Maureen Sisco

Pulmonary Immunotoxic Potentials of Metals Are Governed by Select Physicochemical Properties: Vanadium Agents

The in situ reactions of metal ions/complexes are important in understanding the mechanisms by which environmental and occupational metal particles alter lung immune responses. A better understanding of these reactions in situ will also allow for the improved specificity and controlled toxicity of novel metallocompounds to be used as inhaled diagnostics or therapeutics. Our previous work showed that inhalation of metals (e.g., chromium, vanadium, nickel) caused altered lung immune cell function and host resistance. The data also suggested that the degree of immunomodulation induced depended not only on the amount of metal deposited, but also the compound used. If specificity governs pulmonary immunomodulatory potential, it follows that physicochemical properties inherent to the metal have a role in the elicited effects. We hypothe-size that major determinants of any metal compounds potential are its redox behavior, valency (generally referred to as oxidation state and considered speciation in chemical literature), and/or solubility. In accord with the extensive work carried out with vanadium (chemical symbol V) compounds showing the importance of form used, differences in potential for a range of V agents (pentavalent [VV] insoluble vanadium pentoxide and soluble sodium metavanadate, tetravalent [VIV] vanadyl dipicolinate, and trivalent [VIII] bis(dipicolinato)vanadium) were quantified based on induced changes in local bacterial resistance after host inhalation of each agent at 100 μ g V/m3 (5 hr/d for 5 d). Differences in effect between VV forms indicated that solubility was a critical property in in situ pulmonary immunotoxicity. Among the soluble forms, oxidizing vanadate had the greatest impact on resistance; reducing VIII altered resistance to a lesser extent. Both the VIV and insoluble VV had no effect. When data was analyzed in the context of pre-infection lung V burdens, soluble V agents with different oxidation states induced varying responses, supporting the hypothesis that differences in immunomodulatory potential might be attributed to redox behavior or valency. Our findings both provide a basis for understanding why some metals could be a greater health risk than others (when encountered in equal amounts) and will assist in the design of inhalable metallopharmaceuticals by allowing researchers to preempt selection of certain metal ions or complexes for use in such products.

Immunotoxicity of sulfuric acid aerosol: effects on pulmonary macrophage effector and functional activities critical for maintaining host resistance against infectious diseases.

Despite the widespread occurrence of acidic sulfur oxides in the ambient environment and their potential risks to human health, effects associated with pulmonary immune defenses have been poorly studied. The current in vivo study was designed to provide some insight into this relatively unexplored area by investigating the impact of inhaled sulfuric acid on immune defense mechanisms critical for maintaining pulmonary resistance against infectious diseases. Results of this study demonstrate that repeated inhalation of sulfuric acid reduces the uptake and intracellular killing of pathogenic bacteria by exposed pulmonary macrophages, and depresses the activity/production of important biological modifiers critical for maintaining pulmonary immunocompetence. These findings have important implications for human health, and may contribute to a better understanding of the possible mechanism(s) underlying the epidemiological evidence which suggests an association between total sulfates in the ambient air and increased incidence of acute bronchitis and lower respiratory illness in school-age children.

EFFECTS OF INHALED SULFURIC ACID AEROSOLS ON PULMONARY IMMUNOCOMPETENCE : A COMPARATIVE STUDY IN HUMANS AND ANIMALS

There is increasing concern regarding the potential health effects associated with the inhalation of ambient acid aerosols. Laboratory studies with animals have demonstrated that inhalation exposure to sulfuric acid (H2SO4) can alter airway responsiveness, cause cell damage leading to alveolitis and edema, produce hypertrophy/hyperplasia of epithelial secretory cells in the tracheobronchial tree, and alter nonspecific and specific immune defense mechanisms. While the adverse effects of inhaled H SO upon pulmonary 2 4 immunocompetence in animals appear relatively clear, data concerning effects on host defense in exposed humans are inconclusive and appear to contrast with those observed in animal models. Despite the fact that extrapolation between different species is of major importance in inhalation toxicology, baseline comparative data between species are lacking, which often results in the questionable validity of extrapolation modeling. This interlaboratory study was designed to compare the effects pro...

Effects of metal compounds with distinct physicochemical properties on iron homeostasis and antibacterial activity in the lungs: chromium and vanadium

In situ reactions of metal ions or their compounds are important mechanisms by which particles alter lung immune responses. The authors hypothesized that major determinants of the immunomodulatory effect of any metal include its redox behavior/properties, oxidation state, and/or solubility, and that the toxicities arising from differences in physicochemical parameters are manifest, in part, via differential shifts in lung iron (Fe) homeostasis. To test the hypotheses, immunomodulatory potentials for both pentavalent vanadium (VV; as soluble metavanadate or insoluble vanadium pentoxide) and hexavalent chromium (CrVI; as soluble sodium chromate or insoluble calcium chromate) were quantified in rats after inhalation (5 h/day for 5 days) of each at 100 μg metal/m3. Differences in effects on local bacterial resistance between the two VV, and between each CrVI, agents suggested that solubility might be a determinant of in situ immunotoxicity. For the soluble forms, VV had a greater impact on resistance than CrVI, indicating that redox behavior/properties was likely also a determinant. The soluble VV agent was the strongest immunomodulant. Regarding Fe homeostasis, both VV agents had dramatic effects on airway Fe levels. Both also impacted local immune/airway epithelial cell Fe levels in that there were significant increases in production of select cytokines/chemokines whose genes are subject to regulation by HIF-1 (whose intracellular longevity is related to cell Fe status). Our findings contribute to a better understanding of the role that metal compound properties play in respiratory disease pathogenesis and provide a rationale for differing pulmonary immunotoxicities of commonly encountered ambient metal pollutants.

Interleukin-19: a constituent of the regulome that controls antigen presenting cells in the lungs and airway responses to microbial products.

Background Interleukin (IL)-19 has been reported to enhance chronic inflammatory diseases such as asthma but the in vivo mechanism is incompletely understood. Because IL-19 is produced by and regulates cells of the monocyte lineage, our studies focused on in vivo responses of CD11c positive (CD11c+) alveolar macrophages and lung dendritic cells. Methodology/Principal Findings IL-19-deficient (IL-19-/-) mice were studied at baseline (naïve) and following intranasal challenge with microbial products, or recombinant cytokines. Naïve IL-19-/- mixed background mice had a decreased percentage of CD11c+ cells in the bronchoalveolar-lavage (BAL) due to the deficiency in IL-19 and a trait inherited from the 129-mouse strain. BAL CD11c+ cells from fully backcrossed IL-19-/- BALB/c or C57BL/6 mice expressed significantly less Major Histocompatibility Complex class II (MHCII) in response to intranasal administration of lipopolysaccharide, Aspergillus antigen, or IL-13, a pro-allergic cytokine. Neurogenic-locus-notch-homolog-protein-2 (Notch2) expression by lung monocytes, the precursors of BAL CD11c+ cells, was dysregulated: extracellular Notch2 was significantly decreased, transmembrane/intracellular Notch2 was significantly increased in IL-19-/- mice relative to wild type. Instillation of recombinant IL-19 increased extracellular Notch2 expression and dendritic cells cultured from bone marrow cells in the presence of IL-19 showed upregulated extracellular Notch2. The CD205 positive subset among the CD11c+ cells was 3-5-fold decreased in the airways and lungs of naïve IL-19-/- mice relative to wild type. Airway inflammation and histological changes in the lungs were ameliorated in IL-19-/- mice challenged with Aspergillus antigen that induces T lymphocyte-dependent allergic inflammation but not in IL-19-/- mice challenged with lipopolysaccharide or IL-13. Conclusions/Significance Because MHCII is the molecular platform that displays peptides to T lymphocytes and Notch2 determines cell fate decisions, our studies suggest that endogenous IL-19 is a constituent of the regulome that controls both processes in vivo.

Effects of inhaled ozone on pulmonary immune cells critical to antibacterial responses in situ

The goal of this study was to examine effects from repeated exposure to ozone (O 3) on immune cells involved in cell-mediated antibacterial responses in the lungs. Rats exposed to 0.1 or 0.3 ppm O 3 for 4 h/day, 5 days/wk, for 1 or 3 wk were analyzed for the ability to clear an intrapulmonary challenge with Listeria monocytogenes or had their lungs processed to obtain pulmonary alveolar macrophages (PAM) and lung-associated lymphocytes for analyses of select cell functions and surface marker expression. The results indicate that repeated inhalation exposure to O 3 affected local cell-mediated immunity (CMI) responses as evidenced by effects on clearance of Listeria. However, this modulation was not consistently dependent on exposure concentration or duration. Short-term repeat exposures had more effect on host resistance than did the more prolonged regimen, with rats exposed to 0.1 ppm O 3 most adversely impacted. Clearance patterns suggest modifications in innate resistance following 1 wk of exposure to 0.1 ppm O 3, but no similar effect following a 3-wk regimen. Exposure to 0.3 ppm O 3 appeared to affect both innate and acquired resistance after a 1-wk regimen, but mainly the former after an additional 2 wk of exposure. We conclude that these two mechanisms of resistance are differentially affected by O 3 and that distinct time- and O 3 concentration-dependent adaptation phenomena evolve for each; that is, in situ adaptation to higher levels of O 3 may occur more readily with acquired than with innate/PAM-dependent resistance. A similar pattern of inconsistent effect on PAM and lung-associated lymphocytes was also evident. For example, while 3-wk exposures had a greater effect on PAM reactive oxygen intermediate ROI production, evidence for a significant effect on antibacterial activity was only notable among PAM from rats exposed for 1 wk. Among lung lymphocytes, while 3-wk exposure to 0.1 ppm O 3 led to a significant increase in CD25 expression, there was no corresponding increase in responsivity to concanavalin A (ConA); only among cells from 1-wk-exposed rats did lymphoproliferative responses increase. Though investigations of altered immune cell cytokine receptor expression/binding activity are ongoing, results herein provide further evidence to support our longstanding hypothesis that some well-documented effects of O 3 exposure on human health are quite likely linked to changes in local immune cell (i.e., PAM and lung-associated lymphocytes) functions, with the latter being related to changes in the capacities of these cells to interact with immunoregulatory cytokines.

Impact of coexposure to ozone on the carcinogenic potential of inhaled chromium. I. Effects on retention and on extra- and intracellular distribution

A health hazard to welders is development of lung cancer. It is believed that this is likely due, in part, to the presence in welding fumes of several hexavalent chromium (Cr[VI]) species, whose solubility depends primarily on which process (i.e., manual metal arc verus metal-inert gas) is used. However, inhalation of Cr alone is uncommon in this setting. Thus, an examination of potential contributions from other coinhalants in creating or enhancing conditions whereby inhaled fume-associated Cr (primarily the insoluble forms) may initiate cancer is critical to increasing our understanding and preventing this particular occupational disease. One major chemical species formed and released during welding is ozone (O 3 ). Though implications of adverse pulmonary effects from individual exposure to Cr or O 3 have been investigated, those from simultaneous exposure are unclear. To begin to address whether the carcinogenic potential of insoluble Cr[VI] agents might be enhanced in hosts inhaling mixtures of Cr and O 3 versus Cr alone, analyses of total lung Cr burden, Cr retention in lung epithelium and interstitium, and potential shifts in lung cell distribution of Cr from the cytoplasm to nuclei were undertaken in F-344 rats exposed nose-only (5 h/d, 5 d/wk for up to 48 wk) to an extrapolated occupationally relevant level of Cr (360 w g Cr/m 3 as calcium chromate) alone and in combination with 0.3 ppm O 3 . Overall, there was only a nominal effect from O 3 on Cr retention or on distribution of Cr particles among extracellular sites and within lung cells. However, there were O 3 -related effects upon mechanisms for clearing the Cr from the deep lung, specifically at the levels of particle uptake and postphagocytic/endocytic processing by macrophages. This O 3 exposure-related shift in normal pulmonary clearance might potentially increase the health risk in workers exposed to other insoluble or poorly soluble carcinogenic Cr compounds.

OZONE DIFFERENTIALLY MODULATES AIRWAY RESPONSIVENESS IN ATOPIC VERSUS NONATOPIC GUINEA PIGS

While acute exposures to ozone (O 3) can alter airway responsiveness, effects from long-term exposures at low concentrations are less clear. This study assessed whether such exposures could induce nonspecific hyperresponsiveness in nonatopic (nonsensitized) guinea pigs and/or could exacerbate the pre-existing hyperresponsive state in atopic (sensitized) animals, and whether gender was a factor modulating any effect of O 3. Responsiveness was measured during and following exposures to 0.1 and 0.3 ppm O 3 for 4 h/day, 4 days/wk for 24 wk in male and female nonsensitized animals, those sensitized to allergen (ovalbumin) prior to initiation of O 3 exposures, and those sensitized concurrently with exposures. Ozone did not produce hyperresponsiveness in nonsensitized animals, but did exacerbate hyperresponsiveness to both specific and nonspecific bronchoprovocation challenges in sensitized animals, an effect that persisted through at least 4 wk after exposures ended. Gender was not a factor modulating response to O 3. Induced effects on responsiveness were not associated with numbers of eosinophils in the lungs nor with any chronic pulmonary inflammatory response, but were correlated with antigen-specific antibodies in blood. This study supports a role for chronic O 3 exposure in the exacerbation of airways dysfunction in a certain segment of the general population, namely, those demonstrating atopy.

Acute high-level exposure to WTC particles alters expression of genes associated with oxidative stress and immune function in the lung.

Abstract First responders (FR) present at Ground Zero in the first 72 h after the World Trade Center (WTC) collapsed have progressively exhibited significant respiratory injuries. The few toxicology studies performed to date evaluated effects from just fine (< 2.5 µm) WTC dusts; none examined health effects/toxicities from atmospheres bearing larger particle sizes, despite the fact the majority (> 96%) of dusts were > 10 µm and most FR likely entrained dusts by mouth breathing. Using a system that generated/delivered supercoarse (10–53 µm) WTC dusts to F344 rats (in a manner that mimicked FR exposures), this study sought to examine potential toxicities in the lungs. In this exploratory study, rats were exposed for 2 h to 100 mg WTC dust/m3 (while under isoflurane [ISO] anesthesia) or an air/ISO mixture; this dose conservatively modeled likely exposures by mouth-breathing FR facing ≈750–1000 mg WTC dust/m3. Lungs were harvested 2 h post-exposure and total RNA extracted for subsequent global gene expression analysis. Among the >  1000 genes affected by WTC dust (under ISO) or ISO alone, 166 were unique to the dust exposure. In many instances, genes maximally-induced by the WTC dust exposure (relative to in naïve rats) were unchanged/inhibited by ISO only; similarly, several genes maximally inhibited in WTC dust rats were largely induced/unchanged in rats that received ISO only. These outcomes reflect likely contrasting effects of ISO and the WTC dust on lung gene expression. Overall, the data show that lungs of rats exposed to WTC dust (under ISO) – after accounting for any impact from ISO alone – displayed increased expression of genes related to lung inflammation, oxidative stress, and cell cycle control, while several involved in anti-oxidant function were inhibited. These changes suggested acute inflammogenic effects and oxidative stress in the lungs of WTC dust-exposed rats. This study, thus, concludes that a single very high exposure to WTC dusts could potentially have adversely affected the respiratory system – in terms of early inflammatory and oxidative stress processes. As these changes were not compared with other types of dusts, the uniqueness of these WTC-mediated effects remains to be confirmed. It also still remains to be determined if these effects might have any relevance to chronic lung pathologies that became evident among FR who encountered the highest dust levels on September 11, 2001 and the 2 days thereafter. Ongoing studies using longer-range post-exposure analyses (up to 1-year or more) will help to determine if effects seen here on genes were acute, reversible, or persistent, and associated with corresponding histopathologic/biochemical changes in situ.

A novel system to generate WTC dust particles for inhalation exposures

First responders (FRs) present at Ground Zero within the critical first 72 h after the World Trade Center (WTC) collapse have progressively exhibited significant respiratory injury. The majority (>96%) of WTC dusts were >10 μm and no studies have examined potential health effects of this size fraction. This study sought to develop a system to generate and deliver supercoarse (10–53 μm) WTC particles to a rat model in a manner that mimicked FR exposure scenarios. A modified Fishing Line generator was integrated onto an intratracheal inhalation (ITIH) system that allowed for a bypassing of the nasal passages so as to mimic FR exposures. Dust concentrations were measured gravimetrically; particle size distribution was measured via elutriation. Results indicate that the system could produce dusts with 23 μm mass median aerodynamic diameter (MMAD) at levels up to ≥1200 mg/m3. To validate system utility, F344 rats were exposed for 2 h to ≈100 mg WTC dust/m3. Exposed rats had significantly increased lung weight and levels of select tracer metals 1 h after exposure. Using this system, it is now possible to conduct relevant inhalation exposures to determine adverse WTC dusts impacts on the respiratory system. Furthermore, this novel integrated Fishing Line–ITIH system could potentially be used in the analyses of a wide spectrum of other dusts/pollutants of sizes previously untested or delivered to the lungs in ways that did not reflect realistic exposure scenarios.