Equine Physiology
Cultured Cell Studies:
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- Rats 1: Physiol Res 2000;49(6):733-6 Ozone-induced micronuclei frequency in rat alveolar Type II cells. Chorvatovicova D, Hoet PH, Tatrai E, Kovacikova Z. Institute of Experimental Pharmacology, Slovak Academy of Sciences, Bratislava, Slovak Republic. The effect of ozone, a ubiquitous air pollutant, was tested on cultured pulmonary epithelial type II cells isolated from rats. After 40-hour culture, the cells were exposed for 6 h to 400 ppb of ozone or air. The number of micronucleated cells was counted after the exposure. In each group, 17000 cells were evaluated. The number of micronucleated cells was significantly increased in the ozone-exposed group (12.24 per 1000 cells) compared to the control group (5.00 per 1000 cells). The results showed the mutagenic effect of ozone exposure on alveolar type II cells, manifested in the increased frequency of their micronuclei. PMID: 11252542 [PubMed - in process]
1: Am J Physiol 1995 Oct;269(4 Pt 1):L527-35 Neutrophils enhance removal of ozone-injured alveolar epithelial cells in vitro. Cheek JM, McDonald RJ, Rapalyea L, Tarkington BK, Hyde DM. Department of Veterinary Anatomy, Physiology, and Cell Biology, University of California, Davis 95616, USA. After acute exposure to oxidant gases in vivo, migration and accumulation of inflammatory cells in pulmonary epithelium coincides with epithelial cell necrosis. The present study was designed to test quantitatively the hypothesis that quiescent neutrophils enhance the removal of oxidant-injured pulmonary epithelial cells after exposure to ozone in vitro. Primary isolated rat alveolar type II cells were cultured as monolayers, using serum-free medium. After exposure to 0.1-0.5 ppm ozone for 0.5 h, apical sides of monolayers were administered either fresh nutrient medium only or medium containing quiescent human neutrophils. Monolayer bioelectric properties and cellular uptake of vital dye were recorded from 5 to 48 h after ozone exposure. Ozone dose-dependent increases in monolayer permeability were associated with proportionally higher numbers of injured epithelial cells. However, the direction and magnitude of neutrophil effects on monolayer permeability after ozone exposure were dependent on ozone concentration. Furthermore, neutrophil-treated monolayers exposed to 0.1 ppm ozone had significantly fewer attached cells positive for uptake of vital dye relative to monolayers exposed to the low level of ozone only; this effect was ablated with increasing ozone concentration. These data suggest that at high levels of ozone neutrophils may exacerbate injury to oxidant-impaired epithelial cells, whereas the presence of neutrophils after exposure to ambient concentrations of ozone may expedite the restoration of epithelial barrier function. We conclude that, by enhancing the removal of injured cells, neutrophils may facilitate the repair of centriacinar epithelium after ozone exposure in vivo. PMID: 7485526 [PubMed - indexed for MEDLINE]
1: Toxicol Appl Pharmacol 1994 Mar;125(1):59-69 Ozone injury to alveolar epithelium in vitro does not reflect loss of antioxidant defenses. Cheek JM, Buckpitt AR, Li C, Tarkington BK, Plopper CG. Department of Veterinary Anatomy, University of California, Davis 95616. The objective of this study was to characterize an in vitro model of oxidant gas toxicity, using primary cultures of alveolar type II cells maintained in serum-free medium, by evaluating (1) epithelial barrier function, (2) the stability of cellular antioxidant defenses, and (3) the response of alveolar epithelial barrier properties to ozone exposure. Antioxidant enzyme activities and glutathione levels were measured in rat type II cells that were freshly isolated, cultured for 1 day in serum-supplemented medium, and subsequently grown in serum-free nutrient medium. After measurement of peak bioelectric properties on Day 4 in primary culture, alveolar epithelial monolayers were exposed to ozone at various concentrations and lengths of exposure. Ozone-induced alterations in monolayer bioelectric properties and impairment of cellular organization were used to evaluate oxidant injury. The primary effect of ozone exposure was a dose-dependent increase in monolayer permeability, which resulted from damage to intercellular junctions and/or loss of epithelial integrity. Extensive and persistent permeability increases correlated with focal areas of epithelial degradation. The focal nature of ozone injury to alveolar epithelium in vitro suggests that individual cell susceptibility to oxidant stress may account for the overall decrement in barrier function. However, this sensitivity does not result from overall loss of antioxidant defenses associated with cell culture, as these monolayers (when cultured in serum-free medium) maintained their antioxidant enzyme activities and glutathione content at levels found in freshly isolated cells. We conclude that the sensitivity of these monolayers to ozone injury in vitro reflects a disproportionate degree of oxidant stress on cell membranes relative to intracellular antioxidant defenses, i.e., cellular susceptibility to oxidant injury may depend on the ratio of the surface area of the cell to its cytoplasmic volume. PMID: 8128496 [PubMed - indexed for MEDLINE]
1: Exp Lung Res 1990 Nov-Dec;16(6):561-75 NO2 decreases paracellular resistance to ion and solute flow in alveolar epithelial monolayers. Cheek JM, Kim KJ, Crandall ED. Seaver Cardiopulmonary Laboratory, Will Rogers Institute Pulmonary Research Program, Cornell University Medical College, New York, New York. Primary cultured monolayers of rat alveolar epithelial cells grown on tissue culture-treated Nuclepore filters were exposed to 2.5 ppm nitrogen dioxide (NO2) for 2-20 min. Changes in monolayer bioelectric properties and solute permeabilities were subsequently measured. Exposure to NO2 produced a dose-dependent decrease in monolayer transepithelial electrical resistance (Rt), whereas monolayer short-circuit current was unaffected. Post-exposure monolayer permeability to 14C-sucrose (which primarily crosses alveolar epithelium via the paracellular pathway) increased markedly. That for 3H-glycerol (which permeates through both paracellular and transcellular pathways) increased to a lesser extent. Partial recovery of Rt and solute permeabilities was noted by 48-h post-exposure. The time courses of the decrease in Rt and increase in solute permeabilities were similar. These results suggest that NO2 primarily impairs passive alveolar epithelial barrier functions in vitro, probably by altering intercellular junctions, and does not appear to directly affect cell membrane active ion transport processes. When correlated with results obtained from experimental approaches, studies of in vitro alveolar epithelial monolayers may facilitate investigations of dosimetry, sites, and mechanisms of oxidant injury in the lung. PMID: 2081503 [PubMed - indexed for MEDLINE]
1: Res Rep Health Eff Inst 1987 Oct;(13):3-19 Effects of nitrogen dioxide on alveolar epithelial barrier properties. Crandall ED, Cheek JM, Shaw ME, Postlethwait EM. Division of Pulmonary and Critical Care Medicine, Cornell University. This study analyzed the effects of nitrogen dioxide (NO2) on alveolar epithelial permeability and transport properties. Primary cultured monolayers of rat Type II pneumocytes, cultured on both nonporous and porous surfaces, were used as models of isolated alveolar epithelium for in vitro exposure to nitrogen dioxide. The effects of nitrogen dioxide exposure for monolayers cultured on nonporous substrata were monitored by observing the changes in the net volume of fluid under the monolayer; for cells cultured on porous substrata, alterations in tissue bioelectric properties were noted. As a first step, primary cultured monolayers of rat Type II pneumocytes plated on nonporous plastic Petri dishes were used to investigate the effects of nitrogen dioxide on alveolar epithelial barrier properties. Such monolayers form fluid filled domes that are thought to result from active solute transport from medium to substratum, with water following passively. We used dome formation as a transport marker. Five-day-old cultures were directly exposed to 30 ppm NO2 in 5 percent CO2 in air at 25 degrees C, by cyclically tilting culture plates from side to side, so that both halves of the monolayer were exposed during each cycle. Exposures consisted of 10 cycles of four minutes each (two minutes per side), for a cell exposure time of 20 minutes. Control plates were simultaneously exposed to 5 percent CO2 in air under identical conditions. One day after the exposure, nitrogen dioxide-exposed monolayers exhibited significant decreases in dome density and individual dome volume, compared to the controls. By 48 hours post-exposure, differences between nitrogen dioxide-exposed and control monolayers were less, but remained significant. These results showed that short-term sublethal exposures to nitrogen dioxide produce a decrease in dome formation in Type II alveolar epithelial cell monolayers. This finding is most likely due to a decrease in the active transepithelial sodium transport rate, or an increase in the permeability of cell membranes or tight junctions, or both. Addition of vitamin E-containing liposomes to the culture media 24 hours pre-exposure did not affect the nitrogen dioxide-induced decrease in dome formation, indicating that under these circumstances no protective effect was provided by the antioxidant.(ABSTRACT TRUNCATED AT 400 WORDS) PMID: 3269254 [PubMed - indexed for MEDLINE]
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