Inhalation represents the most prevalent route of exposure with Thorium-232 compounds (Th-nitrate/Th-dioxide)/Th-containing dust in real occupational scenario. The present study investigated the mechanism of Th response in normal human alveolar epithelial cells (WI26), exposed to Th-nitrate or colloidal Th-dioxide (1–100 μg/ml, 24–72 h). Assessment in terms of changes in cell morphology, cell proliferation (cell count), plasma membrane integrity (lactate dehydrogenase leakage) and mitochondrial metabolic activity (MTT reduction) showed that Th-dioxide was quantitatively more deleterious than Th-nitrate to WI26 cells. TEM and immunofluorescence analysis suggested that Th-dioxide followed a clathrin/caveolin-mediated endocytosis, however, membrane perforation/non-endocytosis seemed to be the mode of Th internalization in cells exposed to Th-nitrate. Th-estimation by ICP-MS showed significantly higher uptake of Th in cells treated with Th-dioxide than with Th-nitrate at a given concentration. Both Th-dioxide and nitrate were found to increase the level of reactive oxygen species, which seemed to be responsible for lipid peroxidation, alteration in mitochondrial membrane potential and DNA-damage. Amongst HSPs, the protein levels of HSP70 and HSP90 were affected differentially by Th-nitrate/dioxide. Specific inhibitors of ATM (KU55933) or HSP90 (17AAG) were found to increase the Th- cytotoxicity suggesting prosurvival role of these signaling molecules in rescuing the cells from Th-toxicity.

Glyphosate is the most widely used herbicide in the world. In recent years, many studies have demonstrated that exposure to glyphosate-based herbicides (GHBs) was related to the decrease of serum testosterone and the decline in semen quality. However, the molecular mechanism of glyphosate-induced testosterone synthesis disorders is still unclear. In the present study, the effects of glyphosate on testosterone secretion and the role of endoplasmic reticulum (ER) stress in the process were investigated in TM3 cells. The effects of glyphosate at different concentrations on the viability of TM3 cells were detected by CCK8 method. The effect of glyphosate exposure on testosterone secretion was determined by enzyme-linked immunosorbent assay (ELISA). The expression levels of testosterone synthases and ER stress-related proteins were detected by Western blot and Immunofluorescence stain. Results showed that exposure to glyphosate at concentrations below 200 mg/L had no effect on cell viability, while the glyphosate above 0.5 mg/L could inhibit the testosterone secretion in TM3 cells. Treatment TM3 cells with glyphosate at 5 mg/L not only reduced the protein levels of testosterone synthase StAR and CYP17A1, inhibited testosterone secretion, but also increased the protein level of ER stress molecule Bip and promoted the phosphorylation of PERK and eIF2α. Pretreatment cells with PBA, an inhibitor of ER stress, alleviated glyphosate-induced increase in Bip, p-PERK and p-eIF2α protein levels, meanwhile rescuing glyphosate-induced testosterone synthesis disorders. When pretreatment with GSK2606414, a PERK inhibitor, the glyphosate-induced phosphorylation of PERK and eIF2α was blocked, and the glyphosate-inhibited testosterone synthesis and secretion was also restored. Overall, our findings suggest that glyphosate can interfere with the expression of StAR and CYP17A1 and inhibit testosterone synthesis and secretion via ER stress-mediated the activation of PERK/eIF2α signaling pathway in Leydig cells.

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.

Puberty is a critical period for growth and development. This period is sensitive to external stimuli, which ultimately affects the development of nerves and the formation of social behaviour. 17β-Trenbolone (17β-TBOH) is an endocrine disrupting chemicals (EDCs), which had been widely reported in aquatic vertebrates. But there is little known about the effects of 17β-TBOH on mammals, especially on adolescent neurodevelopment. In this study, we found that 17β-TBOH acute 1 h exposure can cause the activation of the dopamine circuit in pubertal male balb/c mice. At present, there is little known about the effects of puberty exposure of endocrine disruptors on these neurons/nerve pathways. Through a series of behavioural tests, exposure to 80 μgkg⁻¹ d⁻¹ of 17β-TBOH during adolescence increased the anxiety-like behaviour of mice and reduced the control of wheel-running behaviour and the response of social interaction behaviour. The results of TH immunofluorescence staining showed that exposure to 17β-TBOH reduced dopamine axon growth in the medial prefrontal cortex (mPFC). In addition, the results of real-time PCR showed that exposure to 17β-TBOH not only down-regulated the expression of dopamine axon development genes, but also affected the balance of excitatory/inhibitory signals in mPFC. In this research, we reveal the effects of 17β-TBOH exposure during adolescence on mammalian behaviour and neurodevelopment, and provide a reference for studying the origin of adolescent diseases.

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.

Biofilms containing pathogenic organisms from the water supply are a potential source of protozoan parasite outbreaks and a significant public health concern. The aim of the present study was to demonstrate the simultaneous and multi-spatial occurrence of waterborne protozoan pathogens (WBPP) in substrate-associated biofilms (SAB) and compare it to surface water (SW) and sediments with bottom water (BW) counterparts using manual filtration and elution from low-volume samples. For scenario purposes, simulated environmental biofilm contamination was created from in-situ grown one-month-old SAB (OM-SAB) that were spiked with Cryptosporidium parvum oocysts. Samples were collected from the largest freshwater reservoirs in Luzon, Philippines and a University Lake in Thailand. A total of 69 samples (23 SAB, 23 SW, and 23 BW) were evaluated using traditional staining techniques for Cryptosporidium, and Immunofluorescence staining for the simultaneous detection of Cryptosporidium and Giardia. WBPP were found in 43% SAB, 39% SW, and 39% BW of the samples tested in the present study with SAB results reflecting SW and BW results. Further highlights were demonstrated in the potential of using low-volume samples for the detection of parasites in source water. Scanning electron microscopy of OM-SAB samples revealed a naturally-associated testate amoeba shell, while Cryptosporidium oocysts spiked samples provided a visual profile of what can be expected from naturally contaminated biofilms. This study provides the first evidence for the simultaneous and multi-spatial occurrence of waterborne protozoan pathogens in low-volume aquatic matrices and further warrants SAB testing along with SW and BW matrices for improved water quality assessment strategies (iWQAS).

Fluoride is capable of inducing neurotoxicity, but its mechanisms remain elusive. This study aimed to explore the roles of endoplasmic reticulum (ER) stress and autophagy in sodium fluoride (NaF)-induced neurotoxicity, focusing on the regulating role of ER stress in autophagy. The in vivo results demonstrated that NaF exposure impaired the learning and memory capabilities of rats, and resulted in histological and ultrastructural abnormalities in rat hippocampus. Moreover, NaF exposure induced excessive ER stress and associated apoptosis, as manifested by elevated IRE1α, GRP78, cleaved caspase-12 and cleaved-caspase-3, as well as defective autophagy, as shown by increased Beclin1, LC3-II and p62 expression in hippocampus. Consistently, the in vitro results further verified the findings of in vivo study that NaF induced excessive ER stress and defective autophagy in SH-SY5Y cells. Notably, inhibition of autophagy in NaF-treated SH-SY5Y cells with Wortmannin or Chloroquine decreased, while induction of autophagy by Rapamycin increased the cell viability. These results were correlated well with the immunofluorescence observations, thus confirming the pivotal role of autophagic flux dysfunction in NaF-induced cell death. Importantly, mitigation of ER stress by 4-phenylbutyrate in NaF-treated SH-SY5Y cells inhibited the expressions of autophagy markers, and decreased cell apoptosis. Taken together, these data suggest that neuronal death resulted from excessive ER stress and autophagic flux dysfunction contributes to fluoride-elicited neurotoxicity. Moreover, the autophagic flux dysfunction was mediated by excessive ER stress, which provided novel insight into a better understanding of fluoride-induced neurotoxicity.

Colon microenvironment and microbiota dysbiosis are closely related to various human metabolic diseases. In this study, a total of 72 healthy female mice were exposed to fluoride (F) (0, 25, 50 and 100 mg/L F⁻) in drinking water for 70 days. The effect of F on intestinal barrier and the diversity and composition in colon microbiota have been evaluated. Meanwhile, the relationship among F-induced colon microbiota alterations and antimicrobial peptides (AMPs) expression and short-chain fatty acids (SCFAs) level also been assessed. The results suggested that F decreased the goblet cells number and glycoprotein expression in colon. And further high-throughput 16S rRNA gene sequencing result demonstrated that F exposure induced the diversity and community composition of colonic microbiota significantly changes. Linear Discriminant Analysis Effect Size (LEfSe) analysis identified 11 predominantly characteristic taxa which may be the biomarker in response to F exposure. F-induced intestinal microbiota perturbations lead to the significantly decreased SCFAs levels in colon. Immunofluorescence results showed that F increased the protein expression of interleukin-17A (IL-17A) and IL-22 (P < 0.01) and disturbed the expression of interleukin-17 receptor A (IL-17RA) and IL-22R (P < 0.05 or P < 0.01). In addition, the increased expression of IL-17A and IL-22 cooperatively enhanced the mRNA expression of AMPs which response to F-induced microbiota perturbations. Collectively, destroyed microenvironment and disturbed AMPs are the primary reason of microbiota dysbiosis in colon after F exposure. Colonic homoeostasis imbalance would be helpful for finding the source of F-induced chronic systemic diseases.

Zearalenone (ZEA), an estrogen-like mycotoxin, is commonly detected in animal feeds including improperly stored grains. It has been well demonstrated that ovarian granulosa cells (GCs) perform vital roles during follicular development, however, the competing endogenous RNA (ceRNA) network in GCs after ZEA exposure remains to be well described. Here, for the first time, we adopted whole-transcriptome sequence technology to explore the molecular mechanism of ZEA toxicology on porcine GCs. The results provide evidence that the cell cycle of porcine GCs is arrested in the G2/M phase after exposure to ZEA. Furthermore, bioinformation analysis found that cell cycle arrest related genes were perturbed, including CDK1, CCNB1, CDC25A, and CDC25C, which was consistent with the results of RT-qPCR, immunofluorescence, and Western Blotting. Based on the whole-transcriptome sequence data, by constructing ceRNA networks related to cell cycle arrest, we observed that ZEA exposure arrested cell cycle progression at the G2/M phase in porcine GCs, and non-coding RNAs (ncRNAs) played an important role in this process via regulating the expressions of cell cycle arrest related genes. Taken together, our data here provides strong data to support that the toxicological mechanism regarding the widely distributed toxicant ZEA acts through ceRNA networks in porcine granulosa cells.

With the development of modern industry, the problem of cadmium (Cd) pollution cannot be ignored and its toxicity has caused great personal injury to humans. Poly (ADP-ribose) polymerase 1 (PARP-1) protein is a research hotspot in recent years, the research we have published shows that 5 μM of Cd-treated NRK-52E cells activated PARP-1, but the specific effects of PARP-1 on DNA damage and cell cycle is unclear. Therefore, the purpose of this study is to reveal the effect of Cd on DNA damage and cell cycle arrest in NRK-52E cells, in addition, to investigate the role of PARP-1 in mediating this effect. Western blotting, comet assay, QRT-PCR, immunofluorescence, and co-immunoprecipitation were used to detect DNA damage and cell cycle-associated protein expression. Flow cytometry was used to assess cell cycle distribution and the apoptosis rates. Results showed that after the increase in treatment time and Cd concentration, the degree of DNA damage was significantly increased, and a transition from G0/G1 to S phase arrest was observed. In addition, inhibition of PARP-1 expression exacerbated cell damage and cell cycle arrest when DNA damage was low, but attenuated cell damage and even cell cycle arrest when DNA damage was severe. These findings in this study indicate that Cd causes DNA damage in NRK-52E cells, leading to cell cycle arrest at different phases depending on the degree of DNA damage. Moreover, PARP-1 plays an important role in mediating this effect, when DNA damage is low, it functions in DNA repair, however, when DNA damage is severe, it aggravates cell damage and induces cell death.