Nina N. Sun

In vitro cytokine release from rat type II pneumocytes and alveolar macrophages following exposure to JP-8 jet fuel in co-culture.

Alveolar type II epithelial cells (AIIE) and pulmonary alveolar macrophages (PAM) are involved in pulmonary toxicity of JP-8 jet fuel exposure. To further elucidate their inflammatory mechanisms, the effect(s) of JP-8 jet fuel on cytokine secretion were examined in a transformed rat AIIE cell line (RLE-6TN) culture alone, primary PAM (from Fischer 344 rats) culture alone, and the co-culture of AIIE and primary PAM. A series of JP-8 jet fuel concentrations (0-0.8 microg/ml), which may actually be encountered in alveolar space of lungs exposed in vivo, were placed in cell culture for 24 h. Cultured AIIE alone secreted spontaneously interleukin (IL)-1beta and -6 [below detectable limits for IL-10 and tumor necrosis factor-alpha (TNF-alpha)], whereas cultured PAM alone secreted IL-1beta, -10, and TNF-alpha, in a concentration-dependent manner. These data suggest that the release of cytokines, not only from PAM but also from AIIE cells, may contribute to JP-8 jet fuel-induced inflammatory response in the alveolar space. However, the co-cultures of AIIE and PAM showed no significant changes in IL-1beta, -6, and TNF-alpha at any JP-8 jet fuel concentration compared to control values. These cytokine levels in co-cultures of AIIE and PAM were inversely related to these of cultured AIIE or PAM alone. Interestingly, IL-10 levels in the co-culture system were concentration-dependently increased up to 1058% at JP-8 concentrations of 0.8 microg/ml, although under detectable limits in cultured AIIE alone and no significant concentration change in cultured PAM alone. It appears that PAM may possibly act via paracrine and/or autocrine pathways to signal AIIE cells to regulate cytokine release.

Immunomodulatory effects of high-dose α-tocopherol acetate on mice subjected to sidestream cigarette smoke

Several recent epidemiological investigations raise serious questions about the health effects of high-dose supplements of Vitamin E (VE) in cigarette smokers. To examine these findings, a total of 96 C57BL/6 mice were randomly assigned to eight groups in a 2 x 4 factorial design (smoke vs. sham smoke and normal diet vs. 3 VE supplements). The mice were exposed to sidestream cigarette smoke (SSCS), at 0.4 mg total particulate matter/m(3) air, from standard research cigarettes (1R4)/day or filtered room air at 30 min/day, 5 days/week, for 9 weeks through a nose-only exposure chamber. The American Institute of Nutrition 93G purified rodent diet was modulated with 75 (regular diet, 1-fold), 1050 (15-fold), 5550 (75-fold), and 11175 (150-fold) IU dl-alpha-tocopherol acetate (alpha-TA)/kg as VE supplementation and provided ad libitum at an average intake rate of 4.11 g diet/mouse/day. This result demonstrated that SSCS exposure results in lung dysfunction, as indicated by a decrease of pulmonary dynamic compliance (C(dyn)) and increase of lung resistance (R(L)), and body weight loss in mice fed with regular diet. These changes accompanied with increases of bronchoalveolar lavage (BAL) concentrations of cytokines interleukin (IL)-1 beta, IL-4 and IFN-gamma, as well as hepatic lipid peroxidation. However, supplemental alpha-TA at the doses of > or = 1050 IU/kg diet prevented the SSCS-induced body weight loss and lung dysfunction. alpha-TA at > or = 5550 IU/kg significantly increased BAL levels of IL-2 and IL-4 in both the sham SSCS and the SSCS groups. Given at 5550 IU alpha-TA/kg, but not higher, mice elevated BAL IL-1 beta level if they were exposed to SSCS. Hepatic lipid peroxidation was decreased in a dose-dependent fashion with different alpha-TA supplements in both the sham SSCS and SSCS groups. Neither SSCS nor alpha-TA had an effect on lung permeability, BAL IL-6, splenic T and B lymphocyte proliferation and their T helper (Th)1 and Th2 cytokines measured among all groups. Data suggest that supplemental alpha-TA may be needed to counteract SSCS-induced oxidative stress, but that potential side effects introduced by high dosage of this synthetic compound should be considered.

Dose-dependent transcriptome changes by metal ores on a human acute lymphoblastic leukemia cell line

The increased morbidity of childhood leukemia in Fallon, Nevada and Sierra Vista, Arizona has prompted great health concern. The main characteristic that these two towns share is the environmental pollution attributed to metal ore from abandoned mining operations. Consequently, we have investigated the transcriptome effects of metal ores from these endemic areas using a human T-cell acute lymphoblastic leukemia cell line (T-ALL). Metal ore from Fallon significantly increased cell growth after 24, 48 and 72 h of incubation at 1.5 mg/mL concentration, as measured by trypan-blue. Sierra Vista ore significantly increased cell growth with 0.15 and 1.5 mg/mL following 72 h of incubation. From human cDNA microarray, results indicate that in total, eight genes, mostly metallothionein (MT) genes, were up-regulated and 10 genes were down-regulated following treatment of the T-ALL cells with 0.15 and 1.5 mg/mL of metal ores at 72 h, in comparison with untreated cells. Twenty-eight metals of both ores were quantified and their presence may be associated with the cell growth rate and dose-dependent activation of transcriptomes in immature T-cells.

Prenatal exposure of mice to tungstate is associated with decreased transcriptome-expression of the putative tumor suppressor gene, DMBT1: Implications for childhood leukemia

Background. Two concurrent, childhood leukemia clusters have been identified in the southwestern United States at Fallon, Nevada, and Sierra Vista, Arizona. Additionally, Fallon, Nevada has also experienced concurrent contamination by atmospheric tungsten particles. The etiology of leukemia is not known. Hypothesized risk factors for leukemia are environmental exposure, genetic predisposition, and viral infection. Additionally, strong evidence supports a prenatal origin. Our objective is to generate testable hypotheses towards elucidating the probable, multi-factorial etiology of leukemia by identifying the exposures unique to Fallon, Nevada, and held in common with Sierra Vista, Arizona, then exposing C57BL/6 mice, while in utero, to these chemicals to ascertain their leukemogenic potential. Utilizing advances in medical geology to analyze tree rings, surface dust, lichens and atmospheric particulate matter, we have identified tungsten and arsenic as potentially relevant to leukemogenesis. Methods. We utilized microarray (Affymetrix 430A 2.0 mouse) and real-time RTPCR of Dmbt1 transcriptome-expression in spleen tissue collected from four-week-old C57BL/6 mouse pups (N = 6–8/group/gender) exposed, while in utero, to tungstate, arsenite, tungstate/arsenite and longitudinal controls at 20% of the normalized exposure a human mother would receive during gestation at mean environmental concentrations. Results. Prenatal exposure to tungstate is associated with a 37 + 1.2-fold (p = 0.012) decrease in DMBT1 transcriptome-expression in mice expressing DMBT1 at high levels. Additionally, prenatal exposure to tungstate/arsenite significantly altered a cytokine–cytokine receptor interaction pathway associated with lymphocyte activation and a network associated with hematological/immunological disease. Conclusion. Because DMBT1 protein products are known to aggregate viruses and possibly regulate immune response, additional research is warranted to determine the potential that prenatal exposure to tungstate or tungstate/ arsenite has to increase susceptibility to viruses and to induce leukemogenesis.

In Vitro Pro-inflammatory Regulatory role of Substance P in Alveolar Macrophages and Type II Pneumocytes after JP-8 Exposure

The effects of JP-8 on pro-inflammatory cytokine interleukin (IL)-1α,β and nitric oxide (NO) secretion as well as the role of substance P (SP) in these processes were examined in cultured alveolar macrophages (AM), type II epithelial cells (AIIE), and AM/AIIE co-cultures. Exposure of AM to JP-8 for 24 hr exhibited release of IL-1α,β, whereas exposure to AIIE showed a concentration-dependent NO overproduction. Data indicate that there are cell-dependent inflammatory mechanisms responsible for the actual level of JP-8 exposure in alveoli. However, treatment with substance P significantly attenuated JP-8 induced the IL-1α,β secretion. This finding was confirmed by using [Sar9 Met (O2)11] SP (10− 10 M), an agonist of substance P, suggesting that substance P may have signal pathway(s) to AM in the inflammatory response mediated by IL-1. Moreover, AM/AIIE co-culture obviously reduced NO overproduction observed in AIIE alone, suggesting that there may be cell interactions or communications between AM and AIIE in response to the JP-8 exposure.

Tissue-specific patterns of neurokinin-1 receptor (NK-1R) gene expression in mice exposed to sidestream cigarette smoke

Neurokinin-1 receptor (NK-1R), a high-affinity plasma membrane-bound receptor for neurokinin substance P, plays important roles in the onset of the pathophysiological responses. To test whether the transcript levels of gene encoding NK-1R in organs are affected by sidestream cigarette smoke (SSCS) exposure, the C57BL/6 mice were randomly assigned to five groups (six/group) in a study of the dose-effect relationship. The mice were exposed to 0 (filtered room air), 2, 4, 8 and 16 mg total particulate matter (TPM) of SSCS/exposure/day, respectively, for seven days through a nose-only exposure chamber (IN-TOX, Albuquerque, NM, USA). The levels of NK-1R mRNA in the lung, heart, liver, kidney and spleen tissues were detected by reverse transcription-polymerase chain reaction (RT-PCR) techniques and normalized against GAPDH expression. NK-1R mRNA in heart tissue showed SSCS-induced dose-dependent downregulation, with minimum expression at a dose of 8.0 mg TPM. Whereas, the levels of NK-1R mRNA in the liver were upregulated to 2.86 and 5.13- fold after exposure to 2.0 and 4.0 mg TPM of SSCS respectively, then returned to 4.19 and 3.93-fold at the exposure doses of 8.0 and 16.0 mg TPM, respectively, when compared to that of the control. In the kidney, SSCS exposure at a dose of 2.0 TPM, but not higher than that level, induced significant elevation of NK-1R mRNA expression. These findings suggest that there are the paracrine and/or autocrine signalling mechanisms through receptor -ligand interactions. No alteration of NK-1R gene expression was observed in the lungs and spleen tissues in this study. The tissue-specific patterns by which SSCS affect NK-1R gene expression in these organs may partially explain dissimilarity of NK-1R activation and the associated toxicity caused by environmental tobacco smoke.

Acute changes in sputum collected from exposed human subjects in mining conditions

Neprilysin (NEP) is a key cell surface peptidase in the maintenance of airway homeostasis and the development of pulmonary disorders. However, little information is available about the effect of particulate matter (PM) on airway NEP. In this controlled human exposure study, changes in induced sputum were measured in 11 subjects at baseline, overshot (OS) mucking, and diesel exhaust (DE) exposure days. Neither OS condition nor DE exposure was found to induce significant changes in total protein, but DE induced significant increases in cell numbers of macrophages and epithelium. Moreover, significant increases in soluble NEP were observed following OS mining dust particulates (0.43 ± 0.06 nmol/μg protein/min; p = .023) and DE exposure (0.40 ± 0.03 nmol/μg protein/min; p = .035) when compared with the baseline control (0.30 ± 0.04 nmol/μg protein/min), with 42% and 31% average net increase, respectively. Pearson’s correlation analyses indicated that sputum NEP activity was significantly associated with personal exposure product (elemental carbon concentration [mg/m3] × time [min]; C × T). The data suggest that changes in NEP activity may be an early, accurate endpoint for airway epithelial injury and provide a new insight into the mechanism of airway effects following particulate exposure.