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BACKGROUND: Primary ciliary dyskinesia (PCD) is a genetic disorder caused by aberrant motile cilia function that results in defective ciliary airway clearance and subsequently to recurrent airway infections and bronchiectasis. QUESTION: How many functional multiciliated airway cells are sufficient to maintain ciliary airway clearance? METHODS: To answer this question we exploited the molecular defects of the X-linked recessive PCD variant caused by pathogenic variants in DNAAF6 (PIH1D3), characterized by immotile cilia in the affected males. We carefully analyzed the clinical phenotype, molecular defect (immunofluorescence and transmission-electron microscopy) and performed in vitro (particle tracking in air-liquid interface cultures) and in vivo (radiolabeled tracer studies) studies to assess ciliary clearance of respiratory cells from females with heterozygous and males with hemizygous pathogenic DNAAF6 variants. RESULTS: PCD males with hemizygous pathogenic DNAAF6 variants displayed exclusively immotile cilia, absence of ciliary clearance and severe PCD symptoms. Due to random or skewed X-chromosome inactivation in six females with heterozygous pathogenic DNAAF6 variants, 54.3%±10 (range 38%-70%) of multiciliated cells were defective. Nevertheless, in vitro and in vivo assessment of the ciliary airway clearance was normal or slightly abnormal. Consistently, heterozygous female individuals showed no or only mild respiratory symptoms. CONCLUSIONS: Our findings indicate that 30%-62% of functioning multiciliated respiratory cells are able to generate either normal or slightly reduced ciliary clearance. Because heterozygous females displayed either no or subtle respiratory symptoms, complete correction of 30% of cells by precision medicine might be able to improve ciliary airway clearance in PCD individuals as well as clinical symptoms.
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BACKGROUND: Primary ciliary dyskinesia (PCD) represents a group of rare hereditary disorders characterised by deficient ciliary airway clearance that can be associated with laterality defects. We aimed to describe the underlying gene defects, geographical differences in genotypes and their relationship to diagnostic findings and clinical phenotypes. METHODS: Genetic variants and clinical findings (age, sex, body mass index, laterality defects, forced expiratory volume in 1â s (FEV1)) were collected from 19 countries using the European Reference Network's ERN-LUNG international PCD Registry. Genetic data were evaluated according to American College of Medical Genetics and Genomics guidelines. We assessed regional distribution of implicated genes and genetic variants as well as genotype correlations with laterality defects and FEV1. RESULTS: The study included 1236 individuals carrying 908 distinct pathogenic DNA variants in 46 PCD genes. We found considerable variation in the distribution of PCD genotypes across countries due to the presence of distinct founder variants. The prevalence of PCD genotypes associated with pathognomonic ultrastructural defects (mean 72%, range 47-100%) and laterality defects (mean 42%, range 28-69%) varied widely among countries. The prevalence of laterality defects was significantly lower in PCD individuals without pathognomonic ciliary ultrastructure defects (18%). The PCD cohort had a reduced median FEV1 z-score (-1.66). Median FEV1 z-scores were significantly lower in CCNO (-3.26), CCDC39 (-2.49) and CCDC40 (-2.96) variant groups, while the FEV1 z-score reductions were significantly milder in DNAH11 (-0.83) and ODAD1 (-0.85) variant groups compared to the whole PCD cohort. CONCLUSION: This unprecedented multinational dataset of DNA variants and information on their distribution across countries facilitates interpretation of the genetic epidemiology of PCD and indicates that the genetic variant can predict diagnostic and phenotypic features such as the course of lung function.
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Estudos de Associação Genética , Genótipo , Fenótipo , Humanos , Masculino , Feminino , Adulto , Criança , Adolescente , Adulto Jovem , Pessoa de Meia-Idade , Europa (Continente) , Sistema de Registros , Dineínas do Axonema/genética , Volume Expiratório Forçado , Pré-Escolar , Síndrome de Kartagener/genética , Síndrome de Kartagener/fisiopatologia , Variação Genética , Mutação , Idoso , Lactente , Proteínas do Citoesqueleto , ProteínasRESUMO
Nasal nitric oxide (nNO) is extremely low in most people with primary ciliary dyskinesia (PCD) and its measurement is an important contributor to making the diagnosis. Existing guidelines and technical standards focus on nNO measurements in older, cooperative children using chemiluminescence analysers. However, measurements of nNO in pre-school-age children (age 2-5â years) may facilitate early diagnosis and electrochemical rather than chemiluminescence analysers are widely used. Pre-schoolers often need different methods to be employed when measuring nNO. Hence, a European Respiratory Society Task Force has developed this technical standard as the first step towards standardising sampling, analysis and reporting of nNO measured as part of the diagnostic testing for PCD in all age groups, including pre-school-age children. Furthermore, we considered both chemiluminescence and electrochemical analysers that are in use worldwide. There was a paucity of quality evidence for electrochemical analysers and sampling methods used in young children, and the Task Force proposes future research priorities to allow updates of this technical standard.
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Transtornos da Motilidade Ciliar , Síndrome de Kartagener , Humanos , Criança , Pré-Escolar , Idoso , Óxido Nítrico/análise , Síndrome de Kartagener/diagnóstico , Testes Respiratórios/métodos , Diagnóstico Precoce , Taxa Respiratória , Transtornos da Motilidade Ciliar/diagnósticoRESUMO
Primary ciliary dyskinesia (PCD) can be defined as a multiorgan ciliopathy with a dominant element of chronic airway disease affecting the nose, sinuses, middle ear, and in particular, the lower airways. Although most patients with PCD are diagnosed during preschool years, it is obvious that the chronic lung disease starts its course already from birth. The many faces of the clinical picture change, as does lung function, structural lung damage, the burden of infection, and of treatment throughout life. A markedly severe neutrophil inflammation in the respiratory tract seems pervasive and is only to a minimal extent ameliorated by a treatment strategy, which is predominantly aimed at bacterial infections. An ever-increasing understanding of the different aspects, their interrelationships, and possible different age courses conditioned by the underlying genotype is the focus of much attention. The future is likely to offer personalized medicine in the form of mRNA therapy, but to that end, it is of utmost importance that all patients with PCD be carefully characterized and given a genetic diagnosis. In this narrative review, we have concentrated on lower airways and summarized the current understanding of the chronic airway disease in this motile ciliopathy. In addition, we highlight the challenges, gaps, and opportunities in PCD lung disease research.
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Transtornos da Motilidade Ciliar , Ciliopatias , Doença Pulmonar Obstrutiva Crônica , Pré-Escolar , Transtornos da Motilidade Ciliar/genética , Transtornos da Motilidade Ciliar/terapia , Genótipo , HumanosRESUMO
Primary ciliary dyskinesia (PCD) presents with symptoms early in life and the disease course may be progressive, but longitudinal data on lung function are scarce. This multinational cohort study describes lung function trajectories in children, adolescents and young adults with PCD. We analysed data from 486 patients with repeated lung function measurements obtained between the age of 6 and 24â years from the International PCD Cohort and calculated z-scores for forced expiratory volume in 1â s (FEV1), forced vital capacity (FVC) and FEV1/FVC ratio using the Global Lung Function Initiative 2012 references. We described baseline lung function and change of lung function over time and described their associations with possible determinants in mixed-effects linear regression models. Overall, FEV1, FVC and FEV1/FVC z-scores declined over time (average crude annual FEV1 decline was -0.07 z-scores), but not at the same rate for all patients. FEV1 z-scores improved over time in 21% of patients, remained stable in 40% and declined in 39%. Low body mass index was associated with poor baseline lung function and with further decline. Results differed by country and ultrastructural defect, but we found no evidence of differences by sex, calendar year of diagnosis, age at diagnosis, diagnostic certainty or laterality defect. Our study shows that on average lung function in PCD declines throughout the entire period of lung growth, from childhood to young adult age, even among patients treated in specialised centres. It is essential to develop strategies to reverse this tendency and improve prognosis.
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Transtornos da Motilidade Ciliar , Humanos , Criança , Adolescente , Adulto Jovem , Adulto , Estudos de Coortes , Capacidade Vital , Volume Expiratório Forçado , PulmãoRESUMO
Objectives: Primary ciliary dyskinesia (PCD) is a rare congenital disease with defective mucociliary clearance causing frequent and often persistent pulmonary infections. Achromobacter species are opportunistic pathogens renowned for the difficulty of effective treatments and deteriorating effects on lung function. We aimed to describe the occurrence, treatment, and rate of successful eradication of Achromobacter species in patients with PCD. Methods: We retrospectively reviewed 18 years of historical microbiological samples and 10 years of electronic health records for PCD patients in Denmark. Results: We included 136 patients. Twenty-six patients had isolates of Achromobacter species. On average, 5% of the cohort had at least one annual isolate. Infections became persistent in 38% with a median length of 6.6 years leading to a significant number of antibiotic treatments. Resistance toward tobramycin and ciprofloxacin was prevalent. Overall, successful eradication was achieved in 62% of patients. We found the course of lung function significantly worse during persistent Achromobacter species infection than during the two preceding years, but not different to the course in unaffected age-matched controls. Conclusion The prevalence of Achromobacter species in patients with PCD is in line with what has been reported in cystic fibrosis and can occur transiently, intermittently, or develop into a serious persistent lung infection associated with long-term antibiotic treatment.
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Achromobacter , Transtornos da Motilidade Ciliar , Fibrose Cística , Antibacterianos/uso terapêutico , Transtornos da Motilidade Ciliar/tratamento farmacológico , Fibrose Cística/complicações , Fibrose Cística/tratamento farmacológico , Humanos , Estudos RetrospectivosRESUMO
Primary ciliary dyskinesia (PCD) is a genetically heterogeneous chronic destructive airway disease. PCD is traditionally diagnosed by nasal nitric oxide measurement, analysis of ciliary beating, transmission electron microscopy (TEM), and/or genetic testing. In most genetic PCD variants, laterality defects can occur. However, it is difficult to establish a diagnosis in individuals with PCD and central pair (CP) defects, and alternative strategies are required because of very subtle ciliary beating abnormalities, a normal ciliary ultrastructure, and normal situs composition. Mutations in HYDIN are known to cause CP defects, but the genetic analysis of HYDIN variants is confounded by the pseudogene HYDIN2, which is almost identical in terms of intron/exon structure. We have previously shown that several types of PCD can be diagnosed via immunofluorescence (IF) microscopy analyses. Here, using IF microscopy, we demonstrated that in individuals with PCD and CP defects, the CP-associated protein SPEF2 is absent in HYDIN-mutant cells, revealing its dependence on functional HYDIN. Next, we performed IF analyses of SPEF2 in respiratory cells from 189 individuals with suspected PCD and situs solitus. Forty-one of the 189 individuals had undetectable SPEF2 and were subjected to a genetic analysis, which revealed one novel loss-of-function mutation in SPEF2 and three reported and 13 novel HYDIN mutations in 15 individuals. The remaining 25 individuals are good candidates for new, as-yet uncharacterized PCD variants that affect the CP apparatus. SPEF2 mutations have been associated with male infertility but have not previously been identified to cause PCD. We identified a mutation of SPEF2 that is causative for PCD with a CP defect. We conclude that SPEF2 IF analyses can facilitate the detection of CP defects and evaluation of the pathogenicity of HYDIN variants, thus aiding the molecular diagnosis of CP defects.
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Proteínas de Ciclo Celular/deficiência , Cílios/química , Transtornos da Motilidade Ciliar/genética , Proteínas dos Microfilamentos/genética , Axonema/química , Axonema/ultraestrutura , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/fisiologia , Transtornos da Motilidade Ciliar/diagnóstico , Transtornos da Motilidade Ciliar/patologia , Códon sem Sentido , Estudos de Coortes , Análise Mutacional de DNA , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Feminino , Heterogeneidade Genética , Homozigoto , Humanos , Mutação com Perda de Função , Masculino , Proteínas dos Microfilamentos/fisiologia , Microscopia Eletrônica de Transmissão , Microscopia de Fluorescência , Depuração Mucociliar/genética , Mutação , Mutação de Sentido Incorreto , Linhagem , Cultura Primária de Células , Situs Inversus/diagnóstico , Situs Inversus/genética , Situs Inversus/patologiaRESUMO
INTRODUCTION: Nasal nitric oxide (nNO) is extremely low in individuals with primary ciliary dyskinesia (PCD) and is recommended as part of early workup. We investigated whether tidal breathing sampling for a few seconds was as discriminative between PCD and healthy controls (HC) as conventional tidal breathing sampling (cTB-nNO) for 20-30 s. METHODS: We performed very rapid sampling of tidal breathing (vrTB-nNO) for 2, 4 and 6 s, respectively. Vacuum sampling with applied negative pressure (vrTB-nNOvac; negative pressure was applied by pinching the sampling tube) for < 2 s resulted in enhanced suction of nasal air during measurement. Feasibility, success rate, discriminatory capacity, repeatability and agreement were assessed for all four sampling modalities. RESULTS: We included 13 patients with PCD, median (IQR) age of 21.8 (12.2-27.7) years and 17 HC, 25.3 (14.5-33.4) years. Measurements were highly feasible (96.7% success rate). Measured NO values with vrTB-nNO modalities differed significantly from TB-nNO measurements (HC: p < 0.001, PCD: p < 0.05). All modalities showed excellent discrimination. The vacuum method gave remarkably high values of nNO in both groups (1865 vs. 86 ppb), but retained excellent discrimination. vrTB-nNO4sec, vrTB-nNO6sec and vrTB-nNOvac showed identical specificity to cTB-nNO (all: 1.0, 95% CI 0.77-1.0). CONCLUSION: vrTB-nNO sampling requires only a few seconds of probe-in-nose time, is feasible, and provides excellent discrimination between PCD and HC. Rapid TB-nNO sampling needs standardisation and further investigations in infants, young children and patients referred for PCD workup.
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Testes Respiratórios , Transtornos da Motilidade Ciliar/diagnóstico , Óxido Nítrico/metabolismo , Adolescente , Adulto , Idoso , Biomarcadores/metabolismo , Estudos de Casos e Controles , Criança , Transtornos da Motilidade Ciliar/metabolismo , Transtornos da Motilidade Ciliar/fisiopatologia , Estudos de Viabilidade , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Nariz , Valor Preditivo dos Testes , Estudo de Prova de Conceito , Reprodutibilidade dos Testes , Fatores de Tempo , Adulto JovemRESUMO
Multiciliated epithelial cells protect the upper and lower airways from chronic bacterial infections by moving mucus and debris outward. Congenital disorders of ciliary beating, referred to as primary ciliary dyskinesia (PCD), are characterized by deficient mucociliary clearance and severe, recurrent respiratory infections. Numerous genetic defects, most of which can be detected by transmission electron microscopy (TEM), are so far known to cause different abnormalities of the ciliary axoneme. However, some defects are not regularly discernable by TEM because the ciliary architecture of the axoneme remains preserved. This applies in particular to isolated defects of the nexin links, also known as the nexin-dynein regulatory complex (N-DRC), connecting the peripheral outer microtubular doublets. Immunofluorescence analyses of respiratory cells from PCD-affected individuals detected a N-DRC defect. Genome-wide exome sequence analyses identified recessive loss-of-function mutations in GAS8 encoding DRC4 in three independent PCD-affected families.
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Proteínas do Citoesqueleto/genética , Dineínas/antagonistas & inibidores , Síndrome de Kartagener/etiologia , Complexos Multiproteicos/antagonistas & inibidores , Mutação/genética , Proteínas de Neoplasias/genética , Nexinas de Proteases/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal , Adulto , Animais , Western Blotting , Criança , Cílios/fisiologia , Dineínas/genética , Exoma/genética , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Síndrome de Kartagener/patologia , Masculino , Proteínas de Membrana , Camundongos , Camundongos Knockout , Microscopia Eletrônica de Transmissão , Microscopia de Fluorescência , Complexos Multiproteicos/genética , Mucosa Nasal/citologia , Mucosa Nasal/metabolismo , Óxido Nítrico/análise , Linhagem , Fenótipo , Prognóstico , Nexinas de Proteases/genética , Sistema Respiratório , Adulto JovemRESUMO
Nasal nitric oxide (NO) discriminates between patients with primary ciliary dyskinesia (PCD) and healthy individuals. We report feasibility of measurement and natural evolution of nasal NO and upon the impact of respiratory tract infection (RTI) on nasal NO in healthy infants (HI), followed from birth until age 2â years, with comparison to nasal NO in infant PCD.Tidal-breathing nasal NO measurements were performed at scheduled visits at 2â weeks old and at 4, 8, 12, 18 and 24â months old, with extra visits during RTIs. Historical nasal NO measurements for infant PCD were included for comparison.Altogether, 224 nasal NO measurements were performed in 44 enrolled infants. Median newborn nasal NO was 46 ppb (interquartile range (IQR) 29-69 ppb), increasing at a rate of 5.4% per month up to 283â ppb (IQR 203-389 ppb) at the age of 2â years. RTIs in 27 out of 44 infants temporarily suppressed nasal NO by 79%. Values for nasal NO in seven infants with PCD ranged from 6-80â ppb. The success rate to accept nasal NO sampling was 223 out of 224 measurements (99.6%).Tidal-breathing nasal NO measurement was indeed feasible in infancy and nasal NO in HI increased significantly up to 2â years of age, in opposition to nasal NO in PCD cases, which stayed low past 2â years of age. RTI episodes caused marked, temporary reductions in nasal NO in HI indistinguishable from that in infant PCD, suggesting that nasal NO should be measured in RTI-free intervals.
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Síndrome de Kartagener/metabolismo , Óxido Nítrico/metabolismo , Infecções Respiratórias/metabolismo , Testes Respiratórios , Estudos de Casos e Controles , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Modelos Lineares , Estudos Longitudinais , Masculino , Estudos Prospectivos , Respiração , Sensibilidade e EspecificidadeRESUMO
A diverse family of cytoskeletal dynein motors powers various cellular transport systems, including axonemal dyneins generating the force for ciliary and flagellar beating essential to movement of extracellular fluids and of cells through fluid. Multisubunit outer dynein arm (ODA) motor complexes, produced and preassembled in the cytosol, are transported to the ciliary or flagellar compartment and anchored into the axonemal microtubular scaffold via the ODA docking complex (ODA-DC) system. In humans, defects in ODA assembly are the major cause of primary ciliary dyskinesia (PCD), an inherited disorder of ciliary and flagellar dysmotility characterized by chronic upper and lower respiratory infections and defects in laterality. Here, by combined high-throughput mapping and sequencing, we identified CCDC151 loss-of-function mutations in five affected individuals from three independent families whose cilia showed a complete loss of ODAs and severely impaired ciliary beating. Consistent with the laterality defects observed in these individuals, we found Ccdc151 expressed in vertebrate left-right organizers. Homozygous zebrafish ccdc151(ts272a) and mouse Ccdc151(Snbl) mutants display a spectrum of situs defects associated with complex heart defects. We demonstrate that CCDC151 encodes an axonemal coiled coil protein, mutations in which abolish assembly of CCDC151 into respiratory cilia and cause a failure in axonemal assembly of the ODA component DNAH5 and the ODA-DC-associated components CCDC114 and ARMC4. CCDC151-deficient zebrafish, planaria, and mice also display ciliary dysmotility accompanied by ODA loss. Furthermore, CCDC151 coimmunoprecipitates CCDC114 and thus appears to be a highly evolutionarily conserved ODA-DC-related protein involved in mediating assembly of both ODAs and their axonemal docking machinery onto ciliary microtubules.
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Dineínas do Axonema/metabolismo , Cílios/patologia , Síndrome de Kartagener/genética , Proteínas Associadas aos Microtúbulos/fisiologia , Mutação/genética , Animais , Dineínas do Axonema/genética , Axonema/genética , Células Cultivadas , Cílios/metabolismo , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Exoma/genética , Feminino , Imunofluorescência , Humanos , Immunoblotting , Imunoprecipitação , Hibridização In Situ , Síndrome de Kartagener/metabolismo , Síndrome de Kartagener/patologia , Masculino , Camundongos , Camundongos Knockout , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Linhagem , Fenótipo , Técnicas do Sistema de Duplo-Híbrido , Peixe-Zebra/genética , Peixe-Zebra/crescimento & desenvolvimento , Peixe-Zebra/metabolismoRESUMO
Primary ciliary dyskinesia (PCD) is a ciliopathy characterized by airway disease, infertility, and laterality defects, often caused by dual loss of the inner dynein arms (IDAs) and outer dynein arms (ODAs), which power cilia and flagella beating. Using whole-exome and candidate-gene Sanger resequencing in PCD-affected families afflicted with combined IDA and ODA defects, we found that 6/38 (16%) carried biallelic mutations in the conserved zinc-finger gene BLU (ZMYND10). ZMYND10 mutations conferred dynein-arm loss seen at the ultrastructural and immunofluorescence level and complete cilia immotility, except in hypomorphic p.Val16Gly (c.47T>G) homozygote individuals, whose cilia retained a stiff and slowed beat. In mice, Zmynd10 mRNA is restricted to regions containing motile cilia. In a Drosophila model of PCD, Zmynd10 is exclusively expressed in cells with motile cilia: chordotonal sensory neurons and sperm. In these cells, P-element-mediated gene silencing caused IDA and ODA defects, proprioception deficits, and sterility due to immotile sperm. Drosophila Zmynd10 with an equivalent c.47T>G (p.Val16Gly) missense change rescued mutant male sterility less than the wild-type did. Tagged Drosophila ZMYND10 is localized primarily to the cytoplasm, and human ZMYND10 interacts with LRRC6, another cytoplasmically localized protein altered in PCD. Using a fly model of PCD, we conclude that ZMYND10 is a cytoplasmic protein required for IDA and ODA assembly and that its variants cause ciliary dysmotility and PCD with laterality defects.
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Cílios/genética , Dineínas/genética , Infertilidade Masculina/genética , Síndrome de Kartagener/genética , Proteínas/genética , Sistema Respiratório/metabolismo , Proteínas Supressoras de Tumor/genética , Animais , Axonema/genética , Axonema/metabolismo , Axonema/patologia , Cílios/metabolismo , Cílios/patologia , Proteínas do Citoesqueleto , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Dineínas/metabolismo , Exoma , Feminino , Regulação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Infertilidade Masculina/metabolismo , Infertilidade Masculina/patologia , Síndrome de Kartagener/metabolismo , Síndrome de Kartagener/patologia , Masculino , Camundongos , Mutação , Linhagem , Estrutura Terciária de Proteína , Proteínas/metabolismo , Sistema Respiratório/patologia , Proteínas Supressoras de Tumor/metabolismoRESUMO
Primary ciliary dyskinesia (PCD) is a genetically heterogeneous recessive disorder caused by several distinct defects in genes responsible for ciliary beating, leading to defective mucociliary clearance often associated with randomization of left/right body asymmetry. Individuals with PCD caused by defective radial spoke (RS) heads are difficult to diagnose owing to lack of gross ultrastructural defects and absence of situs inversus. Thus far, most mutations identified in human radial spoke genes (RSPH) are loss-of-function mutations, and missense variants have been rarely described. We studied the consequences of different RSPH9, RSPH4A, and RSPH1 mutations on the assembly of the RS complex to improve diagnostics in PCD. We report 21 individuals with PCD (16 families) with biallelic mutations in RSPH9, RSPH4A, and RSPH1, including seven novel mutations comprising missense variants, and performed high-resolution immunofluorescence analysis of human respiratory cilia. Missense variants are frequent genetic defects in PCD with RS defects. Absence of RSPH4A due to mutations in RSPH4A results in deficient axonemal assembly of the RS head components RSPH1 and RSPH9. RSPH1 mutant cilia, lacking RSPH1, fail to assemble RSPH9, whereas RSPH9 mutations result in axonemal absence of RSPH9, but do not affect the assembly of the other head proteins, RSPH1 and RSPH4A. Interestingly, our results were identical in individuals carrying loss-of-function mutations, missense variants, or one amino acid deletion. Immunofluorescence analysis can improve diagnosis of PCD in patients with loss-of-function mutations as well as missense variants. RSPH4A is the core protein of the RS head.
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Proteínas do Citoesqueleto/genética , Proteínas de Ligação a DNA/genética , Síndrome de Kartagener/diagnóstico , Proteínas/genética , Adolescente , Adulto , Criança , Pré-Escolar , Proteínas do Citoesqueleto/metabolismo , Análise Mutacional de DNA , Proteínas de Ligação a DNA/metabolismo , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Síndrome de Kartagener/genética , Síndrome de Kartagener/metabolismo , Masculino , Mutação de Sentido Incorreto , Multimerização Proteica , Proteínas/metabolismo , Adulto JovemRESUMO
Primary ciliary dyskinesia (PCD) is a genetically heterogeneous recessive disorder characterized by defective cilia and flagella motility. Chronic destructive-airway disease is caused by abnormal respiratory-tract mucociliary clearance. Abnormal propulsion of sperm flagella contributes to male infertility. Genetic defects in most individuals affected by PCD cause randomization of left-right body asymmetry; approximately half show situs inversus or situs ambiguous. Almost 70 years after the hy3 mouse possessing Hydin mutations was described as a recessive hydrocephalus model, we report HYDIN mutations in PCD-affected persons without hydrocephalus. By homozygosity mapping, we identified a PCD-associated locus, chromosomal region 16q21-q23, which contains HYDIN. However, a nearly identical 360 kb paralogous segment (HYDIN2) in chromosomal region 1q21.1 complicated mutational analysis. In three affected German siblings linked to HYDIN, we identified homozygous c.3985G>T mutations that affect an evolutionary conserved splice acceptor site and that subsequently cause aberrantly spliced transcripts predicting premature protein termination in respiratory cells. Parallel whole-exome sequencing identified a homozygous nonsense HYDIN mutation, c.922A>T (p.Lys307(∗)), in six individuals from three Faroe Island PCD-affected families that all carried an 8.8 Mb shared haplotype across HYDIN, indicating an ancestral founder mutation in this isolated population. We demonstrate by electron microscopy tomography that, consistent with the effects of loss-of-function mutations, HYDIN mutant respiratory cilia lack the C2b projection of the central pair (CP) apparatus; similar findings were reported in Hydin-deficient Chlamydomonas and mice. High-speed videomicroscopy demonstrated markedly reduced beating amplitudes of respiratory cilia and stiff sperm flagella. Like the hy3 mouse model, all nine PCD-affected persons had normal body composition because nodal cilia function is apparently not dependent on the function of the CP apparatus.
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Genes Recessivos , Síndrome de Kartagener/genética , Proteínas dos Microfilamentos/genética , Mutação , Adulto , Sequência de Bases , Cromossomos Humanos Par 1 , Cromossomos Humanos Par 16 , Cílios/genética , Análise Mutacional de DNA/métodos , Feminino , Loci Gênicos/genética , Predisposição Genética para Doença , Haplótipos , Homozigoto , Humanos , Hidrocefalia/genética , Masculino , Dados de Sequência Molecular , Linhagem , Splicing de RNA/genética , Irmãos , Situs Inversus/genéticaRESUMO
Background: Consistent use of reliable and clinically appropriate outcome measures is a priority for clinical trials, with clear definitions to allow comparability. We aimed to develop a core outcome set (COS) for pulmonary disease interventions in primary ciliary dyskinesia (PCD). Methods: A multidisciplinary international PCD expert panel was set up. A list of outcomes was created based on published literature. Using a modified three-round e-Delphi technique, the panel was asked to decide on relevant end-points related to pulmonary disease interventions and how they should be reported. First, inclusion of an outcome in the COS was determined. Second, the minimum information that should be reported per outcome. The third round finalised statements. Consensus was defined as ≥80% agreement among experts. Results: During the first round, experts reached consensus on four out of 24 outcomes to be included in the COS. Five additional outcomes were discussed in subsequent rounds for their use in different subsettings. Consensus on standardised methods of reporting for the COS was reached. Spirometry, health-related quality-of-life scores, microbiology and exacerbations were included in the final COS. Conclusion: This expert consensus resulted in a COS for clinical trials on pulmonary health among people with PCD.
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Background: Primary ciliary dyskinesia (PCD) is an inherited disorder in which dyskinetic cilia cause impaired mucociliary clearance of upper and lower airways. Airway ciliary movement can be indirectly tested in vivo after administration of a radiolabelled tracer to the lower airways for assessment of pulmonary mucociliary clearance or to the nose for assessing nasal mucociliary clearance (NMC). With this study, we investigated NMC as a quantifiable study outcome parameter in patients with PCD. Material and methods: This single centre proof-of-concept study on NMC velocity investigated patients with PCD across different genotypes and nasal nitric oxide (nasal NO) levels. Healthy controls were used for comparison. NMC was determined as velocity in mm·min-1 of a nasally applied 99mTc-albumin colloid tracer. Using a gamma camera, repeated dynamic series of images each lasting 30â s were acquired during a 10-minute period and digitally stored. Results: NMC velocity was investigated in seven patients with PCD (aged 9-31â years) and five adult healthy controls. Mean NMC velocity in healthy controls (8.5â mm·min-1) was significantly higher compared with people with PCD (0.00â mm·min-1, p<0.0001). NMC was completely absent in all included patients with PCD across different PCD genotypes and regardless of nasal NO values. The success rate of the test was 100% in both groups. Conclusion: NMC velocity discriminated highly significantly between patients with PCD and healthy controls. We suggest here a fast and feasible set up for NMC measurements that is easily applicable for any clinical trial involving PCD medication aimed for the nasal compartment, a step before or parallel to conducting clinical trials investigating whole-lung ciliary function in PCD.
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Background: Pulmonary radioaerosol mucociliary clearance (PRMC) is a reliable method for assessing in vivo whole lung mucociliary clearance and has been used at the Danish PCD Centre as a supplementary diagnostic test for primary ciliary dyskinesia (PCD) for more than two decades. This study aimed to investigate genotype-specific differences in PRMC measures and evaluate its potential as an outcome parameter. Material and methods: The study was based on a retrospective analysis of PRMC tests performed over a 24-year period (1999-2022) in individuals referred for PCD work-up and included patients with genetically confirmed PCD and non-PCD controls. Patients inhaled nebulised technetium-albumin-colloid before static and dynamic imaging was obtained. Three parameters were evaluated: 1-h lung retention (LR1), tracheobronchial velocity (TBV) and cough clearance. Results: The study included 69 patients from the Danish PCD cohort, representing 26 different PCD genotypes. Mucociliary clearance by PRMC was consistently absent in most PCD patients, regardless of genotype. However, a single patient with a CCDC103 mutation, preserved ciliary function and normal nasal nitric oxide levels exhibited normal LR1 and low TBV values. Voluntary cough significantly improved clearance, with a median improvement of 11% (interquartile range 4-24%). Conclusion: Absent mucociliary clearance by PRMC should be expected in PCD regardless of genotype but residual ciliary function could result in measurable PRMC. This indicates a potential for PRMC to detect improvements in ciliary function if this can be restored. Addressing involuntary cough and peripheral deposition of radioaerosol is important if PRMC is to be used as an outcome measure in future clinical PCD trials.
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BACKGROUND: The lung clearance index (LCI) derived from the multiple breath inert gas washout (MBW) test reflects global ventilation distribution inhomogeneity. It is more sensitive than forced expiratory volume in 1 s (FEV(1)) for detecting abnormal airway function and correlates closely with structural lung damage in children with cystic fibrosis, which shares features with primary ciliary dyskinesia (PCD). Normalised phase III slope indices S(cond) and S(acin) reflect function of the small conducting and acinar airways, respectively. The involvement of the peripheral airways assessed by MBW tests has not been previously described in PCD. METHODS: A cross-sectional MBW study was performed in 27 children and adolescents with verified PCD, all clinically stable and able to perform lung function tests. LCI, S(cond) (n=23) and S(acin) (n=23) were derived from MBW using a mass spectrometer and sulfur hexafluoride as inert marker gas. MBW indices were compared with present age, age at diagnosis and spirometry findings, and were related to published normative values. RESULTS: LCI, S(cond) and S(acin) were abnormal in 85%, 96% and 78% of patients with PCD and in 81%, 93% and 79%, respectively, of 13/27 subjects with normal FEV(1). LCI and S(acin) correlated significantly while S(cond) did not correlate with any other lung function parameters. None of the lung function measurements correlated with age or age at diagnosis. CONCLUSIONS: PCD is characterised by marked peripheral airway dysfunction. MBW seems promising in the early detection of lung damage, even in young patients with PCD. The relationship of MBW indices to the outcome of long-term disease and their role in the management of PCD need to be assessed.
Assuntos
Síndrome de Kartagener/fisiopatologia , Depuração Mucociliar/fisiologia , Ventilação Pulmonar/fisiologia , Adolescente , Criança , Pré-Escolar , Cílios/ultraestrutura , Estudos Transversais , Fibrose Cística , Feminino , Seguimentos , Volume Expiratório Forçado/fisiologia , Humanos , Lactente , Recém-Nascido , Síndrome de Kartagener/patologia , Pulmão/patologia , Pulmão/fisiopatologia , Masculino , Prognóstico , Testes de Função Respiratória , Índice de Gravidade de DoençaRESUMO
Nasal nitric oxide (nNO) measurements are used in the assessment of patients suspected of having primary ciliary dyskinesia (PCD), but recommendations for performing such measurements have not focused on children and do not include all current practices. To guide the development of a European Respiratory Society-supported technical standard for nNO measurement in children, an international online survey was conducted to better understand current measurement practices among providers involved in PCD diagnostics. 78 professionals responded, representing 65 centres across 18 countries, mainly in Europe and North America. Nearly all centres measured nNO in children and more than half performed measurements before 5â years of age. The test was often postponed in children with signs of acute airway infection. In Europe, the electrochemical technique was more frequently used than chemiluminescence. A similar proportion of centres performed measurements during exhalation against a resistance (49 out of 65) or during tidal breathing (50 out of 65); 15 centres used only exhalation against a resistance and 15 used only tidal breathing. The cut-off values used to discriminate PCD were consistent across centres using chemiluminescence analysers; these centres reported results as an output (nL·min-1). Cut-off values were highly variable across centres using electrochemical devices, and nNO concentrations were typically reported as ppb. This survey is the first to determine real-world use of nNO measurements globally and revealed remarkable variability in methodology, equipment and interpretation. These findings will help standardise methods and training.
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Primary ciliary dyskinesia (PCD) is a rare genetic disorder characterised by impaired mucociliary clearance leading to irreversible lung damage. In contrast to other rare lung diseases like cystic fibrosis (CF), there are only few clinical trials and limited evidence-based treatments. Management is mainly based on expert opinions and treatment is challenging due to a wide range of clinical manifestations and disease severity. To improve clinical and translational research and facilitate development of new treatments, the clinical trial network for PCD (PCD-CTN) was founded in 2020 under the framework of the European Reference Network (ERN)-LUNG PCD Core. Applications from European PCD sites interested in participating in the PCD-CTN were requested. Inclusion criteria consisted of patient numbers, membership of ERN-LUNG PCD Core, use of associated standards of care, experience in PCD and/or CF clinical research, resources to run clinical trials, good clinical practice (GCP) certifications and institutional support. So far, applications from 22 trial sites in 18 European countries have been approved, including >1400 adult and >1600 paediatric individuals with PCD. The PCD-CTN is headed by a coordinating centre and consists of a steering and executive committee, a data safety monitoring board and committees for protocol review, training and standardisation. A strong association with patient organisations and industrial companies are further cornerstones. All participating trial sites agreed on a code of conduct. As CTNs from other diseases have demonstrated successfully, this newly formed PCD-CTN operates to establish evidence-based treatments for this orphan disease and to bring new personalised treatment approaches to patients.