ROLE OF RETINOIC ACID IN ACUTE LUNG INFLAMMATION AND NEUTROPHIL BIOLOGY
| dc.contributor.advisor | Singh, Baljit | en_US |
| dc.contributor.committeeMember | Paterson, Phyllis | en_US |
| dc.contributor.committeeMember | Forsyth, George | en_US |
| dc.contributor.committeeMember | Carr, Anthony | en_US |
| dc.contributor.committeeMember | Singh, Jaswant | en_US |
| dc.creator | CHANNABASAPPA, SHANKARAMURTHY | en_US |
| dc.date.accessioned | 2013-01-03T22:31:42Z | |
| dc.date.available | 2013-01-03T22:31:42Z | |
| dc.date.created | 2012-06 | en_US |
| dc.date.issued | 2012-06-28 | en_US |
| dc.date.submitted | June 2012 | en_US |
| dc.description.abstract | Acute lung injury is a severe clinical condition with high mortality both in animals and human beings. It is characterized by increased vascular permeability, accumulation of activated neutrophils, and diffuse damage to lung parenchyma. Neutrophil-derived toxic metabolites play a pivotal role in pathogenesis of acute lung injury. One of the ways to ameliorate acute lung injury is to modulate neutrophil functions. Retinoids are a group of compounds that include natural vitamin A and its synthetic derivatives. Role of retinoids in modulation of inflammatory and immune response is well established by previous studies. Effects of retinoids are mediated through two families of nuclear receptors; retinoic acid receptors (RAR) and retinoid X receptors (RXR). There are three subtypes in each family, namely; α, β, and γ. There are limited data available on the species-specific expression of various retinoid receptors and the role of retinoic acid in acute lung inflammation. Therefore, I designed a series of studies to evaluate the expression of retinoid receptors in many species and the role of retinoic acid in neutrophil apoptosis and acute lung inflammation. First, I determined the expression of retinoid receptors in intact lungs of cattle (n=2), pigs (n=2), dogs (n=2), mice (n=5), and humans (n=3). Normal and inflamed mouse lungs (n=5/group), normal human lungs (n=3) and inflamed human lungs (n=3) from patients who died of sepsis were also used. The expression was determined with multiple methods such as western blots, immunohistology, and immunoelectron microscopy. Normal lungs from all of the species showed differential expression of retinoid receptor subtypes in airway epithelium, vascular endothelium, alveolar/septal macrophages, and alveolar septum. In cattle pulmonary intravascular macrophages also showed expression of retinoid receptor subtypes. Intranasal treatment with E. coli lipopolysaccharide (LPS, 055:B5, 80 µg) resulted in increased expression of RXRα in airway epithelium of mouse lungs. Having examined the expression of retinoid receptors, I determined the effects of retinoic acid on spontaneous apoptosis of canine neutrophils using flow cytometry, caspase-3 assay, light microscopy, Terminal Transferase dUTP Nick End Labeling (TUNEL) Assay, and electron microscopy. Neutrophils were treated either with E. coli LPS (1 µg/ml) alone or with both LPS and retinoic acid (1 and 5 µM) for 12 hours and 36 hours. Treatment with LPS resulted in reduced number of apoptotic neutrophils compared to untreated cells (p<0.05). Simultaneous treatment with both LPS and retinoic acid abolished this effect. This suggests that retinoic acid reverses LPS-induced delay in spontaneous apoptosis of neutrophils. After studying the effects of retinoic acid on canine neutrophil apoptosis, I used a mouse model of acute lung injury to investigate the effects of retinoic acid on endotoxin-induced acute lung inflammation. Intranasal LPS treatment resulted in robust infiltration of neutrophils into the alveoli. Retinoic acid pre-treatment (10 mg/kg) 30 minutes before LPS treatment resulted in the reduction of total cell counts (p<0.05) and percentage of neutrophils in bronchoalveolar lavage (BAL) fluid at 6 hours after the LPS treatment. Protein content in BAL fluid was also decreased (p<0.05) following pre-treatment with retinoic acid. Lung tissues showed significantly reduced numbers of myeloperoxidase-positive cells in LPS-treated mice pre-treated with retinoic acid. However, retinoic acid had no effect on cytokine and chemokine levels (TNF-α, IL-1β, IL-10 and KC) in BAL fluid. Effect of retinoic acid on canine neutrophil chemotaxis was also studied under in vitro conditions. Activation with E. coli LPS (1 µg/ml) induced chemotaxis of neutrophils towards recombinant human interleukin-8 (p<0.05). However, retinoic acid did not exert inhibitory effects on the LPS-induced chemotaxis of canine neutrophils. Taken together, the data from these studies show that retinoid receptors are expressed in normal and inflamed lungs, and treatment with retinoic acid hastens apoptosis in activated neutrophils and inhibits neutrophil recruitment and permeability in lipopolysaccharide-induced acute lung inflammation. | en_US |
| dc.identifier.uri | http://hdl.handle.net/10388/ETD-2012-06-487 | en_US |
| dc.language.iso | eng | en_US |
| dc.subject | Acute lung inflammation, Retinoids, Retinoid receptors, Neutrophil apoptosis | en_US |
| dc.title | ROLE OF RETINOIC ACID IN ACUTE LUNG INFLAMMATION AND NEUTROPHIL BIOLOGY | en_US |
| dc.type.genre | Thesis | en_US |
| dc.type.material | text | en_US |
| thesis.degree.department | Veterinary Biomedical Sciences | en_US |
| thesis.degree.discipline | Veterinary Biomedical Sciences | en_US |
| thesis.degree.grantor | University of Saskatchewan | en_US |
| thesis.degree.level | Doctoral | en_US |
| thesis.degree.name | Doctor of Philosophy (Ph.D.) | en_US |