Lecture 10 - ECM Homeostasis

Question 1

What enzymes break down proteins?

Answer 1

Proteinases, they play an essential role in control of intra and extracellular processes
Intra - Placed into special compartments, low pHs
Extra - Secreted into the matrix, usually neutral environments

Question 2

What are the different types of proteinases?

Answer 2

Aspartic, Threonine, Cysteine, Serine, Metallo

Serine and Metallo are mainly extracellular

Question 3

How many metalloproteinases are there?

Answer 3

Around 75 members of the metalloproteinase family

Question 4

What are the domains of metalloproteinases?

Answer 4

Characterised by 3 domains:
Pro/Pre-Domain, this prevents the enzyme working in the cell, the domain needs to be cleaved by a separate enzyme once secreted from the cell to activate it
Catalytic Domain - Binds Zn2+
Substrate Specific Domain - Targets the protein that the enzyme is going to cleave

Question 5

What are the three main types of metalloproteinases?

Answer 5

MMP (Matrix Metalloproteinases) - Key ECM modifiers: Collegenases, Gelatinises, Stromelysins, Membrane-type
ADAM (A disintegrin-like and metalloproteinase) - Mostly membrane bound
ADAMTS (ADAM with thrombospondin motif) - Secreted into ECM like MMPS, Pro-collagenases, Aggrecanases, also involved in ECM breakdown (catabolism)

Question 6

What are Disintegrins?

Answer 6

A family of small proteins that act as potent inhibitors for platelet aggregation and integrin-dependent cell adhesion

Question 7

What does the ECM do?

Answer 7

Support and strength: Basal lamina, Bone/cartilage – made up of ECM
Cellular communication: Hormones, Growth factors, Cytokines
Cell migration, polarity and shape:
Embryonic development, Angiogenesis, Wound repair, Tumour development

Question 8

What are thrombospondins?

Answer 8

Adhesive glycoproteins that mediates cell-cell and cell-matrix interactions

Question 9

What are metalloprotienases?

Answer 9

Key ECM modifiers, they catabolise the ECM, and can lead to the release of molecules which are held within the matrix e.g. hormones and cytokines
They are Zn2+ dependent molecules
They are secreted into the matrix as inactive pro-enzymes
They are activated by removal of the pro region, this region will have a specific sequence that will be recognised enzymes

Question 10

What can inhibit metalloproteinases?

Answer 10

TIMPS - Tissue inhibitors of metalloproteins, family of small secreted proteins which slot into active sites of metalloproteinases catalytic domains 
Alpha2 macroglobulin (in blood, ubiquitous proteinase inhibitor)

Question 11

Cleavage of ECM:

Answer 11

By matrix metalloproteinases
Leads to the production of catabolic fragments
MMPs cut at very specific sequences which can be seen in the genes of the aggrecan (aggrecan cleavage sites)

Question 12

What does cleavage of proteoglycans produce and how?

Answer 12

Neo-epitopes
MMPs cut at very specific sites producing these net-epitopes
An epitope is something an antibody recognises
They are called net-epitopes as before proteoglycan is cleaved by MMP the antibody will not recognise the structure, but once cleaved the net-epitope (neo - new) is produced, which can now be recognised by monoclonal antibodies
For each MMP the epitopes produced will be the exact same as the MMPs cut very specifically

Question 13

Other than MMPs what else can produce ECM fragments/breakdown?

Answer 13

Loading stress also produces fragments of ECM components, these fragments stimulate increased ECM synthesis, which restores healthy matrix

Question 14

When is ECM synthesis/breakdown essential?

Answer 14

Embryonic development:
Cells leave the embryo and become differentiated during, so cells move by interacting with the matrix, also the matrix tells cells where they are and can hold them in a specific place until they reach a certain point in development
The matrix can also store/present growth factors
Wound healing:
MMP-1 required for keratinocyte migration (found in epidermal of the skin)
MMP-7 influences neutrophil migration, and releases VEGF and TNF-alpha allowing angiogenesis
Prevention of tumour development:
If a tumour-prone cell starts to grow out of control, it can be maintained by the matrix in its specific location, however if the matrix is being broken down faster than its being synthesised then the tumour can get out and metastasise

Question 15

How is metalloproteinase activity controlled?

Answer 15

They are first produced by transcription, which can be controlled at the level of gene expression
Then they are either released from the cell, or a few can work on the cell
Some need to be matured - activated by other proteinases e.g. Pro/Pre-Domain removed
They can also be inhibited by TIMPS and alpha 2 globulin

Question 16

How does recognition of ECM break down products lead to homeostatic control?

Answer 16

ECM fragments are recognised by integrins and expressed by many ECM cells, signalling results in increased ECM synthesis
ECM fragments are also recognised by PRR (pattern recognition receptors) expressed on the surface of many ECM, signalling results in inflammation
PRR can also signal to the immune system where the pathogen has entered, as this will cause damage to the ECM, so PRR deal with the pathogen via the innate immune response, and deals with the damage to the ECM

Question 17

What does PRR signal to the immune system?

Answer 17

Where the pathogen has entered
PRR also signal for the immune system to produce cytokines, which can either cause breakdown or synthesis of ECM - both breakdown and synthesis leads to the production of ECM