sabah f iqbal, md 1,2, angela s benton, bas 1, alan m watson, phd, 1,2 mary c rose, phd 1,2, robert...
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DEXAMETHASONE SYNCHRONIZES ASTHMATIC RESPIRATORY EPITHELIUM REPAIR IN VITRO
Sabah F Iqbal, MD 1,2, Angela S Benton, BAS 1, Alan M Watson, PhD, 1,2 Mary C Rose, PhD 1,2, Robert J Freishtat, MD, MPH 1,2
1Children’s National Medical Center, Washington, DC2George Washington University, Washington, DC
Disclosure Statement The authors have documented that they
have nothing to disclose.
Background: Asthma Increasing prevalence and severity
14.7 million annual lost school days* 196,000 hospital admissions** 730,000 ED visits**
*National Health Interview Survey, 2002**National Hospital Medical Care Survey, 2002
1980 2002
35/1000 83/1,000*
The role of glucocorticoids (GC)
Inflammation
Remodeling
?
Circadian Rhythm
Hypothesis
• Injured asthmatic respiratory epithelium is characterized by asynchronous regeneration and will resynchronize in the presence of glucocorticoids
Normal Cell Cycle
Experimental Design: ALI
Primary differentiated HBE cells at an air-liquid interface (ALI) were used (Mattek, Ashland, MA)
‘Gold standard’ model in airway epithelial research as it mimics the human airway
Heterogeneous mixture of ciliated, goblet and basal cells
Mucus layer
Stimuli
Goblet cell Ciliate
d cellCollage
n
Experimental Design:Timeline
-26 -24 -2 0 22 24 46 48
+/- DEX
Mechanical Wound
Photo
+/- DEX
+/- DEX
+/- DEX
Asthma - DEX
Normal - DEX
Normal + DEX
Asthma + DEX
0 48 hours
Bright –field microscopy (16X)
1mm
Experimental analysis Flow cytometry
BrdU (bromodeoxyuridine) Cellular proliferation/mitosis
7-AAD (7-amino-actinomycin-D) DNA label/cell cycle phase
Normal Cell Cycle
G2
G1
Cell cycle analysis by flow cytometry
Summary: Proliferating normal cells were more
likely to be found in G1 of the cell cycle Normal cell cycling
Proliferating asthmatic cells were more evenly distributed among G1 and S Consistent with asynchronous cell cycling
DEX, given at doses analogous to endogenous steroids, re-synchronized the cell cycle
Discussion Chronotherapeutic trials (Pincus et al, 1997)
Single late-afternoon dose of GC improves lung function, symptoms, and inflammation
Rethinking glucocorticoids
Inflammation
Remodeling
?
Future directions DEX at pharmacologic doses may
correct for the asynchronous processes and allow normal rapid healing
Use nasal epithelium cells from pediatric patients with asthma and in controls
Acknowledgements: K12 Mentoring Committee:
Mary Rose Ph.D. Robert Freishtat M.D. Eric Hoffman Ph.D. Anamaris Colberg-Poley Ph.D. Stephen Teach M.D.
Angela Benton Alan Watson Ph.D. Teresa Hawley Ph.D. Funded by the National Institutes of Health -
National Heart, Lung, and Blood Institute grant number K12HL090020