ToxSci Advance Access originally published online on February 14, 2007
Toxicological Sciences 2007 97(1):181-188; doi:10.1093/toxsci/kfm017
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Long- and Short-Term Changes in the Neuroimmune-Endocrine Parameters following Inhalation Exposures of F344 Rats to Low-Dose Sarin
Immunology Division, Lovelace Respiratory Research Institute, 2425 Ridgecrest Dr SE, Albuquerque, New Mexico 87108
1 To whom correspondence should be addressed. Fax: (505) 348-4986. E-mail: msopori{at}lrri.org.
Received February 9, 2007; accepted February 12, 2007
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Inhalation of subclinical doses of sarin suppresses the antibody-forming cell (AFC) response, T-cell mitogenesis, and serum corticosterone (CORT) levels, and high doses of sarin cause lung inflammation. However, the duration of these changes is not known. In these studies, rats were exposed to a subclinical dose of sarin (0.4 mg/m3/h/day) for 1 or 5 days, and immune and inflammatory parameters were assayed up to 8 weeks before sarin exposure. Our results showed that the effects of a 5-day sarin exposure on the AFC response and T-cell receptor (TCR)-mediated Ca2+ response disappeared within 2–4 weeks after sarin exposure, whereas the CORT and adrenocorticotropin hormone (ACTH) levels remained significantly decreased. Pretreatment of rats with chlorisondamine attenuated the effects of sarin on the AFC and the TCR-mediated Ca2+ response, implicating the autonomic nervous system (ANS) in the sarin-induced changes in T-cell function. Moreover, exposure to a single or five repeated subclinical doses of sarin upregulated the mRNA expression of proinflammatory cytokines in the lung, which is associated with the activation of NF
B in bronchoalveolar lavage cells. These effects were lost within 2 weeks of sarin inhalation. Our results suggest that while sarin-induced changes in T cells and cytokine gene expression were short lived, suppression of CORT and ACTH levels were relatively long lived and might represent biomarkers of sarin exposure. Moreover, while the effects of sarin on T-cell function were regulated by the ANS, the decreased CORT levels by sarin might result from its effects on the hypothalamus-pituitary-adrenal axis. Key Words: organophosphates; sarin; neuroimmune modulation; glucocorticoids; hypothalamus-pituitary-adrenal (HPA) axis.
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Sarin (O-isopropyl methylphosphonofluoridate, also known as GB) is an organophosphate nerve agent (Wood, 1951
) that is among the more lethal chemical toxins known to mankind. Because of its high lethality and ease of production and delivery, sarin is a potent weapon in the hands of terrorists and rogue nations. Sarin has been effectively used in the terrorist attacks in Japan and during the 1980's Iraq-Iran War. Like other organophosphorous nerve agents, sarin irreversibly inhibits acetylcholinesterase (AChE) (Grob and Harvey, 1958). Inhibition of AChE by organophosphates causes an instant flooding of acetylcholine at the synapse, leading to cholinergic crisis. Sarin exposure causes circulatory, respiratory, and long- and short-term neurological damage (Abou-Donia, 1993, 2003; Kadar et al., 1995; Nakajima et al., 1999; Niven and Roop, 2004; Sidell, 1994), and most fatalities in animals and humans result from acute respiratory failure. High doses of sarin inhalation in rats cause lung inflammation, which in the surviving animals is resolved within 2–3 weeks after exposure (Pant et al., 1993). We have reported that in rats inhalation of subclinical doses of sarin, which do not cause overt signs of cholinergic toxicity or detectable changes in the brain AChE activity, induces the expression of proinflammatory cytokines in the brain and suppresses the T-cell-dependent antibody response and T-cell mitogenesis (Henderson et al., 2002; Kalra et al., 2002); however, the mechanism and the duration of these effects have not been clearly defined.
There is increasing evidence that the immune, endocrine, and the nervous systems communicate with each other through hormones, neurotransmitters, and cytokines (Blalock, 1994; Felten et al., 1987; Sopori et al., 1998; Tracey, 2005). Many neuroactive substances, such as nicotine, cannabinoids, and opiates, suppress the immune system through the central and peripheral mechanisms (Friedman et al., 2006; Sopori et al., 1998). In a number of instances, the immunosuppressive effects of these agents are associated with the activation of the hypothalamus-pituitary-adrenal (HPA) axis (Sarnyai et al., 2001). Activation of the HPA axis stimulates the production of the adrenocorticotropin hormone (ACTH) from the pituitary and cortisol/corticosterone (CORT) from the adrenals (Meier, 1996; Turnbull and Rivier, 1999; Webster et al., 2002); however, exposure to subclinical doses of sarin causes a dramatic decrease in serum CORT levels (Kalra et al., 2002). It is not clear whether the sarin-induced reduction in CORT levels reflects adrenal toxicity or suppression of the HPA axis. The lymphoid tissues are also innervated with noradrenergic postganglionic sympathetic fibers (Felten and Felten, 1994). Lymphocytes express adrenoceptors, and norepinephrine released at the nerve termini inhibits T-cell responses (Fuchs et al., 1988; Madden, 2003; Madden et al., 1995). We have reported that pretreatment of rats with the ganglionic blocker chlorisondamine (CHL) moderates the effects of sarin on T-cell mitogenesis, but the effects CHL on the antigen receptor-mediated T-cell responses were not addressed (Kalra et al., 2002). In this study, we examined the effects of repeated inhalation of low-dose sarin on the HPA axis, the T-cell receptor (TCR)-mediated responses, and the lung inflammation. The duration of these effects after the sarin exposure was determined.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Animals.
Pathogen-free, 8-week-old male Fischer 344 rats were purchased from Harlan Sprague-Dawley Farms (Indianapolis, IN). Food and water were provided ad libitum to the animals. Approximately 10-week-old animals were used for sarin inhalation. All studies were conducted at Lovelace Respiratory Research Institute (LRRI), a facility fully accredited by the Association for the Assessment and Accreditation of Laboratory Animal Care International.
CHL treatment.
The ganglionic blocker CHL (Tocris, Ballwin, MO) was injected sc into rats (10 mg/kg bw) 7 days prior to sarin exposure. This concentration of CHL blocks the behavioral responses to neuroactive substances for several months (Reuben et al., 1998).
Sarin exposure.
Animals were exposed to subclinical doses of sarin by inhalation as described previously (Henderson et al., 2002). Briefly, sarin, dissolved in isopropyl alcohol, was obtained from the U.S. Army. The sarin nose-only exposure system developed at LRRI was used to expose rats to vehicle or sarin at 0.4 mg/m3/h/day for 1 or 5 days according to a previously published protocol (Henderson et al., 2002). This concentration of sarin produced no detectable histopathological changes in the brain or overt symptoms of behavioral neurotoxicity.
Isolation of tissues and cells.
Blood was collected by cardiac puncture, and the clotted blood was centrifuged to obtain serum. Spleen cell suspensions were made as described (Singh et al., 2000). Briefly, spleens were pressed through stainless steel mesh, and red blood cells were lysed by treatment with NH4Cl solution. After washing, cells were counted and suspended in complete medium (RPMI 1640 containing 10% fetal calf serum, 2mM glutamine, 50mM 2-ME, and 10 µg/ml gentamicin).
Isolation of lungs and bronchoalveolar lavage cells.
Bronchoalveolar lavage (BAL) was collected as described previously (Langley et al., 2004). Briefly, the lungs were removed from the animals at sacrifice. The left side of the lung was tied off at the bronchi with clips, and the right lung lobe was lavaged twice with 3-ml aliquots of sterile saline solution through a tracheal cannula. The lavage was pooled and centrifuged and the cell pellet (BAL cells) collected. The right lung was used for RNA preparation for PCR analysis, and the left lung was fixed at constant hydrostatic pressure in 10% neutral-buffered formalin (Langley et al., 2004) and used for histopathology.
Antibody-forming cell assay.
To determine the antibody-forming cell (AFC) response, vehicle- and sarin-treated animals were injected iv with 5 x 108 sheep red blood cells (SRBC) (Colorado Serum Company, Denver, CO) 2 days after the beginning of vehicle/sarin inhalation. The animals continued to be exposed to sarin or vehicle for three additional days and were sacrificed 24 h after the last treatment (i.e., 4 days after immunization with SRBC). The primary direct AFC response was determined by the Cunningham and Szenberg method as described (Singh et al., 2000). Briefly, spleen cells were mixed with 2% SRBC and 25 µl of guinea pig complement (Accurate Chemicals, Westbury, NY) in 140 µl complete medium. Aliquots were distributed on Cunningham slides and incubated for 45 min at 37°C. Results are expressed as AFC/106 spleen cells.
Electrophoretic mobility shift assay for NFB.
Nuclear extracts were made from BAL cells from vehicle- and sarin-treated animals essentially as described (Chaturvedi et al., 2000). Briefly, 2 x 106 BAL cells were lysed on ice using hypotonic cell lysis buffer. The nuclear pellet was extracted with the high-salt nuclear extraction buffer, and the suspension was centrifuged to yield the nuclear extract supernatant. For the NFB electrophoretic mobility shift assay (EMSA), an aliquot of the nuclear extract containing 8 µg protein was incubated with 32P-end-labeled 22-mer double-stranded NFB oligonucleotide (Promega, Madison, WI) with a NFB consensus sequence: 5'-AGTTGAGGGGACTTTCCCAGG C-3' and 3'-TCAACTCCCCTGAAAGGGTCCG-5' for 30 min at 37°C. The resulting DNA-protein complex was resolved on a 5% native polyacrylamide gel (Upstate Biotech, Charlottesville, VA). For the supershift assay, nuclear extracts were incubated for 15 min at 37°C with anti-p50 antibody prior to the addition of a 32P-labled NFB probe. Samples were resolved by gel electrophoresis (as above), and the radiolabeled bands on the dried gels were visualized by X-ray film (Eastman-Kodak, Rochester, NY).
Assay for intracellular ionized calcium concentration.
Intracellular ionized calcium concentration [Ca2+]i levels were determined by spectrofluorometry as described (Kalra et al., 2002). Briefly, spleen cells were loaded with acetoxymethyl ester of indo-1 (Sigma, St Louis, MO). Indo-1-labeled spleen cells were treated with mouse antirat 
ß-TCR mAb followed by goat antimouse IgG (second Ab). Changes in the [Ca2+]i were determined in a PTI Deltascan fluorometer (Photon Technology International, South Brunswick, NJ).
Assay for serum CORT levels.
All animals assigned for determining the blood CORT and ACTH levels were sacrificed between 7:00 and 8:30 A.M. Serum CORT levels were determined by the CORT RIA-kit (ICN Biochemicals, Orangeburg, NY) according to the manufacturer's instructions.
Assay for plasma ACTH levels.
Briefly, blood samples were drawn via heart puncture, and the blood was collected in prechilled EDTA tubes. The samples were centrifuged and 1000 KIU of aprotinin (Sigma) was added to each milliliter of plasma. The samples were frozen and stored at – 80°C until use. ACTH was assayed by an I125-radioimmunoassay kit (MP Biomedicals, Orangeburg, NY) according to the manufacturer's directions. Briefly, ACTH standards (0–1000 pg/ml) and the plasma samples (diluted 1:8 in saline) were incubated with I125-labled anti-ACTH for 16 h at 4°C. After the addition of precipitation solution, samples were centrifuged, supernatants decanted, and the tubes containing the precipitates were counted in a gamma counter. The amount of ACTH in the plasma was deduced from the standard curve.
RT-PCR analysis for TNF-, IL-1ß, and IL-6.
RNA was extracted from the lungs and subjected to semiquantitative reverse transcriptase (RT)-PCR analysis for IL-1ß, TNF-, and IL-6 by previously published methods (Razani-Boroujerdi et al., 2004). Briefly, total RNA was isolated from the lung tissues using the BCP phase-separation reagent (Molecular Research Center, Cincinnati, OH). RNA was precipitated by 2-propanol and washed with 75% ethanol. The RNA pellet was dried for a short time, resuspended in RNase-free water, and quantitated spectrophotometrically. The gene-specific primer sets for proinflammatory cytokines were purchased from Sigma. The primer sets for rat IL-1ß, TNF-, IL-6, and the housekeeping gene (GAPDH) were as follows: IL-1ß: forward (5'-CCAGGATGAGGACCCAAGCA-3') and reverse (5'-TCCCGACCATTGCTGTTTCC-3'), TNF-: forward (5'-ATGAGCACAGAAAGCATGATCCGC-3') and reverse (5'-CCAAAGTAGACCTGCCCGGACTC-3'), IL-6: forward (5'-AAAATCTGCTCTGGTCTTCTGG-3') and reverse (5'-GGTTTGCCGAGTAGACCTCA-3'), and GAPDH: forward (5'-CGGATTTGG CCGTATCGGACGCC-3') and reverse (5'-GCCTTGGCAGCACCAGTGGATGC-3'). RT-PCR was carried out as detailed previously (Henderson et al., 2002). Briefly, cDNA synthesis and predenaturation were performed by SuperScript First-Strand Synthesis Systems for the RT-PCR kit (Invitrogen, Carlsbad, CA). The cDNA was amplified by 35 PCR cycles (denaturation at 94°C for 1 min, annealing at 55°C for 1 min, and extension at 72°C for 1 min), and for the final extension of one cycle of 72°C for 10 min in a Thermal Cycler 9600 (Perkin Elmer, Boston, MA). The PCR products were electrophoresed on 1.8% agarose gel with ethidium bromide staining for visualization. The gel was photographed; the bands were quantified with a Bio-Rad GS-800 scanner (Bio-Rad Inc, Hercules, CA) and Quantity One software (Bio-Rad Inc), and standardized to GAPDH to present the level of gene expression.
Real time-PCR analysis for IL-1ß and TNF-.
Lung mRNA from 5-day vehicle- and sarin-exposed animals was isolated and quantified as described in RT-PCR. Real time-PCR primer and probes were designed with Primer Express 2.0 (ABI, Foster City, CA). PCR was carried out for 40 cycles with denaturation at 95°C for 15 s and annealing at 58°C for 60 s. Linear regression (Prism Software, GraphPad, Inc, San Diego, CA) was used to generate standard curves for each analysis. IL-1ß and TNF- gene expression is represented as the ratio of the gene to GAPDH expression. The primers shown below are for real time-PCR: IL-1ß: forward (5'-GATGGCTGCACTATTCCTAATGC-3'), reverse (5'-AGACTGCCCATTCTCGACAAG-3'), and probe (6FAM-TGCTCCTCACCCACACCGTCAGC); TNF-: forward (5'-ACAAGGCTGCCCCGACTAT-3'), reverse (5'-CTCCTGGTATGAAGTGGCAAATC-3'), and probe (6FAM-CCCCAGGACATGCTAGGGAGCCC); and GAPDH: forward (5'-TGCCCAGTATGATGACATCAAGAAG-3'), reverse (5'-AGCCCAGGATGCCCTTTAGT-3'), and probe (VIC-TGG TGA AGC AGG CGG CCG A).
Statistical analysis.
Student's t-test was used to compare the means between the vehicle- and sarin-treated animals. For comparison of three or more groups, the data were analyzed by one-way ANOVA followed by Dunnett's posttest. These statistical procedures were performed using GraphPad Prism Software (GraphPad Software, Inc).
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Sarin Causes a Long-Term Decline in Serum CORT Levels
Although repeated exposure to low-dose sarin inhibits the immune system, these immunosuppressive effects did not emanate from increased production of CORT; on the contrary, repeated subclinical sarin treatments significantly reduce the serum CORT levels (Kalra et al., 2002
). Because reduced serum CORT level may be a biomarker for cholinergic toxicity (Langley et al., 2004), we examined the changes in serum CORT levels at various times after sarin exposure. Serum CORT levels of sarin-treated animals remained significantly lower than vehicle-exposed levels, even at 8 weeks after the sarin treatment (Fig. 1).
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Sarin Decreases Plasma ACTH Levels
To test whether the sarin-induced changes in serum CORT levels reflected the inhibitory effects of sarin on the HPA axis, we measured the plasma ACTH levels in sarin-exposed animals. Indeed the sarin treatment significantly reduced the plasma ACTH levels within 1 week after sarin treatment, and those levels remained significantly depressed even at 4 weeks before sarin treatment (Fig. 2).
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Kinetics of Sarin-Induced Changes in Immune and Inflammatory Parameters
We have previously shown that within 24 h after a 5-day sarin exposure, antibody response to the T-dependent antigen SRBC and the TCR-induced increase in [Ca2+]i are significantly decreased in spleen cells from sarin-treated animals (Kalra et al., 2002
). To ascertain the life span of the immunological changes associated with repeated exposure to subclinical doses of sarin, animals were exposed to sarin or vehicle, and the anti-SRBC AFC responses of spleen cells were quantitated by the Cunningham-Szenberg technique. The AFC response of sarin-treated animals was significantly lower than control animals at 1 and 2 weeks after sarin exposure (Fig. 3); however, these suppressive effects waned after this period. Similarly, the increase in [Ca2+]i following the ligation of TCR with anti-TCR antibodies remained significantly lower in spleen cells from sarin-exposed than vehicle-exposed animals at 1 and 2 weeks after sarin treatment (Fig. 4). As with the AFC response, the Ca2+ response of sarin-exposed T cells was normalized after 2 weeks before sarin exposure. Thus, compared to the effects on CORT and ACTH, the effects of sarin on T-cell immunity are relatively short lived.
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A single exposure to high-dose sarin induces lung inflammation in rats, which is resolved within 2–3 weeks after the exposure (Levy et al., 2004
; Pant et al., 1993). To determine if a single subclinical dose of sarin also caused lung inflammation, rats were exposed to sarin (0.4 mg/m3) once, and lungs were evaluated for inflammation by histopathology and expression of proinflammatory cytokines at 24 h after the exposure. While under these conditions histopathological examination of vehicle- and sarin-treated lungs did not show any overt signs of lung inflammation, such as a significant increase in BAL cell numbers and leukocytic infiltration into the lung (not shown), RT-PCR analysis of the lung mRNA showed that the expression of IL-1ß, IL-6, and TNF- was significantly increased after sarin exposure (Fig. 5). Repeated exposure to sarin for 5 days did not produce histopathological evidence of lung inflammation (not shown), but real time-PCR analysis of the lung mRNA showed a significant increase in the mRNA expression of IL-1ß and TNF-, which was lost within 2 weeks after sarin exposure (Fig. 6). These results suggest that single or repeated exposure to subclinical doses of sarin includes the molecular but not the cellular indices of lung inflammation.
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Sarin Activates NF
B in BAL CellsTo understand the molecular mechanism for the increase in the expression of proinflammatory cytokines in sarin-treated lungs, we examined the nuclear localization of NF
B in BAL cells from vehicle- and sarin-treated animals. NFB is the most potent transcription factor for the transcription of the proinflammatory cytokines TNF-, IL-1ß, and IL-6 (Blackwell and Christman, 1997). Nuclear extracts were obtained from BAL cells at 24 h after a single low-dose (0.4 mg/m3/h) exposure to sarin. To visualize nuclear localization of NFB, the extracts were subjected to EMSA using NFB p50-specific oligonucleotides and supershifted with NFB p50 subunit-specific antibody. Sarin strongly increased the nuclear content of NFB that was supershifted with anti-p50 antibody (Fig. 7). Thus, it is likely that sarin-induced activation of NFB stimulated the transcription of proinflammatory cytokines in the lung.
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Ganglionic Blocker Attenuates the Suppressive Effects of Sarin on Antigen-Mediated T-Cell Responses
Pretreatment with the ganglionic blocker CHL moderates the inhibitory effects of sarin on T-cell proliferation in response to the mitogen ConA (Kalra et al., 2002
), suggesting that some effects of sarin on T-cell mitogenesis are mediated through the autonomic nervous system (ANS). To ascertain whether the ANS regulates the effects of sarin on the antigen-stimulated responses, animals were pretreated with CHL 1 week before sarin exposure and then immunized with the T-cell-dependent antigen SRBC on day 2 of sarin exposure. After the immunization, exposure to sarin was continued for three additional days. Spleen cells were obtained at 24 h after the last sarin exposure and tested for the development of the anti-SRBC AFC response. While CHL did not affect the AFC response of vehicle-treated (CON) animals, the sarin-induced (SAR) suppression of the AFC response was significantly attenuated in CHL-pretreated animals (SAR + CHL) (Fig. 8A). These results suggest that the effects of sarin on T-cell-dependent antibody response were mediated through the ANS. To examine directly whether sarin affects the TCR-mediated responses through the ANS, spleen cells from sarin-treated (SAR) and SAR + CHL-treated animals were compared with vehicle-treated (CON) spleen cells in their ability to raise [Ca2+]i after the ligation of TCR with anti-TCR plus a second antibody. While CHL pretreatment per se did not affect the TCR-induced Ca2+ response, the inhibitory effect of sarin on the [Ca2+]i was significantly moderated in CHL-treated animals (Fig. 8B). Thus, the effects of sarin on the TCR-mediated responses are also regulated by the ANS.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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In general, cholinergic compounds that cross the blood-brain barrier suppress T-cell function (Langley et al., 2004
). Sarin is a potent cholinergic compound, and we have reported that repeated inhalation of subclinical doses of sarin suppresses T-cell mitogenesis and T-dependent AFC responses (Kalra et al., 2002). Results presented herein indicate that sarin-induced inhibition of T-cell-dependent functions (i.e., the AFC response to SRBC and the TCR-mediated rise in intracellular Ca2+) is lost 2 weeks after sarin exposure. Therefore, effects of sarin on T-cell function are relatively short lived.
Many neuroactive drugs such as nicotine, cannabinoids, and opiates inhibit both adaptive and inflammatory responses (Friedman et al., 2006; Sopori et al., 1998; Tracey, 2005). However, while sarin suppresses T-cell immunity, it acutely induces lung inflammation resulting from neutrophilic and eosinophilic infiltration (Levy et al., 2004; Pant et al., 1993). Our results show that although a single or repeated exposure to a subclinical dose of sarin does not promote significant leukocytic infiltration into the lung, it does increase the expression of proinflammatory cytokines in the lung. Thus, subclinical doses of sarin do not cause overt signs of lung inflammation, but leave molecular imprints of lung inflammation. However, similar to high-dose, sarin-induced cellular inflammation in the lung (Pant et al., 1993), the effects of low-dose sarin on proinflammatory cytokines are relatively short lived and disappear within 2 weeks after sarin exposure. The transcription factor NFB is a potent activator for transcription of several proinflammatory cytokine genes (Blackwell and Christman, 1997), and our results show that sarin promotes the nuclear translocation of NFB in BAL cells. Therefore, increased gene expression of IL-1ß, TNF-, and IL-6 observed in BAL cells from sarin-treated animals may result through activation of NFB. In higher doses, sarin may elevate the transcription and translation of the inflammatory cytokines to the level that results in frank lung inflammation as observed by others (Levy et al., 2004; Pant et al., 1993). Interestingly, while many immunosuppressive drugs suppress both adaptive and inflammatory responses (Friedman et al., 2006; Sopori, 2002), sarin suppresses T-cell immunity while promoting inflammatory responses. However, such differential effects on the adaptive and inflammatory responses are not unique to sarin and have been observed with other immunomodulatory treatments (Moraska et al., 2002).
Increased CORT levels through activation of the HPA axis contribute to immunosuppression caused by many neuroactive substances, including opiates and cannabinoids (Friedman et al., 2006); however, sarin strongly suppresses serum CORT levels (Kalra et al., 2002) and, as reported here, serum CORT levels are depressed for at least 8 weeks after sarin inhalation. While these observations suggest that activation of the HPA axis is not involved in the sarin-induced suppression of the T-cell function, the moderating effect of CHL on sarin-induced AFC and [Ca2+]i responses suggests that sympathetic ANS is intimately involved in the suppression of antigen-mediated T-cell responses. Our results (Kalra et al., 2002) did not identify the mechanism for decreased CORT levels in sarin-treated animals. While the results presented herein do not rule out some degree of adrenal malfunction in sarin-treated rats, it is clear that, in addition to reduction in serum CORT levels, sarin treatment potently decreases plasma ACTH levels. This indicates that the suppression of the HPA axis contributes to the sarin-induced reduction in CORT production. Moreover, while the other indices of sarin-induced immunological changes (i.e., T-cell function and inflammatory cytokines) are normalized within 2 weeks after sarin exposure, ACTH and CORT levels continue to remain depressed for relatively long periods and might represent a biomarker for nerve gas toxicity.
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ACKNOWLEDGMENTS |
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This work was supported by a grant from the U.S. Army (W81XWH-04-C-0071). We thank the LRRI Inhalation team, particularly Richard White, Dean Kracko, and Terry Zimmerman, for their help in exposing animals to sarin. We also thank Ms Vicki Fisher for editorial help. The views, opinions, and/or findings contained in this report are those of the authors and should not be construed as an official Department of the Army position, policy, or decision unless so designated by other documentation.
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