An optimized in vitro model of the respiratory tract wall to study particle cell interactions

As a part of the respiratory tissue barrier, lung epithelial cells play an important role against the penetration of the body by inhaled particulate foreign materials. In most cell culture models, which are designed to study particle-cell interactions, the cells are immersed in medium. This does not reflect the physiological condition of lung epithelial cells which are exposed to air, separated from it only by a very thin liquid lining layer with a surfactant film at the air-liquid interface. In this study, A549 epithelial cells were grown on microporous membranes in a two chamber system. After the formation of a confluent monolayer the cells were exposed to air. The morphology of the cells and the expression of tight junction proteins were studied with confocal laser scanning and transmission electron microscopy. Air-exposed cells maintained monolayer structure for 2 days, expressed tight junctions and developed transepithelial electrical resistance. Surfactant was produced and released at the apical side of the air-exposed epithelial cells. In order to study particle-cell interactions fluorescent 1 microm polystyrene particles were sprayed over the epithelial surface. After 4 h, 8.8% of particles were found inside the epithelium. This fraction increased to 38% after 24 h. During all observations, particles were always found in the cells but never between them. In this study, we present an in vitro model of the respiratory tract wall consisting of air-exposed lung epithelial cells covered by a liquid lining layer with a surfactant film to study particle-cell interactions.

Particles induce apical plasma membrane enlargement in epithelial lung cell line depending on particle surface area dose

Background Airborne particles entering the respiratory tract may interact with the apical plasma membrane (APM) of epithelial cells and enter them. Differences in the entering mechanisms of fine (between 0.1 μm and 2.5 μm) and ultrafine ( ≤ 0.1 μm) particles may be associated with different effects on the APM. Therefore, we studied particle-induced changes in APM surface area in relation to applied and intracellular particle size, surface and number. Methods Human pulmonary epithelial cells (A549 cell line) were incubated with various concentrations of different sized fluorescent polystyrene spheres without surface charge (∅ fine – 1.062 μm, ultrafine – 0.041 μm) by submersed exposure for 24 h. APM surface area of A549 cells was estimated by design-based stereology and transmission electron microscopy. Intracellular particles were visualized and quantified by confocal laser scanning microscopy. Results Particle exposure induced an increase in APM surface area compared to negative control (p < 0.01) at the same surface area concentration of fine and ultrafine particles a finding not observed at low particle concentrations. Ultrafine particle entering was less pronounced than fine particle entering into epithelial cells, however, at the same particle surface area dose, the number of intracellular ultrafine particles was higher than that of fine particles. The number of intracellular particles showed a stronger increase for fine than for ultrafine particles at rising particle concentrations. Conclusion This study demonstrates a particle-induced enlargement of the APM surface area of a pulmonary epithelial cell line, depending on particle surface area dose. Particle uptake by epithelial cells does not seem to be responsible for this effect. We propose that direct interactions between particle surface area and cell membrane cause the enlargement of the APM.

Visualization and quantitative analysis of nanoparticles in the respiratory tract by transmission electron microscopy

ABSTRACT: Nanotechnology in its widest sense seeks to exploit the special biophysical and chemical properties of materials at the nanoscale. While the potential technological, diagnostic or therapeutic applications are promising there is a growing body of evidence that the special technological features of nanoparticulate material are associated with biological effects formerly not attributed to the same materials at a larger particle scale. Therefore, studies that address the potential hazards of nanoparticles on biological systems including human health are required. Due to its large surface area the lung is one of the major sites of interaction with inhaled nanoparticles. One of the great challenges of studying particle-lung interactions is the microscopic visualization of nanoparticles within tissues or single cells both in vivo and in vitro. Once a certain type of nanoparticle can be identified unambiguously using microscopic methods it is desirable to quantify the particle distribution within a cell, an organ or the whole organism. Transmission electron microscopy provides an ideal tool to perform qualitative and quantitative analyses of particle-related structural changes of the respiratory tract, to reveal the localization of nanoparticles within tissues and cells and to investigate the 3D nature of nanoparticle-lung interactions.This article provides information on the applicability, advantages and disadvantages of electron microscopic preparation techniques and several advanced transmission electron microscopic methods including conventional, immuno and energy-filtered electron microscopy as well as electron tomography for the visualization of both model nanoparticles (e.g. polystyrene) and technologically relevant nanoparticles (e.g. titanium dioxide). Furthermore, we highlight possibilities to combine light and electron microscopic techniques in a correlative approach. Finally, we demonstrate a formal quantitative, i.e. stereological approach to analyze the distributions of nanoparticles in tissues and cells.This comprehensive article aims to provide a basis for scientists in nanoparticle research to integrate electron microscopic analyses into their study design and to select the appropriate microscopic strategy.

Deposition and biokinetics of inhaled nanoparticles

Particle biokinetics is important in hazard identification and characterization of inhaled particles. Such studies intend to convert external to internal exposure or biologically effective dose, and may help to set limits in that way. Here we focus on the biokinetics of inhaled nanometer sized particles in comparison to micrometer sized ones.The presented approach ranges from inhaled particle deposition probability and retention in the respiratory tract to biokinetics and clearance of particles out of the respiratory tract. Particle transport into the blood circulation (translocation), towards secondary target organs and tissues (accumulation), and out of the body (clearance) is considered. The macroscopically assessed amount of particles in the respiratory tract and secondary target organs provides dose estimates for toxicological studies on the level of the whole organism. Complementary, microscopic analyses at the individual particle level provide detailed information about which cells and subcellular components are the target of inhaled particles. These studies contribute to shed light on mechanisms and modes of action eventually leading to adverse health effects by inhaled nanoparticles.We review current methods for macroscopic and microscopic analyses of particle deposition, retention and clearance. Existing macroscopic knowledge on particle biokinetics and microscopic views on particle organ interactions are discussed comparing nanometer and micrometer sized particles. We emphasize the importance for quantitative analyses and the use of particle doses derived from real world exposures.

Recent advances into understanding some aspects of the structure and function of mammalian and avian lungs

Recent findings are reported about certain aspects of the structure and function of the mammalian and avian lungs that include (a) the architecture of the air capillaries (ACs) and the blood capillaries (BCs); (b) the pulmonary blood capillary circulatory dynamics; (c) the adaptive molecular, cellular, biochemical, compositional, and developmental characteristics of the surfactant system; (d) the mechanisms of the translocation of fine and ultrafine particles across the airway epithelial barrier; and (e) the particle-cell interactions in the pulmonary airways. In the lung of the Muscovy duck Cairina moschata, at least, the ACs are rotund structures that are interconnected by narrow cylindrical sections, while the BCs comprise segments that are almost as long as they are wide. In contrast to the mammalian pulmonary BCs, which are highly compliant, those of birds practically behave like rigid tubes. Diving pressure has been a very powerful directional selection force that has influenced phenotypic changes in surfactant composition and function in lungs of marine mammals. After nanosized particulates are deposited on the respiratory tract of healthy human subjects, some reach organs such as the brain with potentially serious health implications. Finally, in the mammalian lung, dendritic cells of the pulmonary airways are powerful agents in engulfing deposited particles, and in birds, macrophages and erythrocytes are ardent phagocytizing cellular agents. The morphology of the lung that allows it to perform different functions-including gas exchange, ventilation of the lung by being compliant, defense, and secretion of important pharmacological factors-is reflected in its "compromise design."

In vitro models of the human epithelial airway barrier to study the toxic potential of particulate matter

BACKGROUND: Several epidemiological studies show that inhalation of particulate matter may cause increased pulmonary morbidity and mortality. Of particular interest are the ultrafine particles that are particularly toxic. In addition more and more nanoparticles are released into the environment; however, the potential health effects of these nanoparticles are yet unknown. OBJECTIVES: To avoid particle toxicity studies with animals many cell culture models have been developed during the past years. METHODS: This review focuses on the most commonly used in vitro epithelial airway and alveolar models to study particle-cell interactions and particle toxicity and highlights advantages and disadvantages of the different models. RESULTS/CONCLUSION: There are many lung cell culture models but none of these models seems to be perfect. However, they might be a great tool to perform basic research or toxicity tests. The focus here is on 3D and co-culture models, which seem to be more realistic than monocultures.

Measurement of charged-particle event shape variables in inclusive √(s)=7 TeV proton-proton interactions with the ATLAS detector

The measurement of charged-particle event shape variables is presented in inclusive inelastic pp collisions at a center-of-mass energy of 7 TeV using the ATLAS detector at the LHC. The observables studied are the transverse thrust, thrust minor, and transverse sphericity, each defined using the final-state charged particles' momentum components perpendicular to the beam direction. Events with at least six charged particles are selected by a minimum-bias trigger. In addition to the differential distributions, the evolution of each event shape variable as a function of the leading charged-particle transverse momentum, charged-particle multiplicity, and summed transverse momentum is presented. Predictions from several Monte Carlo models show significant deviations from data.

Hypnotic and opioid anesthetic drug interactions on the CNS, focus on response surface modeling

This chapter will present the conceptual and applied approaches to capture the interaction of anesthetic hypnotic drugs with opioid drugs, as used in the clinical anesthetic state. The graphic and mathematical approaches used to capture hypnotic/opiate anesthetic drug interactions will be presented. This chapter is not a review article about interaction modeling, but focuses on specific drug interactions within a quite narrow field, anesthesia.