Skewed X-chromosome inactivation drives the proportion of DNAAF6-defective airway motile cilia and variable expressivity in primary ciliary dyskinesia.

BACKGROUND: Primary ciliary dyskinesia (PCD) is a rare airway disorder caused by defective motile cilia. Only male patients have been reported with pathogenic mutations in X-linked DNAAF6, which result in the absence of ciliary dynein arms, whereas their heterozygous mothers are supposedly healthy. Our objective was to assess the possible clinical and ciliary consequences of X-chromosome inactivation (XCI) in these mothers. METHODS: XCI patterns of six mothers of male patients with DNAAF6-related PCD were determined by DNA-methylation studies and compared with their clinical phenotype (6/6 mothers), as well as their ciliary phenotype (4/6 mothers), as assessed by immunofluorescence and high-speed videomicroscopy analyses. The mutated X chromosome was tracked to assess the percentage of cells with a normal inactivated DNAAF6 allele. RESULTS: The mothers' phenotypes ranged from absence of symptoms to mild/moderate or severe airway phenotypes, closely reflecting their XCI pattern. Analyses of the symptomatic mothers' airway ciliated cells revealed the coexistence of normal cells and cells with immotile cilia lacking dynein arms, whose ratio closely mirrored their XCI pattern. CONCLUSION: This study highlights the importance of searching for heterozygous pathogenic DNAAF6 mutations in all female relatives of male PCD patients with a DNAAF6 defect, as well as in females consulting for mild chronic respiratory symptoms. Our results also demonstrate that about one-third-ranging from 20% to 50%-normal ciliated airway cells sufficed to avoid severe PCD, a result paving the way for gene therapy.

TTC12 Loss-of-Function Mutations Cause Primary Ciliary Dyskinesia and Unveil Distinct Dynein Assembly Mechanisms in Motile Cilia Versus Flagella.

Cilia and flagella are evolutionarily conserved organelles whose motility relies on the outer and inner dynein arm complexes (ODAs and IDAs). Defects in ODAs and IDAs result in primary ciliary dyskinesia (PCD), a disease characterized by recurrent airway infections and male infertility. PCD mutations in assembly factors have been shown to cause a combined ODA-IDA defect, affecting both cilia and flagella. We identified four loss-of-function mutations in TTC12, which encodes a cytoplasmic protein, in four independent families in which affected individuals displayed a peculiar PCD phenotype characterized by the absence of ODAs and IDAs in sperm flagella, contrasting with the absence of only IDAs in respiratory cilia. Analyses of both primary cells from individuals carrying TTC12 mutations and human differentiated airway cells invalidated for TTC12 by a CRISPR-Cas9 approach revealed an IDA defect restricted to a subset of single-headed IDAs that are different in flagella and cilia, whereas TTC12 depletion in the ciliate Paramecium tetraurelia recapitulated the sperm phenotype. Overall, our study, which identifies TTC12 as a gene involved in PCD, unveils distinct dynein assembly mechanisms in human motile cilia versus flagella.

Otitis media in severe asthma associated with chronic rhinosinusitis with nasal polyps: a new therapeutic target for biologics.

Background: Chronic rhinosinusitis with nasal polyps (CRSwNPs) is associated with otitis media with effusion (OME) in about 25% of cases. The objective of this study was to assess the clinical efficacy of the 4 biologic agents currently available in France for severe asthma (omalizumab, mepolizumab, benralizumab and dupilumab) in 17 patients followed for both asthma and chronic rhinosinusitis with nasal polyps (CRSwNP) and presenting otitis media with effusion (OME) on otoscopy. Methods: It was a multicenter retrospective study performed in 4 academic ENT and respiratory departments in Paris, France, with assessment of the clinical evolution of 17 patients with severe eosinophilic asthma and with chronic refractory OME and CRSwNPs treated by biologic agents. Global evaluation of treatment effectiveness (GETE) on asthma, CRSwNP and OME was classified on a 5-point scale as 1, excellent; 2, good; 3, moderate; 4, poor; or 5, symptoms worsening. Response was defined as an excellent/good score (1 or 2). Results: 17 patients were prescribed a total of 30 biologics. The evolution of OME did not follow that of asthma and CRSwNPs in 15 (88%) and 12 (70%) cases, respectively. Concerning OME, 19/30 (63%) patients were non-responders. Among the 10 patients who successively received ≥ 2 biologic agents, the OME response differed, depending on the considered agent Dupilumab had the highest response rate. Conclusions: Resistant OME, associated with asthma and chronic rhinosinusitis with nasal polyps, can present a disconnected evolution under biologics. CRSwNP-associated OME requires a specific evaluation to define the best treatment.

Performance of an active transcutaneous bone conduction implant, the Osia system, in high frequencies and hearing in noise in users converted from a Baha attract system: a prospective study.

PURPOSE: Hearing rehabilitation through middle ear surgery can be challenging, particularly for patients with chronic otitis media, but new hearing devices offer opportunity to reach this goal. The aim was to compare hearing outcomes and quality of life in patients who were converted from a Baha Attract system to the Osia system. METHODS: Prospective observational study, in which each subject acted as their own control. Six patients who were converted from a Baha Attract to an Osia system performed audiometric and quality of life assessments with different sound processor at fitting, 3, 6 and 12 months. The speech performance in noise with the Osia system at 12 months, expressed as dB SNR, was compared to the baseline condition (Baha 5). RESULTS: The PTA4 hearing thresholds improved from 42.6 ± 11 with Baha 5 Power to 34.8 ± 13.3 with Baha 5 SuperPower and to 25.4 ± 3.5 with the Osia at 12 months, leading to a significant functional gain of 17.2 ± 10.9 dB vs Baha 5 (p < 0.02). Speech understanding in both quiet and noise was clinically improved reaching a mean SNR of less than 1 dB at 12 months with the Osia system. Quality of life outcomes improved by more than 20% at 12-month follow-up. CONCLUSION: Aided hearing thresholds and speech understanding in noise improved when patients were converted from the Baha Attract system to the Osia system. The aided hearing threshold was maintained up to 8 kHz (26 dB) with the Osia system.

[Post-COVID-19 syndrome]. / Syndrome post-COVID-19.

In the aftermath of acute infection with the severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2), a large number of symptoms persist or appear, constituting a real syndrome called "long COVID-19" or "post-COVID- 19" or "post-acute COVID-19 syndrome". Its incidence is very high, half of patients showing at least one symptom at 4-6 months after Coronarovirus infectious disease 2019 (COVID-19). They can affect many organs. The most common symptom is persistent fatigue, similar to that seen after other viral infections. Radiological pulmonary sequelae are relatively rare and not extensive. On the other hand, functional respiratory symptoms, primarily dyspnoea, are much more frequent. Dysfunctional breathing is a significant cause of dyspnoea. Cognitive disorders and psychological symptoms are also very common, with anxiety, depression and post-traumatic stress symptoms being widely described. On the other hand, cardiac, endocrine, cutaneous, digestive or renal sequelae are rarer. The symptoms generally improve after several months, even if their prevalence at two years remains significant. Most of the symptoms are favored by the severity of the initial illness, and the psychic symptoms by the female sex. The pathophysiology of most symptoms is poorly understood. The influence of the treatments used in the acute phase is also important. Vaccination, on the other hand, seems to reduce their incidence. The sheer number of affected patients makes long-term COVID-19 syndrome a public health challenge.

Mutations in DNAH17, Encoding a Sperm-Specific Axonemal Outer Dynein Arm Heavy Chain, Cause Isolated Male Infertility Due to Asthenozoospermia.

Motile cilia and sperm flagella share an evolutionarily conserved axonemal structure. Their structural and/or functional defects are associated with primary ciliary dyskinesia (PCD), a genetic disease characterized by chronic respiratory-tract infections and in which most males are infertile due to asthenozoospermia. Among the well-characterized axonemal protein complexes, the outer dynein arms (ODAs), through ATPase activity of their heavy chains (HCs), play a major role for cilia and flagella beating. However, the contribution of the different HCs (γ-type: DNAH5 and DNAH8 and ß-type: DNAH9, DNAH11, and DNAH17) in ODAs from both organelles is unknown. By analyzing five male individuals who consulted for isolated infertility and displayed a loss of ODAs in their sperm cells but not in their respiratory cells, we identified bi-allelic mutations in DNAH17. The isolated infertility phenotype prompted us to compare the protein composition of ODAs in the sperm and ciliary axonemes from control individuals. We show that DNAH17 and DNAH8, but not DNAH5, DNAH9, or DNAH11, colocalize with α-tubulin along the sperm axoneme, whereas the reverse picture is observed in respiratory cilia, thus explaining the phenotype restricted to sperm cells. We also demonstrate the loss of function associated with DNAH17 mutations in two unrelated individuals by performing immunoblot and immunofluorescence analyses on sperm cells; these analyses indicated the absence of DNAH17 and DNAH8, whereas DNAH2 and DNALI, two inner dynein arm components, were present. Overall, this study demonstrates that mutations in DNAH17 are responsible for isolated male infertility and provides information regarding ODA composition in human spermatozoa.

Mutations in Outer Dynein Arm Heavy Chain DNAH9 Cause Motile Cilia Defects and Situs Inversus.

Motile cilia move body fluids and gametes and the beating of cilia lining the airway epithelial surfaces ensures that they are kept clear and protected from inhaled pathogens and consequent respiratory infections. Dynein motor proteins provide mechanical force for cilia beating. Dynein mutations are a common cause of primary ciliary dyskinesia (PCD), an inherited condition characterized by deficient mucociliary clearance and chronic respiratory disease coupled with laterality disturbances and subfertility. Using next-generation sequencing, we detected mutations in the ciliary outer dynein arm (ODA) heavy chain gene DNAH9 in individuals from PCD clinics with situs inversus and in one case male infertility. DNAH9 and its partner heavy chain DNAH5 localize to type 2 ODAs of the distal cilium and in DNAH9-mutated nasal respiratory epithelial cilia we found a loss of DNAH9/DNAH5-containing type 2 ODAs that was restricted to the distal cilia region. This confers a reduced beating frequency with a subtle beating pattern defect affecting the motility of the distal cilia portion. 3D electron tomography ultrastructural studies confirmed regional loss of ODAs from the distal cilium, manifesting as either loss of whole ODA or partial loss of ODA volume. Paramecium DNAH9 knockdown confirms an evolutionarily conserved function for DNAH9 in cilia motility and ODA stability. We find that DNAH9 is widely expressed in the airways, despite DNAH9 mutations appearing to confer symptoms restricted to the upper respiratory tract. In summary, DNAH9 mutations reduce cilia function but some respiratory mucociliary clearance potential may be retained, widening the PCD disease spectrum.

Topological data analysis reveals genotype-phenotype relationships in primary ciliary dyskinesia.

BACKGROUND: Primary ciliary dyskinesia (PCD) is a heterogeneous inherited disorder caused by mutations in approximately 50 cilia-related genes. PCD genotype-phenotype relationships have mostly arisen from small case series because existing statistical approaches to investigating relationships have been unsuitable for rare diseases. METHODS: We applied a topological data analysis (TDA) approach to investigate genotype-phenotype relationships in PCD. Data from separate training and validation cohorts included 396 genetically defined individuals carrying pathogenic variants in PCD genes. To develop the TDA models, 12 clinical and diagnostic variables were included. TDA-driven hypotheses were subsequently tested using traditional statistics. RESULTS: Disease severity at diagnosis, measured by forced expiratory volume in 1 s (FEV1) z-score, was significantly worse in individuals with CCDC39 mutations (compared to other gene mutations) and better in those with DNAH11 mutations; the latter also reported less neonatal respiratory distress. Patients without neonatal respiratory distress had better preserved FEV1 at diagnosis. Individuals with DNAH5 mutations were phenotypically diverse. Cilia ultrastructure and beat pattern defects correlated closely to specific causative gene groups, confirming these tests can be used to support a genetic diagnosis. CONCLUSIONS: This large scale, multi-national study presents PCD as a syndrome with overlapping symptoms and variations in phenotype according to genotype. TDA modelling confirmed genotype-phenotype relationships reported by smaller studies (e.g. FEV1 worse with CCDC39 mutation) and identified new relationships, including FEV1 preservation with DNAH11 mutations and diversity of severity with DNAH5 mutations.

Deep phenotyping, including quantitative ciliary beating parameters, and extensive genotyping in primary ciliary dyskinesia.

BACKGROUND: Primary ciliary dyskinesia (PCD) is a rare genetic disorder resulting in abnormal ciliary motility/structure, extremely heterogeneous at genetic and ultrastructural levels. We aimed, in light of extensive genotyping, to identify specific and quantitative ciliary beating anomalies, according to the ultrastructural phenotype. METHODS: We prospectively included 75 patients with PCD exhibiting the main five ultrastructural phenotypes (n=15/group), screened all corresponding PCD genes and measured quantitative beating parameters by high-speed video-microscopy (HSV). RESULTS: Sixty-eight (91%) patients carried biallelic mutations. Combined outer/inner dynein arms (ODA/IDA) defect induces total ciliary immotility, regardless of the gene involved. ODA defect induces a residual beating with dramatically low ciliary beat frequency (CBF) related to increased recovery stroke and pause durations, especially in case of DNAI1 mutations. IDA defect with microtubular disorganisation induces a low percentage of beating cilia with decreased beating angle and, in case of CCDC39 mutations, a relatively conserved mean CBF with a high maximal CBF. Central complex defect induces nearly normal beating parameters, regardless of the gene involved, and a gyrating motion in a minority of ciliated edges, especially in case of RSPH1 mutations. PCD with normal ultrastructure exhibits heterogeneous HSV values, but mostly an increased CBF with an extremely high maximal CBF. CONCLUSION: Quantitative HSV analysis in PCD objectives beating anomalies associated with specific ciliary ultrastructures and genotypes. It represents a promising approach to guide the molecular analyses towards the best candidate gene(s) to be analysed or to assess the pathogenicity of the numerous sequence variants identified by next-generation-sequencing.

Patient and otolaryngologist perceptions of telemedicine during COVID-19 pandemic.

BACKGROUND: The rapid spread of the coronavirus disease 2019 and the implementation of quarantine in many European countries led to a swift change in health care delivery. Telemedicine was implemented in many otolaryngological departments to ensure the continuous care. The purpose of this study is to report our experience about telemedicine in 86 patients consulting virtually in our departments. METHODS: A total of 86 patients benefited from telemedicine consultation from April to Mai May 2020. Patients and physicians were invited to fulfill a satisfaction survey over the 3 days after the consultation. RESULTS: Patients consulted in the following fields: laryngology, voice and swallowing (N=15; 17.4%), head and neck or plastic surgery (N=34; 39.5%), rhinology (N=31; 36.1%) and otology (N=6; 7.0%). Practitioners estimated that the clinical examination would not have changed the consultation issue in 73.2% of cases. The realization of delayed clinical examination was rapidly necessary in 9.3% of cases and useless in 33.7% of cases. Five percent of patients estimated that the consultation did not bring reliable conclusion. Although the majority of patient (87.7%) would recommend telemedicine consultation to friend/family in the context of pandemic, only 44.6% would accept to replace office- consultation by telemedicine consultation outside the pandemic. CONCLUSION: Telemedicine appears to be an interesting alternative approach in situation of pandemic and lock-down. Because the patient motivation to further participate to telemedicine appears to be conditioned by the context, efforts are still required to understand the patient perception, satisfaction and fears in view of future implementation outside pandemic.