Human Respiratory Cilia Research Articles

Primary Ciliary Dyskinesia Associated Disease-Causing Variants in CCDC39 and CCDC40 Cause Axonemal Absence of Inner Dynein Arm Heavy Chains DNAH1, DNAH6, and DNAH7.

Disease-causing bi-allelic DNA variants in CCDC39 and CCDC40 are frequent causes of the hereditary disorder of primary ciliary dyskinesia (PCD). The encoded proteins form a molecular ruler complex, crucial for maintaining the 96 nm repeat units along the ciliary axonemes. Defects of those proteins cause a stiff, rapid, and flickery ciliary beating pattern, recurrent respiratory infections, axonemal disorganization, and abnormal assembly of GAS8, CCDC39, and DNALI1. We performed molecular characterization of the defects in the 96 nm axonemal ruler due to disease-causing variants in CCDC39 and CCDC40 and analyzed the effect on additional axonemal components. We identified a cohort of 51 individuals with disease-causing variants in CCDC39 and CCDC40 via next-generation sequencing techniques and demonstrated that the IDA heavy chains DNAH1, DNAH6, and DNAH7 are conspicuously absent within the respiratory ciliary axonemes by immunofluorescence analyses. Hence, we show for the first time that the centrin2 (CETN2) containing IDAs are also affected. These findings underscore the crucial role of CCDC39 and CCDC40 in the assembly and function of IDAs in human respiratory cilia. Thus, our data improve the diagnostics of axonemal ruler defects by further characterizing the associated molecular IDA defects.

Magnesium borate whiskers/La2-XCeXZr2O7 catalysts preparation on cordierite honeycomb ceramic surfaces for soot catalytic oxidation͏

Magnesium borate whiskers were used to prepare a hierarchical microstructure on cordierite ceramic substrate through the precipitation-molten salt method. The La2-XCeXZr2O7 catalyst was then loaded onto the magnesium borate whiskers to form the hierarchical microstructure. The Ce-doped catalysts exhibited a significant improvement in the catalytic activity of soot combustion compared to the La2Zr2O7 catalyst. The study employed various techniques to characterize the structural morphology and phase composition, including scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDS), Fourier transform-infrared spectroscopy (FT-IR), and X-ray diffraction (XRD). The growth of Magnesium borate whiskers (Mg2B2O5) with diameters of 100–200 nm and lengths of 2–5 μm were grown and tightly bonded to the cordierite ceramic substrate to mimic the structure of human respiratory system cilia. This could improve the filtration efficiency of soot in car exhaust. BET analysis showed that the specific surface area of cordierite increased from 0.684 m2/g to 1.363 m2/g after whisker growth. The study confirmed that the whiskers were firmly attached to the cordierite ceramic substrate, as demonstrated by the binding strength experiments. The addition of Ce metal has been found to significantly enhance soot combustion due to the presence of more active oxygen species and superior mobility of lattice oxygen species with more oxygen vacancies. The catalyst's intrinsic redox property was characterized using X-ray photoelectron spectroscopy (XPS) and H2 temperature-programmed reduction (H2-TPR). The semi-quantitative thermogravimetric analysis (TG-DTG) and the temperature-programmed oxidation (TPO) of catalysts showed that Z-La1.9Ce0·1Zr2O7 exhibited the highest catalytic activity for soot combustion, with an ignition temperature of 268 °C, a peak temperature of 418 °C, and a temperature of 90 % soot conversion of 464 °C. Furthermore, cyclic stability experiments demonstrate that the Z-La1.9Ce0·1Zr2O7 catalyst exhibits excellent structural stability and consistent catalytic performance. This distinctive microstructure holds great potential for applications in the field of DPF.

Axonemal structures reveal mechanoregulatory and disease mechanisms

Motile cilia and flagella beat rhythmically on the surface of cells to power the flow of fluid and to enable spermatozoa and unicellular eukaryotes to swim. In humans, defective ciliary motility can lead to male infertility and a congenital disorder called primary ciliary dyskinesia (PCD), in which impaired clearance of mucus by the cilia causes chronic respiratory infections1. Ciliary movement is generated by the axoneme, a molecular machine consisting of microtubules, ATP-powered dynein motors and regulatory complexes2. The size and complexity of the axoneme has so far prevented the development of an atomic model, hindering efforts to understand how it functions. Here we capitalize on recent developments in artificial intelligence-enabled structure prediction and cryo-electron microscopy (cryo-EM) to determine the structure of the 96-nm modular repeats of axonemes from the flagella of the alga Chlamydomonas reinhardtii and human respiratory cilia. Our atomic models provide insights into the conservation and specialization of axonemes, the interconnectivity between dyneins and their regulators, and the mechanisms that maintain axonemal periodicity. Correlated conformational changes in mechanoregulatory complexes with their associated axonemal dynein motors provide a mechanism for the long-hypothesized mechanotransduction pathway to regulate ciliary motility. Structures of respiratory-cilia doublet microtubules from four individuals with PCD reveal how the loss of individual docking factors can selectively eradicate periodically repeating structures.

Zebrafish Motile Cilia as a Model for Primary Ciliary Dyskinesia.

Zebrafish is a vertebrate teleost widely used in many areas of research. As embryos, they develop quickly and provide unique opportunities for research studies owing to their transparency for at least 48 h post fertilization. Zebrafish have many ciliated organs that include primary cilia as well as motile cilia. Using zebrafish as an animal model helps to better understand human diseases such as Primary Ciliary Dyskinesia (PCD), an autosomal recessive disorder that affects cilia motility, currently associated with more than 50 genes. The aim of this study was to validate zebrafish motile cilia, both in mono and multiciliated cells, as organelles for PCD research. For this purpose, we obtained systematic high-resolution data in both the olfactory pit (OP) and the left–right organizer (LRO), a superficial organ and a deep organ embedded in the tail of the embryo, respectively. For the analysis of their axonemal ciliary structure, we used conventional transmission electron microscopy (TEM) and electron tomography (ET). We characterised the wild-type OP cilia and showed, for the first time in zebrafish, the presence of motile cilia (9 + 2) in the periphery of the pit and the presence of immotile cilia (still 9 + 2), with absent outer dynein arms, in the centre of the pit. In addition, we reported that a central pair of microtubules in the LRO motile cilia is common in zebrafish, contrary to mouse embryos, but it is not observed in all LRO cilia from the same embryo. We further showed that the outer dynein arms of the microtubular doublet of both the OP and LRO cilia are structurally similar in dimensions to the human respiratory cilia at the resolution of TEM and ET. We conclude that zebrafish is a good model organism for PCD research but investigators need to be aware of the specific physical differences to correctly interpret their results.

Simulation study on the mass transport in PCL based on the ciliated dynamic system of the respiratory tract

In this paper, a two-dimensional dynamic model of human respiratory cilia is established by immersed boundary-lattice Boltzmann method. In the model, periciliary liquid and mucus layer with different flow characteristics are considered, in which the non-Newtonian fluid in the viscous layer is described by Oldroyd-B model; the area between two layers is dynamically divided by a virtual membrane. By changing the cilium beat amplitude and the depth of periciliary liquid, this paper focuses on the removal of particle impurities by cilium beat. The results show that when the initial position is near the top of cilium, the particle tends to migrate upward to approach the mucus layer. However, if the initial position is near the middle or at lower part of the cilia, most of the particles will not be transferred to the vicinity of mucus layer by the swinging cilium. It is also found that by increasing the beat amplitude of cilia, particles are more likely to approach the mucus layer. By conducting this study, the migration of the particle in periciliary liquid is discussed in detail. The results are significant to help us to know the material transport law of human respiratory tract.

Recessive DNAH9 Loss-of-Function Mutations Cause Laterality Defects and Subtle Respiratory Ciliary-Beating Defects

Dysfunction of motile monocilia, altering the leftward flow at the embryonic node essential for determination of left-right body asymmetry, is a major cause of laterality defects. Laterality defects are also often associated with reduced mucociliary clearance caused by defective multiple motile cilia of the airway and are responsible for destructive airway disease. Outer dynein arms (ODAs) are essential for ciliary beat generation, and human respiratory cilia contain different ODA heavy chains (HCs): the panaxonemally distributed γ-HC DNAH5, proximally located β-HC DNAH11 (defining ODA type 1), and the distally localized β-HC DNAH9 (defining ODA type 2). Here we report loss-of-function mutations in DNAH9 in five independent families causing situs abnormalities associated with subtle respiratory ciliary dysfunction. Consistent with the observed subtle respiratory phenotype, high-speed video microscopy demonstrates distally impaired ciliary bending in DNAH9 mutant respiratory cilia. DNAH9-deficient cilia also lack other ODA components such as DNAH5, DNAI1, and DNAI2 from the distal axonemal compartment, demonstrating an essential role of DNAH9 for distal axonemal assembly of ODAs type 2. Yeast two-hybrid and co-immunoprecipitation analyses indicate interaction of DNAH9 with the ODA components DNAH5 and DNAI2 as well as the ODA-docking complex component CCDC114. We further show that during ciliogenesis of respiratory cilia, first proximally located DNAH11 and then distally located DNAH9 is assembled in the axoneme. We propose that the β-HC paralogs DNAH9 and DNAH11 achieved specific functional roles for the distinct axonemal compartments during evolution with human DNAH9 function matching that of ancient β-HCs such as that of the unicellular Chlamydomonas reinhardtii.

DNAH11 Localization in the Proximal Region of Respiratory Cilia Defines Distinct Outer Dynein Arm Complexes.

Primary ciliary dyskinesia (PCD) is a recessively inherited disease that leads to chronic respiratory disorders owing to impaired mucociliary clearance. Conventional transmission electron microscopy (TEM) is a diagnostic standard to identify ultrastructural defects in respiratory cilia but is not useful in approximately 30% of PCD cases, which have normal ciliary ultrastructure. DNAH11 mutations are a common cause of PCD with normal ciliary ultrastructure and hyperkinetic ciliary beating, but its pathophysiology remains poorly understood. We therefore characterized DNAH11 in human respiratory cilia by immunofluorescence microscopy (IFM) in the context of PCD. We used whole-exome and targeted next-generation sequence analysis as well as Sanger sequencing to identify and confirm eight novel loss-of-function DNAH11 mutations. We designed and validated a monoclonal antibody specific to DNAH11 and performed high-resolution IFM of both control and PCD-affected human respiratory cells, as well as samples from green fluorescent protein (GFP)-left-right dynein mice, to determine the ciliary localization of DNAH11. IFM analysis demonstrated native DNAH11 localization in only the proximal region of wild-type human respiratory cilia and loss of DNAH11 in individuals with PCD with certain loss-of-function DNAH11 mutations. GFP-left-right dynein mice confirmed proximal DNAH11 localization in tracheal cilia. DNAH11 retained proximal localization in respiratory cilia of individuals with PCD with distinct ultrastructural defects, such as the absence of outer dynein arms (ODAs). TEM tomography detected a partial reduction of ODAs in DNAH11-deficient cilia. DNAH11 mutations result in a subtle ODA defect in only the proximal region of respiratory cilia, which is detectable by IFM and TEM tomography.

Effect of TH2 cytokines and interferon gamma on beat frequency of human respiratory cilia

In asthmatic airways secondary ciliary dyskinesia contributes to impaired mucociliary clearance. To investigate underlying mechanisms, we studied the effects of cytokines associated with asthma phenotype on the ciliary beat frequency (CBF) in a cell culture model of ciliated human respiratory epithelial cells. Nasal respiratory epithelial cells of 21 patients were used to prepare multicellular cells (spheroids) in the presence of the T helper (TH) 2 cytokines interleukin (IL)-4, IL-5, IL-9 and IL-13, and the TH1 cytokine interferon gamma (IFN-γ). CBF was determined by high-speed video microscopy. Addition of IL-4 and IL-13 and IL-4 + IL-13 decreased the mean CBF by 17, 21, and 22%, respectively, compared with untreated controls. Addition of IL-5 and IL-9 lead to an increase in mean CBF (20 and 10%, respectively). Lower concentrations of IFN-γ (0.1 and 1 ng/ml) decreased mean CBF and higher concentrations (10 ng/ml) increased CBF by 6%. Addition of IFN-γ to IL-13 reversed the effect of IL-13 on the CBF of spheroids. Cytokines directly influence the ciliary function of respiratory epithelium and contribute to the impaired mucociliary clearance in asthmatic disease. Our study encourages further research to investigate IFN-γ as a treatment option in diseases with impaired mucociliary clearance like asthma.

Immunofluorescence Analysis and Diagnosis of Primary Ciliary Dyskinesia with Radial Spoke Defects.

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.

Mutations in CCDC11, which encodes a coiled-coil containing ciliary protein, causes situs inversus due to dysmotility of monocilia in the left-right organizer.

In vertebrates, establishment of left-right (LR) asymmetry is dependent on cilia-driven fluid flow within the LR organizer. Mutations in CCDC11 disrupt LR asymmetry in humans, but how the gene functions in LR patterning is presently unknown. We describe a patient with situs inversus totalis carrying homozygous loss-of-function mutations in CCDC11. We show that CCDC11 is an axonemal protein in respiratory cilia, but is largely dispensable for their structure and motility. To investigate the role of CCDC11 in LR development, we studied the zebrafish homolog of the gene. Like in human respiratory cilia, loss of Ccdc11 causes minor defects in the motility of zebrafish kidney cilia, although the protein localizes to their axonemes and base. By contrast, Ccdc11 localizes exclusively to the basal bodies of cilia within Kupffer's vesicle, the organ of laterality of teleost fishes, and within the spinal canal. Moreover, the rotational motion of the cilia in these tissues of ccdc11-deficient embryos was strongly impaired. Our findings demonstrate that CCDC11 has a conserved essential function in cilia of the vertebrate LR organizer. To the best of our knowledge, this is the first ciliary component, which has a differential localization and function in different kinds of motile cilia.

Signature Motifs Identify an Acinetobacter Cif Virulence Factor with Epoxide Hydrolase Activity

Endocytic recycling of the cystic fibrosis transmembrane conductance regulator (CFTR) is blocked by the CFTR inhibitory factor (Cif). Originally discovered in Pseudomonas aeruginosa, Cif is a secreted epoxide hydrolase that is transcriptionally regulated by CifR, an epoxide-sensitive repressor. In this report, we investigate a homologous protein found in strains of the emerging nosocomial pathogens Acinetobacter nosocomialis and Acinetobacter baumannii ("aCif"). Like Cif, aCif is an epoxide hydrolase that carries an N-terminal secretion signal and can be purified from culture supernatants. When applied directly to polarized airway epithelial cells, mature aCif triggers a reduction in CFTR abundance at the apical membrane. Biochemical and crystallographic studies reveal a dimeric assembly with a stereochemically conserved active site, confirming our motif-based identification of candidate Cif-like pathogenic EH sequences. Furthermore, cif expression is transcriptionally repressed by a CifR homolog ("aCifR") and is induced in the presence of epoxides. Overall, this Acinetobacter protein recapitulates the essential attributes of the Pseudomonas Cif system and thus may facilitate airway colonization in nosocomial lung infections.

Three dimensional ultrastructure of human respiratory cilia in health and disease

Motile cilia in the respiratory tract beat to clear mucus from the airways, keeping the lung clean and free from infection. In the inherited genetic condition Primary Ciliary Dyskinesia (PCD) cilia beat is ineffective, resulting in chronic cough, recurrent chest infections and rhinosinustitis. The definitive diagnostic test for PCD is electron microscopy of ciliary ultrastructure. There are a number of known ultrastructural defects associated with PCD. We have recently established the first three dimensional model of human respiratory cilia ultrastructure using electron tomography. Electron tomography is an electron microscopy technique which allows increased resolution and visualisation of structures in three dimensions. Our model highlights key features of the axoneme which cannot be resolved using traditional electron microscopy. The aim of this study was to characterise in 3D the ultrastructure of defects associated with PCD using the normal model as a template. Tomograms of transverse and longitudinal sections of cilia were generated from nasal brush biopsies taken from patients with PCD. Key features of the cilium were resolved using sub- volume or sub-tomographic averaging and measured. We show in detail the ultrastructural phenotype of radial spoke, dynein arm and central pair defects allowing insight into the structure of human respiratory tract cilium and primary ciliary dyskinesia.

Generation of a Three-Dimensional Ultrastructural Model of Human Respiratory Cilia

The ultrastructures of cilia and flagella are highly similar and well conserved through evolution. Consequently, Chlamydomonas is commonly used as a model organism for the study of human respiratory cilia. Since detailed models of Chlamydomonas axonemes were generated using cryoelectron tomography, disparities among some of the ultrastructural features have become apparent when compared with human cilia. Extrapolating information on human disease from the Chlamydomonas model may lead to discrepancies in translational research. This study aimed to establish the first three-dimensional ultrastructural model of human cilia. Tomograms of transverse sections (n = 6) and longitudinal sections (n = 9) of human nasal respiratory cilia were generated from three healthy volunteers. Key features of the cilium were resolved using subatomic averaging, and were measured. For validation of the method, a model of the well characterized structure of Chlamydomonas reinhardtii was simultaneously generated. Data were combined to create a fully quantified three-dimensional reconstruction of human nasal respiratory cilia. We highlight key differences in the axonemal sheath, microtubular doublets, radial spokes, and dynein arms between the two structures. We show a decreased axial periodicity of the radial spokes, inner dynein arms, and central pair protrusions in the human model. We propose that this first human model will provide a basis for research into the function and structure of human respiratory cilia in health and in disease.

Sentan: A Novel Specific Component of the Apical Structure of Vertebrate Motile Cilia

Human respiratory and oviductal cilia have specific apical structures characterized by a narrowed distal portion and a ciliary crown. These structures are conserved among vertebrates that have air respiration systems; however, the molecular components of these structures have not been defined, and their functions are unknown. To identify the molecular component(s) of the cilia apical structure, we screened EST libraries to identify gene(s) that are exclusively expressed in ciliated tissues, are transcriptionally up-regulated during in vitro ciliogenesis, and are not expressed in testis (because sperm flagella have no such apical structures). One of the identified gene products, named sentan, was localized to the distal tip region of motile cilia. Using anti-sentan polyclonal antibodies and electron microscopy, sentan was shown to localize exclusively to the bridging structure between the cell membrane and peripheral singlet microtubules, which specifically exists in the narrowed distal portion of cilia. Exogenously expressed sentan showed affinity for the membrane protrusions, and a protein-lipid binding assay revealed that sentan bound to phosphatidylserine. These findings suggest that sentan is the first molecular component of the ciliary tip to bridge the cell membrane and peripheral singlet microtubules, making the distal portion of the cilia narrow and stiff to allow for better airway clearance or ovum transport.

An investigation into the effects of midazolam and propofol on human respiratory cilia beat frequency in vitro.

Patients in intensive care are known to be prone to both upper and lower respiratory tract infection. Respiratory mucus forms a barrier to infection. Mucus transport rate (MTR) depends upon both the physical properties of mucus and the action of respiratory cilia. Patients undergoing anaesthesia are known to have a reduced MTR that may be related to a depressant effect on cilia beat frequency (CBF) by anaesthetic drugs. The aim of this study was to investigate the effects of two commonly used intensive care sedative agents, midazolam and propofol, on CBF using human nasal turbinate explants in vitro. We exposed ciliated tissue from human nasal turbinate explants to midazolam and propofol in supraclinical concentrations (20 microM midazolam and 70 microM propofol) in a controlled and blinded manner for 90 min and measured CBF by the transmitted light technique. After 90 min, mean (SEM) CBF in the group exposed to midazolam and its control group were 13.0 (0.2) Hz and 12.9 (0.3) Hz, respectively. Mean (SEM) CBF in the group exposed to propofol was 13.6 (0.4) Hz and in the control group the value was 12.0 (0.6) Hz. There was no significant change in CBF (midazolam: p = 0.21, propofol: p = 0.31, MANOVA for repeated measures). We have found no effect of midazolam or propofol in supra-clinical concentrations upon CBF in human turbinate explants after a 90-min exposure. This contrasts with previous work that has shown a depressant effect of inhalational anaesthetic agents on CBF.

Comparison of isoflurane with propofol on respiratory cilia

We have investigated the effects of two techniques of clinical anaesthesia on human respiratory cilia by measuring cilia beat frequency of nasal tissue. In a randomized, controlled study, 13 patients undergoing either inhalation anaesthesia with isoflurane or total i.v. anaesthesia with propofol and alfentanil had nasal ciliated epithelial samples removed at the beginning and after 1 h of anaesthesia. Mean cilia beat frequency in the group anaesthetized with isoflurane changed significantly from 11.5 (95% confidence interval (CI) 10.7-12.2) Hz to 9.1 (8.1-10.1) Hz after anaesthesia whereas in the group anaesthetized with propofol and alfentanil there was a change from 11.5 (10.7-12.2) Hz to 11.0 (10.2-11.8) Hz (ns). The difference between the anaesthetic agents on cilia beat frequency was significant (MANOVA, P < 0.01). These data suggest that different anaesthetic agents may impair respiratory defence mechanisms to differing extents.

Displacement of gold marker in immunoelectron microscopy of human respiratory cilia.

Preembedding immunogold electron microscopy was performed to evaluate the position of outer arm dynein heavy chains in normal human respiratory cilia. Anti-dynein antibody (AD2), which is specific for sea urchin sperm flagellar dynein heavy chains, was used as primary antibody. Direct cross-sections of cilia were selected, and the distance between the center of a cilium and the center of a colloidal gold particle attached to the cilium (X) was measured. The distance between the center of a cilium and the farthest edge of an outer dynein arm of the cilium was measured by ordinary electron microscopy (Yo) and by immunoelectron microscopy (Yi). X was significantly longer than Yo and Yi. If it is assumed that the structure of respiratory cilia is dense and that antibodies are located at the outer side of the actual position of the heavy chains, then the average distance difference of approximately 90-120 A may represent the length of two conjugated antibodies. This length should be kept in mind when performing immunoelectron microscopy. The data suggest that AD2 recognizes the outer arm dynein heavy chains of normal human respiratory cilia.

A perfusion system for in vitro measurement of human cilia beat frequency

We have designed and built a perfusion system and perfusion chamber to allow investigation of the effects of anaesthetic agents on human cilia in vitro. Using this system, samples of human respiratory cilia can be maintained in a stable and controlled environment for several hours. We measured cilia beat frequency of nasal respiratory epithelium from 10 healthy volunteers; cilia beat frequency was constant over a 4-h period, and measurements were found to be in good agreement with previously published work [1]. In a separate study we investigated the effect of a sleep dose of propofol on cilia beat frequency in samples from six patients undergoing minor surgery; samples were obtained before and immediately after induction of anaesthesia with propofol 2-3 mg kg-1. There was no statistically significant difference in cilia beat frequency between data obtained before and after induction with propofol.

Immunohistochemical analysis of rat and human respiratory cilia with anti-dynein antibody: comparison between normal cilia and pathological cilia in primary ciliary dyskinesia.

Wistar Imamichi rat and human respiratory cilia were examined with anti-dynein antibody (AD2), which is specific for sea urchin sperm flagellar dynein. AD2-labelled fresh-frozen normal rat and human cilia stained clearly by immunofluorescence and the peroxidase-antiperoxidase (PAP) technique. On immunoelectron microscopy, AD2 labelled the outer dynein arms of normal human cilia. Paraffin-embedded normal human cilia also stained by immunofluorescence, although not always clearly. Neither the cilia of WIC-Hyd male rats, an animal model of Kartagener's syndrome, nor human cilia from patients with primary ciliary dyskinesia (PCD) reacted positively by the immunofluorescence or PAP technique. Western blots of normal rat cilia yielded a single band of about 450 kDa. In conclusion, AD2 recognizes the outer arm dynein heavy chains of healthy cilia and may be useful in diagnosing and classifying PCD light microscopically especially when only paraffin-embedded specimens are available. This approach may be of potential use for better defining and classifying PCD.

Calcitonin gene-related peptide in vitro stimulation of ciliary beat in human upper respiratory cilia.

Calcitonin gene-related peptide (CGRP) is one of the neuropeptides that is released in the human nasal mucosa after trigeminal nerve stimulation. We investigated the in vitro effect of CGRP on the ciliary beat frequency (CBF) of human upper respiratory cilia. Ciliated epithelium of freshly removed adenoidal tissues was used in all experiments. CBF was measured using a computerized photo-electric method. The results showed that CGRP exerted a significant dose-dependent stimulation on CBF in concentrations of 10(-9)-10(-6) M (n = 10), with a maximum of 23%. Our findings indicate that CGRP in vivo may help to protect the airway epithelium by directly stimulating ciliary beat.