Bioactivity guided isolation and identification of phenolic compounds from Citrus aurantium L. with anti-colorectal cancer cells activity by UHPLC-Q-TOF/MS.

Natural plants are rich sources of various bioactive compounds. Consequently, the efficiently isolation of these bioactive components has always attracted considerable attention. Our work aims to demonstrate a framework for bioactivity guided isolation of potential effective compounds from the complex food materials. We demonstrated its application for isolation of phenolic compounds with anti-proliferative activity against colorectal cancer cells (CRCs) from Citrus aurantium L. Firstly, phenolic rich fraction was successfully identified as the main effective components that could simultaneously suppress the growth of CRCs and inhibit Wnt signaling. In order to obtain the bioactive phenolic constituents, a detailed study was performed by optimizing the purification conditions. Two phenolic rich fractions (40% and 60% ethanol elution fractions) were then obtained by AB-8 macroporous resins under optimized condition. Finally, the main components (65 compounds) were tentatively identified from the 40% ethanol eluant by ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF/MS) analysis. Notably, there were five of the phytochemicals (Feruloylagmatine, Haploside C, Sagittatin A, Linderagalactone C and Koparin-2'-methyl ether) which were hitherto unidentified in Citrus aurantium L. fruit. In conclusion, this study showed that under the principle of bioactivity guided strategy, phenolic constituents with potential anti-CRCs activity were isolated from Citrus aurantium L.

Food-derived natural compounds in the management of chronic diseases via Wnt signaling pathway.

Wnt signaling pathway is an evolutionarily conserved pathway that control embryonic development, adult tissue homeostasis, and pathological processes of organisms throughout life. However, dysregulation of the Wnt signaling is associated with the occurrence of chronic diseases. In comparison with the application of chemical drugs as traditional treatment for chronic diseases, dietary agents have unique advantages, such as less side effects, multiple targets, convenience in accessibility and higher acceptability in long-term intervention. In this review, we summarized current progress in manipulating the Wnt signaling using food components and its benefits in managing chronic diseases. The underlying mechanisms of bioactive food components in the management of the disease progression via the Wnt signaling was illustrated. Then, the review focused on the function of dietary pattern (which might act via combination of foods with multiple nutrients or food ingredients) on targeting Wnt signaling at multiple level. The potential caveats and challenges in developing new strategy via modulating Wnt-associated diseases with food-based agents and appropriate dietary pattern are also discussed in detail. This review shed light on the understanding of the regulatory effect of food bioactive components on chronic diseases management through the Wnt signaling, which can be expanded to other specific signaling pathway associated with disease.

Dependence of Graphene Oxide (GO) Toxicity on Oxidation Level, Elemental Composition, and Size.

The mass production of graphene oxide (GO) unavoidably elevates the chance of human exposure, as well as the possibility of release into the environment with high stability, raising public concern as to its potential toxicological risks and the implications for humans and ecosystems. Therefore, a thorough assessment of GO toxicity, including its potential reliance on key physicochemical factors, which is lacking in the literature, is of high significance and importance. In this study, GO toxicity, and its dependence on oxidation level, elemental composition, and size, were comprehensively assessed. A newly established quantitative toxicogenomic-based toxicity testing approach, combined with conventional phenotypic bioassays, were employed. The toxicogenomic assay utilized a GFP-fused yeast reporter library covering key cellular toxicity pathways. The results reveal that, indeed, the elemental composition and size do exert impacts on GO toxicity, while the oxidation level exhibits no significant effects. The UV-treated GO, with significantly higher carbon-carbon groups and carboxyl groups, showed a higher toxicity level, especially in the protein and chemical stress categories. With the decrease in size, the toxicity level of the sonicated GOs tended to increase. It is proposed that the covering and subsequent internalization of GO sheets might be the main mode of action in yeast cells.

Comparative and mechanistic toxicity assessment of structure-dependent toxicity of carbon-based nanomaterials.

The wide application of carbon-based nanomaterials (CNMs) has resulted in the ubiquity of CNMs in the natural environment and they potentially impose adverse consequences on ecosystems and human health. In this study, we comprehensively evaluated and compared potential toxicological effects and mechanisms of seven CNMs in three representative types (carbon blacks, graphene nanoplatelets, and fullerenes), to elucidate the correlation between their physicochemical/structural properties and toxicity. We employed a recently-developed quantitative toxicogenomics-based toxicity testing system with GFP-fused yeast reporter library targeting main cellular stress response pathways, as well as conventional phenotype-based bioassays. The results revealed that DNA damage, oxidative stress, and protein stress were the major mechanisms of action for all the CNMs at sub-cytotoxic concentration levels. The molecular toxicity nature were concentration-dependent, and they exhibited both similarity within the same structural group and distinctiveness among different CNMs, evidencing the structure-driven toxicity of CNMs. The toxic potential based on toxicogenomics molecular endpoints revealed the remarkable impact of size and structure on the toxicity. Furthermore, the phenotypic endpoints derived from conventional phenotype-based bioassays correlated with quantitative molecular endpoints derived from the toxicogenomics assay, suggesting that the selected protein biomarkers captured the main cellular effects that are associated with phenotypic adverse outcomes.

A Slc25a46 Mouse Model Simulating Age-Associated Motor Deficit, Redox Imbalance, and Mitochondria Dysfunction.

The mitochondrial theory of aging postulates that accumulation of mtDNA mutations and mitochondrial dysfunction are responsible for producing aging phenotypes. To more comprehensively explore the complex relationship between aging and mitochondria dysfunction, we have developed a mouse model with Slc25a46 knockout, a nuclear gene described as encoding mitochondrial carriers, by CRISPR/Cas9 gene editing to mimic some typical aging phenotypes in human. Slc25a46-/- mice present segmental premature aging phenotypes characterized by shortened life span of no more than 2 months, obviously defective motor ability, gastrocnemius muscle atrophy, and imbalance of redox level in brain and liver. The underlying mechanism for multiple organ disorder may attribute to mitochondrial dysfunction, which is mainly manifested in the damaged mitochondrial structure (eg, vacuolar structure, irregular swelling, and disorganized cristae) and an age-associated decrease in respiratory chain enzyme (mainly complex I and IV) activity. In summary, our study suggests that the Slc25a46-/- mouse is a valid animal model for segmental aging-related pathologies studies based on mitochondrial theory, generating a new platform to both understand mechanisms between aging and mitochondria dysfunction as well as to design mitochondria-based therapeutic strategies to improve mitochondrial quality, and thereby the overall healthspan.

Bioactivity-Oriented Purification of Polyphenols from Cinnamomum cassia Presl. with Anti-Proliferation Effects on Colorectal Cancer Cells.

Cinnamomum cassia Presl. (CCP) is a popular natural spice possessing various pharmacological properties. We obtained polyphenol-rich fraction (CCP-P) from CCP by bioactivity-oriented purification method and evaluated its Wnt signaling inhibition activity. Firstly, the phenolic components were identified as the main bioactive compounds with anti-colorectal cancer activity. Then, we compared the anti-colorectal cancer activity of CCP extract obtained from different solvent by cell morphology alteration and EdU assay. Ethanol extract showed higher antiproliferative activity compared to water extract on HCT116 cells, with proliferating cells reducing to 41.12 and 21.83% at 156.00 µg GAE/mL, respectively. Next, separation and enrichment of polyphenols from ethanol extract was performed on AB-8 macroporous resins under optimal conditions. Further evaluation of the CCP-P bioactivity revealed that it exerted more potent antiproliferative activity on RKO and HCT116 cells, showing higher selectivity for Wnt-dependent colorectal cancer cells (CRCs). Ten major polyphenols were identified in the CCP-P by UPLC-ESI-MS/MS. In summary, this study presents evidence that CCP-derived polyphenols are promising potential candidates as functional food ingredients against CRC.

Impact of heavy metals on the formation and properties of solvable microbiological products released from activated sludge in biological wastewater treatment.

This study investigated the acute impact of heavy metals on activated sludge with respect to the amount properties of biopolymers and other solvable microbiological products (SMPs) released from the sludge. Ten heavy metals were selected for the evaluation. Under the experimental conditions, exposing activated sludge to different metals led to an increase in SMPs, with a more significant increase in nitrogenous organics than in carbonaceous ones, where Hg2+, Ag+, Cu2+, and Cr6+ led to the highest increase in SMP species, while Cd2+, Ni2+, Mn2+, Pb2+, and Co2+ caused limited increase in the middle and small SMP molecules, and Zn2+ and Cr3+ resulted in a decrease in SMP content. To probe the molecular impact of heavy metals and the association between cellular stress and SMP formation, the toxicity of heavy metals was evaluated using a toxicogenomics assay. Based on a correlation analysis between the increase in SMP and the molecular toxicity index-transcriptional effect level index (TELI) of different genes under corresponding stress conditions, eight genes demonstrated a strong correlation with SMP properties and were pre-assumed to have the most significant influence on the increment in SMPs. We further validated the correlation equation established to predict SMP production based on the molecular disturbance of the eight key biomarkers, using arsenic As3+ and vanadium V5+ as tests, and by quantifying the amount of SMPs released from the activated sludge under the influence of these metals using a TELI-derived equation. In addition, the heavy metals that generated greater amounts of reactive oxygen species also caused larger increases in SMPs.

Toxicity of Single-Walled Carbon Nanotubes (SWCNTs): Effect of Lengths, Functional Groups and Electronic Structures Revealed by a Quantitative Toxicogenomics Assay.

Single-walled carbon nanotubes (SWCNTs) are a group of widely used carbon-based nanomaterials (CNMs) with various applications, which raise increasing public concerns associated with their potential toxicological effect and risks on human and ecosystems. In this report, we comprehensively evaluated the nanotoxicity of SWCNTs with their relationship to varying lengths, functional groups and electronic structures, by employing both newly established quantitative toxicogenomics test, as well as conventional phenotypic bioassays. The objective is to reveal potential cellular toxicity and mechanisms of SWCNTs at the molecular level, and to probe their potential relationships with their morphological, surface, and electronic properties. The results indicated that DNA damage and oxidative stress were the dominant mechanisms of action for all SWCNTs and, the toxicity level and characteristics varied with length, surface functionalization and electronic structure. Distinguishable molecular toxicity fingerprints were revealed for the two SWCNTs with varying length, with short SWCNT exhibiting higher toxicity level than the long one. In terms of surface properties, SWCNT functionalization, namely carboxylation and hydroxylation, led to elevated overall toxicity, especially genotoxicity, as compared to unmodified SWCNT. Carboxylated SWCNT induced a greater toxicity than the hydroxylated SWCNT. The nucleus is likely the primary target site for long, short, and carboxylated SWCNTs and mechanical perturbation is likely responsible for the DNA damage, specifically related to degradation of the DNA double helix structure. Finally, dramatically different electronic structure-dependent toxicity was observed with metallic SWCNT exerting much higher toxicity than the semiconducting one that exhibited minimal toxicity among all SWCNTs.

The response of Escherichia coli to the alkylating agents chloroacetaldehyde and styrene oxide.

DNA damage is ubiquitous and can arise from endogenous or exogenous sources. DNA-damaging alkylating agents are present in environmental toxicants as well as in cancer chemotherapy drugs and are a constant threat, which can lead to mutations or cell death. All organisms have multiple DNA repair and DNA damage tolerance pathways to resist the potentially negative effects of exposure to alkylating agents. In bacteria, many of the genes in these pathways are regulated as part of the SOS reponse or the adaptive response. In this work, we probed the cellular responses to the alkylating agents chloroacetaldehyde (CAA), which is a metabolite of 1,2-dichloroethane used to produce polyvinyl chloride, and styrene oxide (SO), a major metabolite of styrene used in the production of polystyrene and other polymers. Vinyl chloride and styrene are produced on an industrial scale of billions of kilograms annually and thus have a high potential for environmental exposure. To identify stress response genes in E. coli that are responsible for tolerance to the reactive metabolites CAA and SO, we used libraries of transcriptional reporters and gene deletion strains. In response to both alkylating agents, genes associated with several different stress pathways were upregulated, including protein, membrane, and oxidative stress, as well as DNA damage. E. coli strains lacking genes involved in base excision repair and nucleotide excision repair were sensitive to SO, whereas strains lacking recA and the SOS gene ybfE were sensitive to both alkylating agents tested. This work indicates the varied systems involved in cellular responses to alkylating agents, and highlights the specific DNA repair genes involved in the responses.

Genotoxicity Assessment of Drinking Water Disinfection Byproducts by DNA Damage and Repair Pathway Profiling Analysis.

Genotoxicity is considered a major concern for drinking water disinfection byproducts (DBPs). Of over 700 DBPs identified to date, only a small number has been assessed with limited information for DBP genotoxicity mechanism(s). In this study, we evaluated genotoxicity of 20 regulated and unregulated DBPs applying a quantitative toxicogenomics approach. We used GFP-fused yeast strains that examine protein expression profiling of 38 proteins indicative of all known DNA damage and repair pathways. The toxicogenomics assay detected genotoxicity potential of these DBPs that is consistent with conventional genotoxicity assays end points. Furthermore, the high-resolution, real-time pathway activation and protein expression profiling, in combination with clustering analysis, revealed molecular level details in the genotoxicity mechanisms among different DBPs and enabled classification of DBPs based on their distinct DNA damage effects and repair mechanisms. Oxidative DNA damage and base alkylation were confirmed to be the main molecular mechanisms of DBP genotoxicity. Initial exploration of QSAR modeling using moleular genotoxicity end points (PELI) suggested that genotoxicity of DBPs in this study was correlated with topological and quantum chemical descriptors. This study presents a toxicogenomics-based assay for fast and efficient mechanistic genotoxicity screening and assessment of a large number of DBPs. The results help to fill in the knowledge gap in the understanding of the molecular mechanisms of DBP genotoxicity.

A Quantitative Toxicogenomics Assay for High-throughput and Mechanistic Genotoxicity Assessment and Screening of Environmental Pollutants.

The ecological and health concern of mutagenicity and carcinogenicity potentially associated with an overwhelmingly large and ever-increasing number of chemicals demands for cost-effective and feasible method for genotoxicity screening and risk assessment. This study proposed a genotoxicity assay using GFP-tagged yeast reporter strains, covering 38 selected protein biomarkers indicative of all the seven known DNA damage repair pathways. The assay was applied to assess four model genotoxic chemicals, eight environmental pollutants and four negative controls across six concentrations. Quantitative molecular genotoxicity end points were derived based on dose response modeling of a newly developed integrated molecular effect quantifier, Protein Effect Level Index (PELI). The molecular genotoxicity end points were consistent with multiple conventional in vitro genotoxicity assays, as well as with in vivo carcinogenicity assay results. Further more, the proposed genotoxicity end point PELI values quantitatively correlated with both comet assay in human cell and carcinogenicity potency assay in mice, providing promising evidence for linking the molecular disturbance measurements to adverse outcomes at a biological relevant level. In addition, the high-resolution DNA damaging repair pathway alternated protein expression profiles allowed for chemical clustering and classification. This toxicogenomics-based assay presents a promising alternative for fast, efficient and mechanistic genotoxicity screening and assessment of drugs, foods, and environmental contaminants.

Toxicity mechanisms identification via gene set enrichment analysis of time-series toxicogenomics data: impact of time and concentration.

The advance in high-throughput "toxicogenomics" technologies, which allows for concurrent monitoring of cellular responses globally upon exposure to chemical toxicants, presents promises for next-generation toxicity assessment. It is recognized that cellular responses to toxicants have a highly dynamic nature, and exhibit both temporal complexity and dose-response shifts. Most current gene enrichment or pathway analysis lack the recognition of the inherent correlation within time series data, and may potentially miss important pathways or yield biased and inconsistent results that ignore dynamic patterns and time-sensitivity. In this study, we investigated the application of two score metrics for GSEA (gene set enrichment analysis) to rank the genes that consider the temporal gene expression profile. One applies a novel time series CPCA (common principal components analysis) to generate scores for genes based on their contributions to the common temporal variation among treatments for a given chemical at different concentrations. Another one employs an integrated altered gene expression quantifier-TELI (transcriptional effect level index) that integrates altered gene expression magnitude over the exposure time. By comparing the GSEA results using two different ranking metrics for examining the dynamic responses of reporter cells treated with various dose levels of three model toxicants, mitomycin C, hydrogen peroxide, and lead nitrate, the analysis identified and revealed different toxicity mechanisms of these chemicals that exhibit chemical-specific, as well as time-aware and dose-sensitive nature. The ability, advantages, and disadvantages of varying ranking metrics were discussed. These findings support the notion that toxicity bioassays should account for the cells' complex dynamic responses, thereby implying that both data acquisition and data analysis should look beyond simple traditional end point responses.

Comparative and mechanistic genotoxicity assessment of nanomaterials via a quantitative toxicogenomics approach across multiple species.

This study reports a comparative and mechanistic genotoxicity assessment of four engineered nanomaterials (ENMs) across three species, including E. coli, yeast, and human cells, with the aim to reveal the distinct potential genotoxicity mechanisms among the different nanomaterials and their association with physiochemical features. Both the conventional phenotypic alkaline comet test and the newly developed quantitative toxicogenomics assay, that detects and quantifies molecular level changes in the regulation of six DNA damage repair pathways, were employed. The proposed molecular endpoints derived from the toxicogenomics assays, namely TELI (Transcriptional Effect Level Index) and PELI (Protein Effect Level Index), correlated well with the phenotypic DNA damage endpoints from comet tests, suggesting that the molecular genotoxicity assay is suitable for genotoxicity detection. Temporal altered gene or protein expression profiles revealed various potential DNA damage types and relevant genotoxic mechanisms induced by the tested ENMs. nTiO2_a induced a wide spectrum of DNA damage consistently across three species. Three carbon-based ENMs, namely carbon black, single wall carbon nanotube (SWCNT) and fullerene, exhibited distinct, species and ENM property-dependent DNA damage mechanisms. All carbon based ENMs induced relatively weak DNA damage repair response in E. coli, but more severe DNA double strand break in eukaryotes. The differences in cellular structure and defense systems among prokaryotic and eukaryotic species lead to distinct susceptibility and mechanisms for ENM uptake and, thus, varying DNA damages and repair responses. The observation suggested that eukaryotes, especially mammalian cells, are likely more susceptible to genotoxicity than prokaryotes in the ecosystem when exposed to these ENMs.

A quantitative toxicogenomics assay reveals the evolution and nature of toxicity during the transformation of environmental pollutants.

The incomplete mineralization of contaminants of emerging concern (CECs) during the advanced oxidation processes can generate transformation products that exhibit toxicity comparable to or greater than that of the original contaminant. In this study, we demonstrated the application of a novel, fast, and cost-effective quantitative toxicogenomics-based approach for the evaluation of the evolution and nature of toxicity along the electro-Fenton oxidative degradation of three representative CECs whose oxidative degradation pathways have been relatively well studied, bisphenol A, triclosan, and ibuprofen. The evolution of toxicity as a result of the transformation of parent chemicals and production of intermediates during the course of degradation are monitored, and the quantitative toxicogenomics assay results revealed the dynamic toxicity changes and mechanisms, as well as their association with identified intermediates during the electro-Fenton oxidation process of the selected CECs. Although for the three CECs, a majority (>75%) of the parent compounds disappeared at the 15 min reaction time, the nearly complete elimination of toxicity required a minimal 30 min reaction time, and they seem to correspond to the disappearance of identified aromatic intermediates. Bisphenol A led to a wide range of stress responses, and some identified transformation products containing phenolic or quinone group, such as 1,4-benzoquinone and hydroquinone, likely contributed to the transit toxicity exhibited as DNA stress (genotoxicity) and membrane stress during the degradation. Triclosan is known to cause severe oxidative stress, and although the oxidative damage potential decreased concomitantly with the disappearance of triclosan after a 15 min reaction, the sustained toxicity associated with both membrane and protein stress was likely attributed at least partially to the production of 2,4-dichlorophenol that is known to cause the production of abnormal proteins and affect the cell membrane. Ibuprofen affects the cell transporter function and exhibited significantly high membrane stress related to both membrane structure and function. Oxidative degradation of ibuprofen led to a shift in its toxicity profile from mainly membrane stress to one that exhibited not only sustained membrane stress but also protein stress and DNA stress. The information-rich and high-resolution toxicogenomics results served as "fingerprints" that discerned and revealed the toxicity mechanism at the molecular level among the CECs and their oxidation transformation products. This study demonstrated that the quantitative toxicogenomics assay can serve as a useful tool for remediation technology efficacy assessment and provide guidance about process design and optimization for desired toxicity elimination and risk reduction.

Efficient degradation of contaminants of emerging concerns by a new electro-Fenton process with Ti/MMO cathode.

The performance of a new electro-Fenton process with Ti-based mixed metal oxides (Ti/MMO) cathode, a dimensionally stable electrode, is evaluated for degrading contaminants of emerging concerns (CECs) in aqueous solutions. Bisphenol A (422µgL(-1)) is completely degraded in an undivided cell using Ti/MMO as the cathode in the presence of 6.9mgL(-1) Fe(2+) within 20min under conditions of pH 4 and 25mA. Both bisphenol A degradation and H2O2 production increase with decreasing solution pH and increasing current. Ti/MMO cathode is effective for reducing O2 to H2O2 and regenerating Fe(2+) from Fe(3+). OH radicals are validated to be the predominant reactive oxygen species (ROS) contributing to bisphenol A degradation. This new electro-Fenton process is also effective for degrading bisphenol A, triclosan and ibuprofen even at a relatively high concentration of 5mgL(-1). The partial removals of total organic carbon suggest a moderate extent of mineralization. The transformation pathways of the three CECs are proposed based on the intermediates identified by HPLC and GC-MS, showing that CECs are mainly transformed to nontoxic aliphatic acids. This study demonstrates that Ti/MMO can be used as the cathode in the electro-Fenton process for degrading CECs at trace levels in waters.

Analyzing high dimensional toxicogenomic data using consensus clustering.

Rapid development of high-throughput toxicogenomics technologies has created new approaches to screen environmental samples for mechanistic toxicity assessment. However, challenges remain in the analysis, especially clustering of the resulting high-dimensional data. Because of the lack of commonly accepted validation methods, it is difficult to compare clustering results between studies or to identify the key experimental or data features that impact the clustering results. We applied consensus clustering (CC), an approach that clusters the input data repeatedly through iterative resampling, and identifies frequently occurring high-confidence clusters. We used CC to analyze a set of high dimensional transcriptomics data with temporal resolution, which were generated using our E. coli whole-cell array system for a diverse variety of toxicants at different dose concentrations. The CC analysis allowed us to evaluate the clustering results' robustness and sensitivity against a number of conditions that represent the common variations in high-throughput experiments, including noisy data, subsets of treatments, subsets of reporter genes, and subsets of time points. We demonstrated the value of utilizing rich time-series data and underscored the importance of careful selection of sampling times for a given experimental system. The results also indicated that temporal data compression using our proposed Transcriptional Effect Level Index (TELI) concept followed by CC largely conserved the cluster resolution. We also found that for our cellular stress response ensemble-based high-throughput transcriptomics assay platform, the size and composition of the reporter gene set are critical factors that affect the resulting coherency of clusters. Taken together, these results demonstrated that more robust consensus clustering such as CC may be valuable in analyzing high-dimensional toxicogenomic data sets.

A new Transcriptional Effect Level Index (TELI) for toxicogenomics-based toxicity assessment.

This study proposes and demonstrates the potential application of a new Transcriptional Effect Level Index (TELI) to convert the information-rich toxicogenomic data into integrated and quantitative endpoints. A library of transcriptional fusions of green fluorescent protein (GFP) that includes different promoters for 91 stress-related genes in E. coli K12, MG1655 is employed to evaluate the gene expression alteration induced by exposure to four nanomaterials (NMs), nano silver (nAg), nano titanium dioxide anatase (nTiO2_a), nano titanium dioxide rutile (nTiO2_r), and fullerene soot. TELI is determined for each toxicogenomic assay, and it incorporates the number and identity of genes that had altered expression, the magnitude of alteration, and the temporal pattern of gene expression change in response to toxicant exposure. TELI values exhibit a characteristic "sigmoid" shaped toxicity dose-response curve, based on which TELI(MAX) (the maximal value of TELI), TELI50 (concentration that yields half of TELI(MAX)), NOTEL(TELI) (TELI-based no observed transcriptional effect level), and Slope(TELI) (the slope of TELI-dose response curve) are obtained. TELI-based endpoints are compared to currently used endpoints such as EC50 and no observed transcriptional effect level (NOTEL). The agreement of NOTEL(TELI) and NOTEL values validates the concept and application of TELI. Multiple endpoints derived from TELI can describe the dose response behavior and characteristics more completely and holistically than single points such as NOTEL alone. TELI values determined for genes in each stress response category (e.g., oxidative stress, DNA repair) indicate mode of action (MOA)-related comparative transcriptional level toxicity among compounds, and it reveals detailed information of toxic response pathways such as different DNA damage and repair mechanisms among the NMs. This study presents a methodology for converting the rich toxicogenomic information into a readily usable and transferable format that can be potentially linked to regulation endpoints and incorporated into a decision-making framework.

Mechanistic toxicity assessment of nanomaterials by whole-cell-array stress genes expression analysis.

This study performed mechanistic toxicity assessment of nanosilver (nAg) and nanotitanium dioxide anatase (nTiO2_a) via toxicogenomic approach, employing a whole-cell-array library consisting of 91 recombinated Escherichia coli K12 strains with transcriptional GFP-fusions covering most known stress response genes. The results, for the first time, revealed more detailed transcriptional information on the toxic mechanism of nAg and nTiO2_a, and led to a better understanding of the mode of action (MOA) of metal and metal oxide nanomaterials (NMs). The detailed pathways network established for the oxidative stress system and for the SOS (DNA damage) repair system based on the temporal gene expression profiling data revealed the relationships and sequences of key genes involved in these toxin response systems. Both NMs were found to cause oxidative stress as well as cell membrane and transportation damage. Genotoxicity and DNA damage were also observed, although nTiO2_a induced SOS response via previously identified pathway and nAg seemed to induce DNA repair via a pathway different from SOS. We observed that the NMs at lower concentration tend to induce more chemical-specific toxicity response, while at higher concentrations, more general global stress response dominates. The information-rich real-time gene expression data allowed for identification of potential biomarkers that can be employed for specific toxin detection and biosensor developments. The concentration-dependent gene expression response led to the determination of the No Observed Transcriptional Effect Level (NOTEL) values, which can be potentially applied in the regulatory and risk assessment framework as an alternative toxicity assessment end point.