peer reviewed | Metconazole (MEZ), a chiral triazole fungicide, produces enantioselective adverse effects in non-target organisms. Among MEZ's isomers, cis-MEZ displays robust antimicrobial properties. Evaluating MEZ and cis-MEZ's toxicity may mitigate fungicide usage and safeguard non-target organisms. Our study evaluated the toxicity of MEZ and its cis-isomers at concentrations of 0.02, 0.2, 2, and 4 mg L−1. We report stereoselectivity and severe cardiovascular defects in zebrafish, including pericardial oedema, decreased heart rate, increased sinus venous and bulbous arteries distances, intersegmental vessel defects, and altered cardiovascular development genes (hand2, gata4, nkx2.5, tbx5, vmhc, amhc, dll4, vegfaa, and vegfc). Further, MEZ significantly increased oxidative stress and apoptosis in zebrafish, primarily in the cardiac region. Isoquercetin, an antioxidant found in plants, partially mitigates MEZ-induced cardiac defects. Furthermore, MEZ upregulated the Wnt/β-catenin pathway genes (wnt3, β-catenin, axin2, and gsk-3β) and β-catenin protein expression. Inhibitor of Wnt Response-1 (IWR-1) rescued MEZ-induced cardiotoxicity. Our findings highlight oxidative stress, altered cardiovascular development genes, and upregulated Wnt/β-catenin signaling as contributors to cardiovascular toxicity in response to MEZ and cis-MEZ treatments. Importantly, 1R,5S-MEZ exhibited greater cardiotoxicity than 1S,5R-MEZ. Thus, our study provides a comprehensive understanding of cis-MEZ's cardiovascular toxicity in aquatic life. © 2024 Elsevier Ltd

Bisphenol A (BPA) is widely used by manufacturers and in consumer products. Its release in the environment may affect male reproductive function. In this study, we examined the effect of low dose (0.1 mg/kg BW), short term exposure during puberty (PD21-35) on adult rat male reproduction. The results indicated that such exposure reset growth hormone (GH) and follicular stimulating hormone (FSH) homeostasis and resulted in a significantly higher level of serum testosterone without affecting serum luteinizing hormone level. QPCR and Western blot results showed that BPA significantly up-regulated selective genes/proteins in the Leydig cell steroidogenic pathway, including steroidogenic acute regulatory protein, cytochrome P450 11A1, cytochrome P450 17A, and low-density lipoprotein receptor. RNA-Seq analysis of testicular RNAs showed that BPA significantly affected the gene profiles of multiple testicular interstitial populations without affecting germ cells. Also, GO- and KEGG-analysis suggested that IGF1-related PI3K/AKT signaling was activated, which was confirmed by the increased phosphorylation of IRS1, AKT1 and CREB. The results indicated that a low-dose, short-term BPA exposure during puberty affected the adult male rat pituitary (GH and FSH) and testis (testosterone) homeostasis.

Neuroinflammation induced by lead exposure (Pb) is a major cause of neurotoxicity of Pb in the central nervous system (CNS). The NLR family, domain of pyrin containing 3 (NLRP3) involves in various neurological diseases, while the question of whether NLRP3 plays a role in lead-induced neuroinflammation has not yet been reported.Developmental and knockout (KO) NLRP3 mice were used to establish two in vivo models, and BV2 cells were used to establish an in vitro model. Behavioral and electrophysiologic tests were used to assess the neurotoxicity of Pb, and immunofluorescence staining was used to assess neuroinflammation. Real-time PCR and western blot were performed to examine the mRNA and protein levels of inflammatory cytokines and NLRP3 inflammasomes. siRNA technology was used to block NLRP3 expression.Pb exposure led to neural injure and microglial activation in the hippocampus region, while minocycline intervention attenuated Pb-induced neurotoxicity by inhibiting neuroinflammation. Pb increased the expression of NLRP3 and promoted cleavage of caspase-1 in mRNA and protein levels, and minocycline partially reversed the effects of Pb on NLRP3 inflammasomes. Blocking of NLRP3 by KO mice or siRNA attenuated neural alterations induced by Pb, weakened microglial activation in vivo and in vitro as well, without affecting the accumulation of Pb. Pb increased autophagic protein levels and phosphorylation of NF-κB, while suppressing autophagy or NF-κB inhibited Pb's effects on NLRP3.NLRP3 is involved in the regulation of Pb-induced neurotoxicity. These findings expand mechanism research of Pb neurotoxicity and may help establish new prevention strategies for Pb neurotoxicity.

Forchlorfenuron (CPPU) has been used worldwide, to boost size and improve quality of various agricultural products. CPPU and its metabolites are persistent and have been detected frequently in fruits, water, sediments, and organisms in aquatic systems. Although the public became aware of CPPU through the exploding watermelon scandal of 2011 in Zhenjiang, China, little was known of its potential effects on the environment and wildlife. In this study, adverse effects of CPPU on developmental angiogenesis and vasculature, which is vulnerable to insults of persistent toxicants, were studied in vivo in zebrafish embryos (Danio rerio). Exposure to 10 mg CPPU/L impaired survival and hatching, while development was hindered by exposure to 2.5 mg CPPU/L. Developing vascular structure, including common cardinal veins (CCVs), intersegmental vessels (ISVs) and sub-intestinal vessels (SIVs), were significantly restrained by exposure to CPPU, in a dose-dependent manner. Also, CPPU caused disorganization of the cytoskeleton. In human umbilical vein endothelial cells (HUVECs), CPPU inhibited proliferation, migration and formation of tubular-like structures in vitro. Results of Western blot analyses revealed that exposure to CPPU increased phosphorylation of FLT-1, but inhibited phosphorylation of FAK and its downstream MAPK pathway in HUVECs. In summary, CPPU elicited developmental toxicity to the developing endothelial system of zebrafish and HUVECs. This was do, at least in part due to inhibition of the FAK/MAPK signaling pathway rather than direct interaction with the VEGF receptor (VEGFR).

It is very important to explore the potential harm and underlying mechanism of fluoride due to the extensive distribution and the significant health risks of fluoride in environment. The objective of this study to investigate whether fluoride can induce mitochondrial impairment and mitophagy in testicular cells. For this, 40 male mice were randomly divided into four groups treated with 0, 0.6, 1.2, 2.4 mM NaF deionized water, respectively, for 90 days continuously. The results showed that mitophagy was triggered by F in testicular tissues, especially in the Leydig cells by transmission electron microscopy and mitophagy receptor PHB2 locations by immunofluorescence. Furthermore, TM3 Leydig cells line was employed and treated with 0, 0.125, 0.25, and 0.5 mM NaF for 24 h. The mitochondrial function indicators and mitophagy maker PHB2, COX IV and regulator PINK1 in transcript and protein levels in Leydig cells were examined by the methods of qRT-PCR, western blotting, and immunofluorescence co-localization. The results showed that fluoride decreased the mitochondrial membrane potential with a concomitant increase in the number of lysosomes. Meanwhile, fluoride exposure also increased the expressions of PINK1 and PHB2 in TM3 Leydig cells. These results revealed that fluoride could induce mitochondrial impairment and excessive PINK1/Parkin-mediated mitophagy in testicular cells, especially in Leydig cells, which could contribute to the elucidation of the mechanisms of F-induced male reproductive toxicity.

The placenta is essential for sustaining the growth of the fetus. The aim of this study was to investigate the role of the placenta in MCLR-induced significant reduction in fetal weight, especially the changes in placental structure and function. Pregnant mice were intraperitoneally injected with MCLR (5 or 20 μg/kg) from gestational day (GD) 13 to GD17. The results showed MCLR reduced fetal weight and placenta weight. The histological specimens of the placentas were taken for light and electron microscopy studies. The internal space of blood vessels decreased obviously in the placental labyrinth layer of mice treated with MCLR. After the ultrastructural examination, the edema and intracytoplasmic vacuolization, dilation of the endoplasmic reticulum and corrugation of the nucleus were observed. In addition, maternal MCLR exposure caused a reduction of 11β-hydroxysteroid dehydrogenase type 2 (HSD11B2) expression in placentae, a critical regulator of fetal development. Several genes of placental growth factors, such as Vegfα and Pgf and several genes of nutrient transport pumps, such as Glut1 and Pcft were depressed in placentas of MCLR-treated mice, however nutrient transporters Fatp1 and Snat4 were promoted. Moreover, significant increases in malondialdehyde (MDA) revealed the occurrence of oxidative stress caused by MCLR, which was also verified by remarkable decrease in the glutathione levels, total antioxidant capacity (T-AOC) as well as the activity of antioxidant enzymes. Real-time PCR and western blot analysis revealed that GRP78, CHOP, XBP-1, peIF2α and pIRE1 were remarkable increased in placentas of MCLR-treated mice, indicating that endoplasmic reticulum (ER) stress pathway was activated by MCLR. Furthermore, oxidative stress and ER stress consequently triggered apoptosis which contributed to the impairment of placental development. Collectively, these results suggest maternal MCLR exposure results in reduced fetal body weight, which might be associated with ROS-mediated endoplasmic reticulum stress and impairment in placental structure and function.

Epidemiological relationships between pesticide use and male infertility have been suggested for a long time. Etoxazole (ETX), an oxazoline pesticide, has been extensively used for pest eradication. It is considered relatively safe and has low mammalian toxicity because it specifically inhibits chitin synthesis. However, ETX may have toxic effects on the reproductive system. In this study, we examined the effects of ETX on the reproductive system using mouse testis cell lines (TM3 for Leydig cells and TM4 for Sertoli cells) and C57BL/6 male mice. We confirmed that ETX has anti-proliferative effects on the TM3 and TM4 cell lines. Moreover, ETX induced mitochondrial dysfunction and hampers calcium homeostasis. Western blot analysis of MAPK and Akt signaling cascades was performed to demonstrate the mode of action of ETX at a molecular level. Moreover, ETX induced misregulation of genes related to testicular function. Upon oral administration of ETX in C57BL/6 male mice, testis weight was reduced and transcriptional expression related to testis function was altered. These results indicate that ETX induces testicular toxicity by inducing mitochondrial dysfunction and calcium imbalance and regulating gene expression.

Di-(2-ethylhexyl) phthalate (DEHP) is a prevalent environmental contaminant that severely impacts the health of human and animals. Taxifolin (TAX), a plant flavonoid isolated from yew, exerts protective effects on cardiac diseases. Nevertheless, whether DEHP could induce cardiomyocyte hypertrophy and its mechanism remains unclear. This study aimed to highlight the specific molecular mechanisms of DEHP-induced cardiomyocyte hypertrophy and the protective potential of TAX against it. Chicken primary cardiomyocytes were treated with DEHP (500 μM) and/or TAX (0.5 μM) for 24 h. The levels of glucose and adenosine triphosphate (ATP) were detected, and cardiac hypertrophy-related genes were validated by real-time quantitative PCR (qRT-PCR) and Western blot (WB) in vitro. The results showed that DEHP-induced cardiac hypertrophy was ameliorated by TAX, as indicated by the increased cardiomyocyte area and expression of atrial natriuretic peptide (ANP), natriuretic peptides A-like (BNP) and β-myosin heavy cardiac muscle (β-MHC). Furthermore, DEHP induced cardiac hypertrophy via the interleukin 6 (IL-6)/Janus kinase (JAK)/signal transducer and activator of transcription 3 (STAT3) pathway in vitro. In addition, DEHP disrupted mitochondrial function and glycometabolism by activating the insulin-like growth factor 1 (IGF1)/phosphatidylinositol 3-kinase (PI3K) pathway and the peroxisome proliferator activated receptors (PPARs)/PPARG coactivator 1 alpha (PGC-1α) pathway to induce cardiac hypertrophy in vitro. Intriguingly, those DEHP-induced changes were obviously alleviated by TAX treatment. Taken together, cardiac hypertrophy was induced by DEHP via activating the IL-6/JAK/STAT3 signaling pathway, triggering glycometabolism disorder and mitochondrial dysfunction in vitro, can be ameliorated by TAX. Our findings may provide a feasible molecular mechanism for the treatment of cardiomyocyte hypertrophy induced by DEHP.

Microcystis aeruginosa is one of the main species of cyanobacteria that causes water blooms. M. aeruginosa can release into the water several types of microcystins (MCs), which are harmful to aquatic organisms and even humans. However, few studies have investigated the hepatotoxicity of M. aeruginosa itself in zebrafish in environments that simulate natural aquatic systems. The objective of this study was to evaluate the hepatotoxicity of M. aeruginosa in adult zebrafish (Danio rerio) after short-term (96 h) exposure and to elucidate the potential underlying mechanisms. Distinct histological changes in the liver, such as enlargement of the peripheral nuclei and sinusoids and the appearance of fibroblasts, were observed in zebrafish grown in M. aeruginosa culture. In addition, antioxidant enzyme activity was activated and protein phosphatase (PP) activity was significantly decreased with increasing microalgal density. A proteomic analysis revealed alterations in a number of protein pathways, including ribosome translation, immune response, energy metabolism and oxidative phosphorylation pathways. Western blot and real-time PCR analyses confirmed the results of the proteomic analysis. All results indicated that M. aeruginosa could disrupt hepatic functions in adult zebrafish, thus highlighting the necessity of ecotoxicity assessments for M. aeruginosa at environmentally relevant densities.

With the growing production and applications of silica nanoparticles (SiNPs), human exposure to these nanoparticles continues to increase. However, the possible hazards that SiNP exposure may pose to human cardiovascular system and the underlying mechanisms remain unclear. In the present study, the flow cytometry was employed to investigate the potential of four sizes (10, 25, 50, 100 nm) of SiNPs to induce the apoptosis of human umbilical vein endothelial cells (HUVECs) in culture. The apoptotic pathway was also explored through the determination of the protein expression and/or activation of p53, Bcl-2, Bax, caspases-9, -7, -3, and PARP by western blot. The results showed that all the four sizes of SiNPs could significantly elicit apoptosis in HUVECs at the tested concentrations (1, 5, 25 μg/mL), compared with the negative control (p < 0.05, p < 0.01). Moreover, the apoptotic rates were increased with the elevating levels and decreasing sizes of administrative SiNPs, showing both dose- and size-dependent effect relationships. Interestingly, the enhancing phosphorylation of p53 protein (Ser15), decreasing ratio of Bcl-2/Bax protein, and elevating activation of the downstream proteins, caspase-9, -7, -3 and PARP, were also observed with the decreasing sizes of tested SiNPs, indicating that the p53-caspase pathway is the main way of the SiNP-mediated apoptosis in HUVECs and that the size is an important parameter that determines the SiNPs' potential to induce cellular response.