The Respiratory System and CF

Question 1

Airways are a series of branching tubes

Actual business of gas exhange happens at the _____

Answer 1

Acinus

*group term for alveoli and supporting cells

• Respiratory bronchioles
• Alveolar ducts
• Alveolar sacs
• Alveoli

Question 2

Stages of Lung Development

1. _____ (0-6 weeks gestation)
2. _____ (6-16 weeks gestation)
3. _____ (16-26 weeks gestation)
4. _____ (26-36 weeks gestation)
5. _____ (2 years to adult)

Answer 2

1. Embryonic (0-6 weeks gestation)
2. Pseudoglandular (6-16 weeks gestation)
3. Canalicular (16-26 weeks gestation)
4. Sacular/Alveolar (26-36 weeks gestation)
5. Postnatal (2 years to adult)

Question 3

Lung Development - Embryonic Stage

• Lung bud appears day ______
• Ventral _____ of the primitive _____
• _____ origin
• _____ branching of the airways into surrounding _____

Answer 3

• Lung bud appears day 26-28
• Ventral outpouching of the primitive foregut
• Endodermal origin
• Proximal branching of the airways into surrounding mesoderm

Question 4

Lung Development - Pseudoglandular Stage

• Weeks ___
• Airways develop (branch) to the level of the ______ (pre-___ bronchi)
• ___ generations of airways

Answer 4

• Weeks 6-16
• Airways develop (branch) to the level of the terminal bronchioles (pre-acinar bronchi)
• 16 generations of airways

Question 5

Lung Development - Canalicular Stage

• Weeks ___
• _____ region develops
• Thinning of the ____ (allows gas exchange)
• Type _____ develop (develop into 2 types of cells in alveoli)

Answer 5

• Weeks 16-26
• Acinar region develops
• Thinning of the peripheral epithelium (allows gas exchange)
• Type I and II pneumocytes develop (develop into 2 types of cells in alveoli)

Question 6

Lung Development - Sacular/Alveolar Stage

• Weeks ____
• Sacules form into the ____ and ____
• Marked ____ in ____ tissue
• Sacules become ____ ______
• Sacules and alveoli form ______ ______ of ______ by ______ (division)

Answer 6

• Weeks 26-36
• Sacules form into the alveolar ducts and alveoli
• Marked decrease in interstitial tissue
• Sacules become thin walled
• Sacules and alveoli form futher generations of alveoli by septation (division)

*Before 30 weeks, no alveoli, but there are alveolar ducts so gas exchange can occur

Question 7

Functions of the lung

Q) Which functions of the lung are affected in CF?

Answer 7

1) Defence
2) Gas Exchange

Question 8

Q) Physical (airway) vs Cellular (alveolar) defence mechanisms

Answer 8

• Physical (airway)
  
  • Upper airway filter (particularly in nose, filter out large particles
  • Reflexes
    
    • Sneeze
    • Cough
  • Mucociliary escalator*
• Cellular (alveolar)
  
  • Phagocytes e.g. alveolar macrophages
  • Immunological

Question 9

Q) What is the Mucociliary Escalator?

Answer 9

• At airway surface epithelium, Airway surface liquid (ASL) has 2 layers
  
  • Periciliary layer
  • Mucus gel layer
• Mucus produced by secretory cells incl. Goblet cells
• Mucus propelled by cilia
  
  • Beat 10-15 times per sec
  • 5mm per min

Question 10

What parameters are used to measure lung function? (3 points)

Answer 10

• Volume (litres)
• Flow (L/sec) = Volume over time
• Pressure

Question 11

Lung Volumes and Capacities

Summary slide

Answer 11

Question 12

Forced Expiration

Measured using a spirometer

Maximal inspiration followed by fast exhalation to residual volume

Q) What is FEV1 and FVC, and what do they tell us about the lung?

Answer 12

• FEV1 = Forced expiratory volume
  
  • Volume exhaled in first second
  • Tells us about the flow of the airways
• FVC = Forced vital capacity
  
  • Total volume exhaled
  • Tells you about the size of the lungs

Question 13

FEV1 vs FVC graph

Answer 13

Question 14

Normal Flow-volume curve

TLC = total lung capacity

RV = residual volume

Answer 14

Question 15

Intrathoracic airway obstruction

• Obstructive lung disease
• scooped out appearance which reflects obstruction within the airways

Answer 15

Question 16

Pulmonary Diease in CF

• Significant amount of ___ and ____
• _____ exacerbations
  
  • increased cough and sputum production
  • increased ____ - S.O.B
  • Lethargy/poor appetite
  • Reduced _____** (in ___, which is looked out for when testing)
• ______ decline in lung function
• Respiratory failure
• Death/transplatation

Answer 16

• Significant amount of morbidity and mortality
• Pulmonary exacerbations
  
  • increased cough and sputum production
  • increased dyspnoea - S.O.B
  • Lethargy/poor appetite
  • Reduced lung function** (in FEV1, which is looked out for when testing)
• Progressive decline in lung function
• Respiratory failure
• Death/transplatation

Question 17

CFTR and lung disease

• CFTR mutations >>>>> Lung disease
  
  • Exact mechanism unknown (how CF variants cause LD)
• Complex relationship between ___ and ___ that impacts on ___ production and ___ obstruction
• ___ infections may play a role in the initiation of these events

Answer 17

• Complex relationship between host defense and airway microbiology that impacts on sputum production and airflow obstruction
• Viral infections may play a role in the initiation of these events

Question 18

CFTR and Lung Disease

CFTR expressed on several cell types

• (Highly expressed) ____ plasma membrane of ______ _____ cells in airways; regulates ion transport
  
  • ____, ____
• _____ cells of submucosal glands
• Alveolar epithelial type ___ cells (produce ____)
• Cells of the ____ system
  
  • Alveolar m_____
  • N_____

Answer 18

• (Highly expressed) Apical plasma membrane of ciliated epithelial cells in airways; regulates ion transport
  
  • chloride, bicarbonate
• Serous cells of submucosal glands
• Alveolar epithelial type II cells (produce surfactant)
• Cells of the immune system
  
  • Alveolar macrophages
  • Neutrophils

Question 19

Airway Surface Dehydration

Widely held theory based on studies of cultured human airway epithelia (also mouse fibroblasts and dog kidney-cell lines)

Q) Give a brief outline of the theory

Answer 19

Defective CFTR
>
Increased NA+/Cl- absorption
>
Increased H2O absorption
>
Dehydrated ASL

>>
impaired mucociliary clearance

>
Infection and inflammation
=
Lung disease

Question 20

Ion Transport in Normal Lung

CFTR coordinates modulation of ASL by:

1. Inhibiting ENac
2. Secreting Cl-

Describe/elaborate on the mechanisms

**crucial for modulating the viscosity and amount of fluid in that ASL that the cilia are surrounded by

Answer 20

CFTR coordinates modulation of ASL by:

1. Inhibiting ENac
   
   • Reduce Na+ absorbed into cell
   • Water follows Na+
   • ASL stays hydrated
2. Secreting Cl-
   
   • Cl- ions secreted out into ASL

Question 21

Ion transport in CF lung

2 functions of CFTR lost:

1. ENaC not inhibited
2. Cl- ions not secreted

Elaborate/describe mechanisms

Answer 21

2 functions of CFTR lost:

1. ENaC not inhibited
   
   • increased Na+ absorption into cell
   • water follows sodium (not salt)
   • ASL dehydrated
2. Cl- ions not secreted
   
   • Increase Na+, Cl-, H20 absorption
   • ASL volume decreased

Question 22

ASL volume hyperabsorption has important consequences for both the Periciliary liquid (PCL) and mucus layer:

What are the consequences?

Answer 22

• Volume depletion of PCL
  
  • Failure of ciliary beating
• Absence of lubrication
  
  • Adherent mucus plaque
• >>>Promotes chronic infection

Question 23

Airway surface dehydration theory

Some conflicting studies:

• Airway epithelia in newborn pigs, ferrets, and rats with CF do not hyperabsorb sodium
• But…

Answer 23

• Contradictory study of airway epithelia in humans with CF also showed no evidence of increased sodium absorption

Question 24

Reduced pH of ASL

• CFTR also conducts bicarbonate (HCO3-)
  
  • so…

Answer 24

• Loss of CFTR-dependent bicarb secretion
• pH of airway-surface liquid decreases, resulting in impaired antibacterial activity
  
  • lower pH (more acidic environment) = promote bacterial growth

Question 25

Abnormal Mucus * Mucus produced in ____ glands = CFTR \_\_\_\_ * defective _____ \_\_\_\_ alters property of mucus * In porcine model, defective CFTR prevents the ____ of ____ \_\_\_\_ from glands * ____ of mucus to gland ducts, hindering ____ \_\_\_\_ * Contents of degraded _____ (elastases, proteases, DNA and oxidases) increase the _____ of the mucus, further affecting its transportability

Answer 25

* Mucus produced in submucosal glands = CFTR abundant * defective bicarbonate secretion alters property of mucus * In porcine model, defective CFTR prevents the release of mucus strands from glands * tethering of mucus to gland ducts, hindering mucociliary transport * Contents of degraded neutrophils (elastases, proteases, DNA and oxidases) increase the viscosity of the mucus, further affecting its transportability

Question 26

Abnormal Microenvironment Due to an _____ environment, resulting from: * Thick mucus plaques adherent to epithelial surface * increased ____ \_\_\_\_\_ by CF epithelia (cilia fail to beat so beat harder) Lead to higher risk of infection because of: * Ideal environment for growth of certain bacteria * Pseudomonas converts to _____ \_\_\_\_\_ mode of growth

Answer 26

Due to an anaerobic environment, resulting from: * Thick mucus plaques adherent to epithelial surface * increased oxygen consumption by CF epithelia (cilia fail to beat so beat harder) Lead to higher risk of infection because of: * Ideal environment for growth of certain bacteria * Pseudomonas converts to anaerobic biofilm mode of growth

Question 27

CF and Inflammation * CFTR dysfunction promotes ____ \_\_\_\_ infection * Results in \_\_\_\_, predominantly ____ inflammation * CFTR dysfunction may also directly affect airway immunity * _______ immune pathways * Increased production of ________ mediators

Answer 27

* CFTR dysfunction promotes chronic airway infection * Results in exaggerated, predominantly neutrophilic inflammation * CFTR dysfunction may also directly affect airway immunity * Dysregulated immune pathways * Increased production of pro-inflammatory mediators

Question 28

Dysregulated immune pathways in CF (Summary) Loss of CFTR function leads to * Increased activation of nuclear factor-kappa B (NFkB) * Increased ROS production (and decreased antioxidant defense) * ER stress response * Ceramide accumulation/cholesterol dysfunction * Abnormal basal lipid metabolism * Defective interferon signaling

Answer 28

Question 29

Dysregulated immune pathways in CF Loss of CFTR function leads to * Increased activation of nuclear factor-kappa B (NFkB) * * * * *

Answer 29

Increased activity of transcripton factors e.g. Nuclear factor-kappa B (NFkB) * Increase secretion of pro-inflammatory cytokines e.g. IL-8, IL-6, TNFa, IL-1B * Constituently active NFkB * Enhanced IL-8 production

Question 30

Dysregulated immune pathways in CF Loss of CFTR function leads to * * Increased ROS production (and decreased antioxidant defense) * * * *

Answer 30

* CF ASL contains high levels of ROS * Neutrophils major contributor * Released as part of respiratory burst * Airway epithelium * Exacerbated by decreased antioxidant defense * e.g. decreased glutathione metabolism * Oxidative stress invigorates/creates a proinflammatory environment (defective autophagy) and correlates with inflammatory markers (TG2) in CF

Question 31

Dysregulated immune pathways in CF (Summary) Loss of CFTR function leads to * * * ER stress response * * *

Answer 31

ER sequestration of the misfolded F508del * Accumulation of abnormally folded CFTR in ER results in unfolded protein responses * Trigger 'cell stress' and apoptosis

Question 32

Dysregulated immune pathways in CF (Summary) Loss of CFTR function leads to * * * * Ceramide accumulation/cholesterol dysfunction * *

Answer 32

Ceramide accumulation/cholesterol dysfunction * Ceramide accumulation may further augment pulmonary inflammation by * inducing apoptosis with subsequent deposition of DNA in airways * Ceramide is a breakdown product of sphingomyelin found in plasma membrane and in endolysosomal compartments * Results in: * Abnormal lysosome function * Abnormal receptor signalling

Question 33

Dysregulated immune pathways in CF (Summary) Loss of CFTR function leads to * * * * * Abnormal basal lipid metabolism *

Answer 33

Abnormal basal lipid metabolism * Fatty acid metabolism skewed towards * increased production of arachidonic acid (AA) and leukotriene B4 (LTB4) * Decreased docosahexaenoic acid (DHA) and lipoxin A4 (LXA4) * Thus get impaired resolution of inflammation

Question 34

Dysregulated immune pathways in CF (Summary) Loss of CFTR function leads to * * * * * * Defective interferon signaling

Answer 34

Blunting of interferon signaling * Defective type I INF response * Interferes with eradication of bacteria * increase susceptibility to viral infections

Question 35

Innate immunity abnormalities Pathogens in the airway are sensed in the airway by the innate immune system * Innate immune system senses ____ \_\_\_\_ ____ using pattern recognition receptors * ____ \_\_\_\_ (TLRs) * ______ receptors * ___ receptors * ___________________ (NOD)-like receptors

Answer 35

Pathogens in the airway are sensed in the airway by the innate immune system * Innate immune system senses conserved molecular patterns using pattern recognition receptors * Toll-like receptors (TLRs) * Complement receptors * Fc receptors * Nucleotide-binding oligomerization domain family (NOD)-like receptors

Question 36

Toll-like Receptors * Expressed on a variety of cells * especially _____ cells, \_\_\_\_\_\_ * Majority recognise bacterial patterns * TLR4 senses _____ (LPS) * TLR5 senses ____ (predominant in B cepaciae) * May be modifier gene as may change response to PsA and BC * ___ poorly displayed on surface of CF cells and ______________ to stimulation

Answer 36

* Expressed on a variety of cells * especially dendritic cells, neutrophils ​ * Majority recognise bacterial patterns * TLR4 senses lipopolysaccharide (LPS) * TLR5 senses flagellin (predominant in B cepaciae) * May be modifier gene as may change response to PsA and BC * TLR4 poorly displayed on surface of CF cells and fails to respond well to stimulation

Question 37

Neutrophils are first and main inflammatory cell found in CF airways 3 main anti-bacterial weapons * \_\_\_\_\_\_ * ____ release (protease rich) * ___ \_\_\_\_\_ (DNA fibres that entangle, immobilise and kill pathogens)

Answer 37

3 main anti-bacterial weapons * Phagocytosis * Granule release (protease rich) * NET formation (DNA fibres that entangle, immobilise and kill pathogens)

Question 38

Neutrophils are first and main inflammatory cell found in CF airways Release proteases which may be harmful to our underlying cell structure (3 types proteases):

Answer 38

Release: * Serine proteases * e.g. elastase * Physiological role in tissue remodelling, chemotaxis, microbial killing * Are tightly regulated by protease inhibitors e.g. alpha-1-antitrypsin * Matrix mettaloproteases * Calgranulins: proinflammatory proteins * Increased in CF sputum

Question 39

Role of other immune cells * Macrophages * Dedritic cells **ELABORATE for marks**

Answer 39

* Macrophages * increased numbers * TLR * altered function: impaired efferocytosis (removal of dying cells) * Intrinsic effects * Dendritic cells * TLR * reduced APC function * Decreased levels of MHC Class II receptors * \*expression is dependent on TLR signalling

Question 40

Role of other immune cells * T cells * Invaraint natural killer T cells **Evaluate for marks**

Answer 40

* T cells * Skewed towards Th2/Th17 immune response * Counter-regulatory T cells (Tregs) may be reduced in CF * Invaraint NK T cells * Dysregulated in C, those which kils its own cells * Upregulated in absence of CFTR

Question 41

Vicious cycle of CF lung disease and respiratory failure Summary slide

Answer 41

Question 42

Pathology of CF lung disease ## Footnote **Please describe for 3 marks**

Answer 42

* Infection, inflammation and obstruction of airways * lead to dilation and damage of airways * Bronchiectasis * Dilation of airways * Thickened airways obstructed by mucus

Question 43

Histology of CF lung disease

Answer 43

Photomicrograph shows: * endobronchial and peribronchial neutrophilic inflammation

Question 44

Computerised Tomography (CT) Main takeaway??

Answer 44

Dilated bronchioles (bronchiectasis)

Question 45

Pathogens \*Fungi as well - aspergilis (pro-inflammatory) - candida (non-inflammatory)

Answer 45

* S. Aureus * common pathogen envountered in life * P. aeruginosa * converts to anaerobic biofilm mode of growth * Unusual * Not very common pathogen * Increases during childhood, then significantly increases during adulthood

Question 46

FEV1 %predicted vs Age \*if you get 100% you are average

Answer 46

At FEV1 \<30%, have 50% chance of dying in 2 years \>\>\>makes you eligible for lung transplant