Network Toxicology and Molecular Docking to Investigate the Non-AChE Mechanisms of Organophosphate-Induced Neurodevelopmental Toxicity.

Organophosphate pesticides (OPs) are toxic substances that contaminate aquatic environments, interfere with the development of the nervous system, and induce Neurodevelopmental Toxicity (NDT) in animals and humans. The canonical mechanism of OP neurotoxicity involves the inhibition of acetylcholinesterase (AChE), but other mechanisms non-AChE are also involved and not fully understood. We used network toxicology and molecular docking to identify molecular targets and toxicity mechanisms common to OPs. Targets related to diazinon-oxon, chlorpyrifos oxon, and paraoxon OPs were predicted using the Swiss Target Prediction and PharmMapper databases. Targets related to NDT were compiled from GeneCards and OMIM databases. In order to construct the protein-protein interaction (PPI) network, the common targets between OPs and NDT were imported into the STRING. Network topological analyses identified EGFR, MET, HSP90AA1, and SRC as hub nodes common to the three OPs. Using the Reactome pathway and gene ontology, we found that signal transduction, axon guidance, cellular responses to stress, and glutamatergic signaling activation play key roles in OP-induced NDT.

An innovative insecticidal approach based on plant protease inhibitor and Bt protoxins inhibits trypsin-like activity in zebrafish.

The Leucaena leucocephala trypsin inhibitor (LTI) + Bacillus thuringiensis (Bt) protoxins mix has been proposed as a novel larvicide agent in order to control the vector mosquito of dengue virus, Aedes aegypti, in their aquatic breeding sites. However, use of this insecticide formulation has raised concerns about its impacts on aquatic biota. In this context, this work aimed to assess the effects of LTI and Bt protoxins, separately or in combination, in zebrafish, in regard to the evaluation of toxicity at early life stages and to the presence of LTI inhibitory effects on intestinal proteases of this fish. Results showed that LTI and Bt concentrations (250 mg/L, and 0.13 mg/L, respectively), and LTI + Bt mix (250 mg/L + 0.13 mg/L) - 10 times superior to those with insecticidal action - did not cause death nor did it induce morphological changes during embryonic and larval development (3 to 144 h post-fertilization) of zebrafish. Molecular docking analyses highlighted a possible interaction between LTI and zebrafish trypsin, especially through hydrophobic interactions. In concentrations near to those with larvicidal action, LTI (0.1 mg/mL) was able to inhibit in vitro intestinal extracts of trypsin in female and male fish by 83 % and 85 %, respectively, while LTI + Bt mix promoted trypsin inhibition of 69 % in female and 65 % in male ones. These data show that the larvicidal mix can potentially promote deleterious effects to nutrition and survival in non-target aquatic organisms, especially those with trypsin-like dependent protein digestion.

Toxicological assessment of a bioactive extract from Triplaris gardneriana Wedd. seeds using alternative models.

The Triplaris gardneriana Wedd. seeds extract has great therapeutic potential due to numerous biological activities such as antioxidant, antibacterial and anti-inflammatory, which are associated with phenolic content. Although this herbal preparation has shown many benefits, recently their toxicity profile has begun to be explored. In this present study, the toxic effects of T. gardneriana seeds ethanolic extract (EETg) on biological systems of different taxonomical groups and levels of complexity (from cell culture to lower vertebrates) were assessed, through a variety of viability and toxicological assays. It was found that EETg did not impair the Saccharomyces cerevisiae growth at the highest tested concentration (200 µg/mL), and no toxicant evidence was observed in Aedes aegypti larvae or in Drosophila melanogaster adult stage. Contrarily, the extract reduced the viability of undifferentiated Caco-2 cells (250 µg/mL, 40% of viable cells), but did not affect differentiated ones. The embryotoxicity in Danio rerio model showed a LC50 of 7.41 mg/L (95% confidence interval, 4.78 - 11.49 mg/L). EETg did not show signs of toxicity in the majority of the models used, but lethality and malformations in zebrafish embryos occurred. Further analyses are needed to better understand the selective toxicity mechanism of EETg on zebrafish, as well as whether the toxic effects happen in higher vertebrates.

In vitro antibacterial and anti-inflammatory effects of Anacardium occidentale L. extracts and their toxicity on PBMCs and zebrafish embryos.

This study evaluated the in vitro antimicrobial and immunomodulatory action of crude extracts from Anacardium occidentale L. (cashew tree) leaves and bark, and to determine their toxicity to peripheral-blood mononuclear cells (PBMCs) and to zebrafish embryos and larvae. Chemical analysis of extracts was performed by proton nuclear magnetic resonance (1H-NMR). The antibacterial activity was evaluated against selected bacteria strains by determining the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). Cytotoxicity of the extracts was assessed using resazurin method, while the effect on production of ROS by PMN leukocytes was measured by luminol. Embryotoxicity to zebrafish was assessed using the fish embryo acute toxicity test (FET) and quantification of toxicity marker enzymes (AChE, LDH, and GST). 1H-NMR results showed anacardic acid as the main component of the extracts. All bacterial species tested were sensitive to the extracts, with MICs ranging from 312.5 to 10,000 µg/mL. Streptococcus mutans and Escherichia coli were the most susceptible species. The extracts promoted cell viability above 75% at concentrations from 1.25 to 80 µg/mL. Both extracts reduced zymosan-induced ROS (p < 0.05) at concentrations of 1, 8, and 80 µg/mL compared to the control. In vivo, there were embryotoxic effects in zebrafish embryos exposed to both extracts through the presence of lethal and sublethal endpoints. The samples also acted by inhibiting the activities of biomarker enzymes. The A. occidentale L. bark and leaf extracts showed antimicrobial potential and modulated ROS production in vitro, but these also showed embryotoxic effects to zebrafish.

The new psychoactive substances 25H-NBOMe and 25H-NBOH induce abnormal development in the zebrafish embryo and interact in the DNA major groove.

Toxicological effects of 25H-NBOMe and 25H-NBOH recreational drugs on zebrafish embryos and larvae at the end of 96 h exposure period were demonstrated. 25H-NBOH and 25H-NBOMe caused high embryo mortality at 80 and 100 µg mL-1, respectively. According to the decrease in the concentration tested, lethality decreased while non-lethal effects were predominant up to 10 and 50 µg mL-1 of 25H-NBOH and 25H-NBOMe, respectively, including spine malformation, egg hatching delay, body malformation, otolith malformation, pericardial edema, and blood clotting. We can disclose that these drugs have an affinity for DNA in vitro using biophysical spectroscopic assays and molecular modeling methods. The experiments demonstrated that 25H-NBOH and 25H-NBOMe bind to the unclassical major groove of ctDNA with a binding constant of 27.00 × 104 M-1 and 5.27 × 104 M-1, respectively. Furthermore, these interactions lead to conformational changes in the DNA structure. Therefore, the results observed in the zebrafish embryos and DNA may be correlated.