Hazard Assessment of High-Nitrogen Compounds: Coupling Transcriptomics with Multi-Cellular Co-Culture Assays Provides Accurate Systemic Toxicity Screening [TNT]
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE90618 | GEO | 2018/12/01
REPOSITORIES: GEO
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Project description:Abstract - Current in vitro hazard assessment techniques lack the systems context necessary to holistically describe in vivo toxicity. To remedy this shortcoming, we have employed the Integrated discrete Multiple Organ Co-Culture (IdMOC) system in conjunction with global-transcriptomic expression assessment to provide a systems context of interactive organ and toxicity pathway responses. Specifically, IdMOC exposures containing human cells representative of five organ types (kidney, liver, lung, vascular endothelium and heart muscle) were compared to mono-culture exposures, to evaluate toxic effects in a well-studied legacy munition, 2,4,6-trinitrotoluene (TNT) compared to a structurally similar insensitive munition, 2,4-dinitroanisole (DNAN) having sparsely described toxicity. DNAN toxicity, based on cell viability, was significantly lower than TNT for three cell lines: liver, vascular endothelium, and heart. Toxicity pathways enriched in transcriptional analysis of kidney cells exposed to TNT and DNAN through the IdMOC system reflected the literature-based in vivo mammalian toxicity of parent compounds and metabolites where multiple screening values matched chronic dosing levels within 1 order of magnitude. Enriched pathways were primarily unique to each chemical where TNT exposures affected xenobiotic metabolism, oxidative stress, and cell cycle pathways in addition to pathways providing potential screening for hematotoxicity, renal toxicity and urinary cancer. Ten pathway-level responses from IdMOC suggested that DNAN elicited toxicity representative of the primary metabolite, 2,4-dinitrophenol (2,4-DNP). Relative to monoculture, IdMOC results provided a higher number and more robust enrichment of pathways involved in known in vivo toxicity mechanisms for TNT and a profile indicative of in vivo systemic toxicity of DNAN and metabolite 2,4-DNP.
2018-12-01 | GSE90617 | GEO
Project description:Abstract — Within the US military, new insensitive munitions (IMs) are rapidly replacing conventional munitions improving safety from unintended detonation. Toxicity data for IM chemicals are expanding rapidly, however IM constituents are typically deployed in mixture formulations, and very little is known about their mixture toxicology. In the present study we sought to characterize the mixture effects and toxicology of the two predominant IM formulations IMX-101 and IMX-104 in acute (48 h) larval fathead minnow (Pimephales promelas) exposures. IMX-101 consists of a mixture of 2,4-dinitroanisole (DNAN), 3-nitro-1,2,4-triazol-5-one (NTO), and nitroguanidine (NQ) while IMX-104 is composed of DNAN, NTO, and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX). DNAN was the most potent constituent in IMX-101 eliciting an LC50 of 36.1 mg/L, whereas NTO and NQ did not elicit significant mortality in exposures up to 1040 and 2640 mg/L, respectively. Toxic unit calculations indicated that IMX-101 elicited toxicity representative of the component concentration of DNAN within the mixture. Toxicogenomic responses for the individual constituents of IMX-101 indicated unique transcriptional expression and functional responses characteristic of: oxidative stress, impaired energy metabolism, tissue damage and inflammatory responses in DNAN exposures; impaired steroid biosynthesis and developmental cell-signaling in NQ exposures; and altered mitogen-activated protein kinase signaling in NTO exposures. Transcriptional responses to the IMX-101 mixture were driven by the effects of DNAN where expression and functional responses were nearly identical comparing DNAN alone versus the fractional equivalent of DNAN within IMX-101. Given that each individual constituent of the IMX-101 mixture elicited unique functional responses, and NTO and NQ did not interact with DNAN within the IMX-101 mixture exposure, the overall toxicity and toxicogenomic responses within acute exposures to the IMX-101 formulation are indicative of "independent" mixture toxicology. Alternatively, in the IMX-104 exposure both DNAN and RDX were each present at concentrations sufficient to elicit lethality (RDX LC50 = 28.9 mg/L). Toxic-unit calculations for IMX-104 mixture formulation exposures indicated slight synergistic toxicity (ΣTU LC50 = 0.82, 95% confidence interval = 0.73-0.90). Unique functional responses relative to DNAN were observed in the IMX-104 exposure including responses characteristic of RDX exposure. Based on previous transcriptomics responses to acute RDX exposures in fathead minnow larvae, we hypothesize that the potentially synergistic responses within the IMX-104 mixture are related to interactive effects of each DNAN and RDX on oxidative stress mitigation pathways.
2018-03-01 | GSE101889 | GEO
Project description:Abstract — Within the US military, new insensitive munitions (IMs) are rapidly replacing conventional munitions improving safety from unintended detonation. Toxicity data for IM chemicals are expanding rapidly, however IM constituents are typically deployed in mixture formulations, and very little is known about their mixture toxicology. In the present study we sought to characterize the mixture effects and toxicology of the two predominant IM formulations IMX-101 and IMX-104 in acute (48 h) larval fathead minnow (Pimephales promelas) exposures. IMX-101 consists of a mixture of 2,4-dinitroanisole (DNAN), 3-nitro-1,2,4-triazol-5-one (NTO), and nitroguanidine (NQ) while IMX-104 is composed of DNAN, NTO, and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX). DNAN was the most potent constituent in IMX-101 eliciting an LC50 of 36.1 mg/L, whereas NTO and NQ did not elicit significant mortality in exposures up to 1040 and 2640 mg/L, respectively. Toxic unit calculations indicated that IMX-101 elicited toxicity representative of the component concentration of DNAN within the mixture. Toxicogenomic responses for the individual constituents of IMX-101 indicated unique transcriptional expression and functional responses characteristic of: oxidative stress, impaired energy metabolism, tissue damage and inflammatory responses in DNAN exposures; impaired steroid biosynthesis and developmental cell-signaling in NQ exposures; and altered mitogen-activated protein kinase signaling in NTO exposures. Transcriptional responses to the IMX-101 mixture were driven by the effects of DNAN where expression and functional responses were nearly identical comparing DNAN alone versus the fractional equivalent of DNAN within IMX-101. Given that each individual constituent of the IMX-101 mixture elicited unique functional responses, and NTO and NQ did not interact with DNAN within the IMX-101 mixture exposure, the overall toxicity and toxicogenomic responses within acute exposures to the IMX-101 formulation are indicative of "independent" mixture toxicology. Alternatively, in the IMX-104 exposure both DNAN and RDX were each present at concentrations sufficient to elicit lethality (RDX LC50 = 28.9 mg/L). Toxic-unit calculations for IMX-104 mixture formulation exposures indicated slight synergistic toxicity (ΣTU LC50 = 0.82, 95% confidence interval = 0.73-0.90). Unique functional responses relative to DNAN were observed in the IMX-104 exposure including responses characteristic of RDX exposure. Based on previous transcriptomics responses to acute RDX exposures in fathead minnow larvae, we hypothesize that the potentially synergistic responses within the IMX-104 mixture are related to interactive effects of each DNAN and RDX on oxidative stress mitigation pathways.
2018-03-01 | GSE101888 | GEO
Project description:Abstract — Within the US military, new insensitive munitions (IMs) are rapidly replacing conventional munitions improving safety from unintended detonation. Toxicity data for IM chemicals are expanding rapidly, however IM constituents are typically deployed in mixture formulations, and very little is known about their mixture toxicology. In the present study we sought to characterize the mixture effects and toxicology of the two predominant IM formulations IMX-101 and IMX-104 in acute (48 h) larval fathead minnow (Pimephales promelas) exposures. IMX-101 consists of a mixture of 2,4-dinitroanisole (DNAN), 3-nitro-1,2,4-triazol-5-one (NTO), and nitroguanidine (NQ) while IMX-104 is composed of DNAN, NTO, and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX). DNAN was the most potent constituent in IMX-101 eliciting an LC50 of 36.1 mg/L, whereas NTO and NQ did not elicit significant mortality in exposures up to 1040 and 2640 mg/L, respectively. Toxic unit calculations indicated that IMX-101 elicited toxicity representative of the component concentration of DNAN within the mixture. Toxicogenomic responses for the individual constituents of IMX-101 indicated unique transcriptional expression and functional responses characteristic of: oxidative stress, impaired energy metabolism, tissue damage and inflammatory responses in DNAN exposures; impaired steroid biosynthesis and developmental cell-signaling in NQ exposures; and altered mitogen-activated protein kinase signaling in NTO exposures. Transcriptional responses to the IMX-101 mixture were driven by the effects of DNAN where expression and functional responses were nearly identical comparing DNAN alone versus the fractional equivalent of DNAN within IMX-101. Given that each individual constituent of the IMX-101 mixture elicited unique functional responses, and NTO and NQ did not interact with DNAN within the IMX-101 mixture exposure, the overall toxicity and toxicogenomic responses within acute exposures to the IMX-101 formulation are indicative of "independent" mixture toxicology. Alternatively, in the IMX-104 exposure both DNAN and RDX were each present at concentrations sufficient to elicit lethality (RDX LC50 = 28.9 mg/L). Toxic-unit calculations for IMX-104 mixture formulation exposures indicated slight synergistic toxicity (ΣTU LC50 = 0.82, 95% confidence interval = 0.73-0.90). Unique functional responses relative to DNAN were observed in the IMX-104 exposure including responses characteristic of RDX exposure. Based on previous transcriptomics responses to acute RDX exposures in fathead minnow larvae, we hypothesize that the potentially synergistic responses within the IMX-104 mixture are related to interactive effects of each DNAN and RDX on oxidative stress mitigation pathways.
2018-03-01 | GSE101887 | GEO
Project description:Abstract — Within the US military, new insensitive munitions (IMs) are rapidly replacing conventional munitions improving safety from unintended detonation. Toxicity data for IM chemicals are expanding rapidly, however IM constituents are typically deployed in mixture formulations, and very little is known about their mixture toxicology. In the present study we sought to characterize the mixture effects and toxicology of the two predominant IM formulations IMX-101 and IMX-104 in acute (48 h) larval fathead minnow (Pimephales promelas) exposures. IMX-101 consists of a mixture of 2,4-dinitroanisole (DNAN), 3-nitro-1,2,4-triazol-5-one (NTO), and nitroguanidine (NQ) while IMX-104 is composed of DNAN, NTO, and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX). DNAN was the most potent constituent in IMX-101 eliciting an LC50 of 36.1 mg/L, whereas NTO and NQ did not elicit significant mortality in exposures up to 1040 and 2640 mg/L, respectively. Toxic unit calculations indicated that IMX-101 elicited toxicity representative of the component concentration of DNAN within the mixture. Toxicogenomic responses for the individual constituents of IMX-101 indicated unique transcriptional expression and functional responses characteristic of: oxidative stress, impaired energy metabolism, tissue damage and inflammatory responses in DNAN exposures; impaired steroid biosynthesis and developmental cell-signaling in NQ exposures; and altered mitogen-activated protein kinase signaling in NTO exposures. Transcriptional responses to the IMX-101 mixture were driven by the effects of DNAN where expression and functional responses were nearly identical comparing DNAN alone versus the fractional equivalent of DNAN within IMX-101. Given that each individual constituent of the IMX-101 mixture elicited unique functional responses, and NTO and NQ did not interact with DNAN within the IMX-101 mixture exposure, the overall toxicity and toxicogenomic responses within acute exposures to the IMX-101 formulation are indicative of "independent" mixture toxicology. Alternatively, in the IMX-104 exposure both DNAN and RDX were each present at concentrations sufficient to elicit lethality (RDX LC50 = 28.9 mg/L). Toxic-unit calculations for IMX-104 mixture formulation exposures indicated slight synergistic toxicity (ΣTU LC50 = 0.82, 95% confidence interval = 0.73-0.90). Unique functional responses relative to DNAN were observed in the IMX-104 exposure including responses characteristic of RDX exposure. Based on previous transcriptomics responses to acute RDX exposures in fathead minnow larvae, we hypothesize that the potentially synergistic responses within the IMX-104 mixture are related to interactive effects of each DNAN and RDX on oxidative stress mitigation pathways.
2018-03-01 | GSE101886 | GEO
Project description:Abstract — Within the US military, new insensitive munitions (IMs) are rapidly replacing conventional munitions improving safety from unintended detonation. Toxicity data for IM chemicals are expanding rapidly, however IM constituents are typically deployed in mixture formulations, and very little is known about their mixture toxicology. In the present study we sought to characterize the mixture effects and toxicology of the two predominant IM formulations IMX-101 and IMX-104 in acute (48 h) larval fathead minnow (Pimephales promelas) exposures. IMX-101 consists of a mixture of 2,4-dinitroanisole (DNAN), 3-nitro-1,2,4-triazol-5-one (NTO), and nitroguanidine (NQ) while IMX-104 is composed of DNAN, NTO, and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX). DNAN was the most potent constituent in IMX-101 eliciting an LC50 of 36.1 mg/L, whereas NTO and NQ did not elicit significant mortality in exposures up to 1040 and 2640 mg/L, respectively. Toxic unit calculations indicated that IMX-101 elicited toxicity representative of the component concentration of DNAN within the mixture. Toxicogenomic responses for the individual constituents of IMX-101 indicated unique transcriptional expression and functional responses characteristic of: oxidative stress, impaired energy metabolism, tissue damage and inflammatory responses in DNAN exposures; impaired steroid biosynthesis and developmental cell-signaling in NQ exposures; and altered mitogen-activated protein kinase signaling in NTO exposures. Transcriptional responses to the IMX-101 mixture were driven by the effects of DNAN where expression and functional responses were nearly identical comparing DNAN alone versus the fractional equivalent of DNAN within IMX-101. Given that each individual constituent of the IMX-101 mixture elicited unique functional responses, and NTO and NQ did not interact with DNAN within the IMX-101 mixture exposure, the overall toxicity and toxicogenomic responses within acute exposures to the IMX-101 formulation are indicative of "independent" mixture toxicology. Alternatively, in the IMX-104 exposure both DNAN and RDX were each present at concentrations sufficient to elicit lethality (RDX LC50 = 28.9 mg/L). Toxic-unit calculations for IMX-104 mixture formulation exposures indicated slight synergistic toxicity (ΣTU LC50 = 0.82, 95% confidence interval = 0.73-0.90). Unique functional responses relative to DNAN were observed in the IMX-104 exposure including responses characteristic of RDX exposure. Based on previous transcriptomics responses to acute RDX exposures in fathead minnow larvae, we hypothesize that the potentially synergistic responses within the IMX-104 mixture are related to interactive effects of each DNAN and RDX on oxidative stress mitigation pathways.
2018-03-01 | GSE101870 | GEO
Project description:Abstract: Natural communities of microbes inhabiting amphibian skin, the skin microbiome, are critical to supporting amphibian health and disease resistance. To enable the pro-active health assessment and management of amphibians on Army installations and beyond, we investigated the effects of acute (96h) munitions exposures to Rana pipiens (leopard frog) tadpoles and the associated skin microbiome, integrated with RNAseq-based transcriptomic responses in the tadpole host. Tadpoles were exposed to the legacy munition 2,4,6-trinitrotoluene (TNT), the new insensitive munition (IM) formulation, IMX-101, and the IM constituents nitroguinidine (NQ) and 1-methyl-3-nitroguanidine (MeNQ). The 96h LC50 values and 95% confidence intervals were 2.6 (2.4, 2.8) for ΣTNT and 68.2 (62.9, 73.9) for IMX-101, respectively. The NQ and MeNQ exposures caused no significant impacts on survival in 96h exposures even at maximum exposure levels of 3,560 and 5,285 mg/L, respectively. However, NQ and MeNQ, as well as TNT and IMX-101 exposures, all elicited changes in the tadpole skin microbiome profile, as evidenced by significantly increased relative proportions of the Proteobacteria with increasing exposure concentrations, and significantly decreased alpha-diversity in the NQ exposure. The potential for direct and indirect effects of munitions exposures on the skin microbiome were observed. A direct effect of munitions on microbial flora included the observation of increased relative abundance of the munitions-tolerant, Aeromonadaceae and Pseudomonadaceae, in the NQ exposure. Potential indirect effects on the tadpole skin microbiome resulting from tadpole-host responses to munitions included transcriptional responses indicative of potential changes in skin mucus-layer properties as well as altered production of antimicrobial peptides and innate immune factors. Additional insights into the tadpole host’s transcriptional response to munitions exposures indicated that TNT and IMX-101 exposures each elicited significant enrichment of pathways involved in type-I and type-II xenobiotic metabolism mechanisms where dose-responsive increases in expression were observed. Significant enrichment and increased transcriptional expression of heme and iron binding functions in the TNT exposures was likely connected with known mechanisms of TNT toxicity including hemolytic anemia and methemoglobinemia. The significant enrichment and dose-responsive decrease in transcriptional expression of cell cycle pathways in the IMX-101 exposures was consistent with previous observations in fish, while significant enrichment of immune-related function in response to NQ exposure indicated potential immune suppression at the highest NQ exposure concentration. Finally, the MeNQ exposures elicited significantly decreased transcriptional expression of keratin 16, type I, a gene likely involved in keratinization processes in amphibian skin. Overall, munitions showed the potential to alter tadpole skin microbiome composition and affect transcriptional profiles in the amphibian host, some indicative of potentially impacted host health and immune status, each of which suggest potential implications for munitions exposure on disease susceptibility.
2020-10-01 | GSE154924 | GEO
Project description:The use of a systems biology approach to analyze common and specific mechanisms of liver toxicity induced by munitions compounds TNT, 2,6-DNT, 2,4-DNT, 4A-DNT, and 2A-DNT The munitions compound 2,4,6-trinitrotoluene (TNT), its environmental degradation products 2-amino-4,6-dinitrotoluene (2A-DNT) and 4-amino-2,6-dinitrotoulene (4A-DNT), and two other munitions, 2,4-dinitrotoluene (2,4-DNT) and 2,4-dinitrotoluene (2,6-DNT) contaminate contaminate land, water and retired ammunitions plants. The release of these compounds to the environment is due to military activities and a series of manufacturing processes. Although toxicity has been characterized for these compounds, little is known of their mechanism of action. Here we describe to use an integrative systems biology approach including toxicology, pathology, transcriptomics, metabolomics, gene function classification, pathway analysis and gene network modeling to try to understand the mechanisms of toxicity of these compounds.
2009-12-25 | GSE19628 | GEO
Project description:Abstract — Previous toxicological investigations of the insensitive munition (IM), 3-nitro-1,2,4-triazol-5-one (NTO), demonstrated histopathological and physiological impacts in mammalian testes. The implications of these findings for fish was unknown, therefore we investigated the effects of chronic (21 day) exposures to NTO and an NTO-containing IM formulation called IMX-101 (composed of 2,4-dinitroanisole (DNAN), nitroguanidine (NQ), and NTO) in adult male fathead minnows to assess if impacts on testes were conserved. The NTO exposure caused no significant mortality through the maximum exposure concentration (720 mg/L, measured), however NTO elicited testicular impacts causing significant asynchrony in spermatogenesis and necrosis in secondary spermatocytes at the two highest exposure concentrations (383 mg/L and 720 mg/L) and testicular degeneration at the highest exposure. Microarray-based transcriptomics analysis identified significant enrichment of steroid metabolism pathways and mTORC-signal control of spermatogonia differentiation in NTO exposures each having logical connections to observed asynchronous spermatogenesis. Additionally, NTO impaired transcriptional expression for genes supporting sperm structural and flagellar development including sperm-associated antigen 6 (Spag6). These functional transcriptomic responses are logical contributors underlying impacted reproductive physiology in NTO exposures that ultimately lead to reductions in spermatozoa. In contrast to NTO, the IMX-101 formulation elicited significant mortality at the two highest exposure concentrations of 25.2 and 50.9 mg/L (DNAN nominal + NTO measured + NQ measured). Unlike NTO and NQ, the DNAN component of the IMX-101 formulation underwent significant transformation in the 21d exposure. From previous investigations, neither NTO or NQ caused mortality in fish at >1000 mg/L suggesting that mortality in the present study arose from DNAN / DNAN-attributable transformation products. The 12.6 mg/L IMX-101 exposure caused significant sublethal impacts on testes including sperm necrosis, interstitial fibrosis, and Sertoli-like cell hyperplasia. Transcriptional profiles for IMX-101 indicated significant enrichment on multiple signaling pathways supporting spermatogenesis, mitosis / meiosis, and flagellar structure, all logically connected to observed sperm necrosis. Additionally, pronounced transcriptional increases within the PPARα-RXRα pathway, a known DNAN target, has been hypothesized to correspond to Sertoli cell hyperplasia, presumably as a compensatory response to fulfill the nurse-function of Sertoli cells during spermatogenesis. Overall, the transcriptional results indicated unique molecular responses for NTO and IMX-101. Regarding chemical hazard, NTO impacted testes and impaired spermatogenesis, but at high exposure concentrations (≥ 192 mg/L), whereas the IMX-101 formulation, elicited mortality and impacts on reproductive physiology likely caused by DNAN and its transformation products present at concentrations well below the NTO-component concentration within the IMX-101 mixture formulation. see previous row.
2019-07-01 | GSE127878 | GEO
Project description:Abstract — Previous toxicological investigations of the insensitive munition (IM), 3-nitro-1,2,4-triazol-5-one (NTO), demonstrated histopathological and physiological impacts in mammalian testes. The implications of these findings for fish was unknown, therefore we investigated the effects of chronic (21 day) exposures to NTO and an NTO-containing IM formulation called IMX-101 (composed of 2,4-dinitroanisole (DNAN), nitroguanidine (NQ), and NTO) in adult male fathead minnows to assess if impacts on testes were conserved. The NTO exposure caused no significant mortality through the maximum exposure concentration (720 mg/L, measured), however NTO elicited testicular impacts causing significant asynchrony in spermatogenesis and necrosis in secondary spermatocytes at the two highest exposure concentrations (383 mg/L and 720 mg/L) and testicular degeneration at the highest exposure. Microarray-based transcriptomics analysis identified significant enrichment of steroid metabolism pathways and mTORC-signal control of spermatogonia differentiation in NTO exposures each having logical connections to observed asynchronous spermatogenesis. Additionally, NTO impaired transcriptional expression for genes supporting sperm structural and flagellar development including sperm-associated antigen 6 (Spag6). These functional transcriptomic responses are logical contributors underlying impacted reproductive physiology in NTO exposures that ultimately lead to reductions in spermatozoa. In contrast to NTO, the IMX-101 formulation elicited significant mortality at the two highest exposure concentrations of 25.2 and 50.9 mg/L (DNAN nominal + NTO measured + NQ measured). Unlike NTO and NQ, the DNAN component of the IMX-101 formulation underwent significant transformation in the 21d exposure. From previous investigations, neither NTO or NQ caused mortality in fish at >1000 mg/L suggesting that mortality in the present study arose from DNAN / DNAN-attributable transformation products. The 12.6 mg/L IMX-101 exposure caused significant sublethal impacts on testes including sperm necrosis, interstitial fibrosis, and Sertoli-like cell hyperplasia. Transcriptional profiles for IMX-101 indicated significant enrichment on multiple signaling pathways supporting spermatogenesis, mitosis / meiosis, and flagellar structure, all logically connected to observed sperm necrosis. Additionally, pronounced transcriptional increases within the PPARα-RXRα pathway, a known DNAN target, has been hypothesized to correspond to Sertoli cell hyperplasia, presumably as a compensatory response to fulfill the nurse-function of Sertoli cells during spermatogenesis. Overall, the transcriptional results indicated unique molecular responses for NTO and IMX-101. Regarding chemical hazard, NTO impacted testes and impaired spermatogenesis, but at high exposure concentrations (≥ 192 mg/L), whereas the IMX-101 formulation, elicited mortality and impacts on reproductive physiology likely caused by DNAN and its transformation products present at concentrations well below the NTO-component concentration within the IMX-101 mixture formulation. see previous row.
2019-06-21 | GSE127877 | GEO