Miguel Armengot

Respiratory Syncytial Virus Inhibits Ciliagenesis in Differentiated Normal Human Bronchial Epithelial Cells: Effectiveness of N-Acetylcysteine

Persistent respiratory syncytial virus (RSV) infections have been associated with the exacerbation of chronic inflammatory diseases, including chronic obstructive pulmonary disease (COPD). This virus infects the respiratory epithelium, leading to chronic inflammation, and induces the release of mucins and the loss of cilia activity, two factors that determine mucus clearance and the increase in sputum volume. These alterations involve reactive oxygen species-dependent mechanisms. The antioxidant N-acetylcysteine (NAC) has proven useful in the management of COPD, reducing symptoms, exacerbations, and accelerated lung function decline. NAC inhibits RSV infection and mucin release in human A549 cells. The main objective of this study was to analyze the effects of NAC in modulating ciliary activity, ciliagenesis, and metaplasia in primary normal human bronchial epithelial cell (NHBEC) cultures infected with RSV. Our results indicated that RSV induced ultrastructural abnormalities in axonemal basal bodies and decreased the expression of β-tubulin as well as two genes involved in ciliagenesis, FOXJ1 and DNAI2. These alterations led to a decrease in ciliary activity. Furthermore, RSV induced metaplastic changes to the epithelium and increased the number of goblet cells and the expression of MUC5AC and GOB5. NAC restored the normal functions of the epithelium, inhibiting ICAM1 expression, subsequent RSV infection through mechanisms involving nuclear receptor factor 2, and the expression of heme oxygenase 1, which correlated with the restoration of the antioxidant capacity, the intracellular H2O2 levels and glutathione content of NHBECs. The results presented in this study support the therapeutic use of NAC for the management of chronic respiratory diseases, including COPD.

Eosinophil degranulation patterns in nasal polyposis: An ultrastructural study

Background Eosinophils are possibly the most important inflammatory cells in the pathogenesis of rhinosinusitis with nasal polyposis. Eosinophil degranulation is the mechanism by which these cells exert their inflammatory action. Knowledge of eosinophil state and degranulation mode therefore may help us to better understand this disease. A study is made of eosinophil state and degranulation mode using transmission electron microscopy (TEM), attempting to establish correlations with certain clinical variables considered to be of importance in patients with nasal polyposis. Methods A prospective TEM study was made to examine 582 eosinophils under 5000× magnification, classifying them according to their state and degranulation mode. The cells originated from 36 cases of nasal polyposis and were catalogued according to the clinical–radiological presentation of the disorder, the presence of asthmatic disease, or acetilsalicilic acid (ASA) triad syndrome (nasal polyposis, asthma, and nonsteroidal anti-inflammatory drug intolerance), and the degree of nasal tissue eosinophilia of the case of origin. Results A total of 30.75% of the eosinophils were inactive, 41.75% exhibited piecemeal degranulation (PMD), 27.5% exhibited cytolysis, and 0.34% were in apoptosis. The degranulation mode was significantly correlated to the clinical and histological parameters studied. Thus, cytolysis tended to be less pronounced and PMD greater, in the cells from cases with intense eosinophilia or severe polyposis (high clinical stage and ASA triad). Conclusion Cytolysis and PMD are the principal degranulation modes of eosinophils in nasal polyposis—apoptosis being very infrequent. Nasal polyposis shows a correlation between eosinophil degranulation mode and the clinical and radiological stage and the degree of tissue eosinophilia of the case of origin.

Ultrastructural Patterns of Primary Ciliar Dyskinesia Syndrome

Clinical presentation, ciliary ultrastructure, and nasal mucociliary transport by a radioisotopic technique were analyzed in 14 Kartagener syndrome patients. In this study the most common pattern was the absence of outer and inner dynein arms in 57% of cases. Also reported are 14% patients with short inner dynein arms. A total of 29% of the patients showed normal dynein arms. Mucociliary stasis was observed in 13 cases. Primary ciliary dyskinesia syndrome and Kartagener syndrome are clinically homogeneous and morphologically heterogeneous. The authors conclude that a typical clinical presentation with an altered mucociliary transport obtained by radioisotopic technique is diagnostic although ciliary ultrastructure is normal.

Effect of total nasal obstruction on nocturnal oxygen saturation.

Background Nasal obstruction is considered a risk factor for sleep-disordered breathing (SDB), although the relationship has not been established clearly. This study explores the repercussion of total nasal packing on arterial oxygen saturation during the nocturnal resting hours and its implication in SDB. Methods Forty patients were subjected to total nasal packing after nasal surgery or epistaxis. In all cases the oxygen desaturation index (ODI) was monitored with and without nasal packs, and the results were compared. Results Thirty-seven patients (92.5%) showed poorer oxygen saturation after nasal occlusion. Of these, 19 patients (47.5%) reached pathological desaturation (ODI ≥ 12). Desaturation was greater among obese subjects. Conclusion Total nasal packing causes significant nocturnal oxygen desaturation. This must be taken into account in the postoperative nasal packing of patients with respiratory failure, obesity, or SDB.

Concha bullosa mucocele with invasion of the orbit.

We describe the case of a patient with a large concha bullosa mucocele invading the orbit. The wall of the mucocele was lined by a well-preserved ciliary epithelium with a ciliary density similar to that of the healthy sinus mucosa. Ciliary ultrastructure was normal. Endonasal microsurgery yielded good results, and no recurrence has been observed after 18 months. The thickness of the middle turbinate is variable and often exhibits vacuolar structures. When pneumatized, this gives origin to a so-called concha bullosa.1 This lesion was first described by Zuckerlandl in 1893.2 For a number of years concha bullosa has been related to the pathogenesis of inflammatory disorders of the paranasal sinuses. Few cases have been reported dealing with concha bullosa as a cause of disease. In the past, a case of aspergilloma was reported within a concha bullosa3; however, a triple-index bibliographic search of the past 3 years yielded only a single case of a concha bullosa mucopyocele.4 This study reports a patient with a giant mucocele within a concha bullosa that destroyed the anterior ethmoid bone and invaded the orbit. CASE REPORT

Role of adenylate kinase type 7 expression on cilia motility: possible link in primary ciliary dyskinesia.

Background Adenylate kinase 7 (AK7) mediates the reaction 2ADP ↔ ATP + AMP, providing energy for the beating of cilia. A study recently showed that AK7 expression may be correlated with the primary ciliary dyskinesia (PCD) phenotype in mice. In this study, we characterized AK7 expression in vitro in an air–liquid interface (ALI) model and in middle nasal turbinate biopsy specimens from a cohort of patients with PCD to elucidate whether AK7 expression is correlated with ciliary malfunction. Methods AK7 expression was measured by real-time reverse-transcription polymerase chain reaction and Western blotting. In vitro differentiated nasal human epithelial cell siRNA experiments were performed to investigate the effect of AK7 expression on ciliary beat frequency (CBF). Ciliary motility and ultrastructure were evaluated in a cohort of 29 patients with PCD (PCD, n = 17; Kartageners syndrome, n = 12) and 26 healthy control donors. Results AK7 expression was mainly located on the apical surface of differentiated nasal ALI cells, and targeted suppression of the AK7 gene decreased CBF by 41%. AK7 expression was diminished significantly in patients with PCD (0.54 ± 0.1-fold; p < 0.05) compared with healthy controls (1.1 ± 0.08-fold). Furthermore, AK7 expression was correlated with CBF in patients with PCD (r = 0.5; p = 0.009). Conclusion AK7 expression was correlated with CBF in vitro and in nasal biopsy specimens from patients with PCD, which may have contributed to the ciliary malfunction observed in our patients with PCD.

Cytological and tissue eosinophilia correlations in nasal polyposis.

Background Eosinophils are possibly the most important inflammatory cells in the pathogenesis of chronic rhinosinusitis with nasal polyposis. Eosinophilic nasal mucosal infiltration is characteristic of nasal polyposis. The aim of this work was to study the possibility of correlations between cytological and tissue eosinophilia in patients with nasal polyposis and with the stage of the disease. If this correlation exists, nasal cytology will be very useful in diagnosis and management of patients with nasal polyposis. Methods We studied 40 patients affected idiopathic nasal polyposis and 12 normal group controls. Qualitative measurements of eosinophils in nasal exudates and in tissue biopsy specimens were performed and the results were compared. Results Nasal cytology and tissue biopsy showed a statistical correlation in the normal as well as in the pathological group. Clinical staging polyposis also correlated with cytological and tissue eosinophilia. Conclusions Nasal cytological eosinophilia is a simple, fast, nontraumatic, and inexpensive method in the evaluation of patients with nasal polyposis. Eosinophilic infiltration of the nasal mucosa is characteristic of nasal polyposis. It correlates with the stage of the disease and tends to be higher in patients with asthma, nasal polyposis, and nonsteroidal anti-inflammatory drug intolerance (acetil salicilic acid triad).

Nasal Ciliary Beat Frequency and Beat Pattern in Retinal Ciliopathies

PURPOSE The cilium in photoreceptors appears ultrastructurally very similar to the nasal ciliated epithelium. The purpose of this study was to evaluate the nasal ciliary beat frequency and beat pattern in patients with retinitis pigmentosa (RP) and Usher syndrome type II and compare it with that of healthy control subjects. METHODS A prospective, comparative control study. Fresh samples of nasal mucosa were obtained from 13 patients with typical forms of RP, and from 4 patients with Usher syndrome type II. The nasal ciliary beat frequency (CBF) and beat pattern were determined using high-resolution digital high-speed video imaging (DHSV). The control group included 32 fresh nasal mucosa samples from 32 healthy volunteers without any other confounding diseases. RESULTS The nasal CBF was lower in patients with Usher syndrome than in control subjects (Mann-Whitney U test, P = 0.01). The nasal CBF was 9.28 ± 0.4 (mean ± SD) Hz in patients with Usher syndrome and 10.82 ± 1.39 Hz in patients of the control group. No significant difference was observed in the nasal CBF between the RP (10.59 ± 1.54 Hz) and control group (Mann-Whitney U test, P = 0.64). Normal ciliary beat pattern was observed in all the patients and healthy volunteers. CONCLUSIONS The nasal CBF is diminished in patients with Usher syndrome type II, whereas it remains normal in simplex RP patients. These results add evidence to the fact that Usher syndrome could be a primary ciliary disorder.

Correlations between clinical and histological aspects in nasal polyposis

INTRODUCTION The histological study of nasal polyps does not reveal any specific lesions but eosinophilic infiltration nasal mucosae seems to be characteristic of nasal polyposis. The aim of this work is to study possible links between certain histological and clinical aspects in nasal polyposis. Furthermore, we attempt to compare the quantification of tissue eosinophilia according to the number of eosinophils per field with the percentage figure obtained with respect to the total of inflammatory cells. MATERIAL AND METHOD We have studied 40 patients with idiopathic bilateral nasal polyposis, assessing the correlations between various clinical aspects such as their endoscopic and radiological status, association with asthma and intolerance to NSAIDs, against histological aspects of nasal polyps such as the frequency of metaplasia, fibrosis and the degree of eosinophilic infiltration. A group of 12 healthy subjects allowed comparison of our results with healthy nasal mucosa. RESULTS Tissue eosinophilia correlates statistically with clinical staging and tends to be higher in patients with ASA triad. The quantitative measurement of tissue eosinophilia (number of eosinophils per field) correlates with the percentage figure obtained (with respect to the total number of inflammatory cells in the infiltrate). CONCLUSIONS Eosinophil infiltration of the nasal mucosa is, together with oedema, the most constant histological characteristic of nasal polyposis and seems to be an important factor in the clinical behaviour of sinonasal polyposis. Quantitative measurement of tissue eosinophilia is easier and quicker to perform and equivalent to percentage evaluation.

Study of USH1 Splicing Variants through Minigenes and Transcript Analysis from Nasal Epithelial Cells

Usher syndrome type I (USH1) is an autosomal recessive disorder characterized by congenital profound deafness, vestibular areflexia and prepubertal retinitis pigmentosa. The first purpose of this study was to determine the pathologic nature of eighteen USH1 putative splicing variants found in our series and their effect in the splicing process by minigene assays. These variants were selected according to bioinformatic analysis. The second aim was to analyze the USH1 transcripts, obtained from nasal epithelial cells samples of our patients, in order to corroborate the observed effect of mutations by minigenes in patient’s tissues. The last objective was to evaluate the nasal ciliary beat frequency in patients with USH1 and compare it with control subjects. In silico analysis were performed using four bioinformatic programs: NNSplice, Human Splicing Finder, NetGene2 and Spliceview. Afterward, minigenes based on the pSPL3 vector were used to investigate the implication of selected changes in the mRNA processing. To observe the effect of mutations in the patient’s tissues, RNA was extracted from nasal epithelial cells and RT-PCR analyses were performed. Four MYO7A (c.470G>A, c.1342_1343delAG, c.5856G>A and c.3652G>A), three CDH23 (c.2289+1G>A, c.6049G>A and c.8722+1delG) and one PCDH15 (c.3717+2dupTT) variants were observed to affect the splicing process by minigene assays and/or transcripts analysis obtained from nasal cells. Based on our results, minigenes are a good approach to determine the implication of identified variants in the mRNA processing, and the analysis of RNA obtained from nasal epithelial cells is an alternative method to discriminate neutral Usher variants from those with a pathogenic effect on the splicing process. In addition, we could observe that the nasal ciliated epithelium of USH1 patients shows a lower ciliary beat frequency than control subjects.