Lecture 23 - Mechanobiology 1

Question 1

What is mechanical signalling?

Answer 1

A cell isn’t floating - it is surrounded by layers. It also sites in the extracellular matrix, which can vary in its chemical composition. It can also vary in its physical properties - e.g STIFFNESS & ELASTICITY can vary. Bone, for example, is very hard, which means cells within it experience a hard environment, which differs compared to the brain for example, which has a soft environment.

Question 2

What is important to maintain cellular differentiation?

Answer 2

Environment - it van even act as a driver?

Question 3

What does a stiff environment lead to?

Answer 3

promotes bone cell differentiation (osteoblasts)

Question 4

What does a soft environment lead to?

Answer 4

leads to adipocytes or neurons

Question 5

What is mechanobiology?

Answer 5

the study of how physical forces and changes in cell or tissue mechanics contribute to development, physiology and disease

Question 6

What is mechano-transduction?

Answer 6

the conversion of a physical force to a biochemical response (aka mechano-signalling)

Question 7

What is mechano-sensing?

Answer 7

when a protein or cellular structure responds a physical cue to initiate mechano-transduction

Question 8

What is the hierarchy when it comes to mechano-transduction?

Answer 8

• Mechano-sensing
• Signal transduction
• Signal integration at nucleus
• Cellular response

Question 9

What occurs in mechano-sensing?

Answer 9

• cells test their environment
• adhesion receptors, membrane proteins probe ECM

Question 10

What occurs in signal transduction?

Answer 10

• mechanical signal transduced along a linked network
• cytoskeleton is often the force conduit (pathway)

Question 11

What occurs in signal integration at the nucleus?

Answer 11

• accumulation of signals over time
• chromatin rearrangement (e.g. stiffness = flat nucleus = affected chromatin arrangement = affected transcription = change in cellular response), nuclear pore opening

Question 12

What occurs in the cellular response?

Answer 12

• from microseconds to minutes
• cell shape, fate, motility, growth

Question 13

What is an example of mechanotransduction in action?

Answer 13

Blood pressure autoregulation

Question 14

What is myogenic tone?

Answer 14

Change in blood vessel diameter in response to pressure (a mechanical cue - force which is exerted on cells)

Question 15

How does mechano-transduction in action autoregulate blood pressure?

Answer 15

Increase in pressure, leads an increase in arterial diameter. However, an increase in calcium leads to constriction of the vessels. This ensures there’s an equal blood supply to the tissues (an even flow of blood). This can be regulated by this mechanism.

Question 16

How do cells change their cytoskeleton upon fluid flow?

Answer 16

• Fluid flow is usually above epithelial layers.
• Strain (lack of fluid flow) leads to an affect of the cytoskeleton - lots of filaments of actin

Question 17

How does auditory mechano-transduction and hearing work?

Answer 17

• The movement of fluid (or gel) will be recognised, usually by ion channels, and it is converted to a hearing response

Question 18

How does a lung on chip function?

Answer 18

• resembles in-vitro lung
• epithelial are adjacent to endothelial cells, forming capillaries. These 2 cell types are adjacent
• When alveoli changes size (stretching and relaxing) this can be important for the physiological activity of the cells. This should be represented in-vitro model. The topology should also be reflected - e.g. the endothelial cells adjacent. There is also a membrane in-between. The mechanical forces should also be reflected - e.g. air flow entering the alveoli. In the chip, the air is pumped over the epithelial layer. The blood flow (fluid flow) travels over the endothelial layer. Breathing can also be represented, as a vaccum can be applied to the chambers - stretching the membrane. This device is called a PDMS device and it is flexible and translucent (allows for imaging).

Question 19

What is TER (trans-epithelial resistance)?

Answer 19

a measure of electrical resistance across the monolayer was taken. The tighter the monolayer, the higher the resistance. The formation of the monolayer (only liquid interface), the TER is increasing. At the air-liquid interface, the resistance is much higher. This tells us that having a more physiological parameters allows the epithelium to tighten much faster and to a higher degree.

In order to explore the integrity of a monolayer, dyes can be used. The rate of which the dye is transported can be quantified and measured.

Change of cell size can be measured by analyzing the movement of a quantum dot.

Question 20

What can an infection often lead to?

Answer 20

An infection often leads to an inflammatory response of epithelium. This response leads to TNF (tumour necrosis factor) production

Question 21

What is TNF?

Answer 21

a pro-inflammatory cytokine

Question 22

What is ICAM?

Answer 22

ICAM is a cell adhesion molecule, which is not usually expressed in epithelium. When TNF is applied to epithelium, the expression of ICAM is produced.

Under controlled conditions (with them labelled to monitor them), there is no attachment of neutrophils to the endothelial layer. However, under inflammatory conditions, there are many neutrophils, which accumulated.

Live imaging of neutrophils show that neutrophils are migrating from lower channel, then penetrate the endothelial layer (lower channel), travel through the pore, and travel to the epithelial layer (upper channel).

Question 23

What is the advantage of using organ on chip to study lung inflammation?

Answer 23

Exploring how drugs effect this process can be facilitated through the chip, however you wouldn’t be able to do so in a petri dish in-vitro experiment.

Question 24

How does bacteria found on epithelial layer initiate an inflammatory response?

Answer 24

The inflammatory response leads to the production of ICAM, which then leads to the neutrophils migrating to the epithelial layer an reaching the bacteria and phagocytose them.

Question 25

Explain how the uptake of nanoparticles requires mechanical stretching

Answer 25

• Application of glass particles onto epithelial cells will trigger an inflammatory response.
• Under control (no nanoparticles) and control with stretching - low ICAM expression
• Addition of nanoparticles without stretching led to an increase
• However, the largest increase is with nanoparticles and stretching (stretching representing breathing). This means that the inflammatory response is amplified. This can only be seen in the chip organs and not on a petri-dish