[Show abstract][Hide abstract] ABSTRACT: To determine whether a recently developed murine model of fungus-induced sinonasal inflammation demonstrated alterations in ciliary activity and expression of inflammatory cytokines.
A prospective randomized controlled study of rhinosinusitis after fungal antigenic sensitization was performed with intraperitoneal aspergillus antigen injection followed by intranasal antigen challenge for 4 weeks. Saline solution was used in a parallel fashion for control animals.
Six mice were used to validate the model. Additional 15 mice were used for ciliary beat frequency (CBF) analysis and cytokine expression with multiplex technology. Mean values for degree of inflammation, secretory hyperplasia, CBF, and cytokine expression were compared.
Histologic analyses demonstrated dense chronic inflammation in aspergillus-challenged animals versus sparse inflammatory cells in controls. Significant differences in mean of aspergillus-challenged versus control animals were observed in degree of inflammation (P < 0.01), secretory hyperplasia (P < 0.01), CBF (P < 0.00002), IL-1alpha (P < 0.0002), IL-1beta (P < 0.0003), IL-4 (P < 0.02), TNF-alpha (P < 0.02), and RANTES (P < 0.01).
Alteration in baseline CBF accompanied by increased expression of specific inflammatory cytokines was observed in aspergillus-challenged mice.
Full-text · Article · Nov 2008 · Otolaryngology Head and Neck Surgery
[Show abstract][Hide abstract] ABSTRACT: Retinoic acid (RA) has been shown to enhance ciliary ultrastructure in regenerated sinus mucosa compared with controls. However, the functional status of the regenerated cilia has not been studied. Our objective was to evaluate the ciliary beat frequency (CBF) of regenerated sinus mucosa treated with topical RA in a rabbit model.
Twelve rabbits underwent bilateral surgical stripping of the maxillary sinuses, followed by treatment with RA gel in the right side and an inert gel control in the left side. The rabbits were then killed at either 2 or 4 weeks, and CBF analysis of the regenerated mucosa was performed. Three unoperated rabbits were used to establish normative CBF data.
Functional cilia were recovered from 11/12 RA-treated sinuses and 12/12 gel control sinuses. At 2 weeks postoperatively, the RA-treated sinuses showed an average CBF of 19.78 Hz, which was statistically comparable with the normal unoperated controls (p < 0.26). The inert gel-treated mucosa showed a CBF of 29.24 Hz, which was significantly elevated compared with normals (p < 0.05). At 4 weeks, ciliary activity persisted, but both RA-treated sinuses and gel controls showed elevated CBF compared with normals (p < 0.03).
Topical RA placed in a demucosalized maxillary sinus yields functional cilia. RA appears to have a normalizing effect on CBF early in the mucosal wound healing process compared with control. This effect appears to be mitigated in later stages of wound healing. RA may be beneficial in enhancing morphological and functional aspects of regenerating cilia.
No preview · Article · May 2008 · American Journal of Rhinology
[Show abstract][Hide abstract] ABSTRACT: Transient receptor potential (TRP) channels are a novel class of nonvoltage gated membrane cation channels that can be activated by mechanical stimulation and temperature change. Recently, TRP vanilloid type 4 (TRPV4) has been implicated in detecting viscosity changes in fallopian tube epithelial cells and inducing a compensatory response in ciliary activity and, as such, represents a possible molecular trigger for modulating respiratory ciliary activity. Thus, the goal of this study was to establish the expression pattern of TRPV4 in human sinonasal mucosa and determine whether expression is altered in chronic rhinosinusitis (CRS).
Sinus mucosal biopsy specimens were obtained from patients with CRS, CRS with nasal polyps (NPs), and healthy controls. TRPV4 mRNA and protein expression were confirmed by reverse transcriptase polymerase chain reaction (RT-PCR) and immunoblot analysis, respectively. TRPV4 gene expression was measured next using quantitative RT-PCR. Immunofluorescence was performed on sinus mucosal explants and respiratory epithelial air-liquid interface cultures to localize cellular expression.
TRPV4 mRNA and protein were expressed in all samples. There was a statistically significant increase (p < 0.05) in TRPV4 gene expression in nonpolypoid CRS patients, but no difference in CRS with NP. Dual label immunofluorescence showed TRPV4 expression to be mutually exclusive of ciliated cells.
Although TRPV4 represents an ideal molecular trigger for ciliary modulation, absent expression of the channel in ciliated cells precludes this function. However, altered expression of the channel in CRS and presumed expression of TRPV4 in secretory cells of the mucosa indicate a potential role in mucus homeostasis and CRS pathogenesis.
No preview · Article · Jan 2008 · American Journal of Rhinology
[Show abstract][Hide abstract] ABSTRACT: The sense of smell can be damaged by airborne xenobiotics, including aerosolized heavy metals, reflecting the direct exposure of its receptors to the outside environment.
To determine whether professional welders working in confined spaces exhibit olfactory dysfunction. To determine whether such dysfunction, if present, is related to indices of metal exposure associated with welding, as well as measures of neurologic and neuropsychological function.
The University of Pennsylvania Smell Identification Test (UPSIT) and a battery of neurologic and neuropsychological tests were administered to 43 welders who worked for 1 to 2 years on the San Francisco/Oakland Bay Bridge. Blood levels of Mn, Fe, Cu, and Pb were obtained.
Relative to matched controls, the welders had significantly lower UPSIT scores, with a mean (SEM) of 29.51 (0.90) for welders and 36.55 (0.88) for controls. Eighty-eight percent scored below their individually matched controls. As in idiopathic Parkinson disease, the welders' olfactory test scores were unrelated to a broad spectrum of neurologic and neuropsychological test measures, as determined by principal components analysis. Although blood levels of Mn were associated with the time spent working on the bridge, workers with the highest Mn blood levels exhibited better olfactory function than those with the lowest Mn blood levels. The basis of this paradox, which has been observed previously, is unknown.
Professional welders may be at risk for loss of smell function, although such loss seems to be unrelated to neurologic and neuropsychological test performance.
[Show abstract][Hide abstract] ABSTRACT: Air-liquid interface models using murine tracheal respiratory epithelium have revolutionized the in vitro study of pulmonary diseases. This model is often impractical because of the small number of respiratory epithelial cells that can be isolated from the mouse trachea. We describe a simple technique to harvest the murine nasal septum and grow the epithelial cells in an air-liquid interface. The degree of ciliation of mouse trachea, nasal septum, and their respective cultured epithelium at an air-liquid interface were compared by scanning electron microscopy (SEM). Immunocytochemistry for type IV beta-tubulin and zona occludens-1 (Zo-1) are performed to determine differentiation and confluence, respectively. To rule out contamination with olfactory epithelium (OE), immunocytochemistry for olfactory marker protein (OMP) was performed. Transepithelial resistance and potential measurements were determined using a modified vertical Ussing chamber SEM reveals approximately 90% ciliated respiratory epithelium in the nasal septum as compared with 35% in the mouse trachea. The septal air-liquid interface culture demonstrates comparable ciliated respiratory epithelium to the nasal septum. Immunocytochemistry demonstrates an intact monolayer and diffuse differentiated ciliated epithelium. These cultures exhibit a transepithelial resistance and potential confirming a confluent monolayer with electrically active airway epitheliumn containing both a sodium-absorptive pathway and a chloride-secretory pathway. To increase the yield of respiratory epithelial cells harvested from mice, we have found the nasal septum is a superior source when compared with the trachea. The nasal septum increases the yield of respiratory epithelial cells up to 8-fold.
[Show abstract][Hide abstract] ABSTRACT: Air-liquid interface cultures using murine tracheal respiratory epithelium have revolutionized the in vitro study of airway diseases. However, these cultures often are impractical because of the small number of respiratory epithelial cells that can be isolated from the mouse trachea. The ability to study ciliary physiology in vitro is of utmost importance in the research of chronic rhinosinusitis (CRS). Our hypothesis is that the murine nasal septum is a better source of ciliated respiratory epithelium to develop respiratory epithelial air-liquid interface models.
Nasal septa and tracheas were harvested from 10 BALB/c mice. The nasal septa were harvested by using a simple and straightforward novel technique. Scanning electron microscopy was performed on all specimens. Cell counts of ciliated respiratory epithelial cells were performed at one standard magnification (1535x). Comparative analysis of proximal and distal trachea, midanterior and midposterior nasal septal epithelium, was performed.
Independent cell counts revealed highly significant differences in the proportion of cell populations (p < 0.00001). Ciliated cell counts for the trachea (106.9 +/- 28) were an average of 38.7% of the total cell population. Nasal septal ciliated epithelial cells (277.5 +/- 16) comprised 90.1% of the total cell population.
To increase the yield of respiratory epithelial cells harvested from mice, we have found that the nasal septum is a far superior source when compared with the trachea. The greater surface area and increased concentration of ciliated epithelial cells has the potential to provide an eightfold increase in epithelial cells for the development of air-liquid interface cultures.
No preview · Article · Sep 2007 · American Journal of Rhinology
[Show abstract][Hide abstract] ABSTRACT: There is increasing evidence that bacterial biofilms are present within the sinonasal mucosa of patients with medically recalcitrant chronic rhinosinusitis. The antibiotic concentrations needed to eradicate biofilms are much higher than those commonly used and pose an increased risk for systemic side effects. Topical medications are an alternative approach to deliver high concentrations to the mucosa while limiting systemic side effects. This is the first study to evaluate the effects of a topical antibiotic on Pseudomonas biofilms in an animal model of sinusitis.
Pseudomonas sinusitis was established in nine rabbits. An irrigation catheter was placed within the lumen of the maxillary sinus, and the rabbits were irrigated with 7 days of normal saline or varying concentrations of tobramycin diluted in normal saline. Bacterial cfu were determined for nasal lavage fluid as well as physically disrupted mucosa, and scanning electron microscopy (SEM) of the mucosal surface was performed at the end of the study.
Increasing concentrations of topical tobramycin resulted in the eradication of viable bacteria within the lumen of the sinus but did not eradicate Pseudomonas attached to the mucosa. SEM detected biofilms within the mucosa even in those rabbits treated with high concentrations of topical tobramycin.
Bacterial biofilms attached to the sinonasal mucosa are resistant to topical saline and tobramycin irrigation. SEM was useful in the identification of biofilms, but did not provide a quantitative evaluation for efficacy of treatment.
[Show abstract][Hide abstract] ABSTRACT: Topical antimicrobials are commonly used to treat Pseudomonas sinusitis after endoscopic sinus surgery. Despite their popularity, there is limited data in the literature to support its use. This study is the first to examine the effects of topical tobramycin in an animal model of Pseudomonas sinusitis.
Pseudomonas aeruginosa sinusitis was induced in the maxillary sinus of New Zealand white rabbits. Study groups received various concentrations of topical tobramycin instilled through a single lumen catheter embedded within the maxillary sinus for 7 consecutive days. A control group received normal saline irrigation. Bacterial counts were measured from the nasal lavage after irrigation. The sinonasal complex was analyzed and a histological grading system was used to score the degree of infection at study end.
Nasal lavage bacterial counts of the saline control group remained persistently elevated throughout the study. Introduction of increasing concentrations of topical tobramycin caused the bacterial counts to fall to zero by study end. There was a dose-dependent histological response in which increasing concentrations of tobramycin led to a marked reduction but not total elimination of inflammation within the mucosa and underlying bone.
As opposed to normal saline irrigations, topical tobramycin led to a significant improvement in the degree of infection within an animal model of P. aeruginosa sinusitis. Despite the elimination of bacteria within the nasal lavage, there was a suggestion of a residual indolent infection as evidenced by persistent inflammation within the mucosa and underlying bone.
No preview · Article · Jul 2007 · American Journal of Rhinology
[Show abstract][Hide abstract] ABSTRACT: Although multiple etiologies contribute to the development of chronic rhinosinusitis (CRS), a common pathophysiological sequelae is ineffective sinonasal mucociliary clearance, leading to stasis of sinonasal secretions, with subsequent infection and/or persistent inflammation. Proper therapeutic intervention typically restores mucociliary activity, suggesting that the pathophysiological process(es) responsible for CRS-associated mucostasis may be reversible. We previously demonstrated a blunted response of CRS sinonasal cilia after purinergic stimulation. This study investigated whether the blunted ciliary response is unique to purinergic stimulation and addressed whether the blunted effect is primarily caused by local CRS-associated mediators or inherent genetic defects in ciliary function.
A dual temperature-controlled perfusion chamber, differential interference contrast microscopy, and high-speed digital video were used to analyze both basal as well as cholinergic, adrenergic, and purinergic stimulation of cilia in human sinonasal mucosal explants. Additionally, enzymically dissociated sinonasal ciliated cells were maintained ex vivo in submersion, on glass coverslips, and assessed daily for purinergic ciliary beat frequency stimulation.
Cholinergic and adrenergic stimulation generally were blunted in mucosal explants obtained from CRS patients. Ex vivo maintenance of samples demonstrated that the majority of CRS samples developed a stimulatory phenotype within 36 hours of culturing.
CRS is a common debilitating disease principally affecting sinonasal epithelial function with a resultant diminution of mucociliary transport. Presently, little is known about how this disease process affects the sinonasal epithelial ciliated cells. Our data suggest that ciliary response to environmental insults is blunted in a reversible manner in CRS patients.
No preview · Article · May 2007 · American Journal of Rhinology
[Show abstract][Hide abstract] ABSTRACT: Mucociliary clearance is a critical host defense mechanism of the airways. Effective mucociliary clearance requires appropriate mucus production and coordinated ciliary activity. The important role of these two components is best demonstrated in disorders such as primary ciliary dyskinesia and cystic fibrosis, both of which lead to lifelong recurrent respiratory tract infections. We review the methods used to analyze mucociliary clearance.
Utilization of microdialysis probes has improved temporal resolution of mucociliary clearance in murine airways, availing many genetic mouse models to critical mucociliary clearance analysis, while improved fixation technique for transmission electron microscopy has allowed for detailed resolution of the airway surface liquid. High-speed digital video analysis has improved quantification of ciliary beat frequency while advancements in air-liquid interface culturing techniques have generated in-vitro models to investigate mucociliary clearance.
Advancements in techniques for analysis of mucociliary clearance have improved our understanding of the interaction between the respiratory epithelium and the airway surface liquid, resulting in the ability to study pathologic processes involving mucociliary clearance in great detail.
No preview · Article · Mar 2007 · Current Opinion in Allergy and Clinical Immunology
[Show abstract][Hide abstract] ABSTRACT: Rhinosinusitis after functional endoscopic sinus surgery often is recalcitrant to conventional medical management. Topical therapies have been popularized as a method to deliver powerful medications to diseased mucosa while limiting systemic toxicity. The aim of this study was to develop an animal model that will provide objective data in studying the efficacy of topical medications and establish a platform for translation to human clinical trials.
The maxillary sinus ostium of white rabbits was obstructed with a pledget through an antrostomy created in the anterior face of the maxilla. The sinus was inoculated with Pseudomonas aeruginosa (PAO1) and 7 days later the antrostomy was reopened, the ostial obstruction was removed, and a single lumen catheter was placed. Normal saline was irrigated through the catheter for 7 days in one group of rabbits while a control group received no irrigation. At the end of 7 days, the rabbits were euthanized, analyzed under light microscopy, and bacterial counts of the nasal lavage were determined.
Sinusitis was confirmed in all rabbits 7 days after inoculation. Purulence, mucosal, and underlying bony inflammation persisted in both the control and the saline irrigation groups at study end. Nasal lavage bacterial counts were persistently elevated throughout the study period, indicative of bacterial viability.
An animal model has been created for the study of topical medications in sinusitis. A novel catheter delivery system within an unoccluded maxillary sinus is described that will aid in efficacy studies of topical medications in the management of recalcitrant rhinosinusitis.
No preview · Article · Jan 2007 · American Journal of Rhinology
[Show abstract][Hide abstract] ABSTRACT: Inverted papilloma (IP) is a benign but locally aggressive sinonasal tumor. Those localized to the maxillary sinus or medial maxillary wall have classically been removed through an external approach. In recent years, endoscopic removal has been advocated as an effective, minimally invasive approach. Successful endoscopic management is based on accurate intraoperative identification and complete resection of the tumor. The surgical management of the bone underlying the surface of an IP is less clearcut. Controversy exists as to whether the bony undersurface of an IP should be removed. In this article, histopathologic specimens and preoperative radiologic studies are prospectively examined to better understand the involvement of the bone underlying an inverted papilloma.
A prospective study of patients with maxillary sinus IP treated with endoscopic or endoscopic-assisted resection over a 12-month period was conducted. Intraoperatively, the site of tumor attachment was identified, and a 1- to 2-cm wedge of bone with overlying tumor was removed en bloc and analyzed under light microscopy. Preoperative radiographic studies were analyzed with respect to bony changes in the area of the tumor pedicle.
On histopathologic analysis, the bony surface underlying the IP was irregular with multiple bony crevices in all nine cases. In two cases (22% of specimens), an isolated rest of normal-appearingsalivary gland and/or mucosal tissue was seen embedded in the underlying bone. Radiographic osteitic bony changes were seen at site of tumor attachment in 100% of cases.
The irregularity of the bony surface may hinder complete tumor removal because microscopic rests of mucosa can be hidden within the bony crevices. Intraoperative removal of the bony surface at the site of tumor attachment may ensure a more complete removal.
No preview · Article · Oct 2006 · The Laryngoscope