Estudio del estrés oxidativo en el epitelio nasal ciliado de pacientes con Discinesia Ciliar Primaria

Antecedentes: La Discinesia Ciliar Primaria (DCP) es una enfermedad rara con una prevalencia estimada de 1:20.000 nacidos vivos. Se caracteriza por una alteración en el patrón de motilidad de los cilios y flagelos que al provocar un déficit en el aclaramiento mucociliar, causa infecciones e inflamación crónica de las vías aéreas. Un proceso inflamatorio ineficiente aumenta la producción de especies reactivas del oxígeno y nitrógeno (ERO y ERN), lo que incrementa el riesgo de padecer distintas enfermedades. En la DCP se ha demostrado que el estrés oxidativo está aumentado en el condensado de aire exhalado de pacientes pediátricos pero, hasta el momento, nunca se había determinado en el tejido afectado. Nuestra hipótesis de partida fue que los pacientes con DCP tienen un estado pro-oxidativo en las células del epitelio respiratorio. Objetivos: Estudiar el estado oxidativo de las células del epitelio nasal de pacientes con DCP, compararlo con el de voluntarios sanos, y determinar el papel que juega en la fisiopatología de la enfermedad. Métodos: Se midieron los niveles de ERO, RNS, glutatión reducido (GSH), calcio intracelular (Ca2+), potencial de membrana plasmático, y daño oxidativo en lípidos y proteínas. Además, se analizó la función mitocondrial y la apoptosis de las células del epitelio nasal de los sujetos de estudio mediante citometría de flujo. En el estudio participaron 2 grupos: 35 voluntarios sanos y 35 pacientes diagnosticados de DCP. Resultados: Los pacientes con DCP tienen niveles de apoptosis, NO, ONOO-, O2- total, H2O2 mitocondrial y O2- mitocondrial, en las células del epitelio nasal, inferiores a los de los individuos sanos. No existen diferencias en el resto de parámetros medidos, entre pacientes y controles. En el grupo de pacientes tampoco se aprecian diferencias en ninguno de los parámetros comparados ni entre adultos y niños, ni en función del patrón de movilidad ciliar; las mujeres con DCP tienen menores niveles de H2O2 y O2- mitocondriales que los varones con la enfermedad; los pacientes con atelectasias o con situs inversus, tienen niveles peróxidos intracelulares superiores a los de los pacientes que no presentan estas anomalías; y aquellos con rinosinusitis crónica tienen niveles más bajos de ONOO- que los que no la tienen. No se han encontrado diferencias en los demás síntomas. Se ha diseñado también un árbol de clasificación de los individuos como “paciente” o “control sano” en función de los niveles de ciertos parámetros del perfil oxidativo medidos en sus células del epitelio nasal.Background: Primary ciliary dyskinesia (PCD) is a rare disease with an estimated prevalence of 1:20.000 births. It is characterized by an alteration of the mobility pattern of cilia and flagella, causing lack of mucociliary clearance and subsequent infection and chronic inflammation of the airways. An inefficient inflammatory process increases the production of reactive oxygen and nitrogen species (ROS and RNS), which derives in a higher risk of diseases. In PCD, a study demonstrated that oxidative stress is increased in exhaled breath condensate from paediatric patients. Although oxidative stress had not been determined in the affected tissue yet, we hypothesized that patients with PCD have a pro-oxidative status in respiratory epithelial cells. Aims: To study the oxidative status of nasal epithelial cells from patients with PCD, compare it with healthy controls, and determine their role on the disease pathophysiology. Methods: Levels of ROS, RNS, glutathione (GSH), intracellular Ca2+, plasma membrane potential, and oxidative damage in lipids and proteins were measured. In addition, apoptosis and the mitochondrial function were analysed by flow cytometry in nasal epithelial cells. Two groups took part in the study: 35 healthy subjects and 35 patients diagnosed of PCD. 7 Results: Patients with PCD have levels of apoptosis, NO, ONOO-, total O2-, mitochondrial H2O2 and mitochondrial O2- in their nasal epithelial cells lower than those of healthy individuals. There are no differences between the groups of patients and controls in the rest of the parameters measured. When comparing within the group of patients, there are no differences between adults and children in any compared parameter, nor is there any difference depending on the different ciliary mobility pattern; women with PCD have lower levels of mitochondrial H2O2 and O2- than men with the disease; patients with atelectasis, and those with situs inversus have higher intracellular peroxides levels than those who do not; patients with chronic rhinosinusitis have lower levels of ONOO- than those who do not, and no differences have been found depending on the other symptoms. A tree has been designed to classify individuals as patient or healthy control based on the levels of certain parameters of the oxidative profile measured in their nasal epithelial cells

High-Speed Video Microscopy for Primary Ciliary Dyskinesia Diagnosis: A Study of Ciliary Motility Variations with Time and Temperature

Primary ciliary dyskinesia (PCD) is a rare disease resulting from a defect in ciliary function that generates, among other issues, chronic upper and lower respiratory tract infections. European guidelines recommend studying ciliary function (pattern (CBP) and frequency (CBF)), together with characteristic clinical symptoms, as one of the definitive tests. However, there is no “gold standard”. The present study aims to use high-speed video microscopy to describe how CBF and CBP alter over time and at different temperatures to reduce the error rate in the diagnosis of PCD. Samples of nasal epithelium from 27 healthy volunteers were studied to assess CBF and CBP at 0, 3, 24, 48, and 72 h, at room temperature and 4 °C. It was observed that CBF increased while CBP became dyskinetic, both at room temperature and at 4 °C, as time passed, especially after 3 h. In order to preserve all ciliary function parameters and to perform a reliable analysis to improve the diagnostic process of PCD, analysis should be performed within the first 3 h of sample collection, preferably in reference centers

Proceedings of the 2nd BEAT-PCD conference and 3rd PCD training school: part 1

Primary ciliary dyskinesia (PCD) is a rare heterogenous condition that causes progressive suppurative lung disease, chronic rhinosinusitis, chronic otitis media, infertility and abnormal situs. 'Better Experimental Approaches to Treat Primary Ciliary Dyskinesia' (BEAT-PCD) is a network of scientists and clinicians coordinating research from basic science through to clinical care with the intention of developing treatments and diagnostics that lead to improved long-term outcomes for patients. BEAT-PCD activities are supported by EU funded COST Action (BM1407). The second BEAT-PCD conference, and third PCD training school were held jointly in April 2017 in Valencia, Spain. Presentations and workshops focussed on advancing the knowledge and skills relating to PCD in: basic science, epidemiology, diagnostic testing, clinical management and clinical trials. The multidisciplinary conference provided an interactive platform for exchanging ideas through a program of lectures, poster presentations, breakout sessions and workshops. Three working groups met to plan consensus statements. Progress with BEAT-PCD projects was shared and new collaborations were fostered. In this report, we summarize the meeting, highlighting developments made during the meeting

Lack of Correlation of Sinonasal and Otologic Reported Symptoms With Objective Measurements Among Patients With Primary Ciliary Dyskinesia: An International Study.

peer reviewedSinonasal and otologic symptoms are common among patients with primary ciliary dyskinesia (PCD) of all ages. We used baseline data from the ENT Prospective International Cohort of PCD patients (EPIC-PCD), the first PCD cohort focused on ENT disease manifestations. We assessed agreement between patient- or parent-reported symptoms and relevant examination findings, and calculated unweighted Cohen’s kappa to adjust for agreement by chance. We included 404 participants, from 12 centres. We found no correlation between patient-reported sinonasal symptoms and relevant clinical examination findings. Otologic symptoms correlated poorly or weakly with otoscopy and audiometry findings, with age and centre identified as determinants of agreement

Proceedings of the 3rd BEAT-PCD Conference and 4th PCD Training School

Abstract Primary ciliary dyskinesia (PCD) is a chronic suppurative airways disease that is usually recessively inherited and has marked clinical phenotypic heterogeneity. Classic symptoms include neonatal respiratory distress, chronic rhinitis since early childhood, chronic otitis media, recurrent airway infections leading to bronchiectasis, chronic sinusitis, laterality defects with and without congenital heart disease including abnormal situs in approximately 50% of the cases, and male infertility. Lung function deteriorates progressively from childhood throughout life. ‘Better Experimental Approaches to Treat Primary Ciliary Dyskinesia’ (BEAT-PCD) is a network of scientists and clinicians coordinating research from basic science through to clinical care with the intention of developing treatments and diagnostics that lead to improved long-term outcomes for patients. BEAT-PCD activities are supported by EU funded COST Action (BM1407). The third BEAT-PCD conference and fourth PCD training school were held jointly in February 2018 in Lisbon, Portugal. Presentations and workshops focussed on advancing the knowledge and skills relating to PCD in: basic science, epidemiology, diagnostic testing, clinical management and clinical trials. The multidisciplinary conference provided an interactive platform for exchanging ideas through a program of lectures, poster presentations, breakout sessions and workshops. Three working groups met to plan consensus statements. Progress with BEAT-PCD projects was shared and new collaborations were fostered. In this report, we summarize the meeting, highlighting developments made during the meeting

Generic instabilities in the relativistic Chapman–Enskog heat conduction law

We address the well-posedness of the Cauchy problem corresponding to the relativistic first-order fluid equations, coupled with the Chapman–Enskog heat-flux constitutive relation. We show that the system of equations that results by considering linear perturbations with respect to a generic time direction is non-hyperbolic, since there are modes that may arbitrarily grow as wave-number increases. Then, using a result provided by Strang (J Differ Equ 2:107–114, 1966), we conclude that the full non-linear first-order theory is also non-hyperbolic, thus admitting an ill-posed initial-value formulation. Unlike Eckart’s theory, these instabilities are not present when the time direction is aligned with the fluid’s direction. However, since in general the fluid velocity is not surface-forming, the instability can only be avoided in the particular case where no rotation is present.Fil: García-Perciante, Ana L.. Universidad Autonoma Metropolitana. Division de Ciencias Sociales y Humanidades. Departamento de Humanidades.; MéxicoFil: Rubio, Marcelo Enrique. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Astronomía Teórica y Experimental. Universidad Nacional de Córdoba. Observatorio Astronómico de Córdoba. Instituto de Astronomía Teórica y Experimental; ArgentinaFil: Reula, Oscar Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Física Enrique Gaviola. Universidad Nacional de Córdoba. Instituto de Física Enrique Gaviola; Argentin

Redox Imbalance in Nasal Epithelial Cells of Primary Ciliary Dyskinesia Patients

Background: Primary Ciliary Dyskinesia (PCD) represents a rare condition marked by an abnormal mobility pattern of cilia and flagella, resulting in impaired mucociliary clearance. This deficiency leads to recurrent infections and persistent inflammation of the airways. While previous studies have indicated heightened oxidative stress levels in the exhaled breath condensate of pediatric PCD patients, the assessment of oxidative stress within the affected respiratory tissue remains unexplored. Aims: To assess the oxidative status of human nasal epithelial cells (NECs) in PCD patients. Methods: Thirty-five PCD patients and thirty-five healthy control subjects were prospectively included in the study. Levels of reactive oxygen species (ROS), reactive nitrogen species (RNS), glutathione (GSH), intracellular Ca2+, plasma membrane potential, and oxidative damage in lipids and proteins were measured. In addition, apoptosis and mitochondrial function were analyzed by flow cytometry in NECs. Results: NECs from PCD patients showed reduced levels of apoptosis (p = 0.004), superoxide anion (O2−, p = 0.018), peroxynitrite (ONOO−, p = 0.007), nitric oxide (NO, p = 0.007), mitochondrial hydrogen peroxide (mtH2O2, p 2−, p = 0.0004) and increased mitochondrial mass (p = 0.009) compared to those from healthy individuals. No significant differences were observed in oxidized proteins (p = 0.137) and the oxidized/reduced lipid ratio (p = 0.7973). The oxidative profile of NEC cells in PCD patients, according to their ciliary motility, recurrent otitis, recurrent pneumonia, atelectasis, bronchiectasis, and situs inversus, showed no statistically significant differences in the parameters studied. Conversely, patients with chronic rhinosinusitis exhibited lower levels of ONOO− than PCD patients without this condition, with no significant differences related to other symptoms. Conclusions: Our findings strongly suggest the presence of a redox imbalance, specifically leaning toward a reductive state, in PCD patients

Understanding Primary Ciliary Dyskinesia: Experience From a Mediterranean Diagnostic Reference Centre

Background: Due to the lack of a gold standard diagnostic test, reference centres with experienced personnel and costly procedures are needed for primary ciliary dyskinesia (PCD) diagnostics. Diagnostic flowcharts always start with clinical symptoms. Therefore, the aim of this work is to define differential clinical criteria so that only patients clinically compatible with PCD are referred to reference centres. Materials and methods: 18 variables from 476 Mediterranean patients with clinically suspicious PCD were collected. After analysing cilia function and ultrastructure, 89 individuals were diagnosed with PCD and 387 had a negative diagnosis. Simple logistic regression analysis, considering PCD as a dependent variable and the others as independent variables, was done. In order to define the variables that best explain PCD, a step-wise logistic regression model was defined. Aiming to classify individuals as PCD or PCD-like patients, based on variables included in the study, a classification and regression tree (CART) was designed. Results and conclusions: Simple logistic regression analysis shows statistically significant association between age at the beginning of their symptomatology, periodicity, fertility, situs inversus, recurrent otitis, atelectasis, bronchiectasis, chronic productive cough, rhinorrea, rhinusinusitis and recurrent pneumonias, and PCD. The step-wise logistic regression model selected situs inversus, atelectasis, rhinorrea, chronic productive cough, bronchiectasis, recurrent pneumonias, and otitis as PCD predictive variables (82% sensitivity, 88% specificity, and 0.92 Area Under the Curve (AUC)). A decision tree was designed in order to classify new individuals based on pansinusitis, situs inversus, periodicity, rhinorrea, bronchiectasis, and chronic wet cough

Understanding Primary Ciliary Dyskinesia : Experience From a Mediterranean Diagnostic Reference Centre

Background: Due to the lack of a gold standard diagnostic test, reference centres with experienced personnel and costly procedures are needed for primary ciliary dyskinesia (PCD) diagnostics. Diagnostic flowcharts always start with clinical symptoms. Therefore, the aim of this work is to define di ff erential clinical criteria so that only patients clinically compatible with PCD are referred to reference centres. Materials and methods: 18 variables from 476 Mediterranean patients with clinically suspicious PCD were collected. After analysing cilia function and ultrastructure, 89 individuals were diagnosed with PCD and 387 had a negative diagnosis. Simple logistic regression analysis, considering PCD as a dependent variable and the others as independent variables, was done. In order to define the variables that best explain PCD, a step-wise logistic regression model was defined. Aiming to classify individuals as PCD or PCD-like patients, based on variables included in the study, a classification and regression tree (CART) was designed. Results and conclusions: Simple logistic regression analysis shows statistically significant association between age at the beginning of their symptomatology, periodicity, fertility, situs inversus, recurrent otitis, atelectasis, bronchiectasis, chronic productive cough, rhinorrea, rhinusinusitis and recurrent pneumonias, and PCD. The step-wise logistic regression model selected situs inversus, atelectasis, rhinorrea, chronic productive cough, bronchiectasis, recurrent pneumonias, and otitis as PCD predictive variables (82% sensitivity, 88% specificity, and 0.92 Area Under the Curve (AUC)). A decision tree was designed in order to classify new individuals based on pansinusitis, situs inversus, periodicity, rhinorrea, bronchiectasis, and chronic wet cough. Keywords: standard diagnosis; reference centres; clinical presentation; cilia; primary ciliary dyskinesi

High-Speed Video Microscopy For Primary Ciliary Dyskinesia Diagnosis: A Study of Ciliary Motility Variations with Time and Temperature

Primary ciliary dyskinesia (PCD) is a rare disease resulting from a defect in ciliary function that generates, among other issues, chronic upper and lower respiratory tract infections. European guidelines recommend studying ciliary function (pattern (CBP) and frequency (CBF)), together with characteristic clinical symptoms, as one of the definitive tests. However, there is no "gold standard". The present study aims to use high-speed video microscopy to describe how CBF and CBP alter over time and at different temperatures to reduce the error rate in the diagnosis of PCD. Samples of nasal epithelium from 27 healthy volunteers were studied to assess CBF and CBP at 0, 3, 24, 48, and 72 h, at room temperature and 4 ◦C. It was observed that CBF increased while CBP became dyskinetic, both at room temperature and at 4 ◦C, as time passed, especially after 3 h. In order to preserve all ciliary function parameters and to perform a reliable analysis to improve the diagnostic process of PCD, analysis should be performed within the first 3 h of sample collection, preferably in reference center