Sensitive Detection of Genotoxic Substances in Complex Food Matrices by Multiparametric High-Content Analysis.

Genotoxic substances widely exist in the environment and the food supply, posing serious health risks due to their potential to induce DNA damage and cancer. Traditional genotoxicity assays, while valuable, are limited by insufficient sensitivity, specificity, and efficiency, particularly when applied to complex food matrices. This study introduces a multiparametric high-content analysis (HCA) for the detection of genotoxic substances in complex food matrices. The developed assay measures three genotoxic biomarkers, including γ-H2AX, p-H3, and RAD51, which enhances the sensitivity and accuracy of genotoxicity screening. Moreover, the assay effectively distinguishes genotoxic compounds with different modes of action, which not only offers a more comprehensive assessment of DNA damage and the cellular response to genotoxic stress but also provides new insights into the exploration of genotoxicity mechanisms. Notably, the five tested food matrices, including coffee, tea, pak choi, spinach, and tomato, were found not to interfere with the detection of these biomarkers under proper dilution ratios, validating the robustness and reliability of the assay for the screening of genotoxic compounds in the food industry. The integration of multiple biomarkers with HCA provides an efficient method for detecting and assessing genotoxic substances in the food supply, with potential applications in toxicology research and food safety.

Sensitive Detection and Differentiation of Biologically Active Ricin and Abrin in Complex Matrices via Specific Neutralizing Antibody-Based Cytotoxicity Assay.

Ricin and abrin are highly potent plant-derived toxins, categorized as type II ribosome-inactivating proteins. High toxicity, accessibility, and the lack of effective countermeasures make them potential agents in bioterrorism and biowarfare, posing significant threats to public safety. Despite the existence of many effective analytical strategies for detecting these two lethal toxins, current methods are often hindered by limitations such as insufficient sensitivity, complex sample preparation, and most importantly, the inability to distinguish between biologically active and inactive toxin. In this study, a cytotoxicity assay was developed to detect active ricin and abrin based on their potent cell-killing capability. Among nine human cell lines derived from various organs, HeLa cells exhibited exceptional sensitivity, with limits of detection reaching 0.3 ng/mL and 0.03 ng/mL for ricin and abrin, respectively. Subsequently, toxin-specific neutralizing monoclonal antibodies MIL50 and 10D8 were used to facilitate the precise identification and differentiation of ricin and abrin. The method provides straightforward and sensitive detection in complex matrices including milk, plasma, coffee, orange juice, and tea via a simple serial-dilution procedure without any complex purification and enrichment steps. Furthermore, this assay was successfully applied in the unambiguous identification of active ricin and abrin in samples from OPCW biotoxin exercises.

Total synthesis and herbicidal activity of natural rubrolides C, E and F.

Rubrolides are natural butyrolactones isolated from the tunicate Ritterella rubra, shows antibacterial, antiviral and plant photosynthesis inhibitory activities. In this study, a facile total synthetic method for preparing the rubrolides from benzaldehyde by a Darzens reaction, aldol reaction and vinylogous aldol condensation in five steps is presented. Three natural rubrolides (E, C and F) were synthesised in the total yields of 25-40%. The bioassay results indicate that rubrolides E, C and F exhibit some herbicidal inhibitory effect against rapeseed, in particular, rubrolide F shows the best herbicidal activities against rapeseed root with the growth inhibitory rate of 72.8%. At greenhouse treatment concentrations of 100 and 500 mg/L, rubrolide F show a positive dose-toxicity correlation towards abutilon plants. Collectively, facile total Synthesis strategy provided the base for further bioactivities study of rubrolides family. Rubrolide F may be act as inhibitor of photosynthesis, and this could be lead structure of new herbicide.

Amanitin-induced variable cytotoxicity in various cell lines is mediated by the different expression levels of OATP1B3.

Amanita phalloides is one of the deadliest mushrooms worldwide, causing most fatal cases of mushroom poisoning. Among the poisonous substances of Amanita phalloides, amanitins are the most lethal toxins to humans. Currently, there are no specific antidotes available for managing amanitin poisoning and treatments are lack of efficacy. Amanitin mainly causes severe injuries to specific organs, such as the liver, stomach, and kidney, whereas the lung, heart, and brain are hardly affected. However, the molecular mechanism of this phenomenon remains not understood. To explore the possible mechanism of organ specificity of amanitin-induced toxicity, eight human cell lines derived from different organs were exposed to α, ß, and γ-amanitin at concentrations ranging from 0.3 to 100 µM. We found that the cytotoxicity of amanitin differs greatly in various cell lines, among which liver-derived HepG2, stomach-derived BGC-823, and kidney-derived HEK-293 cells are most sensitive. Further mechanistic study revealed that the variable cytotoxicity is mainly dependent on the different expression levels of the organic anion transporting polypeptide 1B3 (OATP1B3), which facilitates the internalization of amanitin into cells. Besides, knockdown of OATP1B3 in HepG2 cells prevented α-amanitin-induced cytotoxicity. These results indicated that OATP1B3 may be a crucial therapeutic target against amanitin-induced organ failure.

Divinyl sulfone, an oxidative metabolite of sulfur mustard, induces caspase-independent pyroptosis in hepatocytes.

Sulfur mustard (SM) is a highly toxic blister agent which has been used many times in several wars and conflicts and caused heavy casualties. Ease of production and lack of effective therapies make SM a potential threat to public health. SM intoxication causes severe damage on various target organs, such as the skin, eyes, and lungs. In addition, SM exposure can also lead to hepatotoxicity and severe liver injuries. However, despite decades of research, the molecular mechanism underlying SM-induced liver damage remains obscure. SM can be converted into various products via complex hepatic metabolism in vivo. There are some pieces of evidence that one of the oxidation products of SM, divinyl sulfone (DVS), exhibits even more significant toxicity than SM. Nevertheless, the molecular toxicology of DVS is still hardly known. In the present study, we confirmed that DVS is even more toxic than SM in the human hepatocellular carcinoma cell line HepG2. Further mechanistic study revealed that DVS exposure (200 µM) promotes pyroptosis in HepG2 cells, while SM (400 µM) mainly induces apoptosis. DVS induces gasdermin D (GSDMD) mediated pyroptosis, which is independent of caspases activation but depends on the large amounts of reactive oxygen species (ROS) and severe oxidative stress produced during DVS exposure. Our findings may provide novel insights for understanding the mechanism of SM poisoning and may be helpful to discover promising therapeutic strategies for SM intoxication.