Archives of Toxicology最新文献

The Hedgehog (Hh) signaling pathway is essential for maintaining homeostasis during embryogenesis and in adult tissues. In the liver, dysregulation of this pathway often leads to liver cancer development. Recent studies also suggest that disturbances in the Hh pathway can affect liver metabolism in healthy livers through interactions with other signaling pathways, such as the Wnt/β-catenin pathway. As a result, the Hh pathway has emerged as a promising target for therapeutic intervention. However, little is known about the effects of Hh modulators on healthy hepatocytes. In our study, we investigated the effects of the Hh agonists SAG (300 nM) and triamcinolone acetonide (40 µM), as well as the antagonists RU-SKI 43 (100 nM), cyclopamine (5 µM), budesonide (25 µM), GANT61 (0.5 µM), and vismodegib (1 µM) on healthy mouse and human primary hepatocytes in vitro. We employed toxicological, transcriptomic, proteomic, and functional assays, including proliferation and Seahorse assays. Our results show that these compounds significantly impact metabolic pathways such as lipid and glucose metabolism at both transcriptional and protein levels. Mechanistically, our data suggest the involvement of both canonical and non-canonical Hedgehog pathways, a phenomenon not previously described in hepatocytes. These findings highlight the diverse effects of these compounds on signaling and key metabolic functions in the liver, which emphasizes the need to investigate the hepatic Hh cascade and its metabolic control in depth. As the compounds regulate different aspects of metabolism, they need to be carefully studied in appropriate model systems for specific therapeutic use.

Chlorpyrifos (CPF) is one of the most widely used organophosphorus pesticides all over the world. Unfortunately, long-term exposure to CPF may cause considerable toxicity to organisms. Some evidence suggests that the intestinal microbial community may be involved in regulating the toxicity of CPF. In this study, we explored if the intestinal microbial community is involved in regulating the toxicity of CPF. Adult mice were continuously exposed to CPF (4 mg/kg body weight /day) for 10 weeks with or without a 2-week pretreatment of antibiotics to change the ecological structure of intestinal microorganisms in advance. Pathological changes in the liver and kidneys were examined and the biochemical parameters in serum for liver and kidney functions were detected, and changes in the intestinal microbial community of the mice were measured. The results showed that subchronic exposure to low-dose CPF caused an ecological imbalance in the intestinal flora and caused pathological damage to the liver and kidneys. Serum biochemical indicators for liver function such as alanine aminotransferase and total bile acids contents and renal biochemical indicators such as urea nitrogen and creatinine were disrupted. Changes in intestinal microbial community structure by using antibiotics in advance can effectively alleviate the pathological and functional damage to the liver and kidneys caused by CPF exposure. Further analysis showed that intestinal microorganisms such as Saccharibacteria (TM7), Odoribacter, Enterococcus and AF12 genera may be involved in managing the toxicity of CPF. Together, our results indicated that long-term low-dose CPF exposure could induce hepatotoxicity and nephrotoxicity, and liver and kidney damage may be mitigated by altering the ecology of intestinal microorganisms.

The development of in vitro hepatocyte cell culture systems is crucial for investigating drug-induced liver injury (DILI). One prerequisite for monitoring DILI related immunologic reactions is the extension of primary human hepatocyte (PHH) cultures towards the inclusion of macrophages. Therefore, we developed and characterized an autologous co-culture system of PHH and primary human hepatic macrophages (hepM) (CoC1). We compared CoC1 with a co-culture of the same PHH batch + M0 macrophages derived from THP1 cells (CoC2) in order to represent a donor independent macrophage reaction. Then, we treated the mono- and co-cultures with drugs that cause DILI—menadione (MEN, 1 or 10 µM, 3 h), diclofenac (DIC, 0.5 or 5 mM, 6 h), or acetaminophen (APAP, 0.5 or 5 mM, 6 h)—and assessed culture stability, cell activity, macrophage differentiation, cytokine production and cell viability. Without drug treatment, CoC1 was the most stable over a culture time of up to 60 h. Cytokine array analysis revealed a proinflammatory profile of PHH mono-cultures due to isolation stress but showed different influences of hepM and M0 on the cytokine profile in the co-cultures. MEN, DIC and APAP treatment led to donor-dependent signs of cell stress and toxicity. HepM can either promote or reduce the DILI effects donor dependently in CoC1. CoC2 are slightly less sensitive than CoC1 in representing DILI. In summary, we present a new autologous co-culture system that can mimic DILI in a donor-dependent manner. This cellular system could be useful for new drug testing strategies and reducing animal testing.

Alcoholic liver disease (ALD) and non-alcoholic fatty liver disease (NAFLD) are two major chronic liver diseases worldwide. The triggers for fatty liver can be derived from external sources such as adipose tissue, the gut, personal diet, and genetics, or internal sources, including immune cell responses, lipotoxicity, hepatocyte death, mitochondrial dysfunction, and extracellular vesicles. However, their pathogenesis varies to some extent. This review summarizes various immune mechanisms and therapeutic targets associated with these two types of fatty liver disease. It describes the gut-liver axis and adipose tissue-liver crosstalk, as well as the roles of different immune cells (both innate and adaptive immune cells) in fatty liver disease. Additionally, mitochondrial dysfunction, extracellular vesicles, microRNAs (miRNAs), and gastrointestinal hormones are also related to the pathogenesis of fatty liver. Understanding the pathogenesis of fatty liver and corresponding therapeutic strategies provides a new perspective for developing novel treatments for fatty liver disease.

Exposure to fibrogenic multi-walled carbon nanotubes (MWCNTs) induces the production of proinflammatory lipid mediators (LMs) in myeloid cells to instigate inflammation. The molecular underpinnings of LM production in nanotoxicity remain unclear. Here we report that PU.1, an ETS domain-containing master regulator of hematopoiesis, critically regulates the induction of arachidonate 5-lypoxygenase (Alox5) and the production of LMs. MWCNTs (Mitsui-7) at 2.5 or 10 µg/mL induced the expression of Alox5 in murine and human macrophages at both mRNA and protein levels, accompanied by marked elevation of chemotactic LM leukotriene B4 (LTB4). Induction is comparable to those by potent M1 inducers. Carbon black, an amorphous carbon material control, did not increase Alox5 expression or LTB4 production at equivalent doses. MWCNTs induced the expression of a heterologous luciferase reporter under the control of the murine Alox5 promoter. Deletional analysis of the 2 kb promoter uncovered multiple inhibitory and activating activities. The proximal 250 bp region had the largest activation that was further increased by MWCNTs. The Alox5 promoter contains four PU box-like enhancers. PU.1 bond to each of the enhancers constitutively, which was further increased by MWCNTs. Knockdown of PU.1 using specific small hairpin-RNA blocked the basal and induced expression of Alox5 and the production of LTB4 as well as prostaglandin E2. The results demonstrate a critical role of PU.1 in mediating MWCNTs-induced expression of Alox5 and production of proinflammatory LMs, revealing a molecular framework where the hematopoietic transcription factor PU.1 is activated to orchestrate multiple proinflammatory responses to sterile particulates.

An abundant amount of colon cancers is diagnosed every year, accounting for 9% of malignant tumors. Even with the progress of relevant research, the 5-year survival rate for colon cancer is still less than 60%, indicating that improving the prognosis of colon cancer is still a challenge that needs to be overcome. This study employed the algorithm “scissor” to integrate the single-cell sequencing data and bulk transcriptome data with prognosis information to predict prognosis-associated cells (PAC). Summary-data-based Mendelian randomization (SMR) analysis was conducted using expression quantitative trait loci data and GWAS data to identify genes having causal associations with prognosis phenotype in colon cancer patients and five traditional two-sample Mendelian randomization methods were utilized to confirm the results. Finally, our findings were validated based on two independent external validation datasets, GSE17536 and GSE39582. The real-world tissue dataset with corresponding immunohistochemical (IHC) experiments was utilized to confirm our findings. We determined that the majority of PACs were fibroblasts. On top of that, this study identified ADAMDEC1 as a gene that has a significant causal association with overall survival. ADAMDEC1, highly expressed in highly differentiated fibroblasts, was ascertained its high expression was linked with a better prognosis of patients with colon cancer by the related bulk transcriptome analysis. Our dataset presented that higher IHC scores were associated with a better prognosis for colon cancer, further validating our results. This study has identified ADAMDEC1 as a prognostic protective factor for patients with colon cancer, providing clues for clinical trials and drug experimental target research.

Copper (Cu) is an essential trace element involved in fundamental physiological processes in the human body. Even slight disturbances in the physiological Cu homeostasis are associated with the manifestation of neurodegenerative diseases. While suggesting a crucial role of Cu in the pathogenesis, the exact mechanisms of Cu neurotoxicity involved in the onset and progression of neurological diseases are far from understood. This study focuses on the molecular and cellular mechanisms of Cu-mediated neurotoxicity in human brain cells. First, the cytotoxic potential of Cu was studied in fully differentiated, human neurons (LUHMES cells). Lysosomal integrity was considerably affected following incubation with 420 µM CuSO₄ for 48 h. Further mechanistic studies revealed mitochondria and neuronal network as most susceptible target organelles (already at 100 µM CuSO₄, 48 h), while the generation of reactive oxygen species turned out to be a rather later consequence of Cu toxicity. Besides Cu, the homeostasis of other elements might be involved and are likely to contribute to the pathology of Cu-mediated neurological disorders. Besides Cu, also effects on the cellular levels of magnesium, calcium, iron, and manganese were observed in the neurons, presumably aggravating the consequences of Cu neurotoxicity. In conclusion, insights in the underlying mode of action will foster the development of treatment strategies against Cu-mediated neurological diseases. Particularly, the interplay of Cu with other elements might provide a powerful diagnostic tool and might be used as therapeutic approach.

The hematopoietic system is the primary target of benzene exposure. Whether peripheral blood miRNA can serve as sensitive biomarkers for benzene-induced hematopoietic damage has attracted considerable attention. This study focuses on exploring the role of miR-451a and miR-486-5p in benzene-induced erythroid damage and assessing their potential as biomarkers of benzene-induced hematotoxicity. Animal experiments and human studies were conducted to reveal expression patterns of miR-451a and miR-486-5p in bone marrow and peripheral blood after benzene exposure, along with their correlations with erythrocyte indices. In C57BL/6J mice exposed to benzene, the expression levels of miR-451a and miR-486-5p in bone marrow decreased, which also positively correlated with red blood cell count (RBC), hemoglobin (Hb), and hematocrit (HCT). Conversely, in peripheral blood of C57BL/6J mice, the expression levels of the two miRNAs increased and showed a negative correlation with the three erythroid indices. Subsequent validation in bone marrow samples of chronic benzene poisoning patients and peripheral blood of workers from petrochemical plant confirmed significant correlations between miR-451a and miR-486-5p expression levels and red blood cell parameters. Furthermore, receiver operator characteristic (ROC) curve analyses revealed that miR-451a emerged as a potential biomarker for benzene-induced hematotoxicity, exhibiting superior discriminatory power compared to miR-486-5p and conventional erythroid indices. Additionally, in vitro experiments using K562 cells revealed differential regulatory effects of benzene metabolite hydroquinone (HQ) on miR-451a expression based on erythroid differentiation status. These findings emphasized the important role of miR-451a and miR-486-5p in benzene-induced erythrogenesis disruption, offering valuable insights for biomarker development and therapeutic interventions.

Shiga toxin is the leading cause of food poisoning in the world. It is structurally similar to the plant type II ribosome-inactivating proteins (RIPs) and retains N-glycosidase activity. It acts specifically by depurinating the specific adenine A4605 of human 28S rRNA, ultimately inhibiting translation. Recent outbreaks and increasing demands for lab-scale meat assert the necessity for producing toxoids. In the current study, we have carried out the comparative structural and functional analysis of Shiga with ricin for N-glycosidase activity. Primary structural analysis indicates that Shiga is more flexible than ricin and one active site residue Gly121 (ricin), has been mutated to Ser (Shiga). Tertiary structure analysis confirms the conserved active site residue confirmation. Further, molecular dynamic studies indicate that the mutated Ser residue of Shiga imparts flexibility besides interacting with the conserved GAGA loop of 28s rRNA and contributes free energy of −5.39 kcal/mol. We have observed a decreasing trend line of average free binding energy with an average of −23 kcal/mol. The residue interaction network indicates that Arg is the key residue that protonates and initiates the N-glycosidase activity. Overall, these structural studies provide molecular insights into the N-glycosidase mechanism and serve as a prospect for the development of toxoids.

For many years, polybrominated diphenyl ethers (PBDEs) were used as flame retardants in a large number of consumer products. Even though international law meanwhile prohibits the production and usage of PBDEs, these persistent and bioaccumulative chemicals still leak into the environment, and are frequently detected in wildlife and humans. Population-based studies reveal positive correlations between human PBDE exposure and various adverse health effects, emphasizing that a better understanding of the mode of action of these polybrominated chemicals is urgently needed. Therefore, we investigated the effect of two widespread PBDEs, namely BDE-47 and BDE-99, on epidermal growth factor receptor (EGFR) activity in human cells. Recent studies showed that the EGFR is not only orchestrating cellular functions, but also serves as a cell-surface receptor for dioxins, phenolic benzotriazoles and related organic pollutants. Results from in silico docking analyses, AlphaLISA-based receptor binding studies and SDS-PAGE/Western blot analyses revealed that BDE-47 and BDE-99 inhibit the growth factor-triggered activation of EGFR by binding to its extracellular domain. In keratinocytes, PBDEs also inhibit amphiregulin-induced and EGFR-mediated DNA synthesis as well as the EGFR-triggered trans-repression of the aryl hydrocarbon receptor signaling pathway. Our data identify EGFR as a cell-surface receptor for PBDEs and shed light on a novel mode of action of these ubiquitous and persistent chemicals. This finding may contribute to an improved hazard assessment of PBDEs and structurally related flame retardants.