C2-HC13

Question 1

Properties of the ECM:

Answer 1

Extracellular matrix (ECM):

- A complex and intricate network of macromolecules.
- Similar macromolecules in different tissues, but the composition and organization are amazingly diverse.
- Dynamic: ECM properties and composition can change with age, pathophysiology.
- It can be compared with cooking: you have some ingredients, but can make different things out of it.

Question 2

What are two materials to mimic the ECM?

Answer 2

○ There have been some attempts to recreate ECM outside of the body (biomimetic ECM, synthetic ECM) > this can take many different forms.
§ Matrigel is a very common ECM mimic, very popular in organoid studies.
§ Synthetic scaffold: are built from scratch, from the peptide bases and design your own biomimetic ECM. It can be constructed as desired, tunability of the ECM (of the mechanoproperties) is very advantagous.

Question 3

The higher/lower the modulus, the stiffer the material?

Answer 3

The higher the modulus, the stiffer the material

Question 4

3 elements that are crucial for mechanical linking:

Answer 4

Elements that are crucial for mechanical linking:

1. Ligand-receptor binding
2. Receptor linkage to actin cytoskeleton and force transduction
3. Intracellular signal transduction

Question 5

Properties of ligand receptor binding: integrin

Answer 5

Integrin:

- A key cell-matric adhesion protein (transmembrane protein)
- Ectodomains (means there is an inside and outside) binds ECM glycoproteins (e.g., fibronectin, collagen, laminin), cellular receptors (e.g., VCAM-1), intercellular cell adhesion molecules (e.g., ICAM-1, ICAM-3).
- Cytoplasmic tail binds to integrin activators (e.g., talin, kindlin) and inhibitors (e.g., filamin).

Integrin regulation:
○ Integrin can be in active or inactive state.
○ Activation increase integrin affinity for ligands (makes binding sites more accessible)
§ Left: inactive & right: active.
§ Top: extracellular & bottom: intracellular.
§ Why is it not active all the time? Then the cell is realy sticky and can bind to anything, anywhere and you dont want that.
○ Integrin activation recruits proteins and initiates formation of adhesion complex.

Question 6

focal adhesions (FAs):

Answer 6

Formation and maturation of focal adhesions; once you have multiple adhesion complexes and forces generation > these can accumulate to form large complexes, called focal adhesions (FAs).

What are focal adhesions?
○ Clusters of integrins bound to ECM
○ Cytoplasmic domains attach to cytoskeleton connecting exterior forces to internal signals
○ Forms along actin stress fibers

• Focal adhesion in mechanosensing:
  ○ FAs provides cells with the necessary force transmission pathways (‘grips’) to “feel” their microenvironment through actin-myosin contractions.
  § FA acts as a grip; so that cells can sense the stiffness of the substrate. Cells sense this by pulling the substrate. They need to attach to the substrate in order to feel the stiffness.
  ○ Maturity will increase as the substrate become stiffer. Glass is really stiff, so it has a lot of FA.

Question 7

Possible mechanosensors:

Answer 7

Possible mechanosensors: whole cell, cytoskeleton, adhesion complexes, focal adhesion, adhesion/ adaptors proteins, integrins.

Question 8

Mechanoresponse=

Mechano transduction=

Answer 8

Mechanoresponse= cells make decisions based on what they sense. Types of mechanoresponses; replication, differentiation, migration, apoptosis, etc.
Mechano transduction= the way cells transduce the signal they sense into something they can respond to.

Question 9

2 methods in which cells can remodel the matrix:

Answer 9

Cells actively remodel the matrix > cells move, degrade, deposit matrices.

- Physical remodelling: cells can recruit (pull) fibers towards themselves > changes in the density and topography of the ECM.
- Chemical remodelling: cells can secrete their own ECM, produce agents that crosslink or degrade (proteolysis) in the ECM > changes in the architecture of the ECM. The most important one is cancer invasion. 

Everything happens very locally.

Question 10

Contact guidance:

Answer 10

Cell orientation can be achieved via ‘contact guidance’= the ability of cells to align with the anisotropy of the microenvironment.

Question 11

Manipulating cellular environment:

- Cells control the matrix by ...
- Matrix controls cell through....

Answer 11

Manipulating cellular environment:

- Cells control the matrix by applying traction forces and by producing matrix synthesis and degrading products as well as their inhibitors in response to mechanical cues --> regulation of matrix via cellular mechanical conditioning.
- Matrix controls cell -shape, motility, structure, and function - through its 3D structure and focal adhesion organisation --> regulation of (stem) cell function via engineered micro environments.