cystic fibrosis final poster
TRANSCRIPT
Amber Craghead ’15, Miriana Youkhana ’15 & Mariah Allen ‘15 Lake Forest College, Department of Biology: December 2014
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!Cystic Fibrosis (CF) is a lifelong, autosomal recessive genetic disorder defined by mutations within the cystic fibrosis membrane conductance regulator (CFTR), which is on chromosome 7 CFTR is an ABC transporter: requires ATP
Statistics • 30,000 people with CF in US [2] • Over 10 million Americans are carriers of CF [2] • 1,000 children born each year with CF [2]
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!!!!!!!!!!!• Previous studies found that airway surface layer (ASL) must be hydrated
to clear the mucus !!
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Gap in Knowledge
Hypothesis
References
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Future Studies
Methods
Acknowledgements
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Results
Discussion
Lake Forest College
Does the use of amiloride with HS therapy provide long lasting benefits to CF patients or does it reverse the effects of HS therapy? How does HS provide long
lasting benefits?
Examine potential role of salt intake in order to examine the link between high salt diets and inhalation therapy !Role of aquaporins in dehydrating the ASL [1] !Investigate HS therapy and antibiotic treatments against Pseudomonas aeruginosa to potentially find a better treatment for CF [4] !
Figure 1. In the normal CFTR channel, chloride ions are moved to the outside of the cell (1a). In a mutant CFTR channel, chloride ions are not able to move to the outside of the cell, causing a sticky mucus to form on the airway surface layer (1a,1b). This causes an increase in Na+ inside the cells, H2O follows, causing dehydration on the airway surface layer (1b). This results in CF which causes formations of thick mucus in the lungs, blocking airways (1c). CF causes the symptoms presented in figure 1d.
1a. b.
Symptoms
wheezing, shortness of breath
salty-tasting skin
excessive coughing
pneumonia, bronchitis
swelling in the belly
1c.
H0: Amiloride combined with HS therapy provides long lasting benefits in CF patients. !H1: Amiloride negates the long-lasting benefits of HS therapy in CF patients. !
•Cl-
Na+ ASL
x1d.
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This study focuses on amiloride negating the beneficial effects of HS therapy. HS therapy alone may provide long lasting benefits for treating patients with CF.
Figure 2. HS saline was found to increase ASL volume (middle panel) possibly through aquaporins. HS plus amiloride causes inhibition of epithelial Na+ channels, therefore increasing ASL. Amiloride eventually negates the benefits of HS therapy. This can possibly be attributed to amiloride inhibiting the aquaporin channels.
Focus 1: Effect of amiloride and HS on CF-hBE cells
Figure 4. The effect on CF-hBE by amiloride and HS on short circuit currents, transepithelial conductance, and capacitance. At 17 minutes, amiloride is added. At 23 minutes, 12 volumes of isosmotic solution was added to wash out amiloride. At 45 minutes, hypertonic solution was added. At 66 minutes, amiloride was added again.
Focus 3: Prolonged inhibition of INa with HS therapy
Introduction
Figure 6. Na+ recovery following CF-hBE exposure to HS conditions with and without amiloride. We would like to thank Dr. Ann B. Maine and the Department of Biology at Lake Forest College for their assistance in conducting this
research.
Focus 2: Effectiveness of HS therapy on CF-hBE
Figure 5. Near “thin-film” conditions: exposure to HS versus exposure to isosmotic solutions. HS produced 80% more inhibition than isosmotic conditions in INa+ at 30 minutes of incubation
•Figure 3. Experimental setup: 1. Ussing chamber 2. Continuously perfused Ussing chamber 3. Near “thin-film” conditions
3a.
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3b.
• Previously, amiloride was expected to decrease INa and was commonly used as a treatment for CF [1]
• Effects of hypertonic saline (HS) on sodium transport were found to be long-lasting and rapid [1]
• Amiloride reduced the inhibition of Na+ transport when combined with HS [1]
• hBE-cells exposed to HS alone produced increased inhibition of Na+ transport [1]
• Exposure to amiloride accelerated the recovery of ion transport following the HS-induced inhibition of sodium transport [1]
• Possible explanation of amiloride negating the effects of HS may be attributed to amiloride acting as a protective agent against HS-induced inhibition of Na+ transport [1]
• Therefore, understanding the effects of amiloride on HS-induced inhibition of sodium transport can result in developing effective aerosol therapies for CF patients [1]
Figure 7. Epithelial cells of healthy vs. CF patients
[1] Rasgado-Flores, H. et al. (2013). Effect of apical hyperosmotic sodium challenge and amiloride on sodium transport in human bronchial epithelial cells from cystic fibrosis donors. Am. J. Physiol. Cell Physiol (305):C1114-C1122. doi: 10.1152/ajpcell.00166.2013. [2] Rasgado-Flores, H. (2013). The remarkable treatment for cystic fibrosis: salt. DePaul University PowerPoint Presentation. [3] Liedtke, C. (2013). Understanding the cellular mechanism for inhaled saline therapy for patients with cystic fibrosis. Focus on ‘Effect of apical hyper osmotic sodium challenge and amiloride on sodium transport in human bronchial epithelial cells from cystic fibrosis donors’. Am J Physiol Cell Physiol. (305): C1096-C1095. doi: 10.1152/ajpcell.00250.2013. [4] Zemke, A. et al. (2014). Nitrite modulates bacterial antibiotic susceptibility and biofilm formation in association with airway epithelial cells. Free Radical Biology and Medicine. (77): 307-316. [5] http://www.biopta.com/uploads/Image/Ussing-Chamber-Set-Up.png [6] http://cyfbin.weebly.com/uploads/1/7/9/9/17995513/457139_orig.jpg
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Hypertonic saline therapy: removal of amiloride to provide long lasting effects for cystic fibrosis patients