P. Gehr

In vivo determination of surface tension in the horse trachea and in vitro model studies.

We measured the surface tension in the trachea of the non-anaesthetised horse from the spreading behaviour of fluid drops, using videotracheoscopy. To do this, we placed small oil drops onto the tracheal wall with a thin Teflon tubing inserted into a videocolonoscope used in humans. Either 5 ml of saline (control) or 5 ml of bovine lipid extract surfactant (BLES) at 4 mg/ml were administered. Tracheal surface tension was 31.9 +/- 0.54 mN/m (Mean +/- SEM, n = 30) in the control experiments and 24.5 +/- 0.51 mN/m (Mean +/- SEM, n = 21) in the entire trachea after the administration of BLES. These values were determined from calibration curves relating film surface tension to the relative diameter of test fluid droplets. In the calibration experiments, the test fluid droplets were placed onto a surfactant film at various surface tensions in either a modified Langmuir-Wilhelmy balance or a captive bubble surfactometer. The spreading behaviour of a given test fluid droplet in the model studies did not only depend on the film surface tension but also on the thickness of the aqueous layer below the surfactant film. Hence, the computed surface tensions in the trachea depend on the choice of which in vitro model is applied.

Efficiency of airway macrophage recovery by bronchoalveolar lavage in hamsters: a stereological approach

Macrophages play a central role in the defence of the respiratory tract against deposited particles. In addition to the well-studied alveolar macrophages, airway macrophages have been recognized as an important clearance factor. Bronchoalveolar lavage (BAL) has been used for functional and morphological investigations of macrophages in vitro, assuming that all macrophages are removed with equal probability from the lung surface. Airway macrophages have been found in close contact with the epithelial cells. These macrophages may not be easily removed by lavage, and they might constitute a functionally different macrophage population. We have tested the hypothesis that there exists a population of macrophages in the conducting airways that resists removal by lavage. We lavaged the lungs of four hamsters and fixed the lungs, thereafter, by intravascular perfusion. The number of macrophages in the intrapulmonary conducting airways was estimated with an unbiased stereological technique, the fractionator, and compared to the number of macrophages in the airways of four hamsters whose lungs had not been lavaged prior to fixation. This in situ study revealed that, in hamster lungs, 42% of the airway macrophages were not removed by BAL and that about 5% of all macrophages in the BAL fluid were airway macrophages. Additionally, ultrastructural alterations of the airway epithelium were found. It is concluded that there exists a population of airway macrophages that resists lavage. This is an aspect which has to be considered in studies performed with macrophages obtained by BAL, since they could represent a functionally different macrophage population.

Mucus quality on horse tracheal epithelium: microscopic grading based on transparency

The aim of this ex-vivo study on excised tracheas of healthy horses was to characterise the microscopic heterogeneity of mucus quality by a visual grading system based on transparency and to determine whether differences in mucus quality, assessed by a visual grading system, influence tracheal mucus velocity (TMV). Small pieces of each trachea were mounted into a humidified chamber under a microscope. Mucus quality was visually subdivided into four grades (MG) and ciliary beat frequency and TMV were determined. Mucus on excised horse tracheal epithelium does not form a homogenous layer. We observed flakes and streams of a great heterogeneity, which by the characteristic of transparency can be qualified and quantified. Visual characterisation of mucus was able to explain a considerable part of TMV variation. Therefore, it can be considered as a suitable non-invasive method for the evaluation of mucus quality and transport effectiveness.

SECONDARY LYSOSOMES AS AN INTEGRAL PART OF THE CYTOSKELETON : A MORPHOLOGICAL STUDY IN RAT KUPFFER CELLS

Rat Kupffer cells contain the three major cytoskeletal components: microfilaments (MF), microtubules (MT), and intermediate filaments (IF) of the vimentin type. Previous cytomagnetometric data obtained from alveolar macrophages and rat Kupffer cells in culture provided evidence that actin filaments contribute to the movements of lysosomes. The lysosomal transport in living cells was affected, when the MFs were selectively disturbed, whereas the depolymerization of the MTs had no effect on the lysosomal movement measured by cytomagnetometric means. Immunofluorescence and ultrastructural studies of isolated and cultured rat Kupffer cells, presented in this paper, will investigate the relationship between lysosomes and the cytoskeleton. The principal filamentous structure in the peripheral cytoplasm of Kupffer cells in a dense meshwork of actin filaments. The dimension of the meshes combined with the dimensions of lysosomes implies the necessity of either (i) disintegration of the actin filament cross-links, (ii) depolarymerization and redistribution of MFs, or (iii) a displacement of actin filaments by the lysosomes during the organelle transport. The presence of microtubules in cytoplasmic protrusions and their track from the periphery to the perinuclear region during interphase might play a role in the transport mechanism of lysosomes, the more so because microtubules could often be demonstrated in closest association with lysosomes even in the first phase of endocytosis. The distribution pattern of vimentin, found as a dense interconnected framework surrounding the lysosomes like a basket, could play a role in positioning the organelles. The dynamic functions of MFs and MTs and their multifunctionality led to an adaptive and flexible organization of these filaments which may both be involved in lysosomal motion.(ABSTRACT TRUNCATED AT 250 WORDS)

Stereological estimation of particle retention and clearance in the intrapulmonary conducting airways of the hamster lungs

In order to evaluate the potential health effects from hazardous particles in these airways, their retention and subsequent clearance were assessed quantitatively and the pulmonary structures with which they are associated were investigated. This study has been performed with hamsters, who inhaled particles of 6 μm in diameter, a size known to deposit mainly in the conducting airways

Retention and clearance mechanisms of inhaled particles

Abstract We have investigated structure and function of the airway coating related to retention and clearance of inhaled particles. Studies were performed in hamster, sheep and horse, using light and electron microscopy and stereology. Video-bronchoscopy and video-microscopy were used to measure the surface tension in the airways. These measurements were correlated with surface properties of particles and films investigated in a Langmuir-Wilhelmy balance. Many particles deposited in the intrapulmonary conducting airways were found retained for more than 24 hours in close association with the epithelium. In conclusion, the initial retention critically depends on a surfactant film at the air-mucus interface. Surface properties and airway macrophages play an importand role in particle clearance.

A new methodology for controlled particle inhalation by small rodents.

In order to investigate the deposition, retention, and clearance mechanisms implicated in particle inhalation under standardized conditions, we developed a continuous negative-pressure ventilation system, whereby the breathing pattern in small rodents could be controlled during exposure to aerosols. Using an on-line open-flow set-up, 19 anesthetized, intubated, and paralyzed Syrian golden hamsters, individually contained within a whole-body box, were artificially ventilated under the said continuous negative-pressure conditions, 1 of 5 different combinations of breathing frequency and tidal volume being established. The animals were then exposed to aerosols containing 6-micron diameter polystyrene spheres, and the deposition of particles in the conducting airways was monitored photometrically. During exposure, the level of respiration (mean lung inflation) was stabilized by means of a negative-pressure vent. Breathing frequency and tidal volume, as well as the compliance of the system, remained virtually unchanged during the course of a single experiment, and in each case, a reproducible deposition of particles was achieved. Our findings indicate that tidal volume, but not breathing frequency, has a marked influence on the particle deposition ratio. Breathing frequency exerts opposing and counterbalancing effects on this latter parameter by enhancing the impaction of particles on the one hand, and by decreasing sedimentation on the other.

Der gesunde Respirationstrakt

Im Gegensatz zu den meisten anderen Organsystemen nimmt die Lunge ihre Funktion als Gasaustauschapparat erst mit der Geburt auf. Die Luftraume sind intrauterin mit Flussigkeit gefullt. Fast schlagartig wird wahrend der Geburt diese Flussigkeit durch Atemluft ersetzt, und die innere Lungenoberflache tritt zum ersten Mal mit den Atemgasen in Kontakt. Ahnlich dramatisch erfolgt gleichzeitig die Umstellung der embryonalen auf die postnatalen Durchblutungsverhaltnisse. Der Zeitplan und die Morphologie der Lungenentwicklung sind durch diese Umstande wesentlich gepragt.

Macrophage Intracellular Motility and Its Correlation with Cell-Substrate Adhesion

Cell spreading and cell adhesion are influenced by the nature of the cell membrane and the structure of the cytoskeleton. The cell membrane with its surface glycopolymers, in combination with the cytoskeleton, may control interactions between the cell and its environment. Particularly the membrane-bound cytoskeleton appears to be involved in the organization of the membrane, and the formation of cell-substrate and cell-cell contacts (Traas and Ramaekers, 1986).