scholarly journals Epithelial Cell Culture from Human Adenoids: A Functional Study Model for Ciliated and Secretory Cells

2013 ◽  
Vol 2013 ◽  
pp. 1-13 ◽  
Author(s):  
Claudia González ◽  
Marisol Espinosa ◽  
María Trinidad Sánchez ◽  
Karla Droguett ◽  
Mariana Ríos ◽  
...  
Keyword(s):  
Experimental Model ◽  
Defense Mechanism ◽  
Beat Frequency ◽  
Ciliated Cell ◽  
Functional Study ◽  
Secretory Cells ◽  
Ciliary Activity ◽  
Control Mechanisms ◽  
Mucociliary Transport

Background. Mucociliary transport (MCT) is a defense mechanism of the airway. To study the underlying mechanisms of MCT, we have both developed an experimental model of cultures, from human adenoid tissue of ciliated and secretory cells, and characterized the response to local chemical signals that control ciliary activity and the secretion of respiratory mucinsin vitro.Materials and Methods. In ciliated cell cultures, ciliary beat frequency (CBF) and intracellular Ca2+levels were measured in response to ATP, UTP, and adenosine. In secretory cultures, mucin synthesis and secretion were identified by using immunodetection. Mucin content was taken from conditioned medium and analyzed in the presence or absence of UTP.Results. Enriched ciliated cell monolayers and secretory cells were obtained. Ciliated cells showed a basal CBF of 10.7 Hz that increased significantly after exposure to ATP, UTP, or adenosine. Mature secretory cells showed active secretion of granules containing different glycoproteins, including MUC5AC.Conclusion. Culture of ciliated and secretory cells grown from adenoid epithelium is a reproducible and feasible experimental model, in which it is possible to observe ciliary and secretory activities, with a potential use as a model to understand mucociliary transport control mechanisms.

Mucociliary interaction in vitro: effects of physiological and inflammatory stimuli

1990 ◽  
Vol 68 (4) ◽  
pp. 1421-1426 ◽  
Author(s):  
Z. V. Seybold ◽  
A. T. Mariassy ◽  
D. Stroh ◽  
C. S. Kim ◽  
H. Gazeroglu ◽  
...  
Keyword(s):  
Liquid Velocity ◽  
Beat Frequency ◽  
Platelet Activating Factor ◽  
Ciliary Beat Frequency ◽  
Ciliary Activity ◽  
Base Line ◽  
Mucociliary Transport ◽  
Epithelial Surface ◽  
Surface Liquid

Mucociliary transport in the airways is governed by the interaction between ciliary activity and the depth and rheological properties of the liquids (mucus) covering the epithelial surface. A change in one of these parameters may not predict the direction and magnitude of a concomitant change in mucociliary transport. We therefore determined the effects of physiological (neurotransmitters) and pathological (inflammatory mediators) stimuli on ciliary beat frequency (CBF), surface liquid velocity (SLV), surface liquid depth (SLD), and viscoelasticity of mucus in pieces of sheep trachea (n = 5 for each treatment) mounted in a chamber such that the submucosal side was bathed with Krebs-Henseleit perfusate (KH) and the luminal side was exposed to conditioned air. SLV, SLD, and CBF were measured with a microscope provided with an electronic micrometer and strobe light. Apparent viscosity and shear elastic modulus were measured with a microcapillary method using mucus collected at the downstream end of the preparation. Control CBF, SLV, and SLD were 11.6 +/- 0.4 (SE) Hz, 91 +/- 8 micron/s, and 33 +/- 5 microns, respectively, at base line and did not change during KH perfusion for 100 min. Perfusion with both acetylcholine and epinephrine (10(-5) to 10(-3) M) produced concentration-dependent increases in mean CBF (maximum increases at 10(-3) M of 16 and 9%, P less than 0.05), whereas only acetylcholine increased mean SLV (+56% at 10(-3) M, P less than 0.05). Perfusion with platelet-activating factor (10(-7) to 10(-5) M) decreased both mean CBF and SLV in a dose-dependent fashion (-6 and -63% at 10(-5) M, P less than 0.05), whereas antigen perfusion (1:60 dilution) increased mean CBF (+10%, P less than 0.05) but decreased SLV (-47%, P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Differential effect of anesthetics on mucociliary clearance in vivo in mice

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Kyle S. Feldman ◽  
Eunwon Kim ◽  
Michael J. Czachowski ◽  
Yijen Wu ◽  
Cecilia W. Lo ◽  
...  
Keyword(s):  
Mucociliary Clearance ◽  
Defense Mechanism ◽  
Ex Vivo ◽  
Beat Frequency ◽  
Ciliary Beat Frequency ◽  
Wild Type ◽  
Sulfur Colloid ◽  
Ct System

AbstractRespiratory mucociliary clearance (MCC) is a key defense mechanism that functions to entrap and transport inhaled pollutants, particulates, and pathogens away from the lungs. Previous work has identified a number of anesthetics to have cilia depressive effects in vitro. Wild-type C57BL/6 J mice received intra-tracheal installation of 99mTc-Sulfur colloid, and were imaged using a dual-modality SPECT/CT system at 0 and 6 h to measure baseline MCC (n = 8). Mice were challenged for one hour with inhalational 1.5% isoflurane, or intraperitoneal ketamine (100 mg/kg)/xylazine (20 mg/kg), ketamine (0.5 mg/kg)/dexmedetomidine (50 mg/kg), fentanyl (0.2 mg/kg)/1.5% isoflurane, propofol (120 mg/Kg), or fentanyl/midazolam/dexmedetomidine (0.025 mg/kg/2.5 mg/kg/0.25 mg/kg) prior to MCC assessment. The baseline MCC was 6.4%, and was significantly reduced to 3.7% (p = 0.04) and 3.0% (p = 0.01) by ketamine/xylazine and ketamine/dexmedetomidine challenge respectively. Importantly, combinations of drugs containing fentanyl, and propofol in isolation did not significantly depress MCC. Although no change in cilia length or percent ciliation was expected, we tried to correlate ex-vivo tracheal cilia ciliary beat frequency and cilia-generated flow velocities with MCC and found no correlation. Our results indicate that anesthetics containing ketamine (ketamine/xylazine and ketamine/dexmedetomidine) significantly depress MCC, while combinations containing fentanyl (fentanyl/isoflurane, fentanyl/midazolam/dexmedetomidine) and propofol do not. Our method for assessing MCC is reproducible and has utility for studying the effects of other drug combinations.

Functional cross talk after activation of P2 and P1 receptors in oviductal ciliated cells

2000 ◽  
Vol 279 (3) ◽  
pp. C658-C669 ◽  
Author(s):  
Bernardo Morales ◽  
Nelson Barrera ◽  
Pablo Uribe ◽  
Claudio Mora ◽  
Manuel Villalón
Keyword(s):  
Cross Talk ◽  
Adenosine Receptors ◽  
Beat Frequency ◽  
Ciliated Cell ◽  
Ciliary Beat Frequency ◽  
Ciliary Activity ◽  
Dependent Manner ◽  
Ciliated Cells ◽  
P1 Receptors ◽  
Intracellular Ca

The presence of ATP and adenosine receptors and their role in controlling ciliary activity in oviductal ciliated cells was studied by measuring the ciliary beat frequency (CBF) in oviductal tissue cultures. ATP, adenosine, and related compounds increased the CBF in a dose-dependent manner. We established that P2 receptors of subtype 2Y2 and P1 receptors of subtype A2a mediated the responses to ATP and adenosine, respectively. We found evidence to suggest that stimulation of ciliary activity by ATP requires d- myo-inositol 1,4,5-trisphosphate [Ins(1,4,5) P 3] metabolism, intracellular Ca2+ mobilization, and protein kinase C activation. On the other hand, the adenosine effect is mediated by activation of a Gs protein-dependent pathway that enhances cAMP intracellular levels. To study the interaction between P2 and P1 receptors, cells were stimulated simultaneously with both agonists. We observed a synergistic increase of the CBF even at agonist concentrations (100 nM) that did not produce a significant response when added separately to the culture. Furthermore, a blocker of the cAMP pathway produced a reduction of the ATP response, whereas a blocker of the Ins(1,4,5) P 3 pathway also produced an inhibition of the adenosine response. Our evidence demonstrates that both ATP and adenosine receptors are present in a single ciliated cell and that a mechanism of cross talk could operate in the transduction pathways to control ciliary activity.

scholarly journals nNOS regulates ciliated cell polarity, ciliary beat frequency, and directional flow in mouse trachea

2021 ◽  
Vol 4 (5) ◽  
pp. e202000981
Author(s):  
Anatoly Mikhailik ◽  
Tatyana V Michurina ◽  
Krikor Dikranian ◽  
Stephen Hearn ◽  
Vladimir I Maxakov ◽  
...  
Keyword(s):  
Beat Frequency ◽  
Ciliated Cell ◽  
Ciliary Beat Frequency ◽  
Ciliary Activity ◽  
Ciliated Cells ◽  
No Donors ◽  
Multiciliated Cells ◽  
Human Disorders ◽  
Low Levels ◽  
Ciliary Beat

Clearance of the airway is dependent on directional mucus flow across the mucociliary epithelium, and deficient flow is implicated in a range of human disorders. Efficient flow relies on proper polarization of the multiciliated cells and sufficient ciliary beat frequency. We show that NO, produced by nNOS in the multiciliated cells of the mouse trachea, controls both the planar polarity and the ciliary beat frequency and is thereby necessary for the generation of the robust flow. The effect of nNOS on the polarity of ciliated cells relies on its interactions with the apical networks of actin and microtubules and involves RhoA activation. The action of nNOS on the beat frequency is mediated by guanylate cyclase; both NO donors and cGMP can augment fluid flow in the trachea and rescue the deficient flow in nNOS mutants. Our results link insufficient availability of NO in ciliated cells to defects in flow and ciliary activity and may thereby explain the low levels of exhaled NO in ciliopathies.

scholarly journals Flow of Room Air Leads to Rapid Changes in Mucociliary Transport in the Tracheal Epithelium

2020 ◽  
Author(s):  
Susyn Joan Kelly ◽  
Paul Martinsen ◽  
Stanislav Tatkov
Keyword(s):  
Body Temperature ◽  
Beat Frequency ◽  
Tracheal Epithelium ◽  
Mucociliary Transport ◽  
Negative Effects ◽  
Initial State ◽  
Upper Airways ◽  
Lower Airways ◽  
Humidified Air

Abstract BACKGROUND: Inspired air is heated and humidified in the nose before it reaches lower airways. This mechanism is bypassed during tracheostomy, directly exposing the lower airways to colder and drier air from the environment, which is known to have negative effects on mucociliary transport; however, little is known about how quickly mucociliary transport deteriorates. The purpose of this study was to determine the short-term effect of flowing room air on mucociliary transport in the trachea. In an ovine perfused in vitro tracheal model (N=7) the epithelium was exposed to 25 L/min of flow, heated to lamb body temperature (38 °C) and fully saturated with water vapor as the control, followed by room air (22 °C and 50% relative humidity) for a short duration, until mucociliary transport had visually stopped. Mucus transport velocity (MTV) and cilia beat frequency (CBF), as well as the area of the surface with beating cilia, were continuously measured with video-microscopy.RESULTS: Exposing the tracheal epithelium to air heated to body temperature and fully humidified resulted in stable MTV 9.5 ± 1.1 mm/min and CBF 13.4 ± 0.6 Hz. When exposed to the flow of room air, MTV slowed down to 0.1 ± 0.1 mm/min in 2.0 ± 0.4 seconds followed by a decrease in CBF to 6.7 ± 1.9 Hz, after 2.3 ± 0.8 second. Both MTV and CBF recovered to their initial state when heated and humidified air-flow was re-introduced. CONCLUSIONS: This study demonstrates mucociliary transport can deteriorate within seconds of exposing the tracheal epithelium to flowing room air. The reduction in MTV precedes slowing of CBF. Their relationship is non-linear and a minimum CBF of approximately 6 Hz is required for MTV > 0. Clinically these findings indicate a potential rapid detrimental effect of breathing with non-humidified air via bypassed upper airways.

scholarly journals CILIARY ACTIVITY OF BRONCHIAL EPITHELIUM UNDER EXPERIMENTAL COLD EXPOSURE IN VITRO

2017 ◽  
Vol 1 (66) ◽  
pp. 41-49
Author(s):  
Андрей Одиреев ◽  
Andrey Odireev ◽  
Ксения Килимченко ◽  
Kseniya Kilimchenko ◽  
Николай Безруков ◽  
...  
Keyword(s):  
Cold Stress ◽  
Cold Exposure ◽  
Beat Frequency ◽  
Ciliary Beat Frequency ◽  
Solution Temperature ◽  
Ciliary Activity ◽  
Physiological Level ◽  
Functional Disturbance ◽  
Physiological Conditions

To date, the role of dysfunction of the bronchial ciliated epithelium (BCE) in the formation of mucociliary disorders during cold exposure has not been fully established. The aim of the study was to investigate the nature and severity of changes in the ciliary motility of the BCE under the influence of a cold stimulus in vitro. Ten volunteers with asthma underwent a bronchoscopy with biopsy of the lobar bronchus mucosa. The biopsy specimens were placed in a Hank’s balanced salt solution (HBSS) on a slide, located on the thermal stage, used to simulate the effect of various temperature on the BCE. Ciliary beat frequency (CBF, Hz) was recorded using a microscope, high-sensitivity digital camera and computer with specially developed software. The initial recording of CBF was performed at 24ºC, after which the temperature of HBSS was gradually increased to physiological level and repeated recording was made at the control points (28ºC and 36ºC). Then, the temperature of the solution was lowered and CBF was registered again at 28ºC, 21ºC and 17ºC. The initial CBF of the BCE varied from 6.74 to 3.77 Hz with average of 5.25±1.48 Hz (M±m). There was a statistically significant increase in CBF when the solution was heated: at 28ºC it was 6.33±1.36 Hz (p=0.004) and remained at this level up to 36ºC – 7.14±1.33 Hz (p=0.002). A gradual decrease in the solution temperature produced a reduction in CBF in comparison with the physiological conditions: 28ºC – 6.12±1.31 Hz (p=0.121), 21ºC – 5.27±1.32 Hz (p=0.001) and 17ºC – 3.95±1.18 Hz (p=0.0001). By the end of the experiment, CBF of the BCE decreased almost 2-fold in comparison with the physiological conditions. A mathematical model that characterizes the behavior of BCE cilia under cold stress was developed. Thus, the results of the study demonstrate a marked decrease in the motor activity of the BCE cilia under cold stress, which underlines a significant contribution of the functional disturbance of BCE to the pathophysiological mechanisms of hypersecretory disorders induced by inhalation of cold air.

A neuronal mechanism for the control of latero-frontal ciliary activity in the fingernail clam, Musculium transversum

1983 ◽  
Vol 41 ◽  
pp. 658-659
Author(s):  
Anthony Paparo ◽  
Judith A. Murphy ◽  
Robert Dean
Keyword(s):  
Ciliated Cell ◽  
Ciliary Activity ◽  
Suspension Feeding ◽  
Control Mechanisms ◽  
Nerve Supply ◽  
Ciliated Cells ◽  
Bivalve Molluscs ◽  
Neuronal Mechanism ◽  
Gill Filaments ◽  
Fingernail Clams

Extensive studies have dealt with the transport of particles on the gill of suspension feeding bivalves, but our knowledge of the mechanisms involved is still incomplete. There is, however, physiological evidence that each of the ciliated cell systems in bivalve molluscs may be individually controlled. Three types of ciliated cells are distinguished on the gill of fingernail clams: frontal (FC); latero-frontal (LFC); and lateral (LC).For the most part in studies involving ciliary control mechanisms, LFC are neglected. It is thus the purpose of this study to present data which begin to elucidate a neuronal mechanism for LFC control.Gill preps were isolated with an intact nerve supply in a dish which was placed in a holder fastened to an adjustable stage of a microscope. In fields of view of 50 gill filaments, ciliary motility, chemical perfusion and electrical stimulation were observed and/or performed.

scholarly journals Combination therapy with cystic fibrosis transmembrane conductance regulator modulators augment the airway functional microanatomy

2016 ◽  
Vol 310 (10) ◽  
pp. L928-L939 ◽  
Author(s):  
Susan E. Birket ◽  
Kengyeh K. Chu ◽  
Grace H. Houser ◽  
Linbo Liu ◽  
Courtney M. Fernandez ◽  
...  
Keyword(s):  
Ion Transport ◽  
Functional Imaging ◽  
Clinical Benefit ◽  
Effective Viscosity ◽  
Beat Frequency ◽  
Ciliary Beat Frequency ◽  
Clinical Results ◽  
Mucociliary Transport ◽  
Sweat Chloride

Recently approved therapies that modulate CFTR function have shown significant clinical benefit, but recent investigations regarding their molecular mechanism when used in combination have not been consistent with clinical results. We employed micro-optical coherence tomography as a novel means to assess the mechanism of action of CFTR modulators, focusing on the effects on mucociliary clearance. Primary human airway monolayers from patients with a G551D mutation responded to ivacaftor treatment with increased ion transport, airway surface liquid depth, ciliary beat frequency, and mucociliary transport rate, in addition to decreased effective viscosity of the mucus layer, a unique mechanism established by our findings. These endpoints are consistent with the benefit observed in G551D patients treated with ivacaftor, and identify a novel mechanism involving mucus viscosity. In monolayers derived from F508del patients, the situation is more complicated, compounded by disparate effects on CFTR expression and function. However, by combining ion transport measurements with functional imaging, we establish a crucial link between in vitro data and clinical benefit, a finding not explained by ion transport studies alone. We establish that F508del cells exhibit increased mucociliary transport and decreased mucus effective viscosity, but only when ivacaftor is added to the regimen. We further show that improvement in the functional microanatomy in vitro corresponds with lung function benefit observed in the clinical trials, whereas ion transport in vitro corresponds to changes in sweat chloride. Functional imaging reveals insights into clinical efficacy and CFTR biology that significantly impact our understanding of novel therapies.

Adenosine A3 receptor-mediated potentiation of mucociliary transport and epithelial ciliary motility

2002 ◽  
Vol 282 (3) ◽  
pp. L556-L562 ◽  
Author(s):  
Manako Taira ◽  
Jun Tamaoki ◽  
Kazuyuki Nishimura ◽  
Junko Nakata ◽  
Mitsuko Kondo ◽  
...  
Keyword(s):  
Mucociliary Clearance ◽  
Beat Frequency ◽  
Blue Dye ◽  
Dependent Manner ◽  
Mucociliary Transport ◽  
Concentration Dependent Manner ◽  
Maximal Increase ◽  
Ciliary Motility

To examine the effect of adenosine A3 receptor stimulation on airway mucociliary clearance, we measured transport of Evans blue dye in rabbit trachea in vivo and ciliary motility of epithelium by the photoelectric method in vitro. Mucociliary transport was enhanced dose dependently by the selective A3 agonist N 6-(3-iodobenzyl)-5′- N-methylcarbamoyladenosine (IB-MECA) and to a lesser extent by the less-selective N 6-2-(4-amino-3-iodophenyl)ethyladenosine, whereas the A1 agonist N-cyclopentyladenosine (CPA) and the A2 agonist CGS-21680 had no effect. The effect of IB-MECA was abolished by pretreatment with the selective A3 antagonist MRS-1220 but not by the A1 antagonist 1,3-dipropyly-8-cyclopentylxanthine or the A2 antagonist 3,7-dimethyl-l-propargylxanthine. Epithelial ciliary beat frequency was increased by IB-MECA in a concentration-dependent manner, the maximal increase being 33%, and this effect was inhibited by MRS-1220. The IB-MECA-induced ciliary stimulation was not altered by the Rp diastereomer of cAMP but was greatly inhibited by Ca2+-free medium containing BAPTA-AM. Incubation with IB-MECA increased intracellular Ca2+ contents. Therefore, A3 agonist enhances airway mucociliary clearance probably through Ca2+-mediated stimulation of ciliary motility of airway epithelium.

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