Glottis effects on the cough clearance process simulated with a CFD dynamic mesh and Eulerian wall film model

In this study, we have reproduced the cough clearance process with an Eulerian wall film model. The simulated domain is based on realistic geometry from the literature, which has been improved by adding the glottis and epiglottis. The vocal fold movement has been included due to the dynamic mesh method, considering different abduction and adduction angles and velocities. The proposed methodology captures the deformation of the flexible tissue, considers non-Newtonian properties for the mucus, and enables us to reproduce a single cough or a cough epoch. The cough efficiency (CE) has been used to quantify the overall performance of the cough, considering many different boundary conditions, for the analysis of the glottis effect. It was observed that a viscous shear force is the main mechanism in the cough clearance process, while the glottis closure time and the epiglottis position do not have a significant effect on the CE. The cough assistance devices improve the CE, and the enhancement rate grows logarithmically with the operating pressure. The cough can achieve an effective mucus clearance process, even with a fixed glottis. Nevertheless, the glottis closure substantially improves the CE results.     

The mechanism of mucus clearance in cough

An instability resembling an avalanche is proposed as the mechanism by which mucus is expelled from the respiratory tract during cough. The cough event was simulated in a model airway. In these experiments, air was forced through a channel whose walls were lined with a non-Newtonian material rheologically similar to tracheal mucus. Frames from high-speed cine photographs showed an unstable event which began as an undulation of the free surface and progressed to a catastrophic clearance of the channel. Measurements of the longitudinal pressure gradient support the hypothesis that the clearance event is initiated when the total stress applied to the mucus analog exceeds its finite yield stress. A continuum model predicts that yielding occurs within the bottom layers of the mucus analog. Calculations based upon estimates of tracheal geometry and air flow show that the clearance event studied here would be expected to occur during a cough but not during normal breathing. Experiments also show that a lubricant introduced between the channel walls and the mucus blanket can reduce the air flow rate required to precipitate the clearance.     

The role of mucus sol phase in clearance by simulated cough

Using a simulated cough machine, we analyzed the effect of adding tensio-active liquids as sol phase simulant on the clearance of gel mucus simulant by cough. Polysaccharides crosslinked with sodium tetraborate were used at different concentration as gel mucus simulant. A drop of gel mucus simulant was deposited either directly on the model trachea or on a sol phase layer simulant (2% sodium dodecyl sulfate in water). The clearance of the mucus simulants was quantified by observing the movement of marker particles in the gel layer. The viscoelastic properties of gel mucus simulants were determined by using a viscoelastometer (SEFAM). The adhesive properties were analyzed by means of the platinum ring technique. The wettability of the mucus simulants was quantified by the automatic measurement of the contact angle of the drop of gel on the model trachea. We found that the addition of a sol phase significantly decreased by about 50% the adhesivity and wettability of the gel mucus simulants. This decrease was associated with a marked enhancement of cough clearance, whatever the viscoelastic properties of the gel mucus simulants. These results suggest that the sol phase is essential in bronchial respiratory mucus clearance by the cough mechanism.     

A new paradigm in respiratory hygiene: modulating respiratory secretions to contain cough bioaerosol without affecting mucus clearance

Background

Several strategies and devices have been designed to protect health care providers from acquiring transmissible respiratory diseases while providing care. In modulating the physical characteristics of the respiratory secretions to minimize the aerosolization that facilitates transmission of airborne diseases, a fundamental premise is that the prototype drugs have no adverse effect on the first line of respiratory defense, clearance of mucus by ciliary action.

Methods

To assess and demonstrate the primary mechanism of our mucomodulators (XLs), we have built our evidence moving from basic laboratory studies to an ex-vivo model and then to an in-vivo large animal model. We exposed anesthetized dogs without hypersecretion to different dose concentrations of aerosolized XL "B", XL "D" and XL "S". We assessed: cardio-respiratory pattern, tracheal mucus clearance, airway patency, and mucus viscoelastic changes.

Results

Exposure of frog palate mucus to XLs did not affect the clearance of mucus by ciliary action. Dogs maintained normal cardio-respiratory pattern with XL administration. Tracheal mucociliary clearance in anesthetized dogs indicated a sustained 40% mean increase. Tracheal mucus showed increased filance, and there was no mucus retention in the airways.

Conclusion

The ex-vivo frog palate and the in-vivo mammalian models used in this study, appear to be appropriate and complement each other to better assess the effects that our mucomodulators exert on the mucociliary clearance defence mechanism. The physiological function of the mucociliary apparatus was not negatively affected in any of the two epithelial models. Airway mucus crosslinked by mucomodulators is better cleared from an intact airway and normally functioning respiratory system, either due to enhanced interaction with cilia or airflow-dependent mechanisms. Data obtained in this study allow us to assure that we have complied with the fundamental requirement criteria established in the initial phase of developing the concept of mucomodulation: Can we modulate the physical characteristics of the respiratory secretions to reduce aerosolization without affecting normal mucociliary clearance function, or even better improving it?     

CFD transient simulation of the cough clearance process using an Eulerian wall film model

In this study, a cough cycle is reproduced using a computational methodology. The Eulerian wall film approach is proposed to simulate airway mucus flow during a cough. The reproduced airway domain is based on realistic geometry from the literature and captures the deformation of flexible tissue. To quantify the overall performance of this complex phenomenon, cough efficiency (CE) was calculated, which provided an easily reproducible measurement parameter for the cough clearance process. Moreover, the effect of mucus layer thickness was examined. The relationship between the CE and the mucus viscosity was quantified using reductions from 20 to 80%. Finally, predictions of CE values based on healthy person inputs were compared with values obtained from patients with different respiratory diseases, including chronic obstructive pulmonary disease (COPD) and respiratory muscle weakness (RMW). It was observed that CE was reduced by 50% in patients with COPD compared with that of a healthy person. On average, CE was reduced in patients with RMW to 10% of the average value of a healthy person.     

A new paradigm in respiratory hygiene: increasing the cohesivity of airway secretions to improve cough interaction and reduce aerosol dispersion

Background

Infectious respiratory diseases are transmitted to non-infected subjects when an infected person expels pathogenic microorganisms to the surrounding environment when coughing or sneezing. When the airway mucus layer interacts with high-speed airflow, droplets are expelled as aerosol; their concentration and size distribution may each play an important role in disease transmission. Our goal is to reduce the aerosolizability of respiratory secretions while interfering only minimally with normal mucus clearance using agents capable of increasing crosslinking in the mucin glycoprotein network.

Methods

We exposed mucus simulants (MS) to airflow in a simulated cough machine (SCM). The MS ranged from non-viscous, non-elastic substances (water) to MS of varying degrees of viscosity and elasticity. Mucociliary clearance of the MS was assessed on the frog palate, elasticity in the Filancemeter and the aerosol pattern in a "bulls-eye" target. The sample loaded was weighed before and after each cough maneuver. We tested two mucomodulators: sodium tetraborate (XL"B") and calcium chloride (XL "C").

Results

Mucociliary transport was close to normal speed in viscoelastic samples compared to non-elastic, non-viscous or viscous-only samples. Spinnability ranged from 2.5 ± 0.6 to 50.9 ± 6.9 cm, and the amount of MS expelled from the SCM increased from 47 % to 96 % adding 1.5 μL to 150 μL of XL "B". Concurrently, particles were inversely reduced to almost disappear from the aerosolization pattern.

Conclusion

The aerosolizability of MS was modified by increasing its cohesivity, thereby reducing the number of particles expelled from the SCM while interfering minimally with its clearance on the frog palate. An unexpected finding is that MS crosslinking increased "expectoration".     

Formation of giant liposomes promoted by divalent cations: critical role of electrostatic repulsion.

Spontaneous formation of giant unilamellar liposomes in a gentle hydration process, as well as the adhesion energy between liposomal membranes, has been found to be dependent on the concentration of divalent alkali cations, Ca2+ or Mg2+, in the medium. With electrically neutral phosphatidylcholine (PC), Ca2+ or Mg2+ at 1-30 mM greatly promoted liposome formation compared to low yields in nonelectrolyte or potassium chloride solutions. When negatively charged phosphatidylglycerol (PG) was mixed at 10%, the yield was high in nonelectrolytes but liposomes did not form at 3-10 mM CaCl2. In the adhesion test with micropipette manipulation, liposomal membranes adhered to each other only in a certain range of CaCl2 concentrations, which agreed with the range where liposome did not form. The adhesion range shifted to higher Ca2+ concentrations as the amount of PG was increased. These results indicate that the divalent cations bind to and add positive charges to the lipids, and that membranes are separated and stabilized in the form of unilamellar liposomes when net charges on the membranes produce large enough electrostatic repulsion. Under the assumption that the maximum of adhesion energy within an adhesive range corresponds to exact charge neutralization by added Ca2+, association constants of PC and PG for Ca2+ were estimated at 7.3 M(-1) and 86 M(-1), respectively, in good agreement with literature values.     

Effect of airway wall flexibility on clearance by simulated cough

In our previous study, we investigated the relationship between mucus rheology, depth of mucus layer, and clearance by simulated cough. The purpose of the present study was to examine the effect of airway wall flexibility on the clearance of mucuslike gels. Transient airflows similar to cough were generated by both positive and negative pressure, the latter to mimic the dynamic compression that occurs during real cough. As in the previous study, the trachea was modeled as a trough of rectangular cross section with only the bottom lined with the mucus simulant. Clearance was followed by observing the displacement of marker particles. Since cough clearance is intimately related to wave formation in the mucus blanket, we hypothesized that clearance might be impeded if the wave formation occurred simultaneously in the wall and its lining layer. Thus, in one set of experiments the bottom rigid surface of the model trachea was replaced with a frame over which a flexible membrane could be drawn, whereas in the other set the rigid top was replaced by the frame. We also examined the effect of negative-pressure cough in excised canine tracheae, comparing the case where the tracheal membrane was free to deform vs. the case where it was secured. For the rigid-walled model, clearance by positive or negative pressure, with matched flow pattern, was the same. With the mucus simulant lining the flexible bottom surface, clearance increased with increasing membrane flexibility for negative-pressure cough and decreased for positive-pressure cough.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of phospholipid mixtures and surfactant formulations on rheology of polymeric gels,simulating mucus,at shear rates experienced in the tracheobronchial tree

A surface active layer consisting mainly of phospholipids lines the human conducting airways. Dysfunction of this layer could play a role in the pathogenesis of chronic obstructive airway diseases like asthma and chronic bronchitis. Replacement therapy with exogenous surfactants is being considered in such conditions. The relationship between surfactants and mucus viscosity would be important for such an application. Respiratory mucus is composed of high molecular weight glycoprotein molecules which form temporary cross-links and entanglements to form a gel-like material. The present paper studies the interaction of three therapeutic surfactants — Exosurf, ALEC and Survanta; the main phospholipids of lung surfactant (1,2-dipalmitoyl phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylglycerol (PG)) as well as their binary mixtures (PCPE and PCPG) in a PC:(PE or PG) ratio of 2:3; on the viscosity of mucus gel simulants (MGS — a polymeric gel consisting mainly of gum tragacanth and simulating respiratory mucus). The surfactants were studied with respect to their ability to alter MGS viscosity at shear rates ranging from 0.1498 to 51.2 s⁻¹ in a concentric cylinder viscometer at 37°C. The change in viscosity of the MGS on incubation with surfactant versus shear rate was found be non-Newtonian and to follow a power law model (coefficient of regression R²≥0.9). The shear rates experienced by a surfactant mixture, while passing through the tracheobronchial tree, were then calculated by modelling the tracheobronchial tree as cylindrical branching tubes. The equation governing the flow of a power law fluid through a cylindrical pipe was used to determine the shear experienced by a surfactant infusion as it passes through various mucus lined branches of the tracheobronchial tree. The surfactants were then compared based on their ability to alter MGS viscosity at shear rates corresponding to that of large, medium and small bronchi, as calculated by the study.     

The role of mucus viscoelasticity in cough clearance

The relationships between mucus rheology, depth of mucus layer and clearance by simulated cough were examined in a study employing a model plexiglass trachea lined with gels formed from locust bean gum crosslinked with sodium tetraborate. The viscoelastic properties of the mucus simulants were determined by magnetic rheometry at 100 rad/s and expressed as mechanical impedance (dynamic stress/strain ratio) and loss tangent. Cough was simulated by opening a solenoid valve connecting the model trachea to a pressurized tank, using an upstream flow-constrictive element to shape the flow profile to approximate the pattern seen in a normal adult. Mucus clearance was quantitated by observing the movement of contrasting marker particles placed in the mucus layer. The median particle displacement was defined as the clearance index, Cl. For any initial depth of mucus, Cl increased with driving pressure in the tank, and for a given driving pressure, Cl increased linearly with increasing mucus depth. For a given driving pressure and depth, Cl decreased with increasing mechanical impedance of the mucus. At constant mechanical impedance, Cl increased with increasing loss tangent, in other words, cough clearance was impeded more by elasticity than viscosity. Mucus clearance was associated with transient wave formation in the lining layer. Thus dependence on viscoelasticity is consistent with observations that airflow-mucus interaction and wave formation are impeded by elasticity. The clearance vs. loss tangent relationship for cough is opposite to that found for ciliary clearance (Biorheology 1980, 17, 249), suggesting a natural balance in viscosity and elasticity for mucus to be cleared by both mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)     

The role of mucus gel viscosity, spinnability, and adhesive properties in clearance by simulated cough

We investigated the role of the viscoelastic and adhesive properties of mucus gel simulants on the clearance of mucus by simulated cough. Mucus-like gels with widely varying viscoelastic properties were prepared from polysaccharides crosslinked with sodium borate. Cough was simulated by opening a solenoid valve connecting a model trachea to a pressurized tank. The clearance of gels lining the model trachea was quantified by observing marker particle transport. Viscosity elastic modulus, relaxation time and yield stress were measured with a steady-shear viscoelastometer. Spinnability (thread formation) was determined with a filancemeter. Adhesivity (surface tension) was measured by the platinum ring technique. The viscoelastic and adhesive properties of the mucus gel simulants spanned the ranges observed for bronchial secretions from patients with COPD. The relationship between simulated cough clearance and the viscoelastic and adhesive properties of the gels was analyzed by stepwise linear regression of the non-zero data matrix. The major independent variable relating to clearance was viscosity. Secondary, but highly significant dependences, were also found for spinnability and adhesivity. Elastic modulus, relaxation time and yield stress had no independent effect on cough clearance over the investigated range. The results indicate that, in the absence of airway surface liquid, cough-type clearance relates primarily with mucus gel viscosity. For a given viscosity, clearance is also impaired by spinnability, i.e. the capacity of the mucus to form threads. At constant viscosity and spinnability, clearance is further impaired by an increase in the adhesivity of the mucus. The negative dependence of each of these physical factors can be rationalized in terms of their inhibitory effect on wave formation in the mucus lining layer during high velocity airflow interaction.     

Clearance of mucus by simulated cough

We examined the relationship between mucus rheology, depth of mucus layer, and clearance by simulated cough. A model trachea was constructed of rigid Plexiglas of rectangular cross section (1 X 2 X 35 cm). The bottom of the trachea was lined with mucus simulants, gels prepared from locust bean gum cross-linked with sodium borate. Cough was simulated by opening a solenoid valve connecting the model trachea to a pressurized tank. An upstream flow-constrictive element was used to shape the flow profile of the simulated cough to approximate the pattern seen in a normal adult. Clearance of mucus was quantitated by observing the movement of contrasting marker particles floating in the mucus layer. The median particle displacement per cough maneuver was defined as the clearance index (CI). We found that CI for any initial depth of mucus increased with the driving pressure in the tank. For a given driving pressure, CI increased linearly with increasing mucus depth. For a given driving pressure and depth, CI decreased with increasing mucus cross-link density. For mucus samples with comparable levels of dynamic viscosity, samples with higher elasticity cleared less well. Mucus clearance was associated with transient wave formation in the lining layer.     

The mucus binding of Bifidobacterium lactis Bb12 is enhanced in the presence of Lactobacillus GG and Lact. delbrueckii subsp. bulgaricus

The ability to adhere to mucosal surfaces is related to many probiotic health effects. In the presence of Lactobacillus GG or Lact. bulgaricus, the adhesion of Bifidobacterium lactis Bb12 to a mucus model was more than doubled. Other tested lactobacilli did not affect the adhesion, nor was the adhesion of the lactobacilli influenced by the bifidobacteria. Co-aggregation between Bif. lactis Bb12 and the tested lactobacilli was insignificant and does not explain the observed effect. The results suggest that combinations of probiotics strains may have synergistic adhesion effects. Such specific strain combinations should also be assessed in clinical studies.     

Adhesion of food-borne bacteria to stainless steel is reduced by food conditioning films

Aims:  Preconditioning of stainless steel with aqueous cod muscle extract significantly impedes subsequent bacterial adhesion most likely due to repelling effects of fish tropomyosin. The purpose of this study was to determine if other food conditioning films decrease or enhance bacterial adhesion to stainless steel.
Methods and Results:  Attachment of Pseudomonas fluorescens AH2 to stainless steel coated with water-soluble coatings of animal origin was significantly reduced as compared with noncoated stainless steel or stainless steel coated with laboratory substrate or extracts of plant origin. Coating with animal extracts also decreases adhesion of other food-relevant bacteria. The manipulation of adhesion was not attributable to growth inhibitory effects. Chemical analysis revealed that the stainless steels were covered by homogenous layers of adsorbed proteins. The presence of tropomyocin was indicated by appearance of proteins with similar molecular weight based in sodium dodecyl sulfate–polyacrylamide gel electrophoresis, in several extracts that reduced adhesion but also extracts not containing this protein reduced bacterial adhesion, indicating that several molecular species may be involved in the phenomenon.
Conclusions:  It is a common perception that food materials facilitate bacterial adhesion to surfaces; however, this study demonstrates that aqueous coatings of food origin may actually reduce bacterial adhesion.
Significance and Impact of the Study:  Compounds from food extracts may potentially be used as nontoxic coatings to reduce bacterial attachment to inert surfaces.     

Mucoadhesive nanoparticles may disrupt the protective human mucus barrier by altering its microstructure

Mucus secretions typically protect exposed surfaces of the eyes and respiratory, gastrointestinal and female reproductive tracts from foreign entities, including pathogens and environmental ultrafine particles. We hypothesized that excess exposure to some foreign particles, however, may cause disruption of the mucus barrier. Many synthetic nanoparticles are likely to be mucoadhesive due to hydrophobic, electrostatic or hydrogen bonding interactions. We therefore sought to determine whether mucoadhesive particles (MAP) could alter the mucus microstructure, thereby allowing other foreign particles to more easily penetrate mucus. We engineered muco-inert probe particles 1 μm in diameter, whose diffusion in mucus is limited only by steric obstruction from the mucus mesh, and used them to measure possible MAP-induced changes to the microstructure of fresh human cervicovaginal mucus. We found that a 0.24% w/v concentration of 200 nm MAP in mucus induced a ～10-fold increase in the average effective diffusivity of the probe particles, and a 2- to 3-fold increase in the fraction capable of penetrating physiologically thick mucus layers. The same concentration of muco-inert particles, and a low concentration (0.0006% w/v) of MAP, had no detectable effect on probe particle penetration rates. Using an obstruction-scaling model, we determined that the higher MAP dose increased the average mesh spacing ("pore" size) of mucus from 380 nm to 470 nm. The bulk viscoelasticity of mucus was unaffected by MAP exposure, suggesting MAP may not directly impair mucus clearance or its function as a lubricant, both of which depend critically on the bulk rheological properties of mucus. Our findings suggest mucoadhesive nanoparticles can substantially alter the microstructure of mucus, highlighting the potential of mucoadhesive environmental or engineered nanoparticles to disrupt mucus barriers and cause greater exposure to foreign particles, including pathogens and other potentially toxic nanomaterials.     

On the mechanism of hepatic transendothelial passage of large liposomes

Liposomes of 400 nm in diameter can cross the 100-nm fenestrations in the endothelium of the hepatic sinusoid, provided they contain phosphatidylserine (PS) but not phosphatidylglycerol (PG) [Daemen et al. (1997) Hepatology 26, 416]. We present evidence indicating that (i) the PS effect does not involve a pharmacological action of this lipid on the size of the fenestrations, (ii) fluid-type but not solid-type PS liposomes have access to the hepatocytes and (iii) the lack of uptake of PG liposomes by hepatocytes is not due to a lack of affinity of the hepatocytes for PG surfaces. We conclude that the mechanism responsible for the uptake of large PS-containing liposomes by hepatocytes in vivo involves a mechanical deformation of these liposomes during their passage across the endothelial fenestrations.     

Adhesion of four Bifidobacterium strains to human intestinal mucus from subjects in different age groups

The number of bifidobacteria in faeces and intestinal contents has been shown to be reduced with increasing age of the subject. The adhesion of four Bifidobacterium strains was tested to mucus isolated from subjects of different age. All strains bound significantly less to mucus isolated from elderly subjects, compared to mucus from the other age groups. Two of the tested strains also showed decreased adhesion to mucus isolated from 6-month-old and adult subjects compared to the adhesion to mucus from 2-month-old subjects. The results suggest that reduced adhesion may be a factor involved in the decreasing colonisation of elderly subjects by bifidobacteria.     

Defective remodeling of cardiolipin and phosphatidylglycerol in Barth syndrome

Cardiolipin (CL) and phosphatidylglycerol (PG) are the major polyglycerophospholipids observed in mammalian tissues. CL is exclusively found in the inner mitochondrial membrane and is required for optimal function of many of the respiratory and ATP-synthesizing enzymes. The role of CL in oxidative phosphorylation is, however, not fully understood and although reduced CL content leads to aberrant cell function, no human disorders with a primary defect in cardiolipin metabolism have been described. In this paper we present evidence that patients with the rare disorder X-linked cardioskeletal myopathy and neutropenia (Barth syndrome, MIM 302060) have a primary defect in CL and PG remodeling. We investigated phospholipid metabolism in cultured skin fibroblasts of patients and show that the biosynthesis rate of PG and CL is normal but that the CL pool size is 75% reduced, indicating accelerated degradation. Moreover, the incorporation of linoleic acid, which is the characteristic acyl side chain found in mammalian CL, into both PG and CL is significantly reduced, whereas the incorporation of other fatty acids into these phospholipids is normal. We show that this defect was only observed in Barth syndrome patients' cells and not in cells obtained from patients with primary defects in the respiratory chain, demonstrating that the observed defect is not secondary to respiratory chain dysfunction. These results imply that the G4.5 gene product, which is mutated in Barth syndrome patients, is specifically involved in the remodeling of PG and CL and for the first time identify an essential factor in this important cellular process.     

Cough-enhanced mucus clearance in the normal lung

We studied the effectiveness of cough for clearing mucus in 12 nonsmoking subjects with normal lung function. On 2 separate study days, each subject breathed 6-microns Mass Median Aerodynamic Diameter 99mTc-labeled iron oxide particles under controlled breathing conditions while they were seated in front of a gamma camera. Retention (R) of lung activity was measured over the initial 2 h and again at 24 h after particle inhalation. On the control day the subject sat quietly in front of the camera, while on the cough day each subject performed 60 controlled coughs during the 1st h of retention measurements. By paired analysis, retentions at both 1 and 2 h (R1 and R2, respectively) for the cough measurements were significantly less than control (mean control R1 = 85% vs. mean cough R1 = 72%, P less than 0.002; mean control R2 = 75% vs. mean cough R2 = 65%, P less than 0.02). Retention at 24 h (R24) was not significantly different between cough and control measurements (mean cough R24 = 35% and mean control R24 = 32%). Thus coughing increased the rate at which the radiolabeled particles were cleared from the bronchial airways in these individuals. Follow-up experiments with subjects performing rapid inhalations rather than cough showed similar enhanced particle clearance to that seen with cough. These results suggest that the observed enhancement of mucus clearance by cough (and rapid inhalation) in the normal lung may be due to a stimulation of the mucociliary apparatus rather than via a two-phase gas-liquid flow mechanism.