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BIEN 320 > lec 9-12 > Flashcards

lec 9-12 Flashcards

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1
Q

MCF-10a

A

benign tumor cell

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2
Q

MCF-7

A

invasive tumorgenic

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3
Q

MDA-MB-231

A

highly metastatic

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4
Q

How does stiffness change during metastatic progression

A

Decreases

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5
Q

how does stiffness change in 1)Cytoplasm
2)Nucleus
3)Nucleolus for metastatic cells

A

stiffness decreases for MDA, to different degress for these regions

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6
Q

Metastic cell mvm speed and distance

A

Greater speed and distance in 3D

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7
Q

Traction force metastatic cells

A

Much larger traction force than benign cells

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8
Q

4T1 vs 67NR

A

4T1 metastatic, works harder on stiffer substrates

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9
Q

what happens in epithelial to mesenchymal transition

A
  • Lose apical-basal polarity, cell-cell adhesion
  • Reorganize cytoskeleton
    • migratory and invasive

cells flatten -> area increase, cell and nuclei volume decrease

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10
Q

What happens to forces during EMT

A

Cells apply more forces and do more work as mesenchymal

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11
Q

particles in metastatic cells appear ? this means?

A

More diffusive, meaning the cells are more fluid-like

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12
Q

what does high vs low YR mean

A

Low YR -> YAP inactive (nuclear to cytoplasmic)
High -> YAP active

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13
Q

when does YAP turn on genes

A

when in nucleus, involved in mechanotransduction and cell fate.

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14
Q

How does substrate stiffness affect YAP activity?

A

Alone, substrate stiffness does not predict YAP nuclear localization; contractile work is the actual predictor.

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15
Q

What mechanical factor best correlates with YAP nuclear localization?

A

Contractile work—the amount of mechanical energy a cell exerts on its substrate.

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16
Q

What role does nuclear compression play in YAP activation?

A

Compression of the nucleus stretches nuclear pores, allowing YAP to enter and activate gene expression.

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17
Q

Can nuclear swelling activate YAP?

A

No. Only nuclear compression (not swelling) causes YAP to translocate to the nucleus.

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18
Q

How does cell spreading relate to YAP activation?

A

Larger spreading area → more contractility → more nuclear YAP, due to greater traction force and nuclear compression.

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19
Q

What experimental method measures the mechanical input for YAP activation?

A

Traction Force Microscopy (TFM)—used to measure cell-generated forces on a substrate.

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20
Q

Why is YAP mechanosensing important in cancer biology?

A

In tumors, increased matrix stiffness and cell contractility can abnormally activate YAP, driving proliferation and tumor progression.

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21
Q

what happened to
HeLa and MDA-MB231 cancer cells with synthetic peptide crosslinkers

A

cells became stronger, but

Migrated less in 2D and 3D environments.

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22
Q

What does RT-DC (Real-Time Deformability Cytometry) measure?

A

Cell mechanical phenotype in flow, useful for identifying disease-specific mechanical changes in blood cells.

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23
Q

How does malaria affect RBC mechanics?

A

Malaria-infected RBCs become stiffer.

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24
Q

What mechanical changes occur in blood cells during COVID-19?

A

RBCs stiffen, neutrophils soften, and monocytes enlarge.

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25
Why are platelets ideal for studying cell mechanics?
They are physically simple (no nucleus), strongly contractile, easy to isolate, and relevant to cardiovascular disease.
26
What do Plavix and aspirin target?
Platelet aggregation: Plavix blocks ADP receptors, aspirin inhibits thromboxane production.
27
What triggers pathological clotting?
Endothelial damage exposing collagen + high shear stress.
28
What is fibrin?
A polymer formed from fibrinogen by thrombin; crosslinked by Factor XIII to form an elastic clot scaffold.
29
How extensible is fibrin?
Up to ~600% of its original length; extremely elastic.
30
How do platelets interact with fibrin?
Platelets adhere to and contract fibrin fibers using actomyosin-generated forces, compacting the network.
31
What is stress-stiffening in fibrin networks?
As applied stress increases, fibrin stiffens due to straightening of thermal fluctuations in the polymer network.
32
What effect does more platelets have on a clot?
Greater contraction, more stiffening, and higher elastic modulus.
33
What role does myosin play in clot contraction?
Myosin II generates contractile forces; inhibition by blebbistatin reduces compaction and clot stiffness.
34
What does abciximab do?
Inhibits integrin αIIbβ3, blocking platelet-fibrin adhesion and aggregation.
35
What are the three phases of clot contraction?
P1: Pre-contraction, P2: Active contraction, P3: Final compaction.
36
How do platelet filopodia contribute to clotting?
Filopodia extend/retract to pull on fibrin fibers, gathering and compacting the network.
37
How does contraction affect clot structure?
Increases fibrin density and stiffness; traps RBCs to form the solid clot.
38
Why is clot compaction spatially non-uniform?
The edges contract faster than the core, leading to uneven platelet speed and deformation.
39
What mechanical actions of platelets drive clot formation?
Actomyosin contraction, filopodia-driven fiber pulling, and integrin-mediated adhesion.
40
Where do platelets come from?
Formed by shearing of proplatelets extended by megakaryocytes in the bone marrow sinus.
41
What promotes platelet release in vitro?
Shear stress (e.g., from fluid flow) and microtubule-based sliding and twisting driven by dynein motors.
42
What role does microtubule sliding play in platelets?
Drives elongation and ring formation of proplatelets, contributing to platelet release.
43
How can we mimic bone marrow shear in vitro?
Using a biochip or platelet bioreactor-on-a-chip with fluid shear flow to accelerate platelet production.
44
What is the importance of synthetic platelets?
Enable scalable platelet production for therapy and drug testing; industry sees high investment potential.
45
What happens in thrombocytopenia?
Too few platelets → bleeding, poor clotting, and increased risk of hemorrhage.
46
How is platelet biogenesis engineered in vitro?
By recreating marrow shear and cytoskeletal mechanics to stimulate proplatelet extension and fragmentation.
47
Latrunculin A
depol actin
48
What happens to cell contractility in the absence of FLNa?
Cells (e.g., M2 cells) fail to increase contractility (stiffness, and spreading) in response to substrate stiffness or shear stress.
49
What is FilGAP?
A GTPase-activating protein that inactivates Rac by converting Rac-GTP to Rac-GDP, shifting cells toward Rho activation and contraction.
50
What is the effect of Rac inactivation by FilGAP?
Decreases actin polymerization and spreading, increases myosin-mediated contractility.
51
How is FilGAP binding to FLNa regulated?
Strain on FLNa reduces FilGAP binding, which allows Rac to become active again. This increases spreading and decreases Rho activity.
52
What does FLNa do under strain?
It changes conformation, releasing FilGAP and increasing integrin binding — altering signaling output.
53
What is the effect of deleting FLNa domain 20 (d20)?
It unmasks the integrin binding site, increasing integrin binding without the need for mechanical force.
54
What is FLAC (Fluorescence Loss After photoConversion)?
A technique used to measure protein unbinding rates with high time resolution (better than FRAP). longer fluo -> stronger binding
55
What happens to FLNa networks with increased myosin activity?
They stiffen due to internal stress and strain, mimicking external shear.
56
What causes rupture in actin–FLNa networks?
Unbinding of FLNa crosslinks at high stress, not actin filament breakage.
57
What happens to FilGAP and integrin binding under strain?
FilGAP unbinds faster, integrin binds more strongly. This switches cellular signaling toward adhesion and spreading.
58
What is the role of Rho and Rac in cancer?
- Rho promotes invasion, angiogenesis, and actomyosin contractility. - Rac promotes lamellipodia and mesenchymal migration.
59
How do M2 and A7 cells differ in response to substrate stiffness?
- M2 (no FLNa): poor spreading and weak contractility. - A7 (with FLNa): increased spreading and contractility on stiff substrates.
60
What is mechanosensation dependent on?
Not just on FLNa, but on its dynamic interaction with FilGAP under strain.
61
What controls the speed of cellular contractile response to shear?
Shear stress magnitude — higher shear → faster FLNa unfolding → faster FilGAP release → faster Rho activation.
62
What technique quantifies Rho GTPase activity?
DORA-RhoB FRET sensor — High FRET = Rho-GTP (active); Low FRET = Rho-GDP (inactive).
63
What cellular behavior does strain modulate via FLNa?
It changes binding preferences, shifting balance between spreading (Rac) and contraction (Rho).
64
Why is Filamin called a "molecular strain gauge"?
Because its mechanical deformation alters signaling protein binding, enabling feedback control of cytoskeletal mechanics.
65
Which substrate stiffness is associated with neurogenic differentiation of MSCs?
0.1–1 kPa
66
Which stiffness range promotes myogenic differentiation of MSCs?
8–17 kPa
67
Which stiffness range promotes osteogenic differentiation of MSCs?
25–40 kPa
68
What happens to MyoD expression on myogenic matrices when treated with blebbistatin?
It decreases
69
What does blebbistatin do in MSC differentiation studies?
Blocks stiffness-mediated lineage specification by inhibiting non-muscle myosin II
70
Which transcription factor is associated with osteogenic fate on stiff matrices?
YAP/TAZ
71
What do RNA profiles show for MSCs on soft (0.1–1 kPa) matrices?
High expression of neurogenic markers
72
Which cell shape induced the most osteogenesis in Kilian et al.?
Star shape
73
Which aspect ratio resulted in highest osteogenesis?
0.16736111111111107
74
Which shape favored adipogenesis the most?
Flower shape
75
What is the role of NMMII in mechanosensing?
Transduces matrix stiffness into intracellular tension
76
What effect does nocodazole have on MSC differentiation?
Increases contractility and promotes osteogenesis
77
What is the effect of Cytochalasin D on MSC fate?
Reduces contractility and promotes adipogenesis
78
How does Y-27632 affect MSC differentiation?
Inhibits ROCK, leading to decreased osteogenesis and increased adipogenesis
79
What is the function of α5β1 integrin in mechanotransduction?
Couples the cytoskeleton to the ECM for sensing stiffness
80
What matrix material was used by Engler et al. to control stiffness?
Polyacrylamide gels
81
How does substrate stiffness affect focal adhesions?
Stiffer substrates promote larger focal adhesions
82
How does cell contractility relate to substrate stiffness?
Contractile stress increases with substrate stiffness
83
What is the impact of shape cues on MSC fate?
Shapes with higher aspect ratio and curvature promote osteogenesis
84
What happens when contractility is disrupted?
Shape- and stiffness-mediated differentiation is abrogated

BIEN 320 (6 decks)

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