Faculty Bibliography

Intracellular free Ca2+ ([Ca2+]i) plays a vital role both in maintaining normal cellular function and in cell killing. Few studies have been published regarding its role in ozone (O3)-induced health effects. This study investigated the effect and mechanism of O3 exposure on [Ca2+]i in human tracheal epithelial (HTE) cells. HTE cells grown on Costar Transwell inserts with a liquid-gas interface were exposed to 0, 0.05, 0.1, 0.2 and 0.4 ppm O3 at 37 degrees C for 1 h. After exposure, [Ca2+]i was measured using the fluorescent dye Fluo 3. O3 at 0.4 ppm produced a significant increase in [Ca2+]i, and the increases in [Ca2+]i were blocked by verapamil and 8-(diethylamino)-octyl-3,4,5,-trimethoxybenzoate (TMB-8). These results suggest that the O3-induced [Ca2+]i elevation may involve both Ca2+ release from internal stores and Ca2+ influx across the plasma membrane. Furthermore, both buffer and cell lysate of HTE cells exposed to 0.4 ppm O3 caused a rapid increase in [Ca2+]i of THP-1 human phagocytic monocytes, but the buffer and lysate from air exposed cells did not. These results suggest that O3 exposure causes HTE cells to release a diffusible mediator from the empty Ca(2+)-storing organelle and may be responsible for the sustained and persistent [Ca2+]i elevation in HTE cells exposed to 0.4 ppm O3

Recent studies have demonstrated that additive and synergistic effects on rabbit pulmonary macrophages (PM phi) function can occur after combined exposures to acid aerosols and ozone. This study investigated intracellular pH (pHi) homeostasis and H+ extrusion mechanisms of PM phi from rabbits exposed to sulfuric acid, ozone, and their mixtures. Animals were exposed for 3 h to 125 micrograms/m3 sulfuric acid, 0.1, 0.3, 0.6 ppm ozone, or combinations of acid with each concentration of ozone, and the pHi was determined by a fluorescent dye ratioing technique. Exposure to 125 micrograms/m3 acid reduced pHi and exposure to ozone resulted in a concentration-dependent reduction in pHi. Ozone generally tended to mitigate the effect of the acid aerosol on pHi. Other groups of rabbits were exposed to 50 micrograms/m3 sulfuric acid, 0.6 ppm ozone, or their mixture, for 3 h, and PM phi were again harvested. The pHi of PM phi following exposure to each of the pollutant atmospheres was not different from control. However, H+ extrusion with an imposed internal acid load was found to be significantly depressed following exposure to either sulfuric acid or ozone alone, while the mixture produced a significant interaction

The fumarate salt of methylecgonidine (MEG; anhydroecgonine methylester), a pyrolysis product of cocaine, has previously been shown to antagonize contractions of guinea pig isolated trachea induced by acetylcholine (ACh) and other spasmogenics. We determined the effects of MEG fumarate on ACh-induced bronchoconstriction in vivo. Specific airway conductance (SGaw) was measured in guinea pigs receiving 30-300 mg/kg s.c. MEG fumarate and exposed one hour later to nebulized ACh (0.2-3.2%; by inhalation). MEG fumarate did not induce any changes in SGaw; neither did it antagonize dose-dependent decreases in SGaw induced by ACh. However, tremors, salivation, startle and increased numbers of fecal boli were observed after MEG administration. Thus, unlike antagonism of ACh-induced contractions of guinea pig isolated trachea observed in vitro, MEG fumarate does not antagonize ACh-induced bronchoconstriction in vivo, even at doses which induced changes in grossly-observable behavior. Inhalation of a condensation aerosol of MEG base induced lung damage as evidenced by the presence of blood and higher levels of protein and lactate dehydrogenase in the lung lavage fluid of MEG-treated animals than of control animals. Aerosols of MEG fumarate, on the other hand, did not induce lung damage when inhaled. These results extend previous observations that MEG base may contribute to detrimental pulmonary effects of crack smoking

The pulmonary effects of the cocaine pyrolysis product, methylecgonidine (MEG; anhydroecgonine methyl ester), were assessed in guinea pigs. Specific airway conductance (SGaw), which decreases during bronchoconstriction, was measured in guinea pigs exposed to atmospheres containing a condensation aerosol of MEG free base (13 +/- 1 mg/liter of air), nebulized MEG fumarate (3 and 12% in phosphate buffered saline) or nebulized acetylcholine chloride (0.2 and 0.4% in phosphate buffered saline). A decrease in SGaw to 24.0 +/- 4.2% (mean +/- 2 S.E.M.) of baseline levels was observed in guinea pigs breathing MEG free base. A decrease to 28.4 +/- 4.5% of baseline was observed following administration of 0.4% acetylcholine. No change in SGaw was measured in guinea pigs exposed to 3% MEG fumarate but SGaw was reduced to 69.3 +/- 5.3% of baseline after exposure to 12% MEG fumarate. MEG free base poses an alkaline challenge to the lung, 3% MEG fumarate is neutral (pH approximately 7.4) and 12% MEG fumarate is acidic (pH approximately 4.3); thus, MEG free-base and 12% MEG fumarate might provoke a reflex bronchoconstriction due to direct pulmonary irritant effects. These results suggest that MEG free base produced during crack pyrolysis may play a role in bronchoconstriction observed in crack smokers

BACKGROUND: A number of epidemiologic studies have suggested that every year environmental tobacco smoke (second-hand smoke) is responsible for tens of thousands of deaths, mostly from heart disease, in the United States. Environmental tobacco smoke is composed mainly (80% to 85%) of aged and diluted sidestream smoke. The remainder is exhaled mainstream smoke. Among the thousands of compounds that have been identified in environmental tobacco smoke are a number of carcinogens, including polynuclear aromatic hydrocarbon carcinogens, such as benzo(a)pyrene. We have demonstrated previously that a number of carcinogens, including benzo(a)pyrene, promote plaque development after injection into cockerels. There have been almost no studies showing a direct stimulatory effect of environmental tobacco smoke on plaque development. Recently we demonstrated that cockerels exposed to sidestream smoke for approximately 0.4% of their projected lifespan exhibited accelerated development of arteriosclerotic plaques. In that study, cockerels in specially designed inhalation chambers were exposed to the steady-state sidestream smoke from 5 cigarettes for 6 h/d for 16 weeks. This level of exposure is high but environmentally plausible. Statistically significant increases in plaque size were demonstrated in the smoke-exposed cockerels. METHODS AND RESULTS: In the present study, exposure levels were decreased by a factor of 5. Thirty cockerels were exposed to the steady-state sidestream smoke from 1 cigarette for 6 hours per day for 16 weeks. The smoke was mixed with filtered air. Ten control cockerels were exposed to filtered air only. Levels of smoke surrogates, including carbon monoxide and total suspended particulates, were measured three times a day. Again, there was a statistically significant increase in plaque size in the smoke-exposed cockerels. To place these studies within a context of environmental relevance, levels of carbon monoxide were measured independently over 1 to 3 hours in four bars where there was heavy smoking. Measured carbon monoxide levels were as high or higher in the bars than they were in the exposure chambers during the 1-cigarette sidestream-smoke study. CONCLUSIONS: Experimental exposure to secondhand smoke at levels equal to or even below those routinely encountered by people in smoke-filled environments is sufficient to promote arteriosclerotic plaque development

Biological responses to inhaled acid sulfates result from the deposition of hydrogen ion (H+) on airway surfaces. Thus, effects from sulfuric acid and ammonium bisulfate, the two major ambient species, have been assumed to be the same for a given H+ concentration within the exposure atmosphere, assuming similar respiratory tract deposition patterns. However, recent inhalation studies have indicated that sulfuric acid is disproportionately potent compared to ammonium bisulfate when the H+ content of the exposure atmosphere is considered, suggesting that some factors following inhalation affect the amount of H+ contacting airway surfaces. This study assessed a mechanism potentially underlying this phenomenon, namely, the extent of neutralization by respiratory tract ammonia. This was evaluated using a physical model system designed to mimic transit of these aerosols in the upper respiratory tract of the animal model used in this laboratory, the rabbit. The results suggest that for equal exposure quantities of H+, more acid would be deposited when sulfuric acid is inhaled than when ammonium bisulfate is inhaled. Furthermore, results from a series of in vitro exposures of tracheal epithelial cells to sulfuric acid and ammonium bisulfate aerosols indicated that the biological response is a function of the total mass (ionic) concentration of H+ deliverable to the cells or the total extractable H+ per particle. The results of this study have possible implications for ambient monitoring of particulate-associated strong acidity, suggesting that it may be necessary to specilate such measures into the relative amounts of H+ as sulfuric acid or ammonium bisulfate in order to most accurately relate atmospheric acid levels to observed health effects. In addition, since much of the ambient particulate-associated H+ exists as sulfuric acid/ammonium bisulfate mixtures rather than pure compounds, H(+)-associated health effects from controlled exposure studies of sulfuric acid may not be transferable to ambient population situations on a 1:1 basis. Since any such errors in exposure assessment will necessarily bias downward the strength of H(+)-related health effects associations found via epidemiological studies, failure to address the specification of H+ may cause such studies to underestimate the human health effects of strong acids

Inhaled acids are associated with adverse health effects, a conclusion based largely on studies with particulate-associated acid sulfates. The acidic component of ambient air in some regions, such as California, contains nitric acid (HNO3) vapor, but there is a limited database concerning its biological effects. Furthermore, effects of HNO3 may be modulated by coexposure to other pollutants, such as ozone (O-3). Rabbits were exposed for 4 h/day, 3 days/wk for 4 wk to HNO3 vapor at 0, 50, 150, and 450 mu g/m(3) alone; 0.15 ppm 0(3) alone; or to a mixture of 50 mu g/m(3) HNO3 + 0.15 ppm 0(3). Exposure was followed by assays of biochemical markers in lavage fluid pulmonary macrophage function, and in vitro bronchial responsivity to smooth muscle constrictor challenge. Nitric acid had no effect on viability or numbers of cells recovered, nor on lactate dehydrogenase or total protein in lavage. All acid concentrations reduced both basal levels and stimulated production of superoxide anion by macrophages, while the release/activity of tumor necrosis factor by stimulated macrophages was reduced following exposure to greater than or equal to 150 mu g/m(3) HNO3. Bronchi from rabbits exposed to greater than or equal to 150 mu g/m(3) HNO3 exhibited reduced smooth muscle responsivity in vitro compared to control. Although exposure to the HNO3/O-3 mixture resulted in no interaction for most end points, antagonism was noted for stimulated superoxide production, while synergism was noted for spontaneous superoxide production and bronchial responsivity. Exposure to the mixture resulted in a total abrogation of response to spasmogens in most bronchi examined and a marked attenuation in others. These results indicate that HNO3, when inhaled in vapor phase, may adversely impact upon pulmonary health by affecting target sites throughout the lungs, and that inhalation of an HNO3/O-3 mixture can produce synergistic interaction in affecting some biological parameters

In vivo exposure to sulfuric acid aerosols produces profound effects on pulmonary macrophage (PM phi) phagocytic function and cytokine release and perturbs intracellular pH (pHi) homeostasis. Because pHi influences a multitude of cellular processes, we sought to investigate the mechanism by which acid aerosol exposure affects its regulation. Guinea pigs underwent a single or 5 repeated 3-hr exposures to sulfuric acid aerosol (969 and 974 micrograms/m3 for single and repeated exposures, respectively). PM phi harvested immediately after exposure were incubated in HCO3-free media and their pHi recovery from an intracellular acid load was examined. The overall pHi recovery was depressed after single and multiple exposures to sulfuric acid aerosol. delta pHi (the difference between initial pHi and the one measured at 150 sec) decreased by 15.6 and 23.3% (p < 0.05) for single and repeated exposures, respectively. Initial dpHi/dt (maximum pHi recovery rate) after cytoplasmic acidification diminished by 20.3 and 32.2%, which were not statistically significant (p = 0.08 for repeated exposure). To determine whether the activity of the H(+)-ATPase pump the Na(+)-H+ exchanger was specifically altered by the acid exposures, PM phi were first incubated in Na+ and HCO3-free media with NBD-Cl (7-chloro-4-nitrobenz-2-oxa-1,3-diazol, blocking H(+)-ATPase and leaving only the Na(+)-H+ exchanger in effect) and then challenged with 30 mM NaCl. The pHi recovery of PM phi after Na challenge was significantly reduced in acid aerosol exposed guinea pigs (p < 0.05) compared to controls (for delta pHi, 18.2% lower in single exposure and 22.7% in multiple exposure groups; for initial dpHi/dt, 26.9% lower in single exposure and 22.4% in multiple exposure groups). In contrast, the H(+)-ATPase pump was inconsistently affected as indicated by delta pHi and initial dpHi/dt measured in the presence of MIA (amiloride-5-N-methylisobutyl, inhibiting the Na(+)-H+ exchanger and leaving only the H(+)-ATPase pump in effect). These results suggest that in vivo exposure to sulfuric acid aerosols induces alterations in pHi regulation in guinea pig PM phi attributable to changes in Na(+)-H+ exchanger activity