Hepatic Transcriptomic Patterns in the Neonatal Rat After Pentabromodiphenyl Ether Exposure.

Human exposure to pentabromodiphenyl ether (PBDE) mixture (DE-71) and its PBDE-47 congener can occur both in utero and during lactation. Here, we tested the hypothesis that PBDE-induced neonatal hepatic transcriptomic alterations in Wistar Han rat pups can inform on potential toxicity and carcinogenicity after longer term PBDE exposures. Wistar Han rat dams were exposed to either DE-71 or PBDE-47 daily from gestation day (GD 6) through postnatal day 4 (PND 4). Total plasma thyroxine (T4) was decreased in PND 4 pups. In liver, transcripts for CYPs and conjugation enzymes, Nrf2, and ABC transporters were upregulated. In general, the hepatic transcriptomic alterations after exposure to DE-71 or PBDE-47 were similar and provided early indicators of oxidative stress and metabolic alterations, key characteristics of toxicity processes. The transcriptional benchmark dose lower confidence limits of the most sensitive biological processes were lower for PBDE-47 than for the PBDE mixture. Neonatal rat liver transcriptomic data provide early indicators on molecular pathway alterations that may lead to toxicity and/or carcinogenicity if the exposures continue for longer durations. These early toxicogenomic indicators may be used to help prioritize chemicals for a more complete toxicity and cancer risk evaluation.

Sulforaphane enriched transcriptome of lung mitochondrial energy metabolism and provided pulmonary injury protection via Nrf2 in mice.

Nrf2 is essential to antioxidant response element (ARE)-mediated host defense. Sulforaphane (SFN) is a phytochemical antioxidant known to affect multiple cellular targets including Nrf2-ARE pathway in chemoprevention. However, the role of SFN in non-malignant airway disorders remain unclear. To test if pre-activation of Nrf2-ARE signaling protects lungs from oxidant-induced acute injury, wild-type (Nrf2+/+) and Nrf2-deficient (Nrf2-/-) mice were given SFN orally or as standardized broccoli sprout extract diet (SBE) before hyperoxia or air exposure. Hyperoxia-induced pulmonary injury and oxidation indices were significantly reduced by SFN or SBE in Nrf2+/+ mice but not in Nrf2-/- mice. SFN upregulated a large cluster of basal lung genes that are involved in mitochondrial oxidative phosphorylation, energy metabolism, and cardiovascular protection only in Nrf2+/+ mice. Bioinformatic analysis elucidated ARE-like motifs on these genes. Transcript abundance of the mitochondrial machinery genes remained significantly higher after hyperoxia exposure in SFN-treated Nrf2+/+ mice than in SFN-treated Nrf2-/- mice. Nuclear factor-κB was suggested to be a central molecule in transcriptome networks affected by SFN. Minor improvement of hyperoxia-caused lung histopathology and neutrophilia by SFN in Nrf2-/- mice implies Nrf2-independent or alternate effector mechanisms. In conclusion, SFN is suggested to be as a preventive intervention in a preclinical model of acute lung injury by linking mitochondria and Nrf2. Administration of SFN alleviated acute lung injury-like pathogenesis in a Nrf2-dependent manner. Potential AREs in the SFN-inducible transcriptome for mitochondria bioenergetics provided a new insight into the downstream mechanisms of Nrf2-mediated pulmonary protection.

The Diacetyl-Exposed Human Airway Epithelial Secretome: New Insights into Flavoring-Induced Airways Disease.

Bronchiolitis obliterans (BO) is an increasingly important lung disease characterized by fibroproliferative airway lesions and decrements in lung function. Occupational exposure to the artificial food flavoring ingredient diacetyl, commonly used to impart a buttery flavor to microwave popcorn, has been associated with BO development. In the occupational setting, diacetyl vapor is first encountered by the airway epithelium. To better understand the effects of diacetyl vapor on the airway epithelium, we used an unbiased proteomic approach to characterize both the apical and basolateral secretomes of air-liquid interface cultures of primary human airway epithelial cells from four unique donors after exposure to an occupationally relevant concentration (∼1,100 ppm) of diacetyl vapor or phosphate-buffered saline as a control on alternating days. Basolateral and apical supernatants collected 48 h after the third exposure were analyzed using one-dimensional liquid chromatography tandem mass spectrometry. Paired t tests adjusted for multiple comparisons were used to assess differential expression between diacetyl and phosphate-buffered saline exposure. Of the significantly differentially expressed proteins identified, 61 were unique to the apical secretome, 81 were unique to the basolateral secretome, and 11 were present in both. Pathway enrichment analysis using publicly available databases revealed that proteins associated with matrix remodeling, including degradation, assembly, and new matrix organization, were overrepresented in the data sets. Similarly, protein modifiers of epidermal growth factor receptor signaling were significantly altered. The ordered changes in protein expression suggest that the airway epithelial response to diacetyl may contribute to BO pathogenesis.

Proteomic Analysis of Primary Human Airway Epithelial Cells Exposed to the Respiratory Toxicant Diacetyl.

Occupational exposures to the diketone flavoring agent, diacetyl, have been associated with bronchiolitis obliterans, a rare condition of airway fibrosis. Model studies in rodents have suggested that the airway epithelium is a major site of diacetyl toxicity, but the effects of diacetyl exposure upon the human airway epithelium are poorly characterized. Here we performed quantitative LC-MS/MS-based proteomics to study the effects of repeated diacetyl vapor exposures on 3D organotypic cultures of human primary tracheobronchial epithelial cells. Using a label-free approach, we quantified approximately 3400 proteins and 5700 phosphopeptides in cell lysates across four independent donors. Altered expression of proteins and phosphopeptides were suggestive of loss of cilia and increased squamous differentiation in diacetyl-exposed cells. These phenomena were confirmed by immunofluorescence staining of culture cross sections. Hyperphosphorylation and cross-linking of basal cell keratins were also observed in diacetyl-treated cells, and we used parallel reaction monitoring to confidently localize and quantify previously uncharacterized sites of phosphorylation in keratin 6. Collectively, these data identify numerous molecular changes in the epithelium that may be important to the pathogenesis of flavoring-induced bronchiolitis obliterans. More generally, this study highlights the utility of quantitative proteomics for the study of in vitro models of airway injury and disease.

Hepatic transcriptomic alterations for N,N-dimethyl-p-toluidine (DMPT) and p-toluidine after 5-day exposure in rats.

N,N-dimethyl-p-toluidine (DMPT), an accelerant for methyl methacrylate monomers in medical devices, was a liver carcinogen in male and female F344/N rats and B6C3F1 mice in a 2-year oral exposure study. p-Toluidine, a structurally related chemical, was a liver carcinogen in mice but not in rats in an 18-month feed exposure study. In this current study, liver transcriptomic data were used to characterize mechanisms in DMPT and p-toluidine liver toxicity and for conducting benchmark dose (BMD) analysis. Male F344/N rats were exposed orally to DMPT or p-toluidine (0, 1, 6, 20, 60 or 120 mg/kg/day) for 5 days. The liver was examined for lesions and transcriptomic alterations. Both chemicals caused mild hepatic toxicity at 60 and 120 mg/kg and dose-related transcriptomic alterations in the liver. There were 511 liver transcripts differentially expressed for DMPT and 354 for p-toluidine at 120 mg/kg/day (false discovery rate threshold of 5 %). The liver transcriptomic alterations were characteristic of an anti-oxidative damage response (activation of the Nrf2 pathway) and hepatic toxicity. The top cellular processes in gene ontology (GO) categories altered in livers exposed to DMPT or p-toluidine were used for BMD calculations. The lower confidence bound benchmark doses for these chemicals were 2 mg/kg/day for DMPT and 7 mg/kg/day for p-toluidine. These studies show the promise of using 5-day target organ transcriptomic data to identify chemical-induced molecular changes that can serve as markers for preliminary toxicity risk assessment.

Airway injury in an in vitro human epithelium-fibroblast model of diacetyl vapor exposure: diacetyl-induced basal/suprabasal spongiosis.

Inhalation exposure to diacetyl (DA) is associated with obliterative bronchiolitis (OB) in workers and induces OB-like fibrotic airway lesions in rats. The pathogenesis of OB is poorly understood in part due to complex interactions between airway epithelial, mesenchymal and blood-derived inflammatory cells. DA-induced airway toxicity in the absence of recruited-inflammatory/immune cells was characterized using an air-liquid interface (ALI) model consisting of human airway epithelium with (Epi/FT) and without (Epi) a mesenchymal component. ALI cultures were exposed to 25 mM DA-derived vapors (using vapor cups) for 1 h on day 0, 2 and 4. In some experiments, the tissues were exposed to 2,3-hexanedione (Hex) which is structurally-similar, but much less fibrogenic than DA. Lactate dehydrogenase activity and day 6 histopathologic changes associated with epithelial injury, including basal/suprabasal spongiosis, were increased following exposure of Epi/FT tissues to DA but not control or Hex vapors. IL-1a, IL-6, IL-8, sIL-1Ra, TGFa, MCP-3 and TNFa proteins were increased following DA exposure of Epi/FT tissues; only IL-1a, IL-8, sIL-1Ra and TGFa were increased following exposure of Epi tissues. MMP-1, MMP-3 and TIMP-1 proteins were increased following DA exposure of Epi/FT tissues; whereas MMP-2, MMP-7 and TIMP-2 were decreased, and production was largely dependent upon the presence of sub-epithelial stromal matrix/fibroblasts. Hex-induced protein changes were minimal. This in vitro study demonstrated that exposure of human airways to DA vapors induced epithelial injury (with the histopathologic feature of basal/suprabasal spongiosis) and increased release of pro-inflammatory and pro-fibrotic cytokines/chemokines as well as MMPs/TIMPs in the absence of recruited-inflammatory cells.

Comparative inhalation toxicity of ethyltoluene isomers in rats and mice.

The C9 alkylbenzenes, composed mostly of ethyltoluenes and trimethylbenzenes, comprise 75-90% of the naphtha fraction of crude oil. Occupational and environmental exposure to C9 alkylbenzenes occur via inhalation. We conducted short-term inhalation studies on the ethyltoluene isomers (2-, 3- or 4-) to select one isomer for more comprehensive studies. Male Hsd:Sprague Dawley rats and female B6C3F1/N mice (n = 10) were exposed by nose-only inhalation to 2-, 3- or 4-ethyltoluene (0, 1000 or 2000 ppm) or cumene (a reference compound: 0, 500 or 1000 ppm) 3 h/day, 5 days/week, for 2 weeks. Clinical observations included abnormal gait and delayed righting reflex. Rats and mice exposed to 2000 ppm 2-ethyltoluene and mice exposed to 2000 ppm 4-ethyltoluene were euthanized early in moribund condition; no exposure-related deaths were observed with 3-ethyltoluene or cumene. Histopathology of selected tissues revealed that the nose and liver (rats and mice) and lung (mice only) to be toxicity targets. In the mouse lung, all compounds except 4-ethyltoluene produced bronchial and bronchiolar hyperplasia. In rats and mice, 2-ethyltoluene was the only compound to produce lesions in the nose and liver: in mice, squamous metaplasia and neutrophilic inflammation of the respiratory epithelium and atrophy and degeneration of the olfactory epithelium were observed in the nose and centrilobular hypertrophy and necrosis were observed in the liver. In rats, 2-ethyltoluene exposure produced atrophy of the olfactory epithelium in the nose and centrilobular necrosis in the liver. Based on mortality, body weight effects and histopathology, the 2-ethyltoluene isomer was the most potent isomer.

Comparative pulmonary toxicity of inhaled metalworking fluids in rats and mice.

Metalworking fluids (MWFs) are complex formulations designed for effective lubricating, cooling, and cleaning tools and parts during machining operations. Adverse health effects such as respiratory symptoms, dermatitis, and cancer have been reported in workers exposed to MWFs. Several constituents of MWFs have been implicated in toxicity and have been removed from the formulations over the years. However, animal studies with newer MWFs demonstrate that they continue to pose a health risk. This investigation examines the hypothesis that unrecognized health hazards exist in currently marketed MWF formulations that are presumed to be safe based on hazard assessments of individual ingredients. In vivo 13-week inhalation studies were designed to characterize and compare the potential toxicity of four MWFs: Trim VX, Cimstar 3800, Trim SC210, and Syntilo 1023. Male and female Wistar Han rats or Fischer 344N/Tac rats and B6C3F1/N mice were exposed to MWFs via whole-body inhalation at concentrations of 0, 25, 50, 100, 200, or 400 mg/m3 for 13 weeks, after which, survival, body and organ weights, hematology and clinical chemistry, histopathology, and genotoxicity were assessed following exposure. Although high concentrations were used, survival was not affected and toxicity was primarily within the respiratory tract of male and female rats and mice. Minor variances in toxicity were attributed to differences among species as well as in the chemical components of each MWF. Pulmonary fibrosis was present only in rats and mice exposed to Trim VX. These data confirm that newer MWFs have the potential to cause respiratory toxicity in workers who are repeatedly exposed via inhalation.

Impact of Environmental Enrichment Devices on NTP In Vivo Studies.

The goal of this study was to determine whether the use of nesting material or polycarbonate shelters as enrichment devices would have an impact on end points commonly measured during the conduct of the National Toxicology Program (NTP) 13-week studies. The study design was consistent with the NTP 13-week toxicity studies. Harlan Sprague-Dawley (HSD) rats and their offspring and B6C3F1/N mice were assigned to control (unenriched) and enriched experimental groups. Body weight, food and water consumption, behavioral observations, fecal content, clinical pathology, gross pathology, organ weights, and histopathology were evaluated. Enriched male mice and male and female rats exhibited decreased feed intake without a subsequent decrease in body weight; this may have been the result of the nesting material reducing the effect of cold stress, thereby allowing for more efficient use of feed. There were statistical differences in some hematological parameters; however, these were not considered physiologically relevant since all values were within the normal range. Gross pathology and histopathological findings were background changes and were not considered enrichment-related. Nesting material and shelters were used frequently and consistently and allowed animals to display species-typical behavior. There was no significant impact on commonly measured end points in HSD rats and B6C3F1/N mice given enrichment devices.

Chemical Reactivity and Respiratory Toxicity of the α-Diketone Flavoring Agents: 2,3-Butanedione, 2,3-Pentanedione, and 2,3-Hexanedione.

Occupational exposure to 2,3-butanedione (BD) vapors has been associated with severe respiratory disease leading to the use of potentially toxic substitutes. We compared the reactivity and respiratory toxicity of BD with that of two structurally related substitutes, 2,3-pentanedione (PD) and 2,3-hexanedione (HD). Chemical reactivity of the diketones with an arginine substrate decreased with increasing chain length (BD > PD > HD). Animals were evaluated the morning after a 2-week exposure to 0, 100, 150, or 200 ppm BD, PD, or HD (postexposure) or 2 weeks later (recovery). Bronchial fibrosis was observed in 5/5 BD and 5/5 PD rats at 200 ppm and in 4/6 BD and 6/6 PD rats at 150 ppm in the postexposure groups. Following recovery, bronchial fibrosis was observed in all surviving rats exposed to 200 ppm BD (5/5) or PD (3/3) and in 2/10 BD and 7/9 PD rats exposed to 150 ppm. Bronchial fibrosis was observed only in 2/12 HD-exposed rats in the 200 ppm postexposure group. Patchy interstitial fibrosis affected lungs of recovery groups exposed to 200 ppm PD (3/3) or BD (1/5) and to 150 ppm PD (4/9) or BD (7/10) and correlated with pulmonary function deficits. BD and PD were more reactive and produced more bronchial fibrosis than HD.

Molecular Changes in the Nasal Cavity after N, N-dimethyl-p-toluidine Exposure.

N, N-dimethyl-p-toluidine (DMPT; Cas No. 99-97-8), an accelerant for methyl methacrylate monomers in medical devices, is a nasal cavity carcinogen according to a 2-yr cancer study of male and female F344/N rats, with the nasal tumors arising from the transitional cell epithelium. In this study, we exposed male F344/N rats for 5 days to DMPT (0, 1, 6, 20, 60, or 120 mg/kg [oral gavage]) to explore the early changes in the nasal cavity after short-term exposure. Lesions occurred in the nasal cavity including hyperplasia of transitional cell epithelium (60 and 120 mg/kg). Nasal tissue was rapidly removed and preserved for subsequent laser capture microdissection and isolation of the transitional cell epithelium (0 and 120 mg/kg) for transcriptomic studies. DMPT transitional cell epithelium gene transcript patterns were characteristic of an antioxidative damage response (e.g., Akr7a3, Maff, and Mgst3), cell proliferation, and decrease in signals for apoptosis. The transcripts of amino acid transporters were upregulated (e.g., Slc7a11). The DMPT nasal transcript expression pattern was similar to that found in the rat nasal cavity after formaldehyde exposure, with over 1,000 transcripts in common. Molecular changes in the nasal cavity after DMPT exposure suggest that oxidative damage is a mechanism of the DMPT toxic and/or carcinogenic effects.

Diacetyl induces amphiregulin shedding in pulmonary epithelial cells and in experimental bronchiolitis obliterans.

Diacetyl (DA), a component of artificial butter flavoring, has been linked to the development of bronchiolitis obliterans (BO), a disease of airway epithelial injury and airway fibrosis. The epidermal growth factor receptor ligand, amphiregulin (AREG), has been implicated in other types of epithelial injury and lung fibrosis. We investigated the effects of DA directly on the pulmonary epithelium, and we hypothesized that DA exposure would result in epithelial cell shedding of AREG. Consistent with this hypothesis, we demonstrate that DA increases AREG by the pulmonary epithelial cell line NCI-H292 and by multiple independent primary human airway epithelial donors grown under physiologically relevant conditions at the air-liquid interface. Furthermore, we demonstrate that AREG shedding occurs through a TNF-α-converting enzyme (TACE)-dependent mechanism via inhibition of TACE activity in epithelial cells using the small molecule inhibitor, TNF-α protease inhibitor-1, as well as TACE-specific small inhibitor RNA. Finally, we demonstrate supportive in vivo results showing increased AREG transcript and protein levels in the lungs of rodents with DA-induced BO. In summary, our novel in vitro and in vivo observations suggest that further study of AREG is warranted in the pathogenesis of DA-induced BO.

Severely blunted allergen-induced pulmonary Th2 cell response and lung hyperresponsiveness in type 1 transient receptor potential channel-deficient mice.

Transient receptor potential channels (TRPCs) are widely expressed and regulate Ca²âº entry in the cells that participate in the pathophysiology of airway hyperreactivity, inflammation, and remodeling. In vitro studies point to a role for TRPC1-mediated Ca²âº signaling in several of these cell types; however, physiological evidence is lacking. Here we identify TRPC1 signaling as proinflammatory and a regulator of lung hyperresponsiveness during allergen-induced pulmonary response. TRPC1-deficient (Trpc1(-/-)) mice are hyposensitive to methacholine challenge and have significantly reduced allergen-induced pulmonary leukocyte infiltration coupled with an attenuated T helper type 2 (Th2) cell response. Upon in vitro allergen exposure, Trpc1(-/-) splenocytes show impaired proliferation and T cell receptor-induced IL-2 production. A high number of germinal centers in spleens of Trpc1(-/-) mice and elevated levels of immunoglobulins in their serum are indicative of dysregulated B cell function and homeostasis. Thus we propose that TRPC1 signaling is necessary in lymphocyte biology and in regulation of allergen-induced lung hyperresponsiveness, making TRPC1 a potential target for treatment of immune diseases and asthma.

Bronchial and bronchiolar fibrosis in rats exposed to 2,3-pentanedione vapors: implications for bronchiolitis obliterans in humans.

2,3-Pentanedione (PD) is a component of artificial butter flavorings. The use of PD is increasing since diacetyl, a major butter flavorant, was associated with bronchiolitis obliterans (BO) in workers and has been removed from many products. Because the toxicity of inhaled PD is unknown, these studies were conducted to characterize the toxicity of inhaled PD across a range of concentrations in rodents. Male and female Wistar-Han rats and B6C3F1 mice were exposed to 0, 50, 100, or 200 ppm PD 6 h/d, 5 d/wk for up to 2 wk. Bronchoalveolar lavage fluid (BALF) was collected after 1, 3, 5, and 10 exposures, and histopathology was evaluated after 12 exposures. MCP-1, MCP-3, CRP, FGF-9, fibrinogen, and OSM were increased 2- to 9-fold in BALF of rats exposed for 5 and 10 days to 200 ppm. In mice, only fibrinogen was increased after 5 exposures to 200 ppm. The epithelium lining the respiratory tract was the site of toxicity in all mice and rats exposed to 200 ppm. Significantly, PD also caused both intraluminal and intramural fibrotic airway lesions in rats. The histopathological and biological changes observed in rats raise concerns that PD inhalation may cause BO in exposed humans.

Prostaglandin E2 protects murine lungs from bleomycin-induced pulmonary fibrosis and lung dysfunction.

Prostaglandin E(2) (PGE(2)) is a lipid mediator that is produced via the metabolism of arachidonic acid by cyclooxygenase enzymes. In the lung, PGE(2) acts as an anti-inflammatory factor and plays an important role in tissue repair processes. Although several studies have examined the role of PGE(2) in the pathogenesis of pulmonary fibrosis in rodents, results have generally been conflicting, and few studies have examined the therapeutic effects of PGE(2) on the accompanying lung dysfunction. In this study, an established model of pulmonary fibrosis was used in which 10-12-wk-old male C57BL/6 mice were administered a single dose (1.0 mg/kg) of bleomycin via oropharyngeal aspiration. To test the role of prostaglandins in this model, mice were dosed, via surgically implanted minipumps, with either vehicle, PGE(2) (1.32 µg/h), or the prostacyclin analog iloprost (0.33 µg/h) beginning 7 days before or 14 days after bleomycin administration. Endpoints assessed at 7 days after bleomycin administration included proinflammatory cytokine levels and measurement of cellular infiltration into the lung. Endpoints assessed at 21 days after bleomycin administration included lung function assessment via invasive (FlexiVent) analysis, cellular infiltration, lung collagen content, and semiquantitative histological analysis of the degree of lung fibrosis (Ashcroft method). Seven days after bleomycin administration, lymphocyte numbers and chemokine C-C motif ligand 2 expression were significantly lower in PGE(2)- and iloprost-treated animals compared with vehicle-treated controls (P < 0.05). When administered 7 days before bleomycin challenge, PGE(2) also protected against the decline in lung static compliance, lung fibrosis, and collagen production that is associated with 3 wk of bleomycin exposure. However, PGE(2) had no therapeutic effect on these parameters when administered 14 days after bleomycin challenge. In summary, PGE(2) prevented the decline in lung static compliance and protected against lung fibrosis when it was administered before bleomycin challenge but had no therapeutic effect when administered after bleomycin challenge.

Multisite carcinogenicity and respiratory toxicity of inhaled 1-bromopropane in rats and mice.

Two-year 1-bromopropane (1-BP) inhalation studies were conducted because of the potential for widespread exposure, the lack of chronic toxicity and carcinogenicity data, and the known carcinogenicity of structurally related compounds. Male and female F344/N rats and B6C3F1/N mice were exposed by inhalation to 0, 62.5 (mice only), 125, 250, or 500 (rats only) ppm 1-BP for 6 hr/day, 5 days/week for 105 weeks. Exposure of male and female rats to 1-BP resulted in significantly increased incidences of adenomas of the large intestine and skin neoplasms. In male rats, the incidence of malignant mesothelioma of the epididymis was statistically significantly increased at 500 ppm, but the biological significance of this common lesion is unclear. Incidences of pancreatic islet adenoma in male rats were significantly increased at all concentrations relative to concurrent controls but were within the historical control range for inhalation studies. There was no evidence of carcinogenic activity of 1-BP in male B6C3F1 mice; however, significantly increased incidences of alveolar/bronchiolar neoplasms of the lung were present in female mice. Exposure to 1-BP also resulted in increased incidences of nonneoplastic lesions in the nose of rats and mice, the larynx of rats and male mice, the trachea of female rats and male and female mice, and the lungs of mice. Inflammatory lesions with Splendore Hoeppli (S-H) material were present primarily in the nose and skin of exposed male and female rats, indicating that 1-BP caused immunosuppression.

Modulation of allergic airway inflammation by the oral pathogen Porphyromonas gingivalis.

Accumulating evidence suggests that bacteria associated with periodontal disease may exert systemic immunomodulatory effects. Although the improvement in oral hygiene practices in recent decades correlates with the increased incidence of asthma in developed nations, it is not known whether diseases of the respiratory system might be influenced by the presence of oral pathogens. The present study sought to determine whether subcutaneous infection with the anaerobic oral pathogen Porphyromonas gingivalis exerts a regulatory effect on allergic airway inflammation. BALB/c mice sensitized and subsequently challenged with ovalbumin exhibited airway hyperresponsiveness to methacholine aerosol and increased airway inflammatory cell influx and Th2 cytokine (interleukin-4 [IL-4], IL-5, and IL-13) content relative to those in nonallergic controls. Airway inflammatory cell and cytokine contents were significantly reduced by establishment of a subcutaneous infection with P. gingivalis prior to allergen sensitization, whereas serum levels of ovalbumin-specific IgE and airway responsiveness were not altered. Conversely, subcutaneous infection initiated after allergen sensitization did not alter inflammatory end points but did reduce airway responsiveness in spite of increased serum IgE levels. These data provide the first direct evidence of a regulatory effect of an oral pathogen on allergic airway inflammation and responsiveness. Furthermore, a temporal importance of the establishment of infection relative to allergen sensitization is demonstrated for allergic outcomes.

Whole-body inhalation exposure to 1-bromopropane suppresses the IgM response to sheep red blood cells in female B6C3F1 mice and Fisher 344/N rats.

1-Bromopropane (1-BP) is categorized as a high-production-volume chemical and is currently used in the manufacture of pharmaceuticals, pesticides, and other chemicals. Its usage is estimated to be around 5 million pounds per year, resulting in the potential for widespread exposure in the workplace. Case reports and animal studies have suggested exposure to this compound may cause adverse reproductive and neurological effects. Using a battery of immunological assays, the immunotoxicity of 1-BP after whole body inhalation exposure in both mice and rats was evaluated. Significant decreases in the spleen immunoglobulin (Ig) M response to sheep red blood cells (SRBC) were observed in both mice (125-500 ppm) and rats (1000 ppm) after exposure to 1-BP for 10 wk. In addition, total spleen cells and T cells were significantly decreased after approximately 4 wk of 1-BP exposure in both mice (125-500 ppm) and rats (1000 ppm). No change in natural killer (NK) cell activity was observed. The observed alterations in spleen cellularity, phenotypic subsets, and impairment of humoral immune function across species raise further concern about human exposure to 1-BP and demonstrate the need for additional investigations into potential adverse health effects.

Gene expression studies reveal that DNA damage, vascular perturbation, and inflammation contribute to the pathogenesis of carbonyl sulfide neurotoxicity.

Carbonyl sulfide (COS) is an odorless gas that produces highly reproducible lesions in the central nervous system. In the present study, the time course for the development of the neurotoxicological lesions was defined and the gene expression changes occurring in the posterior colliculus upon exposure to COS were characterized. Fischer 344 rats were exposed to 0 or 500 ppm COS for one, two, three, four, five, eight, or ten days, six hours per day. On days 1 and 2, no morphological changes were detected; on day 3, 10/10 (100%) rats had necrosis in the posterior colliculi; and on day 4 and later, necrosis was observed in numerous areas of the brain. Important gene expression changes occurring in the posterior colliculi after one or two days of COS exposure that were predictive of the subsequent morphological findings included up-regulation of genes associated with DNA damage and G1/S checkpoint regulation (KLF4, BTG2, GADD45g), apoptosis (TGM2, GADD45g, RIPK3), and vascular mediators (ADAMTS, CTGF, CYR61, VEGFC). Proinflammatory mediators (CCL2, CEBPD) were up-regulated prior to increases in expression of the astrocytic marker GFAP and macrophage marker CSF2rb1. These gene expression findings were predictive of later CNS lesions caused by COS exposure and serve as a model for future investigations into the mechanisms of disease in the central nervous system.