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A rapid and sensitive method to determine the characteristics of carcinogens is needed. In this study, we used a microarray-based genomics approach, with a short-term in vivo model, in combination with insights from statistical and mechanistic analyses to determine the characteristics of carcinogens. Carcinogens were evaluated based on the differen...
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... each treatment type (single and multiple), 10 compounds were administered, and the experi- ment was repeated in triplicate. For DEG selection and mechanis- tic studies, a total of 41 data points (three GTX carcinogens, three NGTX carcinogens and their controls) were used, and the rest (24 data points, four test carcinogens) of the data points were used only in hierarchical clustering as a test set ( Figure 1). ...
Citations
... Multiple studies have reported promising results in differentiating between subclasses of carcinogenicity [4,5] and predicting hepatotoxicity of a novel compound using genomic signatures of human in vivo disease states [6][7][8][9][10][11][12][13]. Nevertheless, in vitro assays are not without their limitations. ...
... In recent years toxicogenomic assays have achieved notable success in predicting the hepatotoxicity of a novel compound [4][5][6][7][8][9][10][11][12][13]. However, relating changes in gene expression profiles from these rodent and cell line assays to relevant human outcomes still proves challenging. ...
The liver is the primary site for the metabolism and detoxification of many compounds, including pharmaceuticals. Consequently, it is also the primary location for many adverse reactions. As the liver is not readily accessible for sampling in humans; rodent or cell line models are often used to evaluate potential toxic effects of a novel compound or candidate drug. However, relating the results of animal and in vitro studies to relevant clinical outcomes for the human in vivo situation still proves challenging. In this study, we incorporate principles of transfer learning within a deep artificial neural network allowing us to leverage the relative abundance of rat in vitro and in vivo exposure data from the Open TG-GATEs data set to train a model to predict the expected pattern of human in vivo gene expression following an exposure given measured human in vitro gene expression. We show that domain adaptation has been successfully achieved, with the rat and human in vitro data no longer being separable in the common latent space generated by the network. The network produces physiologically plausible predictions of human in vivo gene expression pattern following an exposure to a previously unseen compound. Moreover, we show the integration of the human in vitro data in the training of the domain adaptation network significantly improves the temporal accuracy of the predicted rat in vivo gene expression pattern following an exposure to a previously unseen compound. In this way, we demonstrate the improvements in prediction accuracy that can be achieved by combining data from distinct domains.
... Alkylating agents are members of a structurally diverse group of DNA damaging compounds which form adducts at ring Nitrogen and extra-cyclic Oxygen atoms of DNA bases (Bodakuntla et al., 2014) N-Nitroso-N-ethyl urea (NEU), an ethylating agent has been described as a very potent transplacental teratogen and carcinogen in rodents (Bodakuntla et al., 2014) Human carcinogens could be genotoxic, which means to directly alter the genetic material of target cells (Mulware, 2013). Other carcinogens are classified as non-genotoxic, in which case are capable of producing cancer by some secondary mechanism and not by direct gene damage (Hernández et al., 2009) (Lee et al., 2013. These secondary mechanisms include the abilities of an agent to alter DNA repair or cause genomic instability, epigenetic alterations, oxidative stress, chronic inflammation, immunosuppression, immortalization or alter cell proliferation, cell death, or nutrient supply (Smith et al., 2016). ...
The effect of carcinogen exposure has been established to alter biochemical and hematological profiles in humans. Secondary metabolites from plant sources like flavonoids, alkaloids, saponins, and others have been found to be potent anticancer agents and also investigated to protect against the effect of environmental carcinogens. Female albino rats were rats exposed to freshly prepared 3 fractionated doses of 50 mg/kg N-Nitroso-N-ethyl urea (carcinogen) dissolved in 1/15 M Phosphate buffer, given intraperitoneally and treated with varying concentrations of Senna alata plant aqueous extracts. At the end of the test period, hematological and biochemical parameters were determined in blood and serum samples. Compared to the control group, the carcinogen exposed rats treated group showed significances in several hematological parameters, including decreases in White blood cell (WBC), Red blood cell (RBC), and Platelet (PLT) counts. Furthermore, in comparison to the control group, the carcinogen exposed rats showed significantly increased blood glucose, serum total cholesterol, Low density lipoprotein (LDL-cholesterol), triacylglycerols levels and High density lipoprotein (HDL-cholesterol) level. The hematological and biochemical parameters in the carcinogen exposed rats treated group were approximately similar to control group. The S. alata extract significantly (P<0.05) restored the hematological and biochemical parameters in N-Nitroso-N-ethyl urea carcinogen exposed rats
... As there are no validated in vitro methods to identify NGTX chemicals, many of those substances remain unidentified (Felter et al. 2020(Felter et al. , 2021. In an attempt to address the limitations of the standard regulatory testing procedures for carcinogenicity assessment, several studies have searched for alternatives with better predictability and enhanced human relevance (Watanabe et al. 2012;Lee et al. 2013;Cohen 2017;Wilde et al. 2018;Batke et al. 2021). The development of human-based in vitro models that specifically and reliably detect NGTX chemicals is indeed increasingly being explored (Jacobs et al. 2016). ...
... Camptothecin (10 μM) was used as a positive control. Each graph presents at least 3 experimental replicates with each experiment comprising 3 technical replicates inflammation, oxidative stress, immune suppression, epigenetic silencing, or other changes in biological processes, leading to aberrant cellular signaling and genomic instability which are predominant to their carcinogenic potential (Lee et al. 2013;Oku et al. 2022). Additionally, many NGTX chemicals are likely to be tumor-promoting compounds (Ohmori et al. 2022). ...
Validated in vitro assays for testing non-genotoxic carcinogenic potential of chemicals are currently not available. Consequently, the two-year rodent bioassay remains the gold standard method for the identification of these chemicals. Transcriptomic and proteomic analyses have provided a comprehensive understanding of the non-genotoxic carcinogenic processes, however, functional changes induced by effects at transcriptional and translational levels have not been addressed. The present study was set up to test a number of proposed in vitro biomarkers of non-genotoxic hepatocarcinogenicity at the functional level using a translational 3-dimensional model. Spheroid cultures of human hepatocytes and stellate cells were exposed to 5 genotoxic carcinogenic, 5 non-genotoxic carcinogenic, and 5 non-carcinogenic chemical compounds and assessed for oxidative stress, mitochondrial dysfunction, endoplasmic reticulum stress, apoptosis, and inflammation. The spheroid model could capture many of these events triggered by the genotoxic carcinogenic chemicals, particularly aflatoxin B1 and hydroquinone. Nonetheless, no clear distinction could be made between genotoxic and non-genotoxic hepatocarcinogenicity. Therefore, spheroid cultures of human liver cells may be appropriate in vitro tools for mechanistic investigation of chemical-induced hepatocarcinogenicity, however, these mechanisms and their read-outs do not seem to be eligible biomarkers for detecting non-genotoxic carcinogenic chemicals.
... During xenobiotic chemical metabolism, cytochrome P450 (CYP) produces oxygen free radicals, which can lead to oxidative stress. In the early stages of exposure to chemicals, P450 is normally expressed [96]. In this study, ACOX3, SMOX, CYP2J6, CYP3A4, CYP3A8, CYP4C1, CYP9E2, and CYP49A1 were found to be significantly up-regulated under single or multiple concentrations of Hg exposure, which indicate that Hg exposure increases the production of ROS and H 2 O 2 . ...
As one of the most toxic elements, mercury (Hg) is a widespread toxicant in aquatic environments. Crayfish are considered suitable for indicating the impact of heavy metals on aquatic crustaceans. Nevertheless, Hg toxicity on Procambarus clarkii is largely unknown. In this research, the acute Hg-induced alterations of biochemical responses, histopathology, hepatopancreatic transcriptome, and intestinal microbiome of Procambarus clarkii were studied. Firstly, Hg induced significant changes in reactive oxygen species (ROS) and malonaldehyde (MDA) content as well as antioxidant enzyme activity. Secondly, Hg exposure caused structural damage to the hepatopancreas (e.g., vacuolization of the epithelium and dilatation of the lumen) as well as to the intestines (e.g., dysregulation of lamina epithelialises and extension of lamina proprias). Thirdly, after treatment with three different concentrations of Hg, RNA-seq assays of the hepatopancreas revealed a large number of differentially expressed genes (DEGs) linked to a specific function. Among the DEGs, a lot of redox metabolism- (e.g., ACOX3, SMOX, GPX3, GLO1, and P4HA1), ion transport- (e.g., MICU3, MCTP, PYX, STEAP3, and SLC30A2), drug metabolism- (e.g., HSP70, HSP90A, CYP2L1, and CYP9E2), immune response- (e.g., SMAD4, HDAC1, and DUOX), and apoptosis-related genes (e.g., CTSL, CASP7, and BIRC2) were identified, which suggests that Hg exposure may perturb the redox equilibrium, disrupt the ion homeostasis, weaken immune response and ability, and cause apoptosis. Fourthly, bacterial 16S rRNA gene sequencing showed that Hg exposure decreased bacterial diversity and dysregulated intestinal microbiome composition. At the phylum level, there was a marked decrease in Proteobacteria and an increase in Firmicutes after exposure to high levels of Hg. With regards to genus, abundances of Bacteroides, Dysgonomonas, and Arcobacter were markedly dysregulated after Hg exposures. Our findings elucidate the mechanisms involved in Hg-mediated toxicity in aquatic crustaceans at the tissue, cellular, molecular as well as microbial levels.
... Non-genotoxic carcinogens include tumor promoters (1,4-dichlorobenzene), endocrinemodifiers (17β-estradiol and endocrine disruptors), receptor-mediators (2,3,7,8tetrachlorodibenzo-p-dioxin), immunosuppressants (cyclosporine), or toxicant/inflammatory agents (metals, such as arsenic and beryllium) [71,72]. Unfortunately, such findings are usually overlooked, given that such data could hardly be explained within the framework of SMT. ...
The currently accepted theory on the influence of DNA mutations on carcinogenesis (the Somatic Mutation Theory, SMT) is facing an increasing number of controversial results that undermine the explanatory power of mutated genes considered as “causative” factors. Intriguing results have demonstrated that several critical genes may act differently, as oncogenes or tumor suppressors, while phenotypic reversion of cancerous cells/tissues can be achieved by modifying the microenvironment, the mutations they are carrying notwithstanding. Furthermore, a high burden of mutations has been identified in many non-cancerous tissues without any apparent pathological consequence. All things considered, a relevant body of unexplained inconsistencies calls for an in depth rewiring of our theoretical models. Ignoring these paradoxes is no longer sustainable. By avoiding these conundrums, the scientific community will deprive itself of the opportunity to achieve real progress in this important biomedical field. To remedy this situation, we need to embrace new theoretical perspectives, taking the cell–microenvironment interplay as the privileged pathogenetic level of observation, and by assuming new explanatory models based on truly different premises. New theoretical frameworks dawned in the last two decades principally focus on the complex interaction between cells and their microenvironment, which is thought to be the critical level from which carcinogenesis arises. Indeed, both molecular and biophysical components of the stroma can dramatically drive cell fate commitment and cell outcome in opposite directions, even in the presence of the same stimulus. Therefore, such a novel approach can help in solving apparently inextricable paradoxes that are increasingly observed in cancer biology.
... Within that context, liver cell models such as HepaRG [17,67] and HepG2 [6,68], as well as comparative approaches with different cell lines [4,57,69], have been used frequently. Moreover, mouse liver [5,70] and rat liver [71,72] have been analyzed. In addition, blood cell models have been used [42,45], including TK6 cells [22,26,30]. ...
Transcript signatures are a promising approach to identify and classify genotoxic and non-genotoxic compounds and are of interest as biomarkers or for future regulatory application. Not much data, however, is yet available about the concordance of transcriptional responses in different cell types or tissues. Here, we analyzed transcriptomic responses to selected genotoxic food contaminants in the human p53-competent lymphoblastoid cell line TK6 using RNA sequencing. Responses to treatment with five genotoxins, as well as with four non-genotoxic liver toxicants, were compared with previously published gene expression data from the human liver cell model HepaRG. A significant overlap of the transcriptomic changes upon genotoxic stress was detectable in TK6 cells, whereas the comparison with the HepaRG model revealed considerable differences, which was confirmed by bioinformatic data mining for cellular upstream regulators or pathways. Taken together, the study presents a transcriptomic signature for genotoxin exposure in the human TK6 blood cell model. The data demonstrate that responses in different cell models have considerable variations. Detection of a transcriptomic genotoxin signature in blood cells indicates that gene expression analyses of blood samples might be a valuable approach to also estimate responses to toxic exposure in target organs such as the liver.
... The tumor suppressor p53 gene is a pro-apoptotic and potent growth-suppressive factor, which protecting organisms from cancer development by playing a vital role in cellular apoptosis [15]. P53 is a powerful transcription factor, controlling expression of genes which regulate its tumor suppression functions, as DNA repair, arrest of cell cycle, senescence, and apoptosis [16]. P53 is stimulated in response action to cellular stresses, such as DNA damage and oxidative stress [17]. ...
Background
Hepatocellular carcinoma (HCC) is among the highest life-threatening malignancies. On both a molecular and histological level, HCC is a highly heterogeneous malignancy. This study was aimed to study the correlation between the molecular expression of some molecular biomarkers ( P53 and Cox-2) and the histopathological alterations in the chemically induced HCC by Diethylnitrosamine (DEN) in Adult female Rats. The liver tumor induction was done by injection of DEN intraperitoneally one, two and three times/week for 2 months by the dose of 50 mg/kg Bw. The histopathological analysis was done and expression level of P53 and cox-2 was detected by quantitative polymerase chain reaction (qRT-PCR) at the end of the experiment.
Results
In this study, Grossly, livers of the groups administered with DEN showed multiple grayish-white macronodules on the outer surface which is dose dependent. Histopathologically, DEN induce multifocal micronodules of hepatocellular carcinoma which characterized by nuclear atypia , clear cell, mitotic figures and necrosis of hepatocytes. P53 mRNA expression to GAPDH , revealed that, there was a statistically significant decrease in HCC groups compared to healthy control group, while Cox-2 mRNA expression was significantly increased in HCC groups than healthy control group.
Conclusions
HCC staging can be achieved by detection the expression of P53 , and Cox-2 as molecular markers as it considers noninvasive, rapid and easy method than the histopathological analysis. Finally, Cox-2 could be a therapeutic candidate for HCC due to the higher expression of Cox-2 in HCC lesions.
... 17 As such, it is pertinent to investigate the mechanisms of QD cytotoxicity. Indeed, to assess the global effect of nanoparticles on target cells, transcriptomic profiling has been undertaken in several studies, as in the case of silver and titanium nanoparticles, 18 genotoxic and non-genotoxic carcinogens, 19 11-nm dimercaptosuccinic acid-coated magnetite nanoparticles, 20 and for fibroblasts. 21 The aim of this study is thus to investigate the effects of CdTe-QD exposure in Chang cancer cells via microarray gene expression profiling to explore their oncogenic potential in terms of causing a more aggressive form of cancer. ...
Cadmium telluride quantum dots (CdTe-QDs) are acquiring great interest in terms of their applications in biomedical sciences. Despite earlier sporadic studies on possible oncogenic roles and anticancer properties of CdTe-QDs, there is limited information regarding the oncogenic potential of CdTe-QDs in cancer progression. Here, we investigated the oncogenic effects of CdTe-QDs on the gene expression profiles of Chang cancer cells. Chang cancer cells were treated with 2 different doses of CdTe-QDs (10 and 25 μg/ml) at different time intervals (6, 12, and 24 h). Functional annotations helped identify the gene expression profile in terms of its biological process, canonical pathways, and gene interaction networks activated. It was found that the gene expression profiles varied in a time and dose-dependent manner. Validation of transcriptional changes of several genes through quantitative PCR showed that several genes upregulated by CdTe-QD exposure were somewhat linked with oncogenesis. CdTe-QD-triggered functional pathways that appear to associate with gene expression, cell proliferation, migration, adhesion, cell-cycle progression, signal transduction, and metabolism. Overall, CdTe-QD exposure led to changes in the gene expression profiles of the Chang cancer cells, highlighting that this nanoparticle can further drive oncogenesis and cancer progression, a finding that indicates the merit of immediate in vivo investigation.
... However, in an in vitro experiment, ethanol extract of A. afra was cytotoxic against U937 and HeLa cancer cells via the decrease in mitochondrial membrane potential after G2/M arrest (Spies et al., 2013;Venables et al., 2016). Some Artemisia species and their active components that have been shown to arrest cells in in vitro experiments with cell lines include, A. sphaerocephala (Wang et al., 2017), A. princeps, cell cycle arrest at G2/M (Lee et al., 2013), A. monosperma, cell cycle arrest at S+G2/Mphase (Whelan and Ryan, 2004) and A. annua (Mizushina et al., 2010). ...
... Some Artemisia species and their active components that were cytotoxic in in vitro experiments with cell lines include, A. sphaerocephala (Wang et al., 2017), A. princeps (Lee et al., 2013) and A. monosperma (Whelan and Ryan, 2004). ...
A battery of short-term in vitro assays for apoptosis and/or cell cycle arrest, cytotoxicity, genotoxicity and antigenotoxicity are used to screen and establish the efficacy of medicinal plants. This study evaluated three concentrations (0.1, 0.2 or 0.4 mg/ml) of methanolic leaf extracts of Artemisia afra and Leucosidea sericea and their individual mixtures with Ethyl methanesulfonate (EMS) (0.15 mg/ml) for induction of those end points using the in vivo Allium cepa assay. Cytotoxicity was measured by the mitotic index, genotoxicity was expressed as the number of aberrant mitotic cells per 100 mitotic cells and modulatory effect (ME) was calculated as: ME = (B-C)-(A-C) / (A-C) and the value, positive or negative, indicated the number of units of the mutagen-induced genotoxicity (A) that equaled the mixture-induced genotoxicity (B). The three concentrations of A. afra extract tested did not induce cell cycle arrest and were not cytotoxic. The 0.4 mg/ml concentration and its mixture with EMS were genotoxic. The concentrations of L. sericea extract tested did not induce cell cycle arrest, were not cytotoxic nor genotoxic to the A. cepa root tip cells. The mixture of either 0.2 or 0.4 mg/ml L. sericea extract with EMS was genotoxic. The mixture of 0.4 mg/ml L. sericea extract with EMS was significantly more (ME = 4.40>2) genotoxic than EMS alone. Leaf extracts of A. afra and L. sericea lacked cell-cycle arrest activity, were non-toxic but lacked antigenotoxic activity against EMS-induced genotoxicity. High concentrations of A. afra were genotoxic whereas high concentrations of L. sericea interacted synergistically with EMS. Chromosomal abnormalities observed included sticky chromosomes, c-mitosis, chromosome largards, chromosome fragments, anaphase and telophase bridges.
... Transcriptomics has emerged as an informative method for evaluating biological functions and pathways targeted by xenobiotics. Transcriptional profiling of gene signatures has been especially useful in identifying mechanisms linked to adverse outcomes and cancer modes of action [15][16][17] . Additional applications in toxicology include modeling transcriptional information for risk assessment [18][19][20] and evaluating chemical mixture interactions 21,22 . ...
