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BMS 346- Epithelial physiology in health and disease > Lecture 3 > Flashcards

Lecture 3 Flashcards

1
Q

Airways Surface Liquid layer

A
  • Clearance mucous

- airways defence against respiratory pathogens

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2
Q

What is the Airways surface layer also known as ??

A

Periciliary layer

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3
Q

What is most important about the ASL/PCL?

A

Height is important for mucous clearance

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4
Q

what happens to the volume of the ASL as you go up the respiratory tract ?

A

As the respiratory structure gets larger the surface area goes down, so the volume decreases as you go up the respiratory tract

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5
Q

What is important about the height of the ASL/PCL?

A

Height of layer too low or high impacts the ability of lungs to move mucous

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6
Q

What are the to types of control of the ASL ??

A
  • Passive – mucous layer acts as a reservoir

* Active – Active ion transport controlling salt level in pcl

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7
Q

how do you test the effect of height on theASL/PCL?

A

Culture human airway epithelial cells - made liquid layer higher optimum

  • cells move it to the optimum - 7 microns
  • optimum in the lab is 14 microns - moving the dish back and forth to imitate movement of breathing in and out
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8
Q

Bumetanide

A

loop diuretic NKCC1 block CL secretion

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9
Q

Amiloride blocks what

A

ENac

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10
Q

Why is bumetanide used ?

A

to see how much Cl secretion is occurring

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11
Q

change in transepithelial potential tell us what

A

mechanism of epithelial transport changes over time- upreg or down reg
- transport properties change

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12
Q

bumetanide / amiloride experiment

A

at 48 hours
bumetanide- cl secretion more than double
amiloride- Na secretion halved

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13
Q

When height of layer is high ?

A

More Na absorption at steady sate more Cl secretion then Na absorption

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14
Q

What happens when height of layer goes up ??

A

if height of layer goes up cells respond by upregulating ENac function to bring height of layer back down

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15
Q

What happens if height of the layer drops ?

A

cells upregulate chloride secretion

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16
Q

Draw the cell model

A

see lecture notes

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17
Q

What is on the apical membrane of an eptihelial channel ?

A

ENac
CFTR
cilia

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18
Q

What is on the basolateral side of the epithelial cell in the airways?

A

3NA+ out 2K+ in
NA+ 2Cl- K + in (NKcc1)
K+ out

19
Q

Epithelial cell function

A

Na recycles across basolateral membrane
High CL in cell – open cftr secrete cl
Potassium channels sets driving force of NA across the cell

20
Q

Respiratory Syncitial virus

A

Respiratory pathogens disturb fluid balance respiratory tract
RSV = nasal congestion
bronchiolitis children
pneumonia adults
Major respiratory pathogen in children
-
RSv inhibits ENac after 1 hour incubation

21
Q

What animal model is used to show the effect of RSV?

A

Mouse trachea using SCC

22
Q

amiloride before exposure to virus

A

deflection - shift represents function of the epithelial Na channel

23
Q

exposure to RSV and the amiloride

A

inhibit ENac - Runny nose liquid

24
Q

3 main pathways of which viruses have an impact on epithelial N channels ?

A

Activation of PKC
Bind Glycoproteins
Bind to Glycolipids

25
Q

M1 model is a cell model for what

A

Principle cell line

26
Q

BIM inhibits what

A

PKC

27
Q

PPMP inhibits what

A

Inhibits binding to glycolipids

28
Q

M1 cell model and trachea experiment to show how RSV disrupts ENac ?

A

M1 cell
Mouse trachea - addition of BIM so pkc isnt active
control vs virus
if RSV working through activating PKC in the presence of BIM it cant do that.
tells us downstream of the virus and between the virus and ENac is the activation of PKC
- 2nd experiment
NA inhibits Glycoproteins - no change between RSV on its own and with Na - inhibition still seen- Glycoproteins therefore not important.
Final Experiment - M! cells -
PPMP inhibits GLycolipids
- inhibition lost - therefore RSV binds to glycolipids which activates PKC and inhibits ENac.

29
Q

How does RSV inhibit ENac?

A

RSV binds to glycolipids which activates PKC and inhibits ENac.

30
Q

Different viruses

A

work by inhibiting ENac but through different pathways

31
Q

influenza

A

inhibits ENac
Influenza kills ~ 36,000 individuals
influenza virion contains lots of glycoproteins- including Matrix protein M1 -
Haemagglutinin – binds to sialic acid residues – actn PKC & transient inhibition ENaC

32
Q

M2 in influenza

A

M2 – forms an acid activated, amantadine inhibited H+ channel, inserted into apical membrane host cell during infection
M2 involved long term regulation
- inhibits ENac

33
Q

overxpression of M2 and GFP in airway cells

A

inhibits ENac

  • impacts on open probability shown from single channel recordings
  • channels open lots in normal
  • Open much less in Overexpressio M2 GFP
  • M2 protein reducing opening probability
  • Reduced amount of ENac in membrane
34
Q

M2 promotes what process in ENac channels

A

Endocytosis - when infected

- proven by test on liddles syndrome - dont endocytose well so difference in ENac current not shown

35
Q

Liddles Mutants

A

Cant be endocytosed easily from the membrane

36
Q

Influenza and M2 mode of action

A

influenza virus- binding to the cell and introducing the M2 protein

37
Q

What remains down stream of the influenza virus

A

ROS and PKC shown using RFP control

  • Green presence of ROS
  • overexpress M2 with RFP - lots of red where green - increasing ROS in cells
  • one of the mechanism where M2 inhibits ENac
38
Q

What is one of the mechanisms M2 uses to inhibit ENac?

A

Increase ROS

39
Q

Glutathione

A

Antioxidant - oxygen scavenger

- reduces some of the inhibition of M2 on ENac

40
Q

block PKC

A

block response to M2

41
Q

How does the virus work ?

A

virus works by partially inserting M2 in membrane stimulate PKC and generate ROS which inhibits ENac

42
Q

Mutations in ENac gives rise to what condition

A

Pseudo hypoaldosteronism

43
Q

PHA 1

A

2 forms - autosomal dominant (Renal form) and autosomal recessive

44
Q

PHA1 autosomal recessive

A
  • frequent lower respiratory tract infections
  • ENac gene mutations in all subunits
  • multiple organs affected
  • permanent runny nose
  • LOF in ENac
  • Struggle to reabsorb sodium
  • Less negative potential
  • Height of ALS higher than optimum – difficult to remove mucos by cilia – increased risk of infection

BMS 346- Epithelial physiology in health and disease (13 decks)

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