pathway - alternative pathway - mannose-binding lectin pathway

Question 28

What is the endpoint of all three complement pathways?

Answer 28

The formation of MAC

Question 29

How does the classical pathway differ from the other two pathways?

Answer 29

The classical pathway is antibody dependent

Question 30

Why are antibodies needed for the classical pathway?

Answer 30

Complement fixation is needed and this only happens because there was an antibody there first and then complement came along -> in order for complement to stick to the surface of something there needs to be an antibody there first

Question 31

What antibodies initiate the classical pathway?

Answer 31

IgM and IgG

Question 32

Which subclasses of IgG initiate the pathway?

Answer 32

3/4 subclasses initiate the pathway

Question 33

Which is better at complement activation, IgG or IgM?

Answer 33

IgM is way better

Question 34

What is the first type of antibody made after an infection?

Answer 34

IgM is the first antibody secreted by plasma cells they may switch to different types such as IgG after

Question 35

What is the first molecule involved in the classical pathway?

Answer 35

C1q

Question 36

What does C1q bind with during the classical pathway?

Answer 36

2x C1r and 2x C1s to form C1qrs

Question 37

Where is C1q found?

Answer 37

It is found floating around in circulation

Question 38

What are the two fragments of antibodies?

Answer 38

Fab (fragment with antigen binding) and Fc (fragment that is crystallisable) fragments

Question 39

How many Fc fragments does C1qrs have to bind to?

Answer 39

C1qrs has to bind to two Fc fragments on antibodies (this is why the classical pathway is complement dependent)

Question 40

What is the evolutionary point of the classical pathway needing two Fc fragments to bind to on antibodies?

Answer 40

This prevents C1qrs activating in circulation when it accidently bumps into an antibody It prevents unwanted inflammation

Question 41

What happens after C1qrs binds to two Fc fragments?

Answer 41

There is a conformational change in shape and one of the C1r's is activated -> this then activates the other C1r -> this activates both of the C1s s

Question 42

Where exactly on the Fc fragment does complement bind?

Answer 42

The Ch2 domain

Question 43

Why is IgM much better at activated C1qrs than IgG?

Answer 43

IgM is a pentamer while IgG is a monomer

Question 44

What happens after C1s s are activated?

Answer 44

C1s can now cleave C4 and C2 to give C4a, C4b, C2a and C2b

Question 45

What does C4b do?

Answer 45

It sticks down on to the surface of the pathogen

Question 46

What does C2a do?

Answer 46

It sticks down to the surface of the pathogen along with C4b to form C42a

Question 47

What happens after C4b2a is formed?

Answer 47

C4b2a is a C3 convertase => C3 is cleaved into C3a and C3b

Question 48

What happens after C3 is cleaved

Answer 48

C3a floats away C3b sticks to the pathogen surface C4b2a3b is formed

Question 49

What happens after C4b2a3b is formed?

Answer 49

C4b2a3b is a c5 convertase => c5 is split into C5a and C5b C5b binds to C6

Question 50

What happens when C5b binds to C6

Answer 50

This initiates the formation of the membrane attack complex

Question 51

How is the MAC formed

Answer 51

C5b, C6 and C7 bind together to form C5b67 C8 then joins to form C5b678 Many C9 molecules then join to create a pore and form C5b6789

Question 52

How does the Mannose binding lectin pathway work?

Answer 52

MBL attaches to mannose/frucose containing polysaccharides on bacteria MBL is then bound by MASP-1 and MASP-2

Question 53

What does MASP stand for?

Answer 53

MBL associated serine proteases

Question 54

How many subunits are there to MBL?

Answer 54

6

Question 55

When can the MBL pathway begin?

Answer 55

Day one of infection

Question 56

How does the alternative pathway work?

Answer 56

C3 can spontaneously split

Question 57

Write a note on the MAC

Answer 57

Membrane attack complex Best at killing gram negative bacteria - low peptidoglycan Gram positive are generally resistant to the MAC

Question 58

What do the Ca s do after they have been released?

Answer 58

They are anaphylatoxins (pro inflammatory molecules)

Question 59

What do anaphylotoxins do?

Answer 59

Activate basophils and mast cells to degranulate resulting in increased vascular permeability and contraction of smooth muscle cells

Question 60

What is C3bs function outside of pathway activation?

Answer 60

it acts as an opsonin

Question 61

What is C4bs function outside of pathway activation?

Answer 61

It acts as an opsonin

Question 62

List the anaphylatoxins

Answer 62

C5a C4a C3a

Question 63

What are the functions of anaphylatoxins (4)

Answer 63

Degranulation of mast cells Smooth muscle contraction Increased vascular permeability C5a causes chemotaxis of leukocytes

Question 64

Write a note on C3 deficiency

Answer 64

Most severe clinical manifestations of complement deficiency Recurrent bacterial infection and immune complex diseases

Question 65

Give two examples of conditions that are caused by regulatory protein deficiencies

Answer 65

Hereditary angioedema Paroxysmal nocturnal haemoglobinuria

Question 66

Write a note on hereditary angioedema (3)

Answer 66

Caused by mutations in the C1 inhibitor Patients experience intermittent episodes of oedema Oedema can be life threatening if airway becomes blocked

Question 67

How does C1 inhibitor work?

Answer 67

C1 inhibitor causes the dissociation of C1r and C1s from C1q This prevents the classical pathway

Question 68

Write a note on paroxysmal nocturnal haemoglobinuria (2)

Answer 68

An acquired disorder where a defect in PIGA results in a deficiency in GPI-anchored proteins including CD59 Without this RBCs are unprotected from lysis by complement

Question 69

How is paroxysmal nocturnal haemoglobinuria treated?

Answer 69

The monoclonal antibody Eculizumab is used Eculizumab binds C5 and halts the complement cascade

Question 70

What is the CH50 assay used for?

Answer 70

It demonstrates the ability of serum complement to lyse sheep RBCs coated with antibody

Question 71

What does a high CH50 mean?

Answer 71

A poor complement activation