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Bio lab 1 > Efferoctyosis > Flashcards

Efferoctyosis Flashcards

(55 cards)

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1
Q

Efferocytosis

A

Removal of STRICTLY apostolic cells by phagocytes

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2
Q

What are we doing in this experiment

A

Using cultured mouse macrophages to investigate the efficiency of macrophage engulfment of plastic beads. Two different effectors will be tested to see if the rate of phagocytosis can be increased

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3
Q

Phagocytosis

A

Macrophage engulfment

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4
Q

Effector

A

A molecule that binds to a protein (often an enzyme) and affects function of protein. These molecules can enhance phagocytosis

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5
Q

What is the purpose of lab this week

A

To demonstrate that macrophages can detect and engulf foreign substances. In this case, the macrophages will engulf plastic beads so we can know that we have a good model system for next week. Also, we will alter the concentration of different effectors to alter efficiency of process. Lastly, we willtitrate the ideal concentration of effector.

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6
Q

Apoptosis

A

Programmed cell death

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7
Q

Necrosis

A

Cell rupture

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8
Q

Steps when the apoptotic cell has clearance

A

Before it becomes necrotic, live macrophage efferocytoses dead cell. No release of cell contents

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9
Q

Resolution

A

When dead cell is consumed by macrophage and there is no release of cell contents

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10
Q

What causes inflammation

A

Cell being necrotic and release contents

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11
Q

Phagocytes

A

Broader term. Cells that engulf and digest foreign invaders (bacteria) and other cells (cancer cells and dead cells)

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12
Q

M1 macrophage is pro inflammatory or pro resolving

A

pro inflammatory

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13
Q

M2 macrophage is pro inflammatory or pro resolving

A

pro resolving

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14
Q

Cytokines

A

Signaling molecules that can be pro or anti inflammatory. They are small protein(s) significant in immune response signaling

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15
Q

M1 has incresased/decreased bactericidal activity, pro/anti inflammatory cytokines and high/low efferocytosis

A

Increased, pro, low

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16
Q

M2 has incresased/decreased bactericidal activity, pro/anti inflammatory cytokines and high/low efferocytosis

A

Decreased, anti, high

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17
Q

How does live macrophage deal with toxins after engulfing dead cell

A

Efflux: move toxins out
Metabolism: break them down

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18
Q

Steps of efficient dead cell clearance

A

Find me signal release from dead cell and phagocyte recruitment
Exposed eat me signal on dead cell and phagocytic receptors recognize signals
Actin filament of phagocyte push out plasma membrane to engulf dead cell
Dead cell digests and phagocyte releases cytokines that are pro inflammatory or pro resolving

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19
Q

How does efferocytosis help in resolution and repair

A

Prevents necrosis, which terminates inflammatory response, encourages body to not attack own cells, activation of pro resolving pathways

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20
Q

Efferoctyosis defective diseases it can cause

A

Autoimmune, atherosclerosis, myocardial infarction, obesity and diabetes

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21
Q

IL

A

Largest kind of cytokine

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22
Q

SPMs

A

Specialized proresolving mediators (effectors). These enhance efferoctyosis

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23
Q

SPMs do what

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A

Resolve tissue inflammation

24
Q

Kind of effectors we’re using

Study These Flashcards
A

Resolvins, IL-10, dexamethasone

25
Cell plasticity
Macro expresses genes for both m1 and m2 macrophages
26
LPS
Pushes macrophages toward m1 by changing gene expression. So then it can engulf bacteria of beads better
27
IL-4
Makes macrophage more like m2 so it can engulf dead cells better
28
Steps to pipette
To set at right volume: Spin dial at top of pipette Steps: Put tip on pipette Open tube Push down on plunger to first notch Put tip in liquid, pull plunger up Take new tube Put pipette near bottom of tube Push plunger to first stop to get rid of liquid then 2nd stop Eject tip in waste
29
How to draw up liquid from well
Tilt dish at 30 degrees, put tips at bottom, draw up liquid
30
EvOS microscope
Identifties problems after plate reader data. Looks at confirmation that only engulfed beads are left
31
Plate reader
Scans for fluorescence at each well by exciting and emitting at wavelengths
32
Phosphatidylserine
Eat me signal
33
Leukotrienes
stimulate the production of proinflammatory cytokines
34
When resolvin:Leuko ratio is high, are there more m1 or m2 macrophages. And how is plaque affected
More resolvin=higher resolution, less inflammation=more m2 macrophages and decreased plaque size
35
Inflammasome
cytosolic proteins (composed of many subunits) responsible for the activation of the inflammatory response
36
Gas6
Bridging molecule between eat me signal and mertk (a scavenger receptor found on surface of macrophage)
37
Mertk
Scavenger receptor on macrophage
38
How does efferocytosis become more defective. By what factor
Age.
39
How does cell engulf dead cell while in plate
The cell pushes out protrusions at its leading edge • Protrusions adhere (anchor) to the “crawling” surface • Rest of cell drags itself forward by traction on the anchorage points Cell stays on plate
40
Do spms and cytokines bind to different cell surface receptors
Yes
41
Purpose of lab
To see how different concentrations of inhibitors affect the inhibition of efferocytosis. Also seeing how combinations of different resolvins enhance efferocytosis
42
CD36
Scavenger receptor
43
Thbs-1
bridging protein for CD36
44
CD47
“Don’t eat me” signal
45
Annexin A1
inhibits prostaglandin synthesis – anti-inflammatory effect
46
FPR2
resolvin receptor
47
IL-10
anti-inflammatory cytokine
48
TNF-alpha
pro-inflammatory cytokine
49
TGF-beta
anti-inflammatory cytokine
50
Rac1
activates actin polymerization/reorganization
51
LC-3
autophagy protein that is recruited to the phagosome (main marker for phagocytosis)
52
Rubicon
negative regulator of autophagy/phagocytosis
53
ATG16L
downstream LAP machinery
54
Control/housekeeping gene for experiment 4
Glyceraldehyde 3-phosphate dehydrogenase (GAPDH)
55
Formula for relative quantification (RQ)
2^(-🔼🔼CT) Triangle means delta CT is threshold cycle . Aka, subtract first 🔼CT from 2nd 🔼CT, switch sign, and raise 2 to this

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