Abstract: Introduction: The paranasal sinuses are air-filled cavities in the bones of the skull surrounding the nose. They communicate with the nose via narrow ducts, the ostia. Despite the fact that the sinuses are non-ventilated hollow organs, it is possible to penetrate the sinus cavities by generating pressure differences between nasal and paranasal cavities [1, 2, 3]. Two physical mechanisms affect the transport of aerosols into non-actively ventilated areas: diffusion and flow enforcement by pressure xD;difference [1]. Flow induction by a pressure gradient was found to be the most important factor carrying particles into non-ventilated areas. Vibrating air flows or humming can be used to generate such pressure gradients. The PARI SINUS⢠(US: VibrENTâ¢) is a novel device that targets drug delivery to the sinuses via a pulsating aerosol [2]. In spite of the widespread use of aerosols in respiratory diseases, only few studies have been performed in order to assess ventilation and aerosol deposition into the sinuses [3]. The objective of this study was to develop a technique to visualize sinus penetration in human volunteers using dynamic 81mKr-gas ventilation imaging in combination with the PARI SINUS pulsating drug delivery system.
Abstract: Der Begriff âultrafeine Partikelâ gilt für Partikel, die kleiner als 100 nm sind, und wird überwiegend im Bereich der Wissenschaften gebraucht, die sich mit der Charakterisierung und Exposition partikulärer Luftverschmutzung und deren Wirkung auf die menschliche Gesundheit befassen. Unter Nanopartikeln werden solche verstanden, die wenigstens in einer Dimension kleiner als 100 nm sind. Während die ultrafeinen Partikel (UFP) der Umweltluftverschmutzung in aller Regel entweder direkt durch Verbrennungsprozesse und anschlieÃende sekundäre Reaktionen in der Gasphase oder durch Gasphasenreaktionen zu komplexen Partikelzusammensetzungen mit vielfältigen Strukturen entstanden sind, werden Nanopartikel gezielt hergestellt, um ihre besonderen chemischen und physikalischen Eigenschaften für gezielte Anwendungen in Industrie, Forschung, Medizin aber auch im alltäglichen Gebrauch zu nutzen. Neuere epidemiologische Studien ergaben Assoziationen zwischen der Anzahlkonzentration von ultrafeinen Partikeln und adversen Gesundheitseffekten auf das Herz-Kreislaufsystem und den Atemtrakt. Die Dosimetrie, beginnend mit der Deposition in den verschiedenen Bereichen des Atemtrakts, der nachfolgenden Retention und Relokation im Atemtrakt sowie die Translokation in die Blutzirkulation und sekundäre Zielorgane sowie die Clearance aus dem Körper ist nur teilweise verstanden. Obwohl erhebliche Fortschritte bei toxikologischen Studien in den letzten Jahren erzielt worden sind, bestehen zusätzlich Unsicherheiten über die Wirkungsweisen ultrafeiner Partikel auf Körperflüssigkeiten, Proteine, Zellen und Gewebe im Atemtrakt und den verschiedenen sekundären Zielorganen, wie dem Herz-Kreislaufsystem, dem Immunsystem und dem zentralen Nervensystem. Diese Unsicherheiten gelten insbesondere für UFP-Konzentrationen, wie sie in der Umwelt anzutreffen sind. Zur Zeit geht man davon aus, daà Partikel die Bildung von Sauerstoff- und Stickstoffradikalen in den Zellen und Geweben auslösen können. Dieser sogenannte âoxidative StreÃâ führt zur Veränderung von Signaltransduktionsketten im Zellkern mit der Folge der Hochregulierung oder Unterdrückung von Zytokinen und Mediatoren, die in ihrer Summe proinflammatorische Prozesse auslösen können. Aus letzteren können sich dann insbesondere bei suszeptiblen Personen wie z.B. Kleinkindern, Kranken, Ãlteren und genetisch disponierten Individuen, Krankheiten verstärken oder neu ausbilden. Die groÃe spezifische Oberfläche ultrafeiner Partikel bietet ein wirkungsvolles Interface zur Induktion von oxidativem Streà in Proteinen und Zellen, wie dies auch für bioverfügbare Ãbergangsmetalle und für biologisch reaktive Organika beschrieben worden ist. Für Anwendungen in Industrie, Forschung, Medizin, aber auch für den alltäglichen Gebrauch werden Nanopartikel gezielt wegen ihrer besonderen Oberflächeneigenschaften hergestellt. Es ist daher nicht auszuschlieÃen, daà diese Nanopartikel auch spezielle Reaktionen im menschlichen Organismus auslösen, die zu Gesundheitsschäden führen können. Bei der Bewertung des Gefährdungspotentials durch neuartige Nanopartikel ist es erforderlich, den gesamten Zyklus von seiner Produktion über die Anwendung bis zu seiner Entsorgung inklusive der unterschiedlichen Expositionsszenarien und Inkorporationspfade zu berücksichtigen.
Abstract: The aim of this study was to determine particle clearance and retention from non-alveolated airways of 14 healthy subjects (HS), 10 subjects with asymptomatic bronchial hyperresponsiveness (BHR), and 23 patients with chronic obstructive pulmonary disease (COPD). Monodisperse iron oxide particles of 1.6 μ m geometric and 3.5 μ m aerodynamic diameter labeled with 99mTc were delivered to the airways by inspiration of small aerosol boli into shallow volumetric lung depths. In each subject the penetration front depth of the aerosol boli was adjusted to 55% of the Fowler dead space of the airways. Particle deposition was enhanced by about 7 seconds of breath-holding after bolus inhalation. Retention of the particles in the airways during the 48 hours after their administration was assessed by measuring the decline in lung activity with a sensitive gamma counter. Particle deposition was not significantly different among study groups. Twenty-four hour particle retention in the airways was not different among study groups. Sixty-one percent of the particles were retained at 24 hours in HS, 58% in BHR, and 64% in COPD. However, subjects with BHR showed accelerated mucociliary clearance compared to healthy subjects, whereas clearance was retarded in COPD patients. This long-term particle retention in the airways has to be taken into account in aerosol toxicology risk assessment and aerosol therapy dose evaluation.
Abstract: Rationale: Little is known about clearance of ultrafine carbon particles from the different regions of the human lung. These particles may accumulate and present a health hazard because of their high surface area. xD;Objectives: Technetium Tc 99m (99mTc)âradiolabeled 100-nm-diameter carbon particles were inhaled by healthy nonsmokers, asymptomatic smokers, and by patients with chronic obstructive pulmonary disease (COPD). xD;Methods: Using a bolus inhalation technique, particle deposition was targeted either to the airways or to the lung periphery, and retention, clearance, and translocation were measured using retained radiotracer imaging. xD;Measurements and Main Results: In vitro studies revealed that mean leaching of soluble 99mTc-radiotracer from the carbon particles was 4.1 (2.6 [SD]) % after 24 hours. Cumulative 99mTc activity in urine at 24 hours was 1.1 (1.3) % of activity deposited in the lungs. In the lung periphery, particle retention was not affected by smoking or pulmonary disease; retention was 96 (3) % after 24 hours. The small amount of clearance could be attributed to leaching of the 99mTc label, suggesting negligible particle clearance. In healthy nonsmokers, retention of particles targeted to the airways was 89 (6) and 75 (10) % after 1.5 and 24 hours, respectively. Radiolabel activity did not accumulate in the liver. xD;Conclusions: Within the limits of detection of our experimental system, most inhaled ultrafine carbon particles are retained in the lung periphery and in the conducting airways without substantial systemic translocation or accumulation in the liver at 48 hours. Repeated exposure may result in significant pulmonary accumulation of ultrafine particles.
Abstract: New drug developments for the treatment of topical lung diseases and systemic uptake can be achieved by using the inhalative route. High, specific and reproducible targeting of these new drugs to specific regions of the lung is desired. Radioaerosol techniques in combination with planar gamma camera imaging are described to achieve an individual dose assessment of inhaled aerosolized drugs and to adjust inhalation devices for individual patient use.
Abstract: Background: Although there is a high incidence of nasal disorders including chronic sinusitis, there is limited success in the topical drug delivery to the nose and the paranasal sinuses. This is caused by the nose being an efficient filter for inhaled aerosol particles and the paranasal sinuses being virtually non ventilated. Method: The objective of this study was to visualize the efficiency of sinus ventilation in a nasal cast using dynamic 81mKr-gas imaging in combination with pulsating airflows. Furthermore, the efficiency of the deposition of radiolabelled aerosol was assessed. Results: Pulsation increased ventilation efficiency of the sinuses more than fivefold and aerosol deposition efficiency more than twentyfold, compared to delivery without pulsation. Furthermore pulsation increased aerosol deposition in the nasal airways by a factor of three. Using pulsating airflow Kr-gas ventilation and aerosol deposition efficiencies increased with increasing sinus volume. Pulsating airflow resulted in a deposition of up to 8% of the nebulized drug within the sinuses compared to 0.2% without pulsation. Conclusions: The study demonstrates the high efficiency of a pulsating airflow in paranasal sinus ventilation and aerosolized drug delivery. This proves that topical drug delivery to the paranasal sinuses in relevant quantities is possible.
Abstract: Ferrimagnetic iron oxide particles were inhaled by 17 healthy volunteers (9 non-smokers, 8 smokers), and the retained particles were magnetized and detected by a SQUID. Stochastic particle transport due to cytoskeletal reorganizations within macrophages (relaxation) and directed particle motion in a weak magnetic twisting field were investigated with respect to viscous and elastic properties of the cytoskeleton. Relaxation and cytoskeletal stiffness were not influenced by cigarette smoking. Relaxation and particle twisting revealed a non-Newtonian viscosity with a pure viscous and a viscoelastic compartment. Viscous and elastic data obtained from relaxation correlated with particle twisting, indicating that the proposed simple model is a reasonable approximation of cytoskeletal mechanical properties.
Abstract: Newly synthesized magnetic nanomaterials possess high DNA binding capacity either itself or in the presence of a positively charged lipid-based Metafectene(TM) reagent or branched polyethylene imine 25 kDa. Polyethylene imine (PEI)-modified nanomaterials are able to deliver nucleic acids in cell culture in duplexes. Magnetofection with triplexes of nanomaterials results in higher transduction efficiencies compared to optimal PEI or Metafectene formulations. 90% transient down-regulation of the target protein in HeLa-green fluorescence protein cells was achieved at short interfering RNA concentrations as low as 8 nM with a formulation of PEI-modified nanoparticles.
Abstract: Epidemiological studies continue to indicate associations between exposure to increased concentrations of ambient fine and ultrafine particles and adverse health effects in susceptible individuals. The ultrafine particle fraction in the ambient atmosphere seems to play a specific role. Yet, the dosimetry (including deposition patterns in the respiratory tract and, particularly, the biokinetic fate of ultrafine particles) is not fully understood. In contrast to fine particles, inhaled ultrafine particles seem to follow different routes in the organism. Cardiovascular effects observed in epidemiological studies triggered the discussion on enhanced translocation of ultrafine particles from the respiratory epithelium towards circulation and subsequent target organs, such as heart, liver, and brain, eventually causing adverse effects on cardiac function and blood coagulation, as well as on functions of the central nervous system. Current knowledge on systemic translocation of ultrafine particles in humans and animal models is reviewed. Additionally, an estimate of accumulating particle numbers in secondary target organs during chronic exposure is extrapolated from long-term translocation data obtained from rats. Toxicological studies aim to provide the biological plausibility of health effects of ultrafine particles and to identify cascades of mechanisms that are causal for the gradual transition from the physiological status towards pathophysiologcal alterations and eventually chronic disease. Considering the interaction between insoluble ultrafine particles and biological systems (such as body fluids, proteins, and cells), there still are gaps in the current knowledge on how ultrafine particles may cause adverse reactions. This paper reviews the current concept of interactions between insoluble ultrafine particles and biological systems.
Abstract: A generator for the production of radiolabelled ultrafine carbonaceous particles for inhalation and clearance studies is described. A Technegas generator in combination with 99mTc labelling is a device used in clinical routine to investigate the ventilation of the human lung. However, under standard operating conditions the Technegas generator does not provide ultrafine particles with a stable radiolabel. The removal of saline from the Tc-eluate by an ion exchange column yielded low leaching rates of the radiolabel from the particles of below 4% within 24 h (12% under standard conditions) and guaranteed non-hygroscopic aerosol properties. Short burning times (2 s; standard 15 s) and immediate dilution of the generated aerosol into a conductive bag yielded stable ultrafine particles in the size range between 40 and 100 nm count median diameter (standard 200 nm), with a geometric standard deviation of 1.6. We adapted the generator to a bolus inhalation system, allowing regional aerosol deposition into different lung regions and appropriate clearance studies. In addition the particles can be labelled by 111In instead of 99mTc, extending the investigation time of clearance studies from 1 day to some weeks.
Abstract: The role of alveolar macrophages in the fate of ultrafine particles in the lung was investigated. Male Wistar-Kyoto rats were exposed to ultrafine gold particles, generated by a spark generator, for 6 h at a concentration of 88 microg/m3 (4 x 10(6)/cm3, 16 nm modal mobility diameter). Up to 7 days, the animals were serially sacrificed, and lavaged cells and lung tissues were examined by transmission electron microscopy. The gold concentration/content in the lung, lavage fluid, and blood was estimated by inductively coupled plasma-mass spectrometry. Gold particles used were spherical and electron dense with diameters of 5-8 nm. The particles were individual or slightly agglomerated. By inductively coupled plasma-mass spectrometry analysis of the lung, 1945 +/- 57 ng (mean +/- SD) and 1512 +/- 184 ng of gold were detected on day 0 and on day 7, respectively, indicating that a large portion of the deposited gold particles was retained in the lung tissue. In the lavage fluid, 573 +/- 67 ng and 96 +/- 29 ng were found on day 0 and day 7, respectively, which means that 29% and 6% of the retained gold particles were lavageable on these days. A low but significant increase of gold (0.03 to 0.06% of lung concentration) was found in the blood. Small vesicles containing gold particles were found in the cytoplasm of alveolar macrophages. In the alveolar septum, the gold particles were enclosed in vesicles observed in the cytoplasm of alveolar type I epithelial cells. These results indicate that inhaled ultrafine gold particles in alveolar macrophages and type I epithelial cells are processed by endocytotic pathways, though the uptake of the gold particles by alveolar macrophages is limited. To a low degree, systemic particle translocation took place.
Abstract: ABSTRACT: Spherical monodisperse ferromagnetic iron oxide particles of 1.9um geometric and 4.2um aerodynamic diameter were inhaled by seven patients with primary ciliary dyskinesia (PCD) using the shallow bolus technique, and compared to 13 healthy non-smokers (NS) from a previous study. The bolus penetration front depth was limiting to the phase1 dead space volume. In PCD patients deposition was 58+/-8 % after 8s breath holding time. Particle retention was measured by the magnetopneumographic method over a period of nine months. Particle clearance from the airways showed a fast and a slow phase. In PCD patients airway clearance was retarded and prolonged, 42+/-12 % followed the fast phase with a mean half time of 16.8+/-8.6 hours. The remaining fraction was cleared slowly with a half time of 121+/-25 days. In healthy NS 49+/-9 % of particles were cleared in the fast phase with a mean half time of 3.0+/-1.6 hours, characteristic of an intact mucociliary clearance. There was no difference in the slow clearance phase between PCD patients and healthy NS. Despite non-functioning cilia the effectiveness of airway clearance in PCD patients is comparable to healthy NS, with a prolonged kinetics of one week, which may primarily reflect the effectiveness of cough clearance. This prolonged airway clearance allows longer residence times of bacteria and viruses in the airways and may be one reason for increased frequency of infections in PCD patients.
Abstract: Der Begriff âultrafeine Partikelâ gilt für Partikel, die kleiner als 100 nm sind, und wird überwiegend im Bereich der Wissenschaften gebraucht, die sich mit der Charakterisierung und Exposition partikulärer Luftverschmutzung und deren Wirkung auf die menschliche Gesundheit befassen. Unter Nanopartikeln werden solche verstanden, die wenigstens in einer Dimension kleiner als 100 nm sind. Während die ultrafeinen Partikel (UFP) der Umweltluftverschmutzung in aller Regel entweder direkt durch Verbrennungsprozesse und anschlieÃende sekundäre Reaktionen in der Gasphase oder durch Gasphasenreaktionen zu komplexen Partikelzusammensetzungen mit vielfältigen Strukturen entstanden sind, werden Nanopartikel gezielt hergestellt, um ihre besonderen chemischen und physikalischen Eigenschaften für gezielte Anwendungen in Industrie, Forschung, Medizin aber auch im alltäglichen Gebrauch zu nutzen. Neuere epidemiologische Studien ergaben Assoziationen zwischen der Anzahlkonzentration von ultrafeinen Partikeln und adversen Gesundheitseffekten auf das Herz-Kreislaufsystem und den Atemtrakt. Die Dosimetrie, beginnend mit der Deposition in den verschiedenen Bereichen des Atemtrakts, der nachfolgenden Retention und Relokation im Atemtrakt sowie die Translokation in die Blutzirkulation und sekundäre Zielorgane sowie die Clearance aus dem Körper ist nur teilweise verstanden. Obwohl erhebliche Fortschritte bei toxikologischen Studien in den letzten Jahren erzielt worden sind, bestehen zusätzlich Unsicherheiten über die Wirkungsweisen ultrafeiner Partikel auf Körperflüssigkeiten, Proteine, Zellen und Gewebe im Atemtrakt und den verschiedenen sekundären Zielorganen, wie dem Herz-Kreislaufsystem, dem Immunsystem und dem zentralen Nervensystem. Diese Unsicherheiten gelten insbesondere für UFP-Konzentrationen, wie sie in der Umwelt anzutreffen sind. Zur Zeit geht man davon aus, daà Partikel die Bildung von Sauerstoff- und Stickstoffradikalen in den Zellen und Geweben auslösen können. Dieser sogenannte âoxidative StreÃâ führt zur Veränderung von Signaltransduktionsketten im Zellkern mit der Folge der Hochregulierung oder Unterdrückung von Zytokinen und Mediatoren, die in ihrer Summe proinflammatorische Prozesse auslösen können. Aus letzteren können sich dann insbesondere bei suszeptiblen Personen wie z.B. Kleinkindern, Kranken, Ãlteren und genetisch disponierten Individuen, Krankheiten verstärken oder neu ausbilden. Die groÃe spezifische Oberfläche ultrafeiner Partikel bietet ein wirkungsvolles Interface zur Induktion von oxidativem Streà in Proteinen und Zellen, wie dies auch für bioverfügbare Ãbergangsmetalle und für biologisch reaktive Organika beschrieben worden ist. Für Anwendungen in Industrie, Forschung, Medizin, aber auch für den alltäglichen Gebrauch werden Nanopartikel gezielt wegen ihrer besonderen Oberflächeneigenschaften hergestellt. Es ist daher nicht auszuschlieÃen, daà diese Nanopartikel auch spezielle Reaktionen im menschlichen Organismus auslösen, die zu Gesundheitsschäden führen können. Bei der Bewertung des Gefährdungspotentials durch neuartige Nanopartikel ist es erforderlich, den gesamten Zyklus von seiner Produktion über die Anwendung bis zu seiner Entsorgung inklusive der unterschiedlichen Expositionsszenarien und Inkorporationspfade zu berücksichtigen.
Abstract: Particulate air pollution is reported to cause adverse health effects in susceptible individuals. Since most of these particles are derived form combustion processes, the primary composition product is carbon with a very small diameter (ultrafine, less than 100 nm in diameter). Besides the induction of reactive oxygen species and inflammation, ultrafine particles (UFP) can cause intracellular calcium transients and suppression of defense mechanisms of alveolar macrophages, such as impaired migration or phagocytosis. Methods. In this study the role of intracellular calcium transients caused by UFP was studied on cytoskeleton related functions in J774A.1 macrophages. Different types of fine and ultrafine carbon black particles (CB and ufCB, respectively), such as elemental carbon (EC90), commercial carbon (Printex 90), diesel particulate matter (DEP) and urban dust (UD), were investigated. Phagosome transport mechanisms and mechanical cytoskeletal integrity were studied by cytomagnetometry and cell viability was studied by fluorescence microscopy. Macrophages were exposed in vitro with 100 and 320 microg UFP/ml/million cells for 4 hours in serum free medium. Calcium antagonists Verapamil, BAPTA-AM and W-7 were used to block calcium channels in the membrane, to chelate intracellular calcium or to inhibit the calmodulin signaling pathways, respectively. Results. Impaired phagosome transport and increased cytoskeletal stiffness occurred at EC90 and P90 concentrations of 100 microg/ml/million cells and above, but not with DEP or UD. Verapamil and W-7, but not BAPTA-AM inhibited the cytoskeletal dysfunctions caused by EC90 or P90. Additionally the presence of 5 % serum or 1 % bovine serum albumin (BSA) suppressed the cytoskeletal dysfunctions. Cell viability showed similar results, where co-culture of ufCB together with Verapamil, W-7, FCS or BSA produced less cell dead compared to the particles only.
Abstract: The relaxation characteristics of magnetic nanoparticles (CoFe2O4) were investigated in J774A.1 macrophages and after voluntary inhalation. In dry form 25% of the particles showed Neel relaxation. Relaxation in macrophages occurred within minutes and could be inhibited by fixation, showing Brownian relaxation and intracellular transport processes. Relaxation in the lung happened similarly, but was dependent on the time after deposition. The particles were cleared from the lung within 2 weeks.
Abstract: Den unterschiedlichen anatomischen Regionen der Lunge müssen unterschiedliche Clearancemechanismen zugeordnet werden. Die meisten der in den Atemwegen abgelagerten Partikel (anorganische und organische Partikel, Bakterien, Viren, Pollen) werden durch den mukoziliären Schleimtransport innerhalb von 24 Stunden aus der Lunge heraustransportiert. In Abhängigkeit von der GröÃe der Partikel gibt es auch in den Atemwegen eine langzeitretinierte Fraktion, die wahrscheinlich von Atemwegsmakrophagen aufgenommen und abgebaut wird. Störungen der Mukoziliarclearance ergeben sich vor allen nach Infektionen sowie, genetisch bedingt, durch Veränderungen des Ionengleichgewichts im Atemwegsepithel (Mukoviszidose) oder der Zilienstruktur (primäre ziliäre Dyskinesie). Die Folgen dieser Störungen können teilweise durch Atemtherapie und durch Mukolytika vermindert werden. Die Elimination deponierter Partikel aus der Lungenperipherie (alveoläre Clearance) erfolgt durch Phagozytose via Alveolarmakrophagen, intrazellulären Abbau (Digestion), Migration und Translokation. Die alveoläre Clearance schwer löslicher Partikel verläuft sehr langsam und ist primär an die Funktion der Alveolarmakrophagen geknüpft. Der Transport der Partikel zum Bronchialsystem ist beim Menschen von untergeordneter Bedeutung, so daà die Elimination der Partikel vornehmlich durch Digestion (Lösung) innerhalb der Makrophagen bestimmt wird. Lange andauernder Zigarettenrauchkonsum und chronische Entzündungen in der Lungenperipherie (Sarkoidose, interstitielle Lungenfibrose) führen zu einer signifikanten Verminderung der alveolären Clearance. Patienten mit chronischer Bronchitis zeigen hingegen nur eine geringe Beeinträchtigung der alveolären Clearance.
Abstract: While epidemiological studies indicate an association between adverse health effects and ambient ultrafine particle concentrations in susceptible individuals, toxicological studies aim to identify mechanisms which are causal for the gradual transition from the physiological status towards patho-physiological disease. Impressive progress has been made in recent years when objectives changed from classical tests like lung function, etc. to endpoints comprising of particle induced oxidative stress, cell signaling and activation, release of mediators initiating inflammatory processes not only in the respiratory tract but also in the cardio-vascular system. Particularly, the large surface area of ultrafine particles provides a unique interface for catalytic reactions of surface-located agents with biological targets like proteins, cells, etc. However, toxicological studies are hampered by a number of immanent complications when simulating long-term exposure of humans in urban environments with inherited and/or acquired susceptibility (e.g., acute exposure studies at high concentrations either in human subjects or animal models). Yet, based on a conservative estimate results available begin to show an adverse health risk for susceptible individuals and support the epidemiological evidence.
Notes: 0894-2684 xD;eng xD;Journal Article xD;United States xD;8809251
Abstract: The erythroid differentiation regulator (EDR) is a highly conserved autocrine factor produced in many tissues. Its haemoglobin synthesis-inducing activity for human and murine erythroleukaernia cell lines had been detected in WEHI-3 conditioned medium. EDR functions were analysed in detail. It is released from cells immediately in response to various stressful conditions and enhances cell survival particularly at a lower concentration range and low cell density. At high cell density and high EDR concentration the opposite effect of an increase in cell death was observed. Its essential function within a tissue is considered to be the maintenance of growth homeostasis. Cells kept in culture for weeks show a decreasing responsiveness to EDR supply. This was also noted in freshly cloned EDR-responsive mouse erythroleukaernia cells pointing to a molecular adaptation process. Human haematopoietic progenitors were amplified 7-fold by EDR when kept at low cytokine levels. At saturating levels progenitors giving rise to at least two lineages in semisolid medium (E-mix) respond to EDR with an average 1.87-fold increase in colony numbers and a bell-shaped dose-response curve. Of the more mature BFU-E compartment a response was observed particularly in cases with low colony numbers. Given the release from irradiated stromal cells and the ability to partly substitute for stromal cells in the Burkitt's lymphoma cell line BL-70, EDR functions as a stromal survival factor for stroma-responsive cells. (C) 2004 Elsevier Ltd. All rights reserved.
Notes: 14th ERS Annual Congress, Glasgow, UK, September 4-8, 2004; Volume 24; Supplement 48; September 2004 xD; xD;Pernilla Wiebert, Jürgen Seitz, Alejandro Sanchez-Crespo, Rolf Falk, Klas Philipson, Wolfgang Kreyling, Winfried Möller, Knut Sommerer, Stig Larsson, Magnus Svartengren: Retention of ultrafine particles in the human lung; S. 236s; AbstractNr. 1549
Abstract: Spherical monodisperse ferromagnetic iron oxide particles of 1.9 micro m geometric and 4.2 micro m aerodynamic diameter were inhaled by 13 healthy non-smokers using the shallow bolus technique. The bolus width was 100 ml and the penetration front depth was 150+/-27 ml. The mean flow rate during inhalation and exhalation was 250 ml/s. The Fowler dead space and the phase1 dead space of the airways was 282+/-49 ml and 164+/-34 ml, respectively. Deposition was below 20 % without breath holding and 51+/-8 % after 8 s breath holding time. We attempted to confine the bolus deposition to the bronchial airways by limiting the bolus front depth to the phase1 dead space volume. Particle retention was measured by the magnetopneumographic method over a period of 9 months. Particle clearance from the airways showed a fast and a slow phase, 49+/-9 % followed the fast phase with a mean half time of 3.0+/-1.6 hours and characterized the mucociliary clearance. The remaining fraction was cleared slowly with a half time of 109+/-78 days. The slow clearance phase was comparable to clearance measurements from the lung periphery of healthy non-smokers, which allowed macrophage-dependent clearance mechanisms of the slow cleared fraction to be taken into account. Despite the fact that part of the slowly cleared particles may originate from peripheral deposition, the data demonstrate that mucociliary clearance does not remove all particles being deposited in the airways and that a significant fraction undergoes long-term retention mechanisms, the origin of which is still under discussion.
Abstract: Ferromagnetic particles with 1.9 µm geometric (4.2 µm aerodynamic) diameter were inhaled by 14 healthy non-smokers and by 5 patients with primary ciliary dyskinesia (PCD), using the shallow bolus technique. The aerosol penetration front depth was within the functional (Fowler) dead space so that the particles solely deposited in airways. Deposition was enhanced by a breath hold at the end of inhalation. Clearance was measured by the magnetopneumographic method. After magnetization the remanent magnetic field of the lung was detected at zero time, 3h, 6h, 24h, 48h, one week, one month, 3, 6, and 9 months after particle deposition. Particle clearance showed two phases. In healthy non-smokers, about 50 % of the deposited particles were cleared rapidly (half time 3.0+/-1.6 hours) due to mucociliary transport. The remaining fraction was cleared with a half time of 109+/-78 days. This half time is similar to that of particle clearance from the lung periphery of healthy non-smokers. In PCD patients the fast clearance phase was delayed. Its half time was several days. However, the half time of the slow clearance phase was not different from that of healthy subjects. Consequently, despite of non-functioning cilia, particles are cleared from the airways of PCD patients. xD;Funding: EC (FIGD-CT-2000-00053)
Abstract: Magnetic twisting cytometry (MTC) is a novel tool to measure cytoskeleton-associated cell functions by the use of ferromagnetic microbeads. Magnetic beads are either incorporated by living cells by phagocytic processes or attached to integrin receptors to the cell membrane. The magnetic beads are magnetized and aligned in a strong magnetic field pulse. The application of twisting forces allows to investigate mechanical properties (stiffness, viscoelasticity) of the cytoskeleton of living cells by analyzing the magnetic cell field. Incorporated magnetic beads undergo intracellular transport processes, which result in a loss of particle alignment and in a decay of the remanent magnetic cell field. This process, called relaxation, depends on the mechanical cytoskeletal properties and can directly visualize the intracellular energy of cellular transport processes. The preparation of spherical monodisperse ferromagnetic beads made it possible to understand the above-described processes using mathematical models. Experimental conditions with many magnetic particles per cell enhances the formation of aggregates because of the attractive forces between magnetic spheres, resulting in a change of magnetic properties and of hydrodynamic behavior. Due to mutual magnetization, the remanent magnetic moment of an aggregate is stronger compared to the same number of single particles. This implies a higher cell field. Additionally the relaxation is retarded because of the change in shape factor and in volume, which also implies a faulty estimation of intracellular transport energy. Magnetic particle twisting is less influenced. In summary, valuable cytomagnetometric measurements have to be done with less than one particle per macrophage to ensure low probability of multiple particles per cell.
Notes: English xD;Article xD;IEEE TRANS NANOBIOSCI
Abstract: Ultrafine Tc-99m-labeled carbon particles were produced by a Technegas Generator. To avoid formation of soluble pertechnetate anion and to control particle size the generation procedure was modified. (1) NaCl originating from the elution solvent was reduced two orders of magnitude by extracting Tc-99m in ethanol. (2) Argon gas was cleaned from water vapour, oxygen and organics by absorber/filter units. (3) To better remove air from the burner chamber the flushing volume of argon was increased threefold. (4) To avoid coagulation the aerosol was diluted into a 60L bag immediately after generation. Particle number concentration, size distribution and Tc-99m leaching was measured. (5) Particle size was adjusted between 30-150nm count median diameters (CMD) by the burning time of the graphite crucible and its temperature; geometric standard deviation 1.6. All particles were chain aggregates. While Tc-99m leaching of particles with 80-100nm CMD was <= 2%, particles with 30nm CMD showed leaching of 5-10% as a result of the relatively higher NaCl fraction per particle. Conclusion: After improving of the operation conditions of the Technegas Generator, the particle size can be adjusted and the Tc-99m leaching can be minimized. This makes the particles suitable for human inhalation and clearance studies.
Abstract: We used the bolus technique to deposit ultrafine particle in the airways in 2 healthy non-smokers and followed particle retention and clearance over 48 hours. 100 nm carbon particles, labelled with 99mTc, were produced by a Technegas generator in an argon atmosphere. The particles were inhaled as a shallow 150 ml bolus. The penetration front depth was 200 ml. The Fowler dead space was 330 ml in both subjects, confirming, that the particles solely deposited in airways. Deposition was enhanced by an 8 sec breath hold. The retention and particle distribution in the lung was followed by a gamma camera and a sensitive lung counter. Additionally urine and faeces were collected. Deposition varied between 20 and 23 %. Clearance showed two phases: 17 % of the deposited particles were cleared rapidly (half-time " 4 hours) and 83 % slowly (half-time at least 10 days). Less than 3 % of the activity was found in the urine and about 25 % of the activity was found in faeces after 48 hours. The data show that ultrafine particles also underlie a long-term retention mechanism in the human airways. Whole body gamma camera scans showed that no significant particle translocation occurred into organs other than the gastro-intestinal tract.
Abstract: Essential cytoskeletal functions of macrophages are migration, phagocytosis of foreign materials, and intracellular transport and digestion The influence of fine and ultrafine test particles (UFP), such as TiO(2), elemental carbon, commercial carbon black, diesel exhaust particulate matter, and urban dust (UrbD), on cytoskeleton- related functions of macrophages, such as phagocytosis, phagosome transport mechanisms, and mechanical cytoskeletal integrity, were studied by flow cytometry and by cytomagnetometry. Additionally, necrosis and apoptosis caused by the test particles was detected. The diameter of the test particles ranged from 12 to 220 nm and the Brunauer-Emmet-Teller specific surface area ranged from 6 to 600 m(2)/g. Primary alveolar macrophages from beagle dogs (BD-AM), obtained by bronchoalveolar lavage, were used as well as macrophages originating from the cell line J774A.1. For cytomagnetometry studies, spherical 1.8- &mgr;m ferromagnetic particles served as probes for cytoskeletal functions and were incubated together with the macrophages 24 h prior to UFP exposure. Macrophages were exposed in vitro with 10-320 &mgr;g UFP/ml/10(6) cells up to 24 h. In all experiments, J774A.1 macrophages were more sensitive than BD-AM to UFP exposure. Cytoskeletal dysfunctions evaluated by cytomagnetometry were an impaired phagosome transport and an increased cytoskeletal stiffness and occurred at concentrations of 100 &mgr;g UFP/ml/10(6) cells and above, in both BD- AM and J774A.1. Only fine TiO(2) did not show any effect. Urban dust (standard reference material 1649a) and diesel exhaust particles (DEP, standard reference material 1650) caused comparable cytoskeletal dysfunctions to elemental carbon with high specific surface area. Cytoskeletal dysfunctions induced by DEP or UrbD could be reduced after washing the particles by dichloromethane. UFP caused an impaired phagocytosis of 1-&mgr;m diameter fluorescent latex beads, inhibited cell proliferation, and decreased cell viability. All recorded cytotoxic parameters showed only weak correlations with the specific surface area or the total number of UFP, which can result from the different types of particles and different surface compositions. UFP cause cytoskeletal toxicity in vitro in macrophages, which can cause cellular dysfunctions, such as impaired proliferation, impaired phagocytic activity, and retarded intracellular transport processes as well as increased cell stiffness and can result in impaired defense ability in the lung.
Abstract: The elimination of deposited particles (inorganic and organic particles, bacteria, viruses) from the periphery of the human lung (alveolar clearance) implies phagocytosis by alveolar macrophages, intracellular digestion, migration and translocation. Alveolar clearance of poorly soluble particles happens very slowly and primarily depends on the function of alveolar macrophages. In humans, the transport of particles to the bronchial tree is of secondary relevance, suggesting that the elimination of particles primarily depends on digestion (dissolution) processes within macrophages. The dissolved material is excreted via urine, if there is no further metabolization within the body. The pathophysiology of the alveolar clearance mechanisms in the human lung can be studied by a magnetic tracer technique (magnetopneumography). Ferromagnetic magnetite test-particles are deposited in the periphery of the lung by controlled inhalation. After magnetization and particle alignment in a strong external magnetic field pulse, the amount of retained particles can be detected by a sensitive magnetic field sensor (SQUID, superconducting quantum interference device). Long lasting cigarette smoking and chronic lung inflammations (sarcoidosis, interstitial lung fibrosis) induce a significant impairment of alveolar clearance capacity, while patients with chronic bronchitis show only a moderate impairment of alveolar clearance.
Abstract: Monodisperse ferrimagnetic microparticles (Fe3O4) with 1.3 microm geometric diameter were inhaled to study alveolar long-term clearance in healthy and diseased human subjects. Nineteen younger (age 20 to 39 years) and 20 older (age 40 to 65 years) healthy volunteers participated in the study as well as 15 patients with sarcoidosis (SAR), 12 patients with idiopathic pulmonary fibrosis (IPF), and 15 patients with chronic obstructive bronchitis (COB). In each group the subjects were divided into never smokers (NS) and active smokers (S). Clearance was measured by magnetopneumography (MPG) for 300 days after inhalation. In COB, 50% of the deposited particles were removed from the lungs after 2 days, indicating high bronchial deposits due to bronchial obstructions. In healthy NS, only 10% of the particles were removed after 2 days and cigarette smoking enhanced the fraction of fast-cleared particles. In subjects who smoked, slow clearance was significantly impaired (P < . 02). Clearance half-lives (in days) for younger, healthy, NS were 124 +/- 66 (mean +/- SD) compared to 220 +/- 74 for S. Similarly for older subjects, the timeswere 162 +/- 120 for NS and 459 +/- 334 for S. The impairment of alveolar clearance due to cigarette smoking increases by 5.7 +/- 1.3 days/pack-year (P < .01). Alveolar clearance was impaired in SAR and in IPF; half-lives were 275 +/- 109 days (P < .05) and 756 +/- 345 days (P < .02), respectively, compared to healthy NS. Most COB patients were ex-smokers, their long-term clearance was 240 +/- 74 days, which is more than healthy NS (P < .01), but less than healthy S and might indicate a recovery of alveolar clearance. In view of studies using totally inert particles like Teflon, we conclude that the lung clearance measured with iron oxide tracer particles primarily reflects clearance by intraphagosomal particle dissolution within alveolar macrophages, which is impaired by cigarette smoke consumption and in patients.
Abstract: Monodisperse ferrimagnetic iron-oxide particles of 1.4 µm geometric diameter were used to study alveolar macrophage functions (phagocytosis, phagosome transport) and cytoskeletal integrity in healthy subjects and in patients with idiopathic pulmonary fibrosis as well as in cultured macrophages. Dysfunctions in phagocytosis, in phagosome transport and cytoskeletal integrity correlated with an impaired alveolar clearance and could be induced in vitro by cytoskeletal drugs.
Abstract: The production of spherical monodisperse iron-oxide microparticles in the size range between 0.8 and 5 µm is described.The particles can be ferrimagnetic (Fe3O4) or non-magnetic (alpha-Fe2O3). The particles were radiolabeled with 99mTc or 111In, and the leakage of the radiolabel within 24 h was 0.1 % in the human lung. The particles can be used to study the motility and integrity of living cells.
Abstract: This review summarises the results and discussions of an UNESCO-MCBN supported symposium on oxidative stress and its role in the onset and progression of diabetes. There is convincing experimental and clinical evidence that the generation of reactive oxygen species (ROI) is increased in both types of diabetes and that the onset of diabetes is closely associated with oxidative stress. Nevertheless there is controversy about which markers of oxidative stress are most reliable and suitable for clinical practice. There are various mechanisms that contribute to the formation of ROI. It is generally accepted that vascular cells and especially the endothelium become one major source of ROI. An important role of oxidative stress for the development of vascular and neurological complications is suggested by experimental and clinical studies. The precise mechanisms by which oxidative stress may accelerate the development of complications in diabetes are only partly known. There is however evidence for a role of protein kinase C, advanced glycation end products (AGE) and activation of transcription factors such as NF kappaB, but the exact signalling pathways and the interactions with ROI remain a matter of discussion. Additionally, results of very recent studies suggest a role for ROI in the development of insulin resistance. ROI interfere with insulin signalling at various levels and are able to inhibit the translocation of GLUT4 in the plasma membrane. Evidence for a protective effect of antioxidants has been presented in experimental studies, but conclusive evidence from patient studies is missing. Large-scale clinical trials such as the DCCT Study or the UKPDS Study are needed to evaluate the long-term effects of antioxidants in diabetic patients and their potential to reduce the medical and socio-economic burden of diabetes and its complications. Copyright (C) 2001 John Wiley & Sons, Ltd.
Abstract: The role of the different cytoskeletal structures like microfilaments (MF), microtubuli (MT), and intermediate filaments (IF) in phagosome motion is unclear. These cytoskeletal units play an important role in macrophage function (migration, phagocytosis, phagosome transport). We investigated ferromagnetic phagosome motions by cell magnetometry. J774A.1 macrophages were incubated with 1.3-microm spherical magnetite particles for 24 h, after which more than 90% of the particles had been phagocytized. Phagosome motions can be caused either by the cell itself (relaxation) or by applying magnetic twisting forces, yielding cell stiffness and viscoelastic properties of the cytoskeleton. Apparent viscosity of the cytoplasm was non-Newtonian and showed a shear-rate-dependent power law behavior. Elastically stored energy does not force the magnetic phagosomes back to their initial orientation: 57% of the twisting shear was not recoverable. Cytoskeletal drugs, like Cytochalasin D (CyD, 2 - 4 microM), Colchicine (CoL, 10 microM), or Acrylamide (AcL, 40 mM) were added in order to disturb the different cytoskeletal structures. AcL disintegrates IF, but affected neither stochastic (relaxation) nor directed phagosome motions. CyD disrupts MF, resulting in a retarded stochastic phagosome motion (relative decay 0.53 +/- 0.01 after 5 min versus 0.34 +/- 0.01 in control), whereas phagosome twisting shows only a small response with a 9% increase of stiffness and a small reduction of recoverable strain. CoL depolymerizes the MT, inducing a moderately accelerated relaxation (relative decay 0.28 +/- 0.01 after 5 min) and a 10% increase of cell stiffness, where the pure viscous shear is increased and the viscoelastic recoil is inhibited by 40%. Combining the two drugs conserves both effects. After disintegrating either MF or MT, phagosome motion and cytoskeletal stiffness reflect the behavior of either MT or MF, respectively. The results verify that the dominant phagosome transport mechanism is MF-associated. MT depolymerization by CoL induces an activation of the F-actin synthesis, which may induce an accelerated relaxation and an increase of stiffness. Cell mechanical properties are not modulated by MF depolymerization, whereas MT depolymerization causes a loss of viscous resistance and a loss of cell elasticity. The mean energy for stochastic phagosome transport is 5*10(-18) Joules and corresponds to a force of 7 pN on a single 1.3-microm phagosome.
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Abstract: Cytomagnetometry is a noninvasive method to investigate intracellular movements of organelles such as phagosomes by introducing magnetic particles into cells by phagocytosis, magnetizing them and measuring the field from the cells. To analyze the results of the cell-field measurement, we introduce a model for intracellular phagosome motion and investigate their behavior in terms of the cell field. The model includes an elastic body and two viscosity components which are ascribed to the filamentous structures surrounding the phagosomes. The magnetic relaxation phenomenon is assumed to derive from the rotationary Brownian motion as in our previous model. Although the model is simple, its behavior is not trivial because it contains a nonlinear term and the Brownian motion term. This model is the simplest one possible having a viscoelastic body and its behavior hence should be investigated thoroughly.
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Abstract: The distribution pattern of inhaled particles is an important factor for the evaluation of health effects. In this study, we morphologically investigated the fate of agglomerated ultrafine particles in macrophages in vitro and in vivo. Metallic silver (Ag) was chosen as a test particle, since it can be easily produced and detected by elemental and morphologic analyses. Ultrafine Ag particles generated by an electric spark generator in an argon atmosphere were collected on PTFE filters. The particles were suspended in distilled water and adjusted to different concentrations (10 [mu]g/ml to 1 mg/ml) with phosphate-buffered saline (PBS). For the in vitro study, J774 macrophage cell suspensions (200,000 cells in 400 [mu]l medium) were plated in small chambers. Six hours later, 100 [mu]l of the silver-PBS suspension was added to each chamber. For the next 9 days, the chamber slides were examined daffy with an inverted microscope in order to detect agglomerated particles in the cell. The medium was changed every day, and Ag in the medium was checked by inductively coupled plasma mass spectrometry (ICP-MS). On days 1, 3, 5, 7, and 9, cells in the chambers were fixed with 2.5% buffered glutaraldehyde and examined ultrastructurally. For the in vivo study using F344 rats, 50 [mu]g Ag particles were instilled intratracheally. On days 1, 4, and 7 following instillation, rats were sacrificed and the lungs were examined morphologically. The Ag content in the lung, liver, and lung-associated lymph nodes was analysed by ICP-MS. In the in vitro study, the dose-dependent presence of agglomerated particles was observed in J774 cells. The size and form of particles remained unchanged throughout the observation period. Electron microscopy with x-ray microanalysis showed that both single and agglomerated Ag particles were observed in the dilated phagolysosome of J774 cells. In the in vivo study, focal accumulation of Ag-particle-laden alveolar macrophages was found. Ag particles were also observed in the alveolar wall. Ag content in the lung was constant between day 1 and day 7, indicating that no rapid particle translocation from the lung to other organs had taken place in this time period. In vitro and in vivo studies suggested that agglomerated Ag particles remained in targets for a given period of time - at least up to 7 days. [Journal Conference Paper; 16 Refs; In English; Summary in English]
Abstract: We measured the viscoelastic properties of the cytoplasm of J774 macrophages with a recently developed microrheometer. Ferromagnetic beads (1.3 microm in diameter) were used to determine the local viscoelastic moduli. Step-force pulses were applied to the magnetic beads and the displacement was observed by single particle tracking. By analyzing the creep response curves in terms of a triphasic mechanical equivalent circuit, we measured the shear elastic modulus, the effective viscosities, and the strain relaxation time. The values of the shear modulus vary by more than an order of magnitude within the cell population (range, 20-735 Pa; average, 343 Pa) and by a factor of 2 within single cells. The effective viscosity of the cytoplasm exhibits a relatively sharp distribution about an average of eta = 210 Pa s (+/- 143 Pa s). We measured the displacement field generated by the local forces by observing the induced motion of nonmagnetic beads. Even at distances of the order of 1 microm, no induced motion was seen, suggesting that the cytoplasm is composed of clusters of densely packed and cross-linked filaments separated by soft regions. In another series of experiments we analyzed the magnetophoretic motion of the ferromagnetic beads at a constant magnetic force. Measuring the bead velocity parallel and perpendicular to the applied force showed that local active forces on the beads varied from 50 to 900 pN.
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Abstract: We have previously reported that interleukin (IL)-1beta decreases responsiveness of cultured human airway smooth muscle (HASM) cells to beta-agonists. The purpose of this study was to determine whether glucocorticoids inhibit this IL-1beta effect. Dexamethasone (Dex; 10 (-6) M) had no effect on concentration-related decreases in cell stiffness in response to isoproterenol (Iso) in control cells as measured by magnetic twisting cytometry but prevented the decreased responsiveness to Iso observed in IL-1beta (20 ng/ml)-treated cells. In addition, Dex had no effect on Iso-stimulated cAMP formation in control cells but prevented the IL-1beta-induced reduction in Iso- stimulated cAMP formation. Similar effects on cell stiffness and cAMP responses were seen after pretreatment with the glucocorticoid fluticasone proprionate (FP). Dex and FP also prevented IL-1beta- induced hyporesponsiveness to PGE(2) stimulation. In contrast, neither IL-1beta nor glucocorticoids had any effect on cell stiffness responses to dibutyryl cAMP. We have previously reported that the IL-1beta effect on beta-adrenergic responsiveness is mediated through cyclooxygenase-2 expression and prostanoid formation. Consistent with these observations, IL-1beta-induced cyclooxygenase-2 expression was virtually abolished by FP at concentrations of 10(-10) M and greater, with a resultant decrease in PGE(2) formation. However, Dex did not inhibit IL-1beta-induced nuclear translocation of nuclear factor-kappaB or activator protein- 1 in HASM cells. In summary, our results indicate that, in HASM cells, glucocorticoids alone do not alter responses to beta-agonists but do inhibit IL-1beta-induced beta-adrenergic hyporesponsiveness. Glucocorticoids mediate this effect by inhibiting prostanoid formation but without altering nuclear factor-kappaB or activator protein-1 translocation.
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Abstract: We have previously reported that pretreatment of cultured human airway smooth muscle (HASM) cells with interleukin-1beta (IL-1beta) results in decreased beta-adrenergic responsiveness. The purpose of this study was to determine whether prostanoids released as a result of cyclooxygenase-2 (COX-2) induction by IL-1beta contribute to this effect of the cytokine. Confluent serum-deprived HASM cells were studied in passages 4-7. IL-1beta (20 ng/ml for 22 h) reduced the ability of the beta-agonist isoproterenol (Iso) to decrease stiffness of HASM cells as measured by magnetic twisting cytometry. The effect of IL-1beta on Iso-induced changes in cell stiffness was abolished by nonselective (indomethacin (Indo), 10(-6) M) and selective (NS-398, 10(-5) M) COX-2 inhibitors. Indo and NS-398 also inhibited both the increased basal cAMP and the decreases in Iso- stimulated cAMP production induced by IL-1beta. IL-1beta (20 ng/ml for 22 h) caused an increase in both basal (15-fold) and arachidonic acid (AA)-stimulated (10-fold) PGE2 release. Indo blocked basal and AA-stimulated PGE2 release in both control and IL-1beta-treated cells. NS-398 also markedly reduced basal and AA-stimulated PGE2 release in IL-1beta-treated cells but had no significant effect on AA-stimulated PGE2 release in control cells. Western blot analysis confirmed the induction of COX-2 by IL-1beta. Exogenously administered PGE2 (10(-7) M, 22 h) caused a significant reduction in the ability of Iso to decrease cell stiffness, mimicking the effects of IL-1beta. Cycloheximide (10 microg/ml for 24 h), an inhibitor of protein synthesis, also abolished the effects of IL-1beta on Iso- induced cell stiffness changes and cAMP formation. In summary, our results indicate that IL-1beta significantly increases prostanoid release by HASM cells as a result of increased COX-2 expression. The prostanoids appear to contribute to beta-adrenergic hyporesponsiveness, perhaps by heterologous desensitization of the beta2 receptor.
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Abstract: Magnetopneumography (MPG) has been used to study long-term particle clearance from human lungs as well as cellular motility of pulmonary macrophages (PMs). This study describes an extension of the method enabling the measurement of mechanical properties of PM cells in vivo. Ferromagnetic microparticles are inhaled and then retained in the alveolar region of the lungs, where they are phagocytized within hours by PMs. The magnetic particles can be rotated in weak magnetic fields, and the response to this twisting shear (force) is detected as a macroscopic magnetic field producing a measure of cytoskeletal mechanics. Cytoplasmic viscosity is very high compared with that of water and is strongly non-Newtonian. Under rotational stresses from 0.4 to 6.4 Pa, it acts like a pseudoplastic fluid showing a characteristic shear rate dependence. The viscosity as well as the stiffness of the cytoskeleton increases with increasing shear stress as seems typical for living tissue and evidence for an intact cytoskeletal matrix. The particle recoil as measured by the amount of recoverable strain following a short twisting force describes a cytoplasmic elasticity that depends on both level and duration of stress. These investigations on the mechanical properties of living human cells are promising and should lead to better understanding of cellular dysfunction in disease as well as pathways for drug administration.
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Abstract: Using magnetic twisting cytometry (MTC), we measured the cytoskeletal stiffness of adherent human airway smooth muscle (HASM) cells. We hypothesized that modulation of actin-myosin interactions by application of contractile agonists would induce changes in cytoskeletal stiffness. In cells plated on high-density collagen, bradykinin (10(-6) M) and histamine (10(-4) M) increased stiffness by 85 +/- 15 and 68 +/- 16%, respectively. Increases in cell stiffness were also consistently observed after acetylcholine, substance P, and KCl. The bronchodilator agonists isoproterenol, prostaglandin E2, forskolin, dibutryl adenosine 3', 5'-cyclic monophosphate, and 8-bromoguanosine 3', 5'-cyclic monophosphate each caused a dose-dependent decrease in cell stiffness in unstimulated as well as bradykinin-treated cells. HASM cells plated on high- density collagen were stiffer than cells plated on low-density collagen (126 +/- 16 vs. 43 +/- 3 dyn/cm2) and developed more pronounced increases in stiffness in response to bradykinin as well as more pronounced decreases in stiffness in response to isoproterenol. These results are consistent with the hypothesis that modulation of actin-myosin interactions by application of contractile agonists causes changes in cytoskeletal stiffness of HASM cells. MTC may be a valuable tool for evaluating the mechanisms of pharmacomechanical coupling in airway smooth muscle cells in culture.
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Abstract: Magnetopneumography was applied to investigate intracellular phagosome motion in alveolar macrophage cells of healthy subjects (non-smokers and smokers). Ingested magnetic microparticles are inhaled and phagocytized by alveolar macrophages within hours. Thereby the particles are transferred into phagolysosomes. After magnetization the particles produce a macroscopic magnetic field of the lungs. Cellular motility causes a decay of the field (relaxation) by stochastic disorientation of the dipole particles (phagolysosomes) in the cells. Our studies have shown that the deposition of magnetite test particles induces a non-specific activation of the macrophage cells with a faster relaxation. This activation vanishes within the first day after particle deposition. This macrophage activation due to dust exposure was not present in smokers. It follows that cigarette smoking either causes a damage of the cellular defense or causes an adaptation of the macrophage cells to the permanent cigarette smoke inhalation.
Abstract: Mechanical loading of cells is of fundamental relevance in physiological processes and induces several functional responses in cells. Integrins, a family of adhesion receptors, which are responsible for the interaction with the extracellular matrix, may play a role in transmission of mechanical signals into cells. The osteogenic cell line U-2 OS expresses different integrin subunits which are uniformly distributed over the cell surface. We applied defined physical forces on individual integrin receptor subunits using paramagnetic microbeads coated with anti-integrin antibodies. Application of an inhomogeneous magnetic field consequently leads to a mechanical stress on the receptor. Intracellular Ca2+ increased when the alpha 2 or the beta 1 integrin subunits were stressed, whereas mechanical loading of the transferrin receptor had a significantly lower effect. This result indicates that forces specifically exerted to individual integrin receptors induce signal transduction pathways.
Notes: MEETING OF THE EUROPEAN RESPIRATORY SOCIETY (ERS), NICE, FRANCE, OCTOBER 1-OCTOBER 5, 1994. EUROPEAN RESPIRATORY JOURNAL; 7 (SUPPL. 18). 1994. 72S.
Abstract: Magnetic microparticles were used to investigate the defence system of the human lungs against foreign material. About 0.5 mg of spherical monodisperse magnetite particles were deposited in the alveolar region of the human lung by voluntary inhalation. After primary magnetization a remanent magnetic field (RMF) of the lung can be measured that allows estimation of the amount of dust retained in the lung. The decay of this RMF, called relaxation, results from a misalignment of the dipole particles due to the activity of pulmonary macrophages. This macrophage activity was characterized by a cell energy Ez. With a secondary magnetization the lung can be remagnetized by rotation of the dipole particles. This allows estimation of the intracellular viscosity and the motility of the alveolar macrophages in vivo. The macrophage cell- line J774 was used to verify the dynamic processes of the magnetic particles within the cells in vitro. In vitro and in vivo relaxation curves of polydisperse and of spherical monodisperse magnetite particles are presented. Thermal relaxation of mono-disperse and polydisperse particles within a viscous standard could be verified with the Brownian rotary diffusion model. Relaxation with monodisperse particles was double exponential in vivo as well as in vitro, suggesting that 2 different viscous compartments of the cytoplasm should be considered. Relaxation in the macrophage cell- line J774 was particle-size-dependent.
Abstract: Magnetic micro-particles were used to investigate the defence system of the human lungs against foreign material. After primary magnetisation a remanent magnetic field (RMF) of the lung can be measured that allows estimation of the amount of dust retained in the lung. After calibration of the system with a lung phantom the magnetic contamination retained in the lungs of dental technicians and welders was estimated at mean values of 22 and 500 mg respectively. In normal controls only 0.3 mg was found. About 0.5 mg of spherical monodisperse magnetite particles was deposited in the alveolar region of the lung by voluntary inhalation. The decay of the RMF, called relaxation, results from a misalignment of the dipole particles due to the activity of pulmonary macrophages. This macrophage activity is characterised by a cellular energy Ez. With a secondary magnetisation the lung can be remagnetised by rotation of the dipole particles. This allows an estimation of the intracellular viscoelasticity and the motility of the alveolar macrophages in vivo. Secondary magnetisation and relaxation curves of spherical monodisperse magnetic particles are presented. Intracellular viscosity was estimated to be n approximately equal to 100 Pa.s at shear rates near 0.01 s-1, the rigidity modulus being v approximately equal to 4-8 Pa. Macrophage activity was described by a cellular energy EZ approximately 5 x 10(-18) J. Additionally, non- magnetic aerosol exposure resulted in a faster relaxation, which was interpreted to be due to activation of the macrophages. The magnetite particles were cleared with a half-time of approximately 110 days.
Abstract: The models, used in magnetopneumography and in cytomagnetometry to estimate the intracellular viscosity and the energy of the cell motiliy are tested with either monodisperse and with polydisperse ferrimagnetic micro particles, suspended in highly viscous Newtonian solutions. Unsing polydisperse particles deviations form an exponential decay were measured. In relaxation and in viscosity measurements the particles behave with an increased hydrodynamic volume.
Abstract: Magnetic microparticles were used to investigate the defense system of the human lungs against foreign material. About 0.5 mg of spherical monodisperse magnetite particles were deposited in the alveolar region of the human lung by voluntary inhalation. After primary magnetization a remanent magnetic field (rmf) of the lung can be measured that allows estimation of the amount of dust retained in the lung. The decay of this rmf, called relaxation, results from a misalignment of the dipole particles due to the activity of pulmonary macrophages. This macrophage activity was characterized by a cell energy E z. With a secondary magnetization the lung can be remagnetized by rotation of the dipole particles. This allows estimation of the intracellular viscosity and motility of the alveolar macrophages in vivo. Secondary magnetization and relaxation curves of spherical monodisperse magnetite particles are presented. Intracelluar viscosity was estimated to be n approximately equal to 100 Paxs by a shear-rate near 0.01 s -1, macrophage activity was E z approximately equal to 5x10 -18 J. Aerosol exposure resulted in a faster relaxation, which was interpreted to be due to activation of the macrophages. The magnetite particles were cleared with a half-time of approximately equal to 110 days.
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Abstract: Ferrimagnetic microparticles of Fe3O4 were used to investigate the intracellular viscoelastic properties of alveolar macrophages in vivo. After primary strong field magnetization and complete relaxation the lung was remagnetized by a weak magnetic field. In this case the balance between magnetic torque and hydrodynamic retarding torque allows to model remagnetization. The deviations between measurements and theory make it necessary to estimate a non-Newtonian intracelluar viscosity. The measurements were analyzed according to the shear rate dependence of intracellular viscosity, giving a viscosity of 130 Pa·s a shear rate of 0.01 s-1, while it increases to above 10E3 Pa·s for shear rates below 0.001 s-1.
Abstract: An improved spinning top aerosol generator was built which allowed the generation of a monodisperse magnetic aerosol of magnetite (1309371) (Fe3O4) particles in different size ranges. This minimized the duration for inhalation of the aerosol needed to deposit the particles in the lungs. The generator was run with compressed nitrogen gas. An air driven spinning top STAK MK I was used for the aerosol generator. The efficiency of the aerosol generator was measured with the use of a laser aerosol spectrometer, where the scattered light of the particles was classified with a multichannel pulse height analyzer. After half an hour running a constant high performance of the generator was detected, with an output concentration near 1000/cubic centimeter. This corresponds to an efficiency of more than 80% and showed that this system works very well near the maximum achievable efficiency.
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Abstract: This study presented relaxation curves following inhalation and deposition of spherical magnetic particles with an aerodynamic diameter of 2.8 micrometers. The particles were magnetized during three different time intervals to test the elastic components of the environment of the particles in the lungs. Seven days after deposition a lung lavage was done with one person to obtain macrophages with particles for further study. The lavaged macrophages were observed under high resolution light and electron microscopy where the particles could be found in the phagolysosomes of the macrophages. Spherical particles were prepared by spraying a colloidal suspension of magnetite (1309371) with a spinning top aerosol generator. The study demonstrated that relaxation was strongly influenced by the magnetizing and clearance time and could reflect the uptake of particles by macrophages and their activity. Theoretical models are available that allow a physical interpretation of the relaxation curve.
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Abstract: Magnetopneumography was utilized to determine the amount of magnetic material in the lungs of dental technicians who worked with alloys containing high proportions of cobalt (7440484), iron (7439896), and nickel (7440020). The amounts determined were compared with those of occupationally nonexposed human referents and welders. The remanent magnetic field of the lung was measured by a superconducting quantum interference device sensor. Magnetization of the lungs was carried out either by the localized field technique (LFT) or by the homogeneous field technique (HFT). The magnetopneumography system was calibrated with a lung model of 4.6 liter volume, prepared from polyurethane foam with a homogeneous mixing of a known amount of magnetite particles. Descriptions were provided on the determination of the magnetic properties of three dust samples, namely, magnetite (1317619), fumes from stainless steel welding, and grinding dust from a dental technician laboratory, as well as on the calibration of the model. The calibration of the lung model, carried out by both the LFT and the HFT, revealed the remanent magnetic field over different positions of the right and left lung. Higher sensitivity was observed with the HFT. Dental technicians and welders exhibited significantly increased remanent magnetic fields over their lungs, as compared with the values for unexposed referents. The calculated amounts of magnetic material remaining in the lungs of dental technicians were 20.9 and 21.7 milligrams (mg) by the LFT and the HFT, respectively, compared with less than 1mg in the lungs of unexposed referents.
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Abstract: A method was developed for the detection of magnetic aerosols in human lungs. This magnetopneumographic system could detect, via superconducting quantum interference device (SQUID) sensors, particle magnetization within the lung after an inhalation exposure. The standard magnetic particles used in studies with this device were ferromagnetic magnetite particles which were spherical in shape. After inhalation of the magnetic particles, subjects were moved under a super conducting magnet, and the region to be measured was magnetized. Then the SQUID sensor was used to detect the change in magnetic flux in a second order gradiometer. Tests were conducted in a magnetically shielded room. For particles with a 3.5 micrometer aerodynamic diameter, under standard respiratory conditions 60 percent of the particles were deposited in the alveolar region. In this system, using localized fields, small areas of the lung could be studied with increased sensitivity associated with the field localization. Results from a particular study involving a group of welders who were exposed to magnetic particles and a nonexposed group exhibited particle levels up to 400 milligrams in the lungs of the welders, with comparison group subjects exhibiting below 1 milligram.
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