Raul Ghiraldelli Miranda, Ivo F Machado, Anabela Pinto Rolo, Daniel Junqueira Dorta, Carlos Manuel Marques Palmeira
{"title":"HepG2 spheroids cultured in alginate microcapsules as a model for exploring mitochondrial and glycolytic metabolism using the Seahorse XFe24 Analyzer.","authors":"Raul Ghiraldelli Miranda, Ivo F Machado, Anabela Pinto Rolo, Daniel Junqueira Dorta, Carlos Manuel Marques Palmeira","doi":"10.1080/15376516.2024.2447740","DOIUrl":"10.1080/15376516.2024.2447740","url":null,"abstract":"<p><p>Mitochondria are affected by chemical substances and play a critical role in drug-induced liver injury (DILI). Chemical substances can have a significant impact on various cellular processes, such as the disruption of oxidative phosphorylation, oxidative stress, and alteration of glucose metabolism. Given the consequences of these effects, it is crucial to understand the molecular pathways of chemical substances in the context of hepatotoxicity to prevent and treat DILI. In this regard, the Seahorse XFe24 Analyzer is a valuable tool for assessing mitochondrial bioenergetics and glucose metabolism. The Mito Stress Test and Glycolytic Rate Assay allow real-time assessment of the metabolic state after chemical exposure. Additionally, HepG2 spheroids have emerged as an important alternative tool for assessing hepatotoxicity, as they provide results that are more comparable to those found in humans than monolayer cultures or animal tests (such as rodent tests). By integrating these two powerful tools, it is possible to bridge the gap between animal and human tests, resulting in more reliable results in the assessment of human hepatotoxicity and DILI. However, because of the high variability in characteristics between 3D cultures (such as spheroids and organoids), XF analyzer assays are not well optimized for use with HepG2 spheroids. Here, we describe a streamlined and optimized protocol for performing the Mito Stress Test and Glycolytic Rate Assay using HepG2 spheroids cultured in alginate microcapsules in the Seahorse XFe24 Analyzer.</p>","PeriodicalId":23177,"journal":{"name":"Toxicology Mechanisms and Methods","volume":" ","pages":"413-421"},"PeriodicalIF":3.2,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142932687","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Yukta S, Milan K L, Kannan Harithpriya, Cai Zong, S Sahabudeen, Gaku Ichihara, K M Ramkumar
{"title":"Disruptive multiple cell death pathways of bisphenol-A.","authors":"Yukta S, Milan K L, Kannan Harithpriya, Cai Zong, S Sahabudeen, Gaku Ichihara, K M Ramkumar","doi":"10.1080/15376516.2024.2449423","DOIUrl":"10.1080/15376516.2024.2449423","url":null,"abstract":"<p><p>Endocrine-disrupting chemicals (EDCs) significantly contribute to health issues by interfering with hormonal functions. Bisphenol A (BPA), a prominent EDC, is extensively utilized as a monomer and plasticizer in producing polycarbonate plastic and epoxy resins, making it one of the highest-demanded chemicals in commercial use. This is the major component used in plastic products, including bottles, containers, storage items, and food serving ware. Exposure of BPA happens through oral, respiratory, transdermal routes and eye contact. As an EDC, BPA disrupts hormonal binding, leading to various health problems, such as cancers, reproductive abnormalities, metabolic syndrome, immune dysfunction, neurological effects, cardiovascular problems, respiratory issues, and obesity. BPA mimics the hormone estrogen but exhibits a weak affinity for estrogen receptors. This weak binding affinity triggers multiple cell death pathways, including necroptosis, pyroptosis, apoptosis, ferroptosis, and autophagy, across different cell types. Numerous clinical, <i>in-vitro</i>, and <i>in-vivo</i> experiments have demonstrated that BPA exposure results in unfavorable health effects. This review highlights the mechanisms of cell death pathways initiated through BPA exposure and the associated negative health consequences. The extensive use of BPA and its frequent detection in environmental and biological models underscore the urgent need for further investigation into its effects and the development of safe alternatives. Addressing the health risks posed by BPA involves a comprehensive approach that includes reducing exposure and finding novel substitutes to lessen its detrimental impact on humans.</p>","PeriodicalId":23177,"journal":{"name":"Toxicology Mechanisms and Methods","volume":" ","pages":"430-443"},"PeriodicalIF":3.2,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143012051","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Zaira López, M Eduardo Cano, Michelle Valdovinos Pérez, Rogelio Rodríguez Rodríguez, Martha Rodríguez Sahagún, Peter Knauth
{"title":"The role of tetrahydroxyquinone solubility on apoptosis induction in human colorectal cells.","authors":"Zaira López, M Eduardo Cano, Michelle Valdovinos Pérez, Rogelio Rodríguez Rodríguez, Martha Rodríguez Sahagún, Peter Knauth","doi":"10.1080/15376516.2024.2443455","DOIUrl":"10.1080/15376516.2024.2443455","url":null,"abstract":"<p><p>Tetrahydroxy-1,4-benzoquinone (THQ) is a highly redox-active substance that generates reactive oxygen species (ROS), which can induce apoptosis in cell culture experiments. The underlying mechanism for ROS production has previously been postulated to be the autoxidation of THQ to rhodizonic acid (RhA). However, our results suggest that the cells detoxify THQ by reducing it to hexahydroxybenzene (HHB), catalyzed by the NADPH-quinone-oxidoreductase (NQO1). Then, HHB undergoes autoxidation back to THQ, closing a redox cycle that continuously generates ROS. Only this continuous mechanism produces enough ROS to trigger apoptosis. The cell's protective measures can effectively eliminate the ROS generated by a single autoxidation of THQ to RhA because RhA is not reduced back to THQ and thus does not close a redox cycle. This also explains why only fresh THQ solutions are cytotoxic, whereas older THQ solutions, which are readily autoxidized to RhA, are not.</p>","PeriodicalId":23177,"journal":{"name":"Toxicology Mechanisms and Methods","volume":" ","pages":"398-406"},"PeriodicalIF":3.2,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142923375","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Imen Benchikh, Kaddour Ziani, Abdelkrim Benalia, Ahmed Abdelhammid Djebbar, Hayat Argoub, Méghit Boumediène Khaled
{"title":"Thirty-day oral exposure to acetamiprid induces biochemical and histological alterations in rat pancreas: protective effects of carnosine supplementation.","authors":"Imen Benchikh, Kaddour Ziani, Abdelkrim Benalia, Ahmed Abdelhammid Djebbar, Hayat Argoub, Méghit Boumediène Khaled","doi":"10.1080/15376516.2024.2435350","DOIUrl":"10.1080/15376516.2024.2435350","url":null,"abstract":"<p><p>Acetamiprid is a neonicotinoid insecticide used against various insect pests. Serious concerns are emerging regarding their adverse effects on non-target organisms and organs. This study aimed to investigate the mechanistic toxic effect of oral administration of acetamiprid at 21.7 and 43.4 mg/kg body weight on the histological structure and pancreatic function of male Wistar rats and the potential effect of carnosine in mitigating this toxicity for 30 consecutive days. Thirty-six animals were divided into six groups: the control group received distilled water, the second group received 200 mg/kg body weight of carnosine, two groups received 21.7 and 43.4 mg/kg of acetamiprid, and two groups received 21.7 and 43.4 mg/kg + 200 kg/kg body weight of acetamiprid and carnosine, respectively. Acetamiprid caused a significant decrease in body weight (<i>p</i> < 0.001), pancreatic somatic index (<i>p</i> < 0.001), and amylase level (<i>p</i> ≤ 0.0001) and increased lipase level (<i>p</i> ≤ 0.0001), blood glucose level (<i>p</i> ≤ 0.0001), histological scores (<i>p</i> ≤ 0.01), and malondialdehyde level (0.01<<i>p</i> < 0.0001). Administration of carnosine led to a slight improvement in the increase of lipase (<i>p</i> ≤ 0.01) and the decrease of amylase (<i>p</i> ≤ 0.001) secretions and prevention of histopathological features induced by acetamiprid. Our results pointed out for the first time the toxic effect of acetamiprid and the preventive effect of carnosine on rat pancreatic structure and function.</p>","PeriodicalId":23177,"journal":{"name":"Toxicology Mechanisms and Methods","volume":" ","pages":"329-339"},"PeriodicalIF":3.2,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142772594","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Uliana De Simone, Francesca Caloni, Patrizia Pignatti, Carlo Gaetano, Carlo Alessandro Locatelli, Teresa Coccini
{"title":"Human stromal cell-based protocol to generate astrocytes: a straightforward <i>in vitro</i> predictive strategy in neurotoxicology.","authors":"Uliana De Simone, Francesca Caloni, Patrizia Pignatti, Carlo Gaetano, Carlo Alessandro Locatelli, Teresa Coccini","doi":"10.1080/15376516.2024.2435351","DOIUrl":"10.1080/15376516.2024.2435351","url":null,"abstract":"<p><p>The inherent adaptability of human mesenchymal stromal cells (hMSCs) to differentiate into neural lineages provides a valuable resource for investigating potential neurotoxicity in humans. By harnessing the ability of hMSCs to transform into astrocytes, we can evaluate the effects of various agents on these vital cells. Our protocol employs hMSCs sourced from umbilical cord tissue, ensuring a readily available supply of high-quality cells. The hMSC-to-neural workflow encompasses six essential steps: hMSC culture, followed by the generation of embryoid bodies (EBs) from these cells on specialized surfaces. Next, EBs and cells are expanded in a growth-promoting medium, directing them toward neural lineages. Subsequent differentiation into immature astrocytes is achieved through the use of specific factors. The process continues with the maturation of EBs/cells into astrocyte-like cells (hALCs) under optimized conditions, culminating in the final development of hALCs in a specialized medium. This methodology yields cells that display astrocyte morphology and express characteristic markers such as GFAP and S100β. The protocol is efficient, requiring roughly 6 weeks to generate hALCs from primary hMSCs without genetic manipulation. The application of hMSCs in evaluating cell damage triggered by neurotoxicants like MeHg and MGO underscores their potential as a valuable component within a more extensive battery of neurotoxicity tests.</p>","PeriodicalId":23177,"journal":{"name":"Toxicology Mechanisms and Methods","volume":" ","pages":"340-355"},"PeriodicalIF":3.2,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142772653","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Saba Khursheed Khan, Joydeep Dutta, Mohd Ashraf Rather, Ishtiyaq Ahmad, Junaid Nazir, Irfan Ahmad Khan, Gohar Bilal Wani
{"title":"Silver nanoparticle toxicity in rainbow trout: insights into physiological and molecular responses.","authors":"Saba Khursheed Khan, Joydeep Dutta, Mohd Ashraf Rather, Ishtiyaq Ahmad, Junaid Nazir, Irfan Ahmad Khan, Gohar Bilal Wani","doi":"10.1080/15376516.2024.2441952","DOIUrl":"10.1080/15376516.2024.2441952","url":null,"abstract":"<p><p>Metallic nanoparticles, with their large surface area to volume ratio, are increasingly important in various life fields, but they can cause varying toxic effects on fish. This study investigates the toxicological effects of silver nanoparticles (AgNPs) on rainbow trout (<i>Oncorhynchus mykiss</i>), focusing on hematological, biochemical, antioxidant, and histopathological changes. Fish were exposed to varying concentrations of AgNPs (0.2, 0.8, and 1.4 mg/L) for 21 days. Hematological analysis revealed significant reductions in red blood cell (RBC) counts, hemoglobin (Hb), and hematocrit (HCT) at higher AgNPs concentrations, while white blood cell (WBC) counts increased, indicating immune system activation. Biochemical assays demonstrated dose-dependent decreases in total protein and albumin, alongside increased cholesterol and triglyceride levels, suggesting impaired liver function and disrupted lipid metabolism. Antioxidant enzyme activity (SOD, CAT, GST) initially increased at lower AgNPs concentrations but declined at higher exposures, indicating oxidative stress. Molecular analysis further supported these findings, with upregulation of oxidative stress-related genes (SOD1, CAT) and inflammatory markers (HSP70, IL-1β). Histopathological examinations revealed necrosis, hyperplasia, and lamellar fusion in the gills, along with significant liver damage, including vacuolation and Kupffer cell proliferation, particularly at the highest exposure. Behavioral assays showed erratic swimming and reduced feeding in fish exposed to higher AgNPs concentrations. This study highlights the dose-dependent toxic effects of AgNPs on rainbow trout and underscores the potential for long-term, possibly irreversible damage at higher exposure levels. These findings emphasize the need for stricter environmental regulations on nanoparticle use to mitigate their ecological impact.</p>","PeriodicalId":23177,"journal":{"name":"Toxicology Mechanisms and Methods","volume":" ","pages":"382-397"},"PeriodicalIF":3.2,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142855232","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Munir Ahmad, Yasir Mahmood, Abdul Ghaffar, Riaz Hussain, Shazia Anjum, Dalia Fouad, Farid Shokry Ataya, Ahrar Khan
{"title":"Pymetrozine induced remarkable hemato-biochemical modifications and genotoxicity in vital organs of bighead carp (<i>Aristichthys nobilis</i>).","authors":"Munir Ahmad, Yasir Mahmood, Abdul Ghaffar, Riaz Hussain, Shazia Anjum, Dalia Fouad, Farid Shokry Ataya, Ahrar Khan","doi":"10.1080/15376516.2024.2440738","DOIUrl":"10.1080/15376516.2024.2440738","url":null,"abstract":"<p><p>Pymetrozine (a pyridine azomethine pesticide) is one of the most commonly and frequently used insecticides. Scanty information is available about the deleterious effects of Pymetrozine on fish especially bighead carp. Hence, the current study investigated chronic toxicological effects of pymetrozine in bighead carp. A total of 80 fish were reared and divided into four groups(A-D) each containing 20 fish. Pymetrozine was given to experimental fish of groups B, C, and D mixed in water at doses of 5, 10, and 15 mg/L respectively for 30 days. Group A remained as control group. On days 10, 20, and 30 of the experiment, blood and other visceral tissues were collected for analysis of genotoxic effects, erythrocytic morphological and nuclear changes, antioxidant enzymes, and oxidative stress profile. The results revealed significantly higher values of various nuclear abnormalities (erythrocyte with micronuclei, red blood cells with condensed and lobed nuclei) and morphological changes (pear shaped erythrocyte, spindle shaped erythrocytes and spherocyte) in erythrocytes of bighead carp. The investigations on status of antioxidant enzymes and oxidative stress indicated higher values of oxidative stress biomarkers and lower values of antioxidant enzymes in visceral organs (brain, liver, gills, and kidneys) of treated fish. The findings on genotoxic potential of pymetrozine revealed a considerably increased frequency of DNA damage in isolated cells of multiple tissues (brain, liver, gills, and kidneys) of experimental fish at higher doses. In conclusion, it may be suggested that pymetrozine induces toxic effects via disruption of physiological mechanisms of multiple visceral organs of bighead carp.</p>","PeriodicalId":23177,"journal":{"name":"Toxicology Mechanisms and Methods","volume":" ","pages":"372-381"},"PeriodicalIF":3.2,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142932691","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Differences in metabolite genotoxicity test results of rat liver S9 microsomes treated with various microsomal enzyme inducers.","authors":"Shigeru Igaki, Kiyohiro Hashimoto, Toshikatsu Matsui, Tadahiro Shinozawa","doi":"10.1080/15376516.2024.2443543","DOIUrl":"10.1080/15376516.2024.2443543","url":null,"abstract":"<p><p>The rat S9 microsome fraction is commonly used to assess compound metabolite formation during <i>in vitro</i> genotoxicity assessments. However, methods using S9 have not been standardized for genotoxicity studies, and different experimental methods are used at various facilities. Therefore, this study investigated whether the differences between the two experimental conditions (1) S9 inducers, phenobarbital + beta-naphthoflavones vs. Aroclor 1254 and (2) the plate incubation vs. preincubation method) in the micro-Ames test would affect the results. Nitrosamine and in-house genotoxicity-positive compounds were used with benzopyrene and 2-aminofluorene as positive control compounds. No differences were observed in the genotoxicity results in the groups treated with the positive control. However, the S9 fraction induced by Aroclor 1254 showed higher cytochrome P450 activity than the fraction induced by phenobarbital and beta-naphthoflavone. The incubation method also affected the results; the nitrosamine compounds showed different genotoxicity activity between the plate incorporation method and preincubation method. In-house aminomethyl quinoline compounds also showed different results depending on the S9 type. These results suggest that different inducers and methods induce various metabolic enzyme activities, which may lead to differences in genotoxicity through distinct metabolite production.</p>","PeriodicalId":23177,"journal":{"name":"Toxicology Mechanisms and Methods","volume":" ","pages":"407-412"},"PeriodicalIF":3.2,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142915647","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Saba Khursheed Khan, Joydeep Dutta, Mohd Ashraf Rather, Ishtiyaq Ahmad, Junaid Nazir, Arun Karnwal
{"title":"Toxicological impact of copper nanoparticles on rainbow trout: hematological, biochemical, antioxidant, and histopathological responses with oxidative gene expression.","authors":"Saba Khursheed Khan, Joydeep Dutta, Mohd Ashraf Rather, Ishtiyaq Ahmad, Junaid Nazir, Arun Karnwal","doi":"10.1080/15376516.2024.2438120","DOIUrl":"10.1080/15376516.2024.2438120","url":null,"abstract":"<p><p>Copper nanoparticles (CuNPs) are increasingly used across various industries due to their catalytic, antimicrobial, and electrical properties. However, their potential toxicity in aquatic environments, particularly to non-target organisms like fish, remains poorly understood. This study investigated the effects of CuNP exposure on rainbow trout (<i>Oncorhynchus mykiss</i>) by comprehensively evaluating hematological, biochemical, antioxidant, molecular, and histopathological parameters. Rainbow trout fingerlings were exposed to varying concentrations of CuNPs (0.2 mg/L, 0.6 mg/L, and 1.0 mg/L) for 7, 14, and 21 days. The results revealed significant dose-dependent declines in hemoglobin (Hb) and red blood cell (RBC) counts, alongside increases in white blood cell (WBC) counts, indicating an immune response to CuNPs-induced stress. Serum biochemistry showed disruptions in albumin, globulin, cholesterol, and triglycerides, suggesting impaired liver function and altered lipid metabolism. Antioxidant enzyme activity, including catalase (CAT), increased significantly, reflecting oxidative stress, while lipid peroxidation (LPO) levels unexpectedly decreased, suggesting possible activation of compensatory mechanisms. Histopathological analysis confirmed severe gill and liver damage, including hypertrophy, hyperplasia, lamellar fusion, necrosis, and cellular degeneration. Molecular analysis showed upregulation of oxidative and, inflammatory genes, and signs of apoptosis. These findings underscore the toxic potential of CuNPs in aquatic environments and highlight the need for careful regulation and environmental monitoring to mitigate their impact.</p>","PeriodicalId":23177,"journal":{"name":"Toxicology Mechanisms and Methods","volume":" ","pages":"356-371"},"PeriodicalIF":3.2,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142802176","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}