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1.
Toxicol Appl Pharmacol ; 409: 115329, 2020 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-33181145

RESUMO

Hydraulic fracturing ("fracking") is a process in which subterranean natural gas-laden rock is fractured under pressure to enhance retrieval of gas. Sand (a "proppant") is present in the fracking fluid pumped down the well bore to stabilize the fissures and facilitate gas flow. The manipulation of sand at the well site creates respirable dust (fracking sand dust, FSD) to which workers are exposed. Because workplace exposures to FSD have exceeded exposure limits set by OSHA, a physico-chemical characterization of FSD along with comprehensive investigations of the potential early adverse effects of FSDs on organ function and biomarkers has been conducted using a rat model and related in vivo and in vitro experiments involving the respiratory, cardiovascular, immune systems, kidney and brain. An undercurrent theme of the overall hazard identification study was, to what degree do the health effects of inhaled FSD resemble those previously observed after crystalline silica dust inhalation? In short-term studies, FSD was found to be less bioactive than MIN-U-SIL® 5 in the lungs. A second theme was, are the biological effects of FSD restricted to the lungs? Bioactivity of FSD was observed in all examined organ systems. Our findings indicate that, in many respects, the physical and chemical properties, and the short-term biological effects, of the FSDs share many similarities as a group but have little in common with crystalline silica dust.

2.
Toxicol Appl Pharmacol ; 409: 115300, 2020 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-33141058

RESUMO

Hydraulic fracturing (fracking) is a process used to recover oil and gas from shale rock formation during unconventional drilling. Pressurized liquids containing water and sand (proppant) are used to fracture the oil- and natural gas-laden rock. The transportation and handling of proppant at well sites generate dust aerosols; thus, there is concern of worker exposure to such fracking sand dusts (FSD) by inhalation. FSD are generally composed of respirable crystalline silica and other minerals native to the geological source of the proppant material. Field investigations by NIOSH suggest that the levels of respirable crystalline silica at well sites can exceed the permissible exposure limits. Thus, from an occupational safety perspective, it is important to evaluate the potential toxicological effects of FSD, including any neurological risks. Here, we report that acute inhalation exposure of rats to one FSD, i.e., FSD 8, elicited neuroinflammation, altered the expression of blood brain barrier-related markers, and caused glial changes in the olfactory bulb, hippocampus and cerebellum. An intriguing observation was the persistent reduction of synaptophysin 1 and synaptotagmin 1 proteins in the cerebellum, indicative of synaptic disruption and/or injury. While our initial hazard identification studies suggest a likely neural risk, more research is necessary to determine if such molecular aberrations will progressively culminate in neuropathology/neurodegeneration leading to behavioral and/or functional deficits.

3.
Toxicol Appl Pharmacol ; 408: 115256, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-33007384

RESUMO

Hydraulic fracturing ("fracking") is a process used to enhance retrieval of gas from subterranean natural gas-laden rock by fracturing it under pressure. Sand used to stabilize fissures and facilitate gas flow creates a potential occupational hazard from respirable fracking sand dust (FSD). As studies of the immunotoxicity of FSD are lacking, the effects of whole-body inhalation (6 h/d for 4 d) of a FSD, i.e., FSD 8, was investigated at 1, 7, and 27 d post-exposure in rats. Exposure to 10 mg/m3 FSD 8 resulted in decreased lung-associated lymph node (LLN) cellularity, total B-cells, CD4+ T-cells, CD8+ T-cells and total natural killer (NK) cells at 7-d post exposure. The frequency of CD4+ T-cells decreased while the frequency of B-cells increased (7 and 27 d) in the LLN. In contrast, increases in LLN cellularity and increases in total CD4+ and CD8+ T-cells were observed in rats following 30 mg/m3 FSD 8 at 1 d post-exposure. Increases in the frequency and number of CD4+ T-cells and NK cells were observed in bronchial alveolar lavage fluid at 7-d post-exposure (10 mg/m3) along with an increase in total CD4+ T-cells, CD11b + cells, and NK cells at 1-day post-exposure (30 mg/m3). Increases in the numbers of B-cells and CD8+ T-cells were observed in the spleen at 1-day post 30 mg/m3 FSD 8 exposure. In addition, NK cell activity was suppressed at 1 d (30 mg/m3) and 27 d post-exposure (10 mg/m3). No change in the IgM response to sheep red blood cells was observed. The findings indicate that FSD 8 caused alterations in cellularity, phenotypic subsets, and impairment of immune function.

4.
Toxicol Appl Pharmacol ; 408: 115280, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-33065154

RESUMO

The pulmonary inflammatory response to inhalation exposure to a fracking sand dust (FSD 8) was investigated in a rat model. Adult male Sprague-Dawley rats were exposed by whole-body inhalation to air or an aerosol of a FSD, i.e., FSD 8, at concentrations of 10 or 30 mg/m3, 6 h/d for 4 d. The control and FSD 8-exposed rats were euthanized at post-exposure time intervals of 1, 7 or 27 d and pulmonary inflammatory, cytotoxic and oxidant responses were determined. Deposition of FSD 8 particles was detected in the lungs of all the FSD 8-exposed rats. Analysis of bronchoalveolar lavage parameters of toxicity, oxidant generation, and inflammation did not reveal any significant persistent pulmonary toxicity in the FSD 8-exposed rats. Similarly, the lung histology of the FSD 8-exposed rats showed only minimal changes in influx of macrophages following the exposure. Determination of global gene expression profiles detected statistically significant differential expressions of only six and five genes in the 10 mg/m3, 1-d post-exposure, and the 30 mg/m3, 7-d post-exposure FSD 8 groups, respectively. Taken together, data obtained from the present study demonstrated that FSD 8 inhalation exposure resulted in no statistically significant toxicity or gene expression changes in the lungs of the rats. In the absence of any information about its potential toxicity, a comprehensive rat animal model study (see Fedan, J.S., Toxicol Appl Pharmacol. 000, 000-000, 2020) has been designed to investigate the bioactivities of several FSDs in comparison to MIN-U-SIL® 5, a respirable α-quartz reference dust used in previous animal models of silicosis, in several organ systems.

5.
Toxicol Appl Pharmacol ; 408: 115281, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-33065155

RESUMO

Cultured murine macrophages (RAW 264.7) were used to investigate the effects of fracking sand dust (FSD) for its pro-inflammatory activity, in order to gain insight into the potential toxicity to workers associated with inhalation of FSD during hydraulic fracturing. While the role of respirable crystalline silica in the development of silicosis is well documented, nothing is known about the toxicity of inhaled FSD. The FSD (FSD 8) used in these studies was from an unconventional gas well drilling site. FSD 8was prepared as a 10 mg/ml stock solution in sterile PBS, vortexed for 15 s, and allowed to sit at room temperature for 30 min before applying the suspension to RAW 264.7cells. Compared to PBS controls, cellular viability was significantly decreased after a 24 h exposure to FSD. Intracellular reactive oxygen species (ROS) production and the production of IL-6, TNFα, and endothelin-1 (ET-1) were up-regulated as a result of the exposure, whereas the hydroxyl radical (.OH) was only detected in an acellular system. Immunofluorescent staining of cells against TNFα revealed that FSD 8 caused cellular blebbing, and engulfment of FSD 8 by macrophages was observed with enhanced dark-field microscopy. The observed changes in cellular viability, cellular morphology, free radical generation and cytokine production all confirm that FSD 8 is cytotoxic to RAW 264.7 cells and warrants future studies into the specific pathways and mechanisms by which these toxicities occur.

6.
Toxicol Appl Pharmacol ; 409: 115284, 2020 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-33068619

RESUMO

Hydraulic fracturing creates fissures in subterranean rock to increase the flow and retrieval of natural gas. Sand ("proppant") in fracking fluid injected into the well bore maintains fissure patency. Fracking sand dust (FSD) is generated during manipulation of sand to prepare the fracking fluid. Containing respirable crystalline silica, FSD could pose hazards similar to those found in work sites where silica inhalation induces lung disease such as silicosis. This study was performed to evaluate the possible toxic effects following inhalation of a FSD (FSD 8) in the lung and airways. Rats were exposed (6 h/d × 4 d) to 10 or 30 mg/m3 of a FSD collected at a gas well, and measurements were performed 1, 7, 27 and, in one series of experiments, 90 d post-exposure. The following ventilatory and non-ventilatory parameters were measured in vivo and/or in vitro: 1) lung mechanics (respiratory system resistance and elastance, tissue damping, tissue elastance, Newtonian resistance and hysteresivity); 2) airway reactivity to inhaled methacholine (MCh); airway epithelium integrity (isolated, perfused trachea); airway efferent motor nerve activity (electric field stimulation in vitro); airway smooth muscle contractility; ion transport in intact and cultured epithelium; airway effector and sensory nerves; tracheal particle deposition; and neurogenic inflammation/vascular permeability. FSD 8 was without large effect on most parameters, and was not pro-inflammatory, as judged histologically and in cultured epithelial cells, but increased reactivity to inhaled MCh at some post-exposure time points and affected Na+ transport in airway epithelial cells.

7.
Toxicol Appl Pharmacol ; 409: 115282, 2020 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-33068622

RESUMO

Hydraulic fracturing ("fracking") is used in unconventional gas drilling to allow for the free flow of natural gas from rock. Sand in fracking fluid is pumped into the well bore under high pressure to enter and stabilize fissures in the rock. In the process of manipulating the sand on site, respirable dust (fracking sand dust, FSD) is generated. Inhalation of FSD is a potential hazard to workers inasmuch as respirable crystalline silica causes silicosis, and levels of FSD at drilling work sites have exceeded occupational exposure limits set by OSHA. In the absence of any information about its potential toxicity, a comprehensive rat animal model was designed to investigate the bioactivities of several FSDs in comparison to MIN-U-SIL® 5, a respirable α-quartz reference dust used in previous animal models of silicosis, in several organ systems (Fedan, J.S., Toxicol Appl Pharmacol. 00, 000-000, 2020). The present report, part of the larger investigation, describes: 1) a comparison of the physico-chemical properties of nine FSDs, collected at drilling sites, and MIN-U-SIL® 5, a reference silica dust, and 2) a comparison of the pulmonary inflammatory responses to intratracheal instillation of the nine FSDs and MIN-U-SIL® 5. Our findings indicate that, in many respects, the physico-chemical characteristics, and the biological effects of the FSDs and MIN-U-SIL® 5 after intratracheal instillation, have distinct differences.

8.
Toxicol Appl Pharmacol ; 406: 115242, 2020 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-32931794

RESUMO

Hydraulic fracturing is used to access oil and natural gas reserves. This process involves the high-pressure injection of fluid to fracture shale. Fracking fluid contains approximately 95% water, chemicals and 4.5% fracking sand. Workers may be exposed to fracking sand dust (FSD) during the manipulation of the sand, and negative health consequences could occur if FSD is inhaled. In the absence of any information about its potential toxicity, a comprehensive rat animal model study (see Fedan et al., 2020) was designed to investigate the bioactivities of several FSDs in comparison to MIN-U-SIL® 5, a respirable α-quartz reference dust used in previous animal models of silicosis, in several organ systems. The goal of this study was to assess the effects of inhalation of one FSD, i.e., FSD 8, on factors and tissues that affect cardiovascular function. Male rats were exposed to 10 or 30 mg/m3 FSD (6 h/d for 4 d) by whole body inhalation, with measurements made 1, 7 or 27 d post-exposure. One day following exposure to 10 mg/m3 FSD the sensitivity to phenylephrine-induced vasoconstriction in tail arteries in vitro was increased. FSD exposure at both doses resulted in decreases in heart rate (HR), HR variability, and blood pressure in vivo. FSD induced changes in hydrogen peroxide concentrations and transcript levels for pro-inflammatory factors in heart tissues. In kidney, expression of proteins indicative of injury and remodeling was reduced after FSD exposure. When analyzed using regression analysis, changes in proteins involved in repair and remodeling were correlated. Thus, it appears that inhalation of FSD does have some prolonged effects on cardiovascular, and, possibly, renal function. The findings also provide information regarding potential mechanisms that may lead to these changes, and biomarkers that could be examined to monitor physiological changes that could be indicative of impending cardiovascular dysfunction.

9.
Toxicol Pathol ; 47(8): 1012-1026, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31645208

RESUMO

Flavorings-related lung disease is a potentially disabling and sometimes fatal lung disease of workers making or using flavorings. First identified almost 20 years ago in microwave popcorn workers exposed to butter-flavoring vapors, flavorings-related lung disease remains a concern today. In some cases, workers develop bronchiolitis obliterans, a severe form of fixed airways disease. Affected workers have been reported in microwave popcorn, flavorings, and coffee production workplaces. Volatile α-dicarbonyl compounds, particularly diacetyl (2,3-butanedione) and 2,3-pentanedione, are implicated in the etiology. Published studies on diacetyl and 2,3-pentanedione document their ability to cause airway epithelial necrosis, damage biological molecules, and perturb protein homeostasis. With chronic exposure in rats, they produce airway fibrosis resembling bronchiolitis obliterans. To add to this knowledge, we recently evaluated airway toxicity of the 3-carbon α-dicarbonyl compound, methylglyoxal. Methylglyoxal inhalation causes epithelial necrosis at even lower concentrations than diacetyl. In addition, we investigated airway toxicity of mixtures of diacetyl, acetoin, and acetic acid, common volatiles in butter flavoring. At ratios comparable to workplace scenarios, the mixtures or diacetyl alone, but not acetic acid or acetoin, cause airway epithelial necrosis. These new findings add to existing data to implicate α-dicarbonyl compounds in airway injury and flavorings-related lung disease.


Assuntos
Poluentes Ocupacionais do Ar/toxicidade , Bronquiolite Obliterante/induzido quimicamente , Aromatizantes/toxicidade , Pneumopatias/induzido quimicamente , Doenças Profissionais/induzido quimicamente , Acetoína/toxicidade , Poluentes Ocupacionais do Ar/química , Bronquiolite Obliterante/patologia , Diacetil/toxicidade , Aromatizantes/química , Humanos , Exposição por Inalação/efeitos adversos , Pneumopatias/patologia , Doenças Profissionais/patologia , Exposição Ocupacional/efeitos adversos , Pentanonas/toxicidade
10.
J Hazard Mater ; 373: 630-639, 2019 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-30953980

RESUMO

Micronized copper azole (MCA) is a lumber treatment improve longevity. In this study, the in vivo response to PM2.5 sanding dust generated from MCA-treated lumber was compared to that of untreated yellow pine (UYP) or soluble copper azole-treated (CA-C) lumber to determine if the MCA was more bioactive than CA-C. Mice were exposed to doses (28, 140, or 280 µg/mouse) of UYP, MCA, or CA-C sanding dust using oropharyngeal aspiration. Bronchoalveolar lavage fluid (BALF) lactate dehydrogenase activity was increased at 1 day post-exposure to 280 µg/mouse of MCA and CA-C compared to UYP. BALF polymorphonuclear cells were increased by MCA and CA-C. There were increases in BALF cytokines in MCA and CA-C-exposed groups at 1 day post-exposure. Lung histopathology indicated inflammation with infiltration of neutrophils and macrophages. Pulmonary responses were more severe in MCA and CA-C-exposed groups at 1 day post-exposure. MCA caused more severe inflammatory responses than CA-C at 1 day post-exposure. These findings suggest that the MCA and CA-C sanding dusts are more bioactive than the UYP sanding dust, and, moreover, the MCA sanding dust is more bioactive in comparison to the CA-C sanding dust. No chronic toxic effects were observed among all observed sanding dusts.


Assuntos
Cobre/toxicidade , Exposição por Inalação/efeitos adversos , Material Particulado/toxicidade , Animais , Líquido da Lavagem Broncoalveolar/química , Líquido da Lavagem Broncoalveolar/imunologia , Cobre/análise , L-Lactato Desidrogenase/análise , Pulmão/patologia , Camundongos , Testes de Toxicidade , Madeira
11.
Toxicol Appl Pharmacol ; 364: 153-163, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30423287

RESUMO

Incorporation of multi-wall carbon nanotubes (MWCNT) into materials has raised concerns about their potential hazards to manufacturing workers. In animal models, airway inflammation and lung fibrosis follow aspiration, instillation, and inhalation exposures to MWCNT. However, the effects of MWCNT on pulmonary function, airway reactivity and airway epithelium function following inhalation exposure has not been studied. We investigated whether inhaled MWCNT affects lung resistance (RL) and dynamic compliance (Cdyn), reactivity to inhaled methacholine (MCh), epithelial regulation of airway reactivity to MCh in vitro, and airway epithelial ion transport. Male rats were exposed by whole body inhalation for 6 h to air or aerosolized MWCNT (0.5, 1 or 5 mg/m3) for one or nine days. Eighteen h after 1 d exposure to 5 mg/m3 MWCNT, basal RL was increased and basal Cdyn was decreased; changes did not persist for 7 d. Reactivity to MCh (RL) was increased and Cdyn responses were decreased at 18 h, but not 7 d after exposure to 1 and 5 mg/m3 MWCNT. The effects of i.t.-instilled MWCNT and nitrogen-doped MWCNT (N-MWCNT) on pulmonary function and reactivity to MCh at doses comparable to deposition after inhalation of 5 mg/m3 at 1 d and 0.5, 1, and 5 mg/m3 MWCNT 9 d-exposures were compared. Both nanoparticles increased airway reactivity (RL); N-MWCNT did not affect Cdyn responses. Lung function and airway reactivity are altered following a single MWCNT inhalation and generally subside over time. Given i.t., MWCNT's and N-MWCNT's effects were comparable, but N-MWCNT evoke smaller changes in Cdyn responses.


Assuntos
Hiper-Reatividade Brônquica/induzido quimicamente , Broncoconstrição/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Nanotubos de Carbono/toxicidade , Nitrogênio/toxicidade , Aerossóis , Resistência das Vias Respiratórias/efeitos dos fármacos , Animais , Hiper-Reatividade Brônquica/metabolismo , Hiper-Reatividade Brônquica/fisiopatologia , Testes de Provocação Brônquica , Broncoconstritores/administração & dosagem , Relação Dose-Resposta a Droga , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Exposição por Inalação , Transporte de Íons , Pulmão/metabolismo , Pulmão/fisiopatologia , Complacência Pulmonar/efeitos dos fármacos , Masculino , Cloreto de Metacolina/administração & dosagem , Nanotubos de Carbono/química , Nitrogênio/química , Permeabilidade , Ratos Sprague-Dawley , Medição de Risco , Fatores de Tempo
12.
Inhal Toxicol ; 29(7): 322-339, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28967277

RESUMO

The effects of acute pulmonary coexposures to silica and diesel particulate matter (DPM), which may occur in various mining operations, were investigated in vivo. Rats were exposed by intratracheal instillation (IT) to silica (50 or 233 µg), DPM (7.89 or 50 µg) or silica and DPM combined in phosphate-buffered saline (PBS) or to PBS alone (control). At one day, one week, one month, two months and three months postexposure bronchoalveolar lavage and histopathology were performed to assess lung injury, inflammation and immune response. While higher doses of silica caused inflammation and injury at all time points, DPM exposure alone did not. DPM (50 µg) combined with silica (233 µg) increased inflammation at one week and one-month postexposure and caused an increase in the incidence of fibrosis at one month compared with exposure to silica alone. To assess susceptibility to lung infection following coexposure, rats were exposed by IT to 233 µg silica, 50 µg DPM, a combination of the two or PBS control one week before intratracheal inoculation with 5 × 105 Listeria monocytogenes. At 1, 3, 5, 7 and 14 days following infection, pulmonary immune response and bacterial clearance from the lung were evaluated. Coexposure to DPM and silica did not alter bacterial clearance from the lung compared to control. Although DPM and silica coexposure did not alter pulmonary susceptibility to infection in this model, the study showed that noninflammatory doses of DPM had the capacity to increase silica-induced lung injury, inflammation and onset/incidence of fibrosis.


Assuntos
Poluentes Ocupacionais do Ar/toxicidade , Pulmão/efeitos dos fármacos , Material Particulado/toxicidade , Quartzo/toxicidade , Emissões de Veículos/toxicidade , Lesão Pulmonar Aguda/induzido quimicamente , Animais , Líquido da Lavagem Broncoalveolar/citologia , Líquido da Lavagem Broncoalveolar/imunologia , Contagem de Células , Citocinas/imunologia , L-Lactato Desidrogenase/metabolismo , Listeria monocytogenes/patogenicidade , Listeriose , Pulmão/imunologia , Pulmão/microbiologia , Pulmão/patologia , Masculino , Ratos Sprague-Dawley , Testes de Toxicidade Aguda
13.
Toxicol Appl Pharmacol ; 326: 1-6, 2017 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-28411035

RESUMO

Welding fume inhalation causes pulmonary toxicity, including susceptibility to infection. We hypothesized that airway epithelial ion transport is a target of fume toxicity, and investigated the effects of fume particulates from manual metal arc-stainless steel (MMA-SS) and gas metal arc-mild steel (GMA-MS) on ion transport in normal human bronchial epithelium (NHBE) cultured in air-interface. MMA-SS particles, more soluble than GMA-MS particles, contain Cr, Ni, Fe and Mn; GMA-MS particles contain Fe and Mn. MMA-SS or GMA-MS particles (0.0167-166.7µg/cm2) were applied apically to NHBEs. After 18h transepithelial potential difference (Vt), resistance (Rt), and short circuit current (Isc) were measured. Particle effects on Na+ and Cl¯ channels and the Na+,K+,2Cl¯-cotransporter were evaluated using amiloride (apical), 5-nitro-2-[(3-phenylpropyl)amino]benzoic acid (NPPB, apical), and bumetanide (basolateral), respectively. MMA-SS (0.0167-16.7µg/cm2) increased basal Vt. Only 16.7µg/cm2 GMA-MS increased basal Vt significantly. MMA-SS or GMA-MS exposure potentiated Isc responses (decreases) to amiloride and bumetanide, while not affecting those to NPPB, GMA-MS to a lesser degree than MMA-SS. Variable effects on Rt were observed in response to amiloride, and bumetanide. Generally, MMA-SS was more potent in altering responses to amiloride and bumetanide than GMA-MS. Hyperpolarization occurred in the absence of LDH release, but decreases in Vt, Rt, and Isc at higher fume particulate doses accompanied LDH release, to a greater extent for MMA-SS. Thus, Na+ transport and Na+,K+,2Cl¯-cotransport are affected by fume exposure; MMA-MS is more potent than GMA-MS. Enhanced Na+ absorption and decreased airway surface liquid could compromise defenses against infection.


Assuntos
Poluentes Ocupacionais do Ar/toxicidade , Brônquios/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Agonistas do Canal de Sódio Epitelial/toxicidade , Canais Epiteliais de Sódio/efeitos dos fármacos , Simportadores de Cloreto de Sódio-Potássio/efeitos dos fármacos , Aço/toxicidade , Soldagem , Brônquios/metabolismo , Brônquios/patologia , Células Cultivadas , Canais de Cloreto/efeitos dos fármacos , Canais de Cloreto/metabolismo , Relação Dose-Resposta a Droga , Impedância Elétrica , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Canais Epiteliais de Sódio/metabolismo , Gases , Humanos , Exposição por Inalação/efeitos adversos , Transporte de Íons/efeitos dos fármacos , L-Lactato Desidrogenase/metabolismo , Potenciais da Membrana , Exposição Ocupacional/efeitos adversos , Simportadores de Cloreto de Sódio-Potássio/metabolismo , Aço Inoxidável/toxicidade , Fatores de Tempo
14.
Am J Pathol ; 186(11): 2887-2908, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27643531

RESUMO

Inhaled diacetyl vapors are associated with flavorings-related lung disease, a potentially fatal airway disease. The reactive α-dicarbonyl group in diacetyl causes protein damage in vitro. Dicarbonyl/l-xylulose reductase (DCXR) metabolizes diacetyl into acetoin, which lacks this α-dicarbonyl group. To investigate the hypothesis that flavorings-related lung disease is caused by in vivo protein damage, we correlated diacetyl-induced airway damage in mice with immunofluorescence for markers of protein turnover and autophagy. Western immunoblots identified shifts in ubiquitin pools. Diacetyl inhalation caused dose-dependent increases in bronchial epithelial cells with puncta of both total ubiquitin and K63-ubiquitin, central mediators of protein turnover. This response was greater in Dcxr-knockout mice than in wild-type controls inhaling 200 ppm diacetyl, further implicating the α-dicarbonyl group in protein damage. Western immunoblots demonstrated decreased free ubiquitin in airway-enriched fractions. Transmission electron microscopy and colocalization of ubiquitin-positive puncta with lysosomal-associated membrane proteins 1 and 2 and with the multifunctional scaffolding protein sequestosome-1 (SQSTM1/p62) confirmed autophagy. Surprisingly, immunoreactive SQSTM1 also accumulated in the olfactory bulb of the brain. Olfactory bulb SQSTM1 often congregated in activated microglial cells that also contained olfactory marker protein, indicating neuronophagia within the olfactory bulb. This suggests the possibility that SQSTM1 or damaged proteins may be transported from the nose to the brain. Together, these findings strongly implicate widespread protein damage in the etiology of flavorings-related lung disease.


Assuntos
Diacetil/efeitos adversos , Aromatizantes/efeitos adversos , Pneumopatias/etiologia , Proteína Sequestossoma-1/metabolismo , Desidrogenase do Álcool de Açúcar/genética , Ubiquitina/metabolismo , Animais , Autofagia , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Humanos , Exposição por Inalação , Pneumopatias/induzido quimicamente , Pneumopatias/metabolismo , Pneumopatias/patologia , Glicoproteínas de Membrana Associadas ao Lisossomo/metabolismo , Camundongos , Camundongos Knockout , Microglia/metabolismo , Microglia/patologia , Bulbo Olfatório/metabolismo , Bulbo Olfatório/patologia , Proteína de Marcador Olfatório/genética , Proteína de Marcador Olfatório/metabolismo , Sistema Respiratório/metabolismo , Sistema Respiratório/patologia , Proteína Sequestossoma-1/genética , Desidrogenase do Álcool de Açúcar/metabolismo
15.
Toxicol Appl Pharmacol ; 289(3): 542-9, 2015 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-26454031

RESUMO

Inhalation of butter flavoring by workers in the microwave popcorn industry may result in "popcorn workers' lung." In previous in vivo studies rats exposed for 6 h to vapor from the flavoring agents, diacetyl and 2,3-pentanedione, acquired flavoring concentration-dependent damage of the upper airway epithelium and airway hyporeactivity to inhaled methacholine. Because ion transport is essential for lung fluid balance,we hypothesized that alterations in ion transport may be an early manifestation of butter flavoring-induced toxicity.We developed a system to expose cultured human bronchial/tracheal epithelial cells (NHBEs) to flavoring vapors. NHBEs were exposed for 6 h to diacetyl or 2,3-pentanedione vapors (25 or ≥ 60 ppm) and the effects on short circuit current and transepithelial resistance (Rt) were measured. Immediately after exposure to 25 ppm both flavorings reduced Na+ transport,without affecting Cl- transport or Na+,K+-pump activity. Rt was unaffected. Na+ transport recovered 18 h after exposure. Concentrations (100-360 ppm) of diacetyl and 2,3-pentanedione reported earlier to give rise in vivo to epithelial damage, and 60 ppm, caused death of NHBEs 0 h post-exposure. Analysis of the basolateral medium indicated that NHBEs metabolize diacetyl and 2,3-pentanedione to acetoin and 2-hydroxy-3-pentanone, respectively. The results indicate that ion transport is inhibited transiently in airway epithelial cells by lower concentrations of the flavorings than those that result in morphological changes of the cells in vivo or in vitro.


Assuntos
Brônquios/efeitos dos fármacos , Diacetil/efeitos adversos , Células Epiteliais/efeitos dos fármacos , Aromatizantes/efeitos adversos , Transporte de Íons/efeitos dos fármacos , Pentanonas/efeitos adversos , Manteiga , Células Cultivadas , Humanos , Exposição por Inalação/efeitos adversos , Cloreto de Metacolina/efeitos adversos , Micro-Ondas , Exposição Ocupacional/efeitos adversos
16.
Physiol Rep ; 2(7)2014 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-25347857

RESUMO

Nerve growth factor (NGF) is overexpressed in patients with inflammatory lung diseases, including virus infections. Airway surface liquid (ASL), which is regulated by epithelial cell ion transport, is essential for normal lung function. No information is available regarding the effect of NGF on ion transport of airway epithelium. To investigate whether NGF can affect ion transport, human primary air-interface cultured epithelial cells were placed in Ussing chambers to obtain transepithelial voltage (-7.1 ± 3.4 mV), short-circuit current (Isc, 5.9 ± 1.0 µA), and transepithelial resistance (750 Ω·cm(2)), and to measure responses to ion transport inhibitors. Amiloride (apical, 3.5 × 10(-5) mol/L) decreased Isc by 55.3%. Apically applied NGF (1 ng/mL) reduced Isc by 5.3% in 5 min; basolaterally applied NGF had no effect. The response to amiloride was reduced (41.6%) in the presence of NGF. K-252a (10 nmol/L, apical) did not itself affect Na(+) transport, but it attenuated the NGF-induced reduction in Na(+) transport, indicating the participation of the trkA receptor in the NGF-induced reduction in Na(+) transport. PD-98059 (30 µmol/L, apical and basolateral) did not itself affect Na(+) transport, but attenuated the NGF-induced reduction in Na(+) transport, indicating that trkA activated the Erk 1/2 signaling cascade. NGF stimulated phosphorylation of Erk 1/2 and the ß-subunit of ENaC. K-252a and PD-98059 inhibited these responses. NGF had no effect on Isc in the presence of apical nystatin (50 µmol/L). These results indicate that NGF inhibits Na(+) transport through a trkA-Erk 1/2-activated signaling pathway linked to ENaC phosphorylation.

17.
Inhal Toxicol ; 26(12): 697-707, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25140454

RESUMO

Spot welding is used in the automotive and aircraft industries, where high-speed, repetitive welding is needed to join thin sections of metal. Epoxy adhesives are applied as sealers to the metal seams. Pulmonary function abnormalities and airway irritation have been reported in spot welders, but no animal toxicology studies exist. Therefore, the goal of this study was to investigate vascular, immune and lung toxicity measures after exposure to these metal fumes in an animal model. Male Sprague-Dawley rats were exposed by inhalation to 25 mg/m³ to either mild-steel spot welding aerosols with sparking (high metal, HM) or without sparking (low metal, LM) for 4 h/d for 3, 8 and 13 d. Shams were exposed to filtered air. Bronchoalveolar lavage (BAL), lung gene expression and ex vivo BAL cell challenge were performed to assess lung toxicity. Lung resistance (R(L)) was evaluated before and after challenge with inhaled methacholine (MCh). Functional assessment of the vascular endothelium in isolated rat tail arteries and leukocyte differentiation in the spleen and lymph nodes via flow cytometry was also done. Immediately after exposure, baseline R(L) was significantly elevated in the LM spot welding aerosols, but returned to control level by 24 h postexposure. Airway reactivity to MCh was unaffected. Lung inflammation and cytotoxicity were mild and transient. Lung epithelial permeability was significantly increased after 3 and 8 d, but not after 13 d of exposure to the HM aerosol. HM aerosols also caused vascular endothelial dysfunction and increased CD4+, CD8+ and B cells in the spleen. Only LM aerosols caused increased IL-6 and MCP-1 levels compared with sham after ex vivo LPS stimulation in BAL macrophages. Acute inhalation of mild-steel spot welding fumes at occupationally relevant concentrations may act as an irritant as evidenced by the increased R(L) and result in endothelial dysfunction, but otherwise had minor effects on the lung.


Assuntos
Poluentes Ocupacionais do Ar/toxicidade , Endotélio Vascular/efeitos dos fármacos , Exposição por Inalação/efeitos adversos , Pulmão/efeitos dos fármacos , Mucosa Respiratória/efeitos dos fármacos , Vasculite/induzido quimicamente , Soldagem , Adesivos/química , Aerossóis , Animais , Células Cultivadas , Endotélio Vascular/imunologia , Endotélio Vascular/fisiopatologia , Fogo , Hematopoese Extramedular/efeitos dos fármacos , Imunidade nas Mucosas/efeitos dos fármacos , Leucócitos/efeitos dos fármacos , Leucócitos/imunologia , Leucócitos/patologia , Pulmão/irrigação sanguínea , Pulmão/imunologia , Pulmão/patologia , Macrófagos Alveolares/efeitos dos fármacos , Macrófagos Alveolares/imunologia , Macrófagos Alveolares/patologia , Masculino , Ratos Sprague-Dawley , Mucosa Respiratória/imunologia , Mucosa Respiratória/patologia , Organismos Livres de Patógenos Específicos , Baço/efeitos dos fármacos , Baço/imunologia , Baço/patologia , Aço/química , Testes de Toxicidade Aguda , Vasculite/imunologia , Vasculite/patologia , Vasculite/fisiopatologia , Soldagem/métodos
18.
Environ Health Insights ; 8(Suppl 1): 63-74, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25861220

RESUMO

INTRODUCTION: Oil spill cleanup workers come into contact with numerous potentially hazardous chemicals derived from the oil spills, as well as chemicals applied for mitigation of the spill, including oil dispersants. In response to the Deepwater Horizon Macondo well oil spill in the Gulf of Mexico in 2010, a record volume of the oil dispersant, COREXIT EC9500A, was delivered via aerial applications, raising concern regarding potential health effects that may result from pulmonary exposure to the dispersant. METHODS: The current study examined the effects on pulmonary functions, cardiovascular functions, and systemic immune responses in rats to acute repeated inhalation exposure of COREXIT EC9500A at 25 mg/m(3), five hours per day, over nine work days, or filtered air (control). At one and seven days following the last exposure, a battery of parameters was measured to evaluate lung function, injury, and inflammation; cardiovascular function; peripheral vascular responses; and systemic immune responses. RESULTS: No significant alterations in airway reactivity were observed at one or seven days after exposure either in baseline values or following methacholine (MCh) inhalation challenge. Although there was a trend for an increase in lung neutrophils and phagocyte oxidant production at one-day post exposure, there were no significant differences in parameters of lung inflammation. In addition, increased blood monocytes and neutrophils, and decreased lymphocyte numbers at one-day post exposure also did not differ significantly from air controls, and no alterations in splenocyte populations, or serum or spleen immunoglobulin M (IgM) to antigen were observed. There were no significant differences in peripheral vascular responsiveness to vasoconstrictor and vasodilator agonists or in blood pressure (BP) responses to these agents; however, the baseline heart rate (HR) and HR responses to isoproterenol (ISO) were significantly elevated at one-day post exposure, with resolution by day 7. CONCLUSIONS: In summary, acute repeated exposure to COREXIT EC9500A did not alter pulmonary function, lung injury/inflammation, systemic immune responses, or vascular tone, but did cause transient chronotropic effects on cardiac function.

19.
Front Physiol ; 4: 287, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24130533

RESUMO

In asthmatic patients, inhalation of hyperosmolar saline or D-mannitol (D-M) elicits bronchoconstriction, but in healthy subjects exercise causes bronchodilation. Hyperventilation causes drying of airway surface liquid (ASL) and increases its osmolarity. Hyperosmolar challenge of airway epithelium releases epithelium-derived relaxing factor (EpDRF), which relaxes the airway smooth muscle. This pathway could be involved in exercise-induced bronchodilation. Little is known of ASL hyperosmolarity effects on epithelial function. We investigated the effects of osmolar challenge maneuvers on dispersed and adherent guinea-pig tracheal epithelial cells to examine the hypothesis that EpDRF-mediated relaxation is associated with epithelial cell shrinkage. Enzymatically-dispersed cells shrank when challenged with ≥10 mOsM added D-M, urea or NaCl with a concentration-dependence that mimics relaxation of the of isolated perfused tracheas (IPT). Cells shrank when incubated in isosmolar N-methyl-D-glucamine (NMDG) chloride, Na gluconate (Glu), NMDG-Glu, K-Glu and K2SO4, and swelled in isosmolar KBr and KCl. However, isosmolar challenge is not a strong stimulus of relaxation in IPTs. In previous studies amiloride and 4,4'-diisothiocyano-2,2'-stilbenedisulfonic acid (DIDS) inhibited relaxation of IPT to hyperosmolar challenge, but had little effect on shrinkage of dispersed cells. Confocal microscopy in tracheal segments showed that adherent epithelium is refractory to low hyperosmolar concentrations that induce dispersed cell shrinkage and relaxation of IPT. Except for gadolinium and erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), actin and microtubule inhibitors and membrane permeabilizing agents did not affect on ion transport by adherent epithelium or shrinkage responses of dispersed cells. Our studies dissociate relaxation of IPT from cell shrinkage after hyperosmolar challenge of airway epithelium.

20.
J Toxicol Environ Health A ; 76(11): 669-89, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23941636

RESUMO

"Popcorn workers' lung" is an obstructive pulmonary disease produced by inhalation of volatile artificial butter flavorings. In rats, inhalation of diacetyl, a major component of butter flavoring, and inhalation of a diacetyl substitute, 2,3-pentanedione, produce similar damage to airway epithelium. The effects of diacetyl and 2,3-pentanedione and mixtures of diacetyl, acetic acid, and acetoin, all components of butter flavoring, on pulmonary function and airway reactivity to methacholine (MCh) were investigated. Lung resistance (RL) and dynamic compliance (Cdyn) were negligibly changed 18 h after a 6-h inhalation exposure to diacetyl or 2,3-pentanedione (100-360 ppm). Reactivity to MCh was not markedly changed after diacetyl, but was modestly decreased after 2,3-pentanedione inhalation. Inhaled diacetyl exerted essentially no effect on reactivity to mucosally applied MCh, but 2,3-pentanedione (320 and 360 ppm) increased reactivity to MCh in the isolated, perfused trachea preparation (IPT). In IPT, diacetyl and 2,3-pentanedione (≥3 mM) applied to the serosal and mucosal surfaces of intact and epithelium-denuded tracheas initiated transient contractions followed by relaxations. Inhaled acetoin (150 ppm) exerted no effect on pulmonary function and airway reactivity in vivo; acetic acid (27 ppm) produced hyperreactivity to MCh; and exposure to diacetyl + acetoin + acetic acid (250 + 150 + 27 ppm) led to a diacetyl-like reduction in reactivity. Data suggest that the effects of 2,3-pentanedione on airway reactivity are greater than those of diacetyl, and that flavorings are airway smooth muscle relaxants and constrictors, thus indicating a complex mechanism.


Assuntos
Hiper-Reatividade Brônquica/induzido quimicamente , Diacetil/toxicidade , Aromatizantes/toxicidade , Pentanonas/toxicidade , Traqueia/efeitos dos fármacos , Ácido Acético/toxicidade , Acetoína/toxicidade , Resistência das Vias Respiratórias/efeitos dos fármacos , Animais , Hiper-Reatividade Brônquica/fisiopatologia , Testes de Provocação Brônquica , Células Cultivadas , Misturas Complexas/toxicidade , Alimentos , Exposição por Inalação , Masculino , Cloreto de Metacolina , Músculo Liso/efeitos dos fármacos , Músculo Liso/fisiopatologia , Ratos , Ratos Sprague-Dawley , Traqueia/fisiopatologia
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