Chemical Research in Toxicology最新文献

Hydroxyindoles are organic compounds characterized by the presence of a hydroxy group attached to an indole ring (six-membered benzene ring fused to a five-membered pyrrole ring). These compounds are naturally occurring and play a role in the synthesis of various medicinal drugs. One notable example is 4-Hydroxyindole (4-HI), which contains a hydroxy group at the fourth position of the indole ring. In a recent study, we tested various hydroxyindole compounds for their antiferroptotic activity, including 3-hydroxyindole, which demonstrated strong resistance to ferroptosis. Ferroptosis is a regulated form of cell death that occurs due to uncontrolled phospholipid peroxidation and is associated with the development of degenerative conditions, such as neurodegenerative diseases. Here, we tested the hypothesis that 4-HI could protect against ferroptosis, similar to other hydroxyindole compounds. To induce ferroptosis, we utilized established modulators, including erastin, RSL3, and FINO2. We assessed cytotoxicity using the calcein AM assay and measured lipid peroxidation caused by ferroptosis inducers with the C11-BODIPY assay. Our results indicated that 4-HI protects various brain-related cell types, including HT-22, N27, and RBE4 cells, from ferroptosis. We also utilized our newly developed cell-free assay, in which combined iron and arachidonic acid were used to oxidize C11-BODIPY, allowing us to investigate the radical scavenging activity of 4-HI. We discovered that 4-HI exhibits antioxidant effects in cell-free assays, suggesting that its protective action against ferroptosis is likely due to its radical-scavenging capabilities. Interestingly, we found that 4-hydroxyindole-3-carbaldehyde, a structural analog of 4-HI, did not effectively prevent ferroptosis. This suggests that the carbaldehyde group, which is an electron-withdrawing group, may reduce the antiferroptotic activity of 4-HI. In summary, 4-HI appears to be a promising inhibitor of ferroptosis, warranting further research to explore its potential in protecting against neurotoxicity and neurodegeneration associated with this type of cell death.

Skin sensitization is a critical endpoint in human safety risk assessment of chemicals. Risk assessment approaches have evolved, and the field has seen a shift toward adopting new approach methods (NAMs) instead of relying solely on animal or human data. While the direct peptide reactivity assay (DPRA) is considered one of the NAMs of key event (KE) 1 within the OECD guideline 497 in combination with other NAMs for predicting skin sensitization hazard or potency, the assay is limited by the lack of activation features for pre-/pro-haptens. To address this, the peroxidase peptide reactivity assay (PPRA) was developed, utilizing horseradish peroxidase (HRP) and H₂O₂ to facilitate the oxidation and activation of test substances. However, limited information is available on the chemical substrate scope and applicability domain of the PPRA. In this study, we investigated the substrate scope of HRP to gain insights into the mechanism of the PPRA. Based on our analysis, the substrates of HRP include substituted phenols (or aromatic alcohols) and aniline (or aromatic amines) as well as their O- or N-alkyl derivatives. By considering the substrate scope of HRP, depletion patterns and mechanisms in the DPRA/PPRA, and the underlying chemistry of the assays, we categorized chemicals into five distinct chemical groups with unique structural features and depletion patterns in the DPRA/PPRA. This study elucidates the relationship between chemical structures, assay results of the DPRA and PPRA, and their applicability for predicting the skin sensitization potential. These findings contribute to a better understanding of the predictive capabilities of the PPRA and provide valuable insights for incorporating PPRA into next-generation risk assessments (NGRAs).

Electronic nicotine delivery systems (ENDS) are now increasingly used, with commercial electronic cigarettes frequently containing high levels of nicotine and menthol, which is a popular flavoring agent. This has raised multiple concerns about the health risks associated with menthol-flavored ENDS. Although menthol and nicotine are known for their individual effects on respiratory health, their combined impact on pulmonary surfactants remains poorly understood. Therefore, this study aimed at understanding the interactions between the primary components of all ENDS liquids (PG and VG), nicotine and menthol flavoring, and the pulmonary surfactant. This in vitro study used 1,2 dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and a stoichiometric mixture of DPPC/1-palmitoyl-2-Oleoyl-sn-glycero-3-phosphocholine (POPC)/2-Oleoyl-1-palmitoyl-sn-glycero-3- phospho-rac-(1-glycerol) sodium salt (POPG)/cholesterol at 48/32/10/10 to mimic the pulmonary surfactant. These systems were probed using a Langmuir–Blodgett Trough and attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy. The results indicate a concentration dependence of the impact of different nicotine concentrations combined with menthol on the surfactant mimics. Our findings also reveal the effect of menthol on the surface pressure. The combination of nicotine and menthol appears to alter the conformational state of the surfactant, proximately altering characteristic vibrational groups. Moreover, different behaviors are unveiled between the two model surfactants, particularly attributed to the complexities of the four surfactants mixture. Further research is suggested to address the mechanisms and implications involved with ENDS flavoring and additives on surfactant molecules in biological systems. Establishing well-informed regulations on ENDS consumption and distribution should be developed.

The use of two-dimensional organomodified nanoclays (ONCs) to improve nanocomposite properties continues to grow. Recent evidence suggests that airborne nanoclays in occupational environments pose an inhalation hazard; however, health risks and the underlying mechanisms remain undefined. In vivo studies evaluating pre- and post-incinerated ONC exposures found that cytotoxicity, inflammation, and fibrotic signaling responses are coating- and incineration status-dependent. We hypothesized that physicochemical property differences associated with coating presence/absence and incineration status of nanoclays will elicit changes in key events (KE) in exposed human small airway epithelial (SAECs) and normal lung fibroblast (NHLF) cells that contribute to pulmonary lung fibrosis. Using multiplex high-throughput screening strategies, SAEC and NHLF cells were acutely exposed (0–20 μg/cm²) to pristine nanoclay (CloisNa), an ONC (Clois30B), their incinerated byproducts (I-CloisNa and I-Clois30B), and crystalline silica (CS), to evaluate how ONC characteristics influence several KE in the pulmonary fibrosis adverse outcome pathway. In vitro exposure to pre-incinerated nanoclay induced organic coating-dependent cytotoxicity in SAECs. CloisNa caused disruption of mitochondrial membrane potential, which coincided with loss in viability in both cell types. Clois30B exposure caused dose-dependent SAEC cytotoxicity, micronuclei formation, and mitochondrial hyperpolarization in SAECs and NHLFs. Incinerated nanoclays were noncytotoxic but elicited a SAEC mitochondrial radical and pro-inflammatory response. Direct in vitro exposure to NHLFs exhibited particle-dependent increased live cell count, reactive oxygen species production, and α-smooth muscle actin expression. Nanoclay-exposed NHLFs (0.6 μg/cm²) possessed elevated collagen I levels while the same mass dose in vivo (300 μg/lung) favored elevated fibronectin and collagen III deposition for CloisNa and CS. In conclusion, organic coating presence and incineration status influenced nanoclays’ effects on cellular interaction, membrane integrity, inflammation, fibroblast activation, and collagen accumulation in exposed cell models. Although pre-incinerated nanoclay exposure promoted collagen accumulation in vitro, it was a poor predictor of in vivo model reticular fiber deposition.

Cadmium (Cd)-induced nephrotoxicity is a well-known phenomenon; however, several observational studies have used various biomarkers to monitor kidney injury in occupationally exposed populations. The markers used in these studies are found to be varied in sensitivity and are site-specific, and experts have the opinion that a single marker cannot predict the degree of kidney injury in human biomonitoring studies. Therefore, the current systematic review consolidates existing evidence to identify the association between Cd exposure and markers of potential sites of renal dysfunction/damage. Thirty (30) studies with 1980 chronic Cd exposure by occupations and 1292 unexposed were included in the review. The pooled mean difference of Cd exposure was as follows: blood Cd, 6.45 (5.18 to 7.71) μg/L; urine Cd, 4.52 (3.54 to 5.5) μg/g creatinine. Cd exposure was associated with impaired glomerular function (higher serum creatinine, serum β2 microglobulin, and lower creatinine clearance rate), tubular reabsorption (higher urinary β2 microglobulin and retinol binding protein), and injury (higher urinary N-acetyl-β-d-glucosaminidase and kidney injury molecule-1). However, the included studies exhibited high levels of heterogeneity. From the data, it is highly evident that biomarkers such as urinary N-acetyl-β-d-glucosaminidase, and retinol binding protein are found to be more sensitive than conventional clinical renal functional markers such as serum creatinine, urinary albumin, and protein levels, which are found to be within acceptable limits among the Cd-exposed group. Considering the rising disease burden of chronic kidney disease of unknown origin, Cd exposure-associated renal dysfunction and damage is a public health concern. Therefore, the review also discussed emerging biomarkers with higher sensitivity for early detection that can be adopted in occupational biomonitoring studies as early markers to prevent/delay the progression of kidney disease among the working population. Prospero Registration ID: CRD42022380923

Smoking is the leading cause of lung cancer. Differences in CYP2A6-catalyzed nicotine metabolism affect smoking dose and intensity, which, in turn, can affect lung cancer risk. CYP2A6 also catalyzes the bioactivation of the tobacco-specific lung carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). To determine the contribution of CYP2A6 to the metabolic activation of NNK, a group of Japanese American and Native Hawaiian smokers with little or no CYP2A6 activity was recruited to smoke [pyridyl-D₄]-NNK-containing cigarettes for a week. [Pyridyl-D₄]-4-hydroxy-4-(3-pyridyl)butanoic acid (D₄-hydroxy acid), the urinary product of NNK α-hydroxylation, the major bioactivation pathway, was quantified in these individuals and in an equal number of smokers with “normal” CYP2A6 activity. In expectation of low D₄-hydroxy acid levels, a sensitive nanoflow LC-MS/MS assay was developed. CYP2A6 activity was measured as the plasma ratio of 3′-hydroxycotinine to cotinine, which is the nicotine metabolite ratio (NMR). The average concentration of D₄-hydroxy acid in 24 h urine samples over 3 days was 20 ± 14 fmol/mL in low NMR (<0.05) smokers (n = 8) versus 33 ± 18 fmol/mL (p = 0.056) in “normal” NMR (>0.3) smokers (n = 8). The total D₄-hydroxy acid excreted by the low NMR group was half that of the higher NMR group (29.1 ± 16.8 versus 59.7 ± 45.3 pmol/24h, p = 0.048). These data support the role of CYP2A6 in the metabolic activation of NNK. However, it is unlikely that more modest differences in CYP2A6 activity, for example, as might be seen across smokers of European ancestry, would significantly impact NNK bioactivation. The influence of CYP2A6 activity on nicotine metabolism and the associated carcinogen uptake is likely the primary influence of CYP2A6 activity on a smoker’s risk of lung cancer, not a modest effect on the metabolic activation of NNK, one of several lung carcinogens in tobacco smoke.

We used liquid chromatography-nanoelectrospray ionization-high resolution tandem mass spectrometry (LC-NSI-HRMS/MS) to quantify DNA adducts released from human oral cell DNA upon neutral thermal hydrolysis followed by acid hydrolysis. The assay was applied to 80 buccal cell samples selected from those collected in the Shanghai Cohort Study, a prospective epidemiology study of 18,244 Chinese men 45–64 years old who resided in Shanghai, China when the samples were collected in 2001–2003. The DNA adducts quantified were 3-methyladenine (3-Me-Ade), 3-ethyladenine (3-Et-Ade), and 7-ethylguanine (7-Et-Gua). The method used hydrolysis of DNA samples containing the stable isotope labeled internal standards, solid phase extraction for adduct enrichment, and analysis by LC-NSI-HRMS/MS. Accuracy and precision of the analytical method were established with detection limits of 10–20 amol on column. Median levels of 3-Me-Ade -187 adducts/10⁹ nucleotides in smokers and 129 adducts/10⁹ nucleotides in nonsmokers; and 7-Et-Gua -49 adducts/10⁹ nucleotides in smokers and 21 adducts/10⁹ nucleotides in nonsmokers─were significantly higher in smokers than in nonsmokers (both P values <0.01). Levels of 3-Et-Ade -50 adducts/10⁹ nucleotides in smokers and 43 adducts/10⁹ nucleotides in nonsmokers - were not significantly different. These results demonstrate the applicability of a highly sensitive LC-NSI-HRMS/MS method for the analysis of human oral cell DNA for adducts released by neutral thermal and acid hydrolysis and show the significant effects of cigarette smoking on levels of 3-Me-Ade and 7-Et-Gua in this DNA. This is apparently the first study to characterize 3-Me-Ade in intact DNA isolated from any human tissue.

The development of alternative methods to animal testing has gained momentum over the years, including the rapid growth of in silico methods, which are faster and more cost-effective. A large number of computational tools have been published, focusing on Read-Across, (quantitative) Structure–Activity Relationship ((Q)SAR) models, and Physiologically Based Pharmacokinetic (PBPK) models. All of these methods play a crucial role in the risk assessment for cosmetics. However, despite the continuous efforts of various working groups, these methods are not always accepted by regulatory authorities around the world due to a lack of standardization and transparency in their development and application. This study aimed to identify in silico tools that can predict key properties relevant to the hazard assessment of cosmetic ingredients, aiming to streamline decision-making and assist toxicologists in efficiently selecting and integrating in silico predictions. Eighty-four in silico tools were identified based on their predictive capabilities, covering physicochemical parameters, toxicological/ecotoxicological endpoints, and toxicokinetic properties using different computational methods, e.g., (Q)SARs; Read-Across. Additional criteria were also considered for QSAR models, helping toxicologists integrate them into risk assessment processes: (1) definition of the Applicability Domain (AD), (2) model performance, and (3) nearest neighbors of the target substance. Based on these criteria, the models were classified as either useful for screening or suitable for a Weight of Evidence (WoE) approach. Finally, this study highlights the growing number of computational tools available for assessing various endpoints relevant to cosmetic safety. The number of tools continues to increase, and regular reviews are necessary. A deeper understanding of these in silico tools will facilitate their use by toxicologists and improve their acceptance for regulatory purposes from different cosmetic authorities.

Glioblastoma (GBM) is a lethal brain tumor with limited therapeutic options. Temozolomide (TMZ), a standard-of-care chemotherapeutic agent, exerts its cytotoxicity by alkylating DNA, which triggers a DNA damage response and depletes ATP and NAD⁺. However, TMZ also releases the byproduct 4-amino-5-imidazole carboxamide (AIC), which is believed to be a benign metabolite. We considered the possibility that AIC from TMZ could enter the de novo purine synthesis pathway, contributing to AMP and NAD⁺ synthesis and thus potentially antagonizing the anticancer activity of TMZ. The purpose of this article is to determine if AIC from TMZ can be incorporated into cellular purines. Using mass spectrometry with isotope-labeled TMZ, we demonstrate that the AIC derived from TMZ is incorporated into AMP and NAD⁺ in glioblastoma cell lines. Further, we performed an analysis of publicly available transcriptomic data from the Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression (GTEx) databases. Our analyses demonstrate that de novo purine synthesis is upregulated in GBM relative to the normal brain. Collectively, our findings demonstrate that a drug metabolite of TMZ, AIC, can be incorporated into de novo purine synthesis, which is upregulated in GBM.