Toxicology Mechanisms and Methods最新文献

Microplastics (MPs) have emerged as a serious global environmental threat due to their resistance to degradation and persistence in ecosystems. Given their potential risks to human health, it is essential to thoroughly investigate the mechanisms of toxicity and associated health consequences. This study examined the toxicological and reproductive effects of varying doses of polyethylene microplastics (PE-MPs) in 120 male and female Sprague Dawley rats. A statistically significant, dose-dependent increase in malondialdehyde levels was observed, along with a reduction in catalase activity. Furthermore, alterations were detected in sexual hormone levels and disruptions were noted in both the Kelch-like ECH-associated protein 1 (Keap1)-Nrf2-ARE (antioxidant response element) and p38 MAPK-Nrf2 signaling pathways. PE-MP exposure also produced marked histopathological changes in the testes and ovaries. These findings indicate that reproductive toxicity from PE-MPs is associated with impairments in the Keap1-Nrf2-ARE and p38 MAPK-Nrf2 pathways. The results underscore the importance of limiting MP exposure to mitigate potential health hazards and provide new data on the potential mechanisms of toxicity of MPs.

Single cell gel electrophoresis (comet assay) is a research tool known for its use in the assessment of DNA damage in which peripheral blood lymphocytes are used as a cellular model. The objective of this study was to adapt the comet assay to fresh normal and tumor breast tissue explants to assess DNA damage. Representative samples were obtained from nine patients with infiltrating ductal adenocarcinoma during the time of surgical intervention for the resection of the tumor. One hundred micrometer thick explants were prepared from normal and tumor breast tissue using a tissue slicer. Explants were harvested into six-well microplates with 4 mL of sterile serum-free Dulbecco's modified Eagle's medium/Ham's F12 medium at 4 °C for immediate processing of the comet assay. The results indicated that the comet assay can be used to analyze DNA damage in tumors and healthy breast tissue in an ex vivo model that allows visualization of the level of DNA damage. Compared to non-tumor cells, DNA damage in breast cancer was significantly increased (p < 0.05); furthermore, DNA damage was higher in samples treated with H₂O₂ (positive control) and confirmed that cells in the explants can interact with the chemicals tested, as well as the functionality of the technique. The comet assay using fresh tissue explants represents an alternative tool with potential to identify DNA damage in tumor tissue and to study new drugs to personalize the use of DNA-damaging therapies.

Cardiotoxicity of traditional Chinese medicines (TCMs) is an important factor in pharmacogenetic cardiovascular diseases, which directly affects the clinical use of TCM. Recently, cinobufagin (CBG), the active substance of Toad venom, has shown as a potential anti-tumor natural product. However, its cardiotoxicity limits the clinical application and the intrinsic mechanism is unclear. Here, we found that CBG induced oxidative stress and cell apoptosis in rat cardiomyocytes (H9c2), which caused cardiomyocyte injury. SET and MYND domain containing 1 (SMYD1) is a histone methyltransferase containing a SET-MYND domain, which is specifically expressed only in cardiomyocytes and skeletal muscle cells. RNA-Seq analysis revealed that the expression of SMYD1 was significantly decreased in the CBG treated H9c2 cells. Overexpression of SMYD1 alleviated CBG-induced myocardial injury, while knockdown of SMYD1 did the opposite. To summary, our study indicated that CBG induced cell apoptosis and oxidative stress via suppressing the expression of SMYD1 and ultimately leaded to myocardial toxicity. This research revealed the side-effects of CBG in myocardial toxicity and provided mechanism basis in specific disease conditions for CBG therapy.

To explore aloe-emodin (AE)'s effects on aristolochic acid I (AAI)-induced toxicity in renal tubular epithelial cells and particularly through its regulation of dual-specificity phosphatase 1 (DUSP1). Human renal proximal tubular epithelial cells (HK-2 cells were treated with different concentrations of AE and then exposed to AAI to induce cell damage. The experimental groups included the control group, AAI group, AAI + AE 10 μM group, AAI + AE 50 μM group, AAI + AE + shNC group, AAI + AE + sh-DUSP1 group, AAI + AE + oe-DUSP1 group, and AAI + AE + oe-NC group. Cell viability was assessed using the MTT assay, and apoptosis was detected by the TUNEL assay. Levels of pro-inflammatory cytokines were measured by ELISA. Mitochondrial function was evaluated. In addition, qRT-PCR and Western blot were used to analyze the expression of DUSP1 and apoptosis-related proteins. AE significantly increased the viability of HK-2 cells after AAI treatment and inhibited apoptosis. In the AAI-treated group, cell viability was significantly reduced, apoptosis increased, levels of pro-inflammatory cytokines were elevated, and mitochondrial function was impaired. In DUSP1 knockdown HK-2 cells, the protective effect of AE was significantly weakened, with decreased cell viability, increased apoptosis, aggravated inflammation, and impaired mitochondrial function. In the DUSP1 overexpression group, the protective effect of AE was further enhanced, indicating that DUSP1 plays a key role in the protective effects mediated by AE. AE can enhance AAI-induced HK-2 cell viability, inhibit cell apoptosis, and reduce inflammation and oxidative stress. The mechanism of AE may be related to the up-regulation of DUSP1 expression.

Snakebite envenoming remains a major global health issue, particularly in underserved regions. To better understand venom composition and toxic effects, crude snake venoms can be fractionated into distinct toxin groups using high-throughput nanofractionation analytics. These fractions can then be assessed using venomics to identify specific toxins, in conjunction with bioassays to evaluate their bioactivity. Traditional in vivo testing in rodents is limited by legal and ethical concerns, the high number of fractions, and the small quantities of toxins in each. In this study, zebrafish larvae were introduced as an efficient alternative model for in vivo analysis of toxins that impair locomotion. A behavioral bioassay was developed using a light-dark challenge to assess locomotion in 5-day post-fertilization (dpf) larvae. A Locomotion Index was created to compare movement during the dark phase between venom-exposed and control groups. Using venom from the spitting cobra Naja nigricollis, two fractions were found to reduce locomotor activity significantly. Microscopic screening also revealed three tissue-toxic fractions, one of which caused bradycardia. Dose-response testing was performed for each toxic fraction. Furthermore, oxygen consumption assays were conducted with venom fractions from N. nigricollis, N.haje, N. siamensis, and N. subfulva, identifying two fractions that elevated oxygen consumption. High-throughput venomics linked these toxic effects to three-finger toxins and phospholipases A₂. This integrative approach offers a scalable, efficient method for in vivo toxicity screening of venom components and shows promise for broader applications in the discovery and profiling of diverse bioactive compounds.

Metabolic dysfunction-associated fatty liver disease (MAFLD) is a risk factor for hepatocellular carcinoma (HCC). Evidence links the nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome/pyroptosis pathway to MAFLD-related fibrosis; however, its role in MAFLD progression to HCC remains poorly understood. This study investigates the temporal activation of the NLRP3 inflammasome/pyroptosis pathway in MAFLD-associated HCC. Rats were assigned to a control group, who received a standard diet with intraperitoneal injections of liquid petrolatum and water, or to experimental groups, subjected to a hepatopathogenic diet, CCl₄, and diethylnitrosamine. Serum liver enzymes, inflammatory cytokines, NLRP3 inflammasome/pyroptosis pathway, fibrosis, and HCC markers were assessed. Hepatosteatosis and serum liver enzymes increased after 3 weeks. Hepatosteatosis was associated with elevated levels of ALT (p = 0.0051), GGT (p < 0.0001), IL-6 (p = 0.0499), TNF-α (p = 0.0020), and IL-1β (p < 0.0001), and the NLRP3 inflammasome/pyroptosis pathway activators (NLRP3, p = 0.0246; ASC, p = 0.0003, caspase-1, p = 0.0003, and GSDMD p = 0.0111). Levels of fibrosis markers, including TGF-β (p < 0.0001), α-SMA (p = 0.0035), and collagen (p < 0.0001), increased after 7 weeks, while those of HCC markers PTGR1 and KRT19 increased 13 weeks onwards (both p < 0.0001). These findings provide the first evidence of the NLRP3 inflammasome/pyroptosis pathway's involvement in MAFLD-associated HCC development. The data strongly suggest that metabolic dysregulation and NLRP3-driven inflammation lead to pyroptosis, triggering ongoing cycles of cellular damage and regeneration and accelerating the transition from MAFLD to HCC.

Isothiazolinones are heterocyclic compounds with potent broad-spectrum antimicrobial properties, commonly used as preservatives in cosmetics, pharmaceuticals, and industrial products. Their antimicrobial efficacy results from interference with microbial protein and nucleic acid synthesis. However, their limited water solubility and instability under alkaline conditions, elevated temperatures, and UV exposure constrain their formulation and use. Despite regulatory restrictions, especially in the EU, due to their allergenic and toxic potential, exposure remains widespread, particularly through industrial and household products. Studies indicate that isothiazolinones can penetrate the skin, bind to cellular components, and induce oxidative stress, apoptosis, and immune responses. Furthermore, emerging data suggest possible endocrine-disrupting and neurotoxic effects. The mechanisms driving these toxicities are still not fully understood. This review examines the physicochemical characteristics, antimicrobial mechanisms, applications, exposure routes, cytotoxic effects, and molecular interactions of isothiazolinones. Deeper mechanistic insight is needed to support the development of safer alternatives and to inform more stringent regulatory frameworks.

Flavonoids, found in fruits and vegetables, can potentially prevent brain diseases. Diosmin (diosmetin-7-O-rutinoside), a flavonoid, exhibits various pharmacological activities, but its impact on calcium ion (Ca²⁺) signaling and the associated mechanisms in human glioblastoma cells remain unclear. This study investigated the effect of diosmin on intracellular Ca²⁺ levels ([Ca²⁺]_i), cell viability, and the participation of Ca²⁺-related pathways in DBTRG-05MG human glioblastoma cells. It also investigated the connection between Ca²⁺ signaling and toxicity in cells treated with diosmin and the Ca²⁺ chelator BAPTA-AM. Research indicates that diosmin (20-60 μM) caused an increase in [Ca²⁺]_i and induced cytotoxicity in a concentration-dependent manner. Furthermore, pre-treating the cells with the BAPTA-AM can intensify the cytotoxic effect. The removal of extracellular Ca²⁺ suppressed the entry of Ca²⁺. Agents that modulate store-operated Ca²⁺ channels, SKF96365 and 2-APB, can inhibit the entry of Ca²⁺ induced by diosmin. Treatment with the endoplasmic reticulum Ca²⁺ pump inhibitor thapsigargin in a Ca²⁺-free environment inhibited the increase in [Ca²⁺]_i caused by diosmin; conversely, treatment with diosmin reduced the increase in [Ca²⁺]_i caused by thapsigargin. Moreover, inhibiting phospholipase C (PLC) with U73122 eliminated the increase in [Ca²⁺]_i triggered by diosmin. In DBTRG-05MG cells, diosmin-induced cell death is associated with Ca²⁺, a process involving the entry of Ca²⁺ through store-operated Ca²⁺ channels and the release of Ca²⁺ from the endoplasmic reticulum, which relies on the PLC. Additionally, BAPTA-AM, a compound with Ca²⁺-chelating properties, shows promise in enhancing diosmin-induced cytotoxicity, and this could represent a significant development in glioblastoma research.

Micro- and nanoplastics (MNPs), pervasive environmental pollutants, contaminate water, soil, air, and the food chain and ultimately accumulate in living organisms. Macrophages are the main immune cells that gather around MNPs and engulf them through the process of phagocytosis. This internalization triggers M1 polarization and the secretion of inflammatory cytokines, including IL-1, IL-18, IL-12, TNF-α, and IFN-γ. Furthermore, MNPs damage mitochondria and lysosomes, causing overactivation of iNOS and excessive production of ROS. This results in cellular stress and induce apoptosis, necroptosis, and, in some cases, metosis in macrophages. The internalization of MNPs also increases the expression of receptors, involving CD36, SR-A, LOX-1, and the macrophage receptor with a collagenous structure (MARCO) while decreasing ABCA-1 and ABCG-1. MNPs in adipose tissue macrophages trigger proinflammatory cytokine secretion, causing adipogenesis, lipid accumulation, insulin resistance, and the secretion of inflammatory cytokines in adipocytes. Various factors influence the rate of MNP internalization by macrophages, including size, charge, and concentration, which affect internalization through passive diffusion. Receptor-mediated phagocytosis of MNPs occurs directly via receptors like T-cell immunoglobulin and mucin domain containing 4 (TIM-4) and MARCO. The attachment of biomolecules, including proteins, antibodies, opsonins, or microbes to MNPs (forming corona structures) promotes indirect receptor-mediated endocytosis, as macrophages possess receptors like TLRs and FcγRIII. MNPs also cause gut dysbiosis, a risk factor for proinflammatory microenvironment and M1 polarization. Here, we review the mechanisms and consequences of MNP macrophage exposure, which is linked to autoimmunity, inflammation, and cardiometabolic syndrome manifestations, including atherosclerosis and obesity, highlighting the immunotoxicity of MNPs.

Endocrine-disrupting chemicals (EDCs), such as bisphenol A (BPA), bisphenol S (BPS), phthalates, and micro- and nanoplastics, present substantial environmental and health hazards because of their potential to disrupt hormonal systems. Micro- and nanoplastics can release EDCs that disrupt reproductive and developmental processes, potentially affecting future generations. BPA, a common plasticizer and resin component, mimics estrogen and disrupts thyroid hormone metabolism, contributing to obesity, diabetes, and cardiovascular issues. BPS, a BPA substitute, exhibits similar endocrine-disrupting properties and persists longer in the environment. Phthalates, which are widely used as plasticizers, are associated with reproductive issues, metabolic conditions, and developmental issues in children. Combined exposure to multiple EDCs can amplify health risks, underscoring the need for further research on the synergistic impacts of these chemicals. This review underscores the urgent need for effective regulatory measures and further investigations into the health impacts of EDCs to mitigate their harmful impacts on the health of humans and the environment.