Cadmium (Cd) is a ubiquitous toxic heavy metal derived mainly from industrial processes. In industrialized societies, individuals are exposed to a plethora of sources of Cd pollution. Cd can trigger serious diseases such as rheumatoid arthritis (RA) and chronic obstructive pulmonary disease (COPD) by the over-activating immune system. As an effector mechanism in innate immunity, neutrophil extracellular traps (NETs) not only play an important role in defending against infection but also lead to tissue damage. However, the role of NETs in Cd-induced lung damage process has not been previously studied. In this study, we aimed to investigate the potential effects of Cd-induced NETs on lung injury in vivo and further to clarify the molecular mechanisms of Cd-induced NETs formation. In vivo, Cd treatment destroyed the structural integrity of lung tissue and significantly increased the levels of NETs in the bronchoalveolar lavage fluid (BALF). The known NETs inhibitor DNase I ameliorated pathologic changes and significantly decreased levels of NETs in BALF, which suggesting the curial role of NETs in Cd-induced lung injury. Further investigation showed that Cd could significantly trigger NETs formation, which is composed of DNA backbone decorated with histones (H3) and neutrophils elastase (NE). The inhibitors of NADPH oxidase, ERK1/2 and p38 MAPK-signaling pathways significantly reduced the formation of NETs, and western blotting analysis also showed that Cd significantly increased the phosphorylation of p38 and ERK1/2 signaling pathways. Above results confirmed that NADPH oxidase, ERK1/2 and p38 MAPK-signaling pathways were related to Cd-induced NETs formation. In conclusion, NETs was involved in Cd-induced lung injury, and the mechanisms of Cd-induced NETs formation was via activating NADPH oxidase, ERK1/2 and p38 MAPK-signaling pathways, which might provide a new perspective in Cd-induced lung injury.

Hydrogen gas (H₂) has been shown as an important factor in plant tolerance to abiotic stresses, but the underlying mechanisms remain unclear. In the present study, the effects of H₂ and its interaction with nitric oxide (NO) on alleviating cadmium (Cd) stress in Brassica campestris seedlings were investigated. NO donor (SNP) or hydrogen-rich water (HRW) treatment showed a significant improvement in growth of Cd-stressed seedlings. Cd treatment upregulated both endogenous NO and H₂ (36% and 66%, respectively), and the increase of H₂ was prior to NO increase. When treated with NO scavenger (PTIO) or NO biosynthesis enzyme inhibitors (L-NAME and Gln), HRW-induced alleviation under Cd stress was prevented. Under Cd stress, HRW pretreatment significantly enhanced the NO accumulation, and together up-regulated the activity of NR (nitrate reductase) and expression of NR. HRW induced lower reactive oxygen species (ROS), higher AsA content, enhanced activity of POD (peroxidase) and SOD (superoxide dismutase) in seedling roots were inhibited by PTIO, L-NAME and Gln. Through proteomic analysis, the level of 29 proteins were changed in response to H₂ and NO-induced amelioration of Cd stress. Nearly half of them were involved in oxidation-reduction processes (about 20%) or antioxidant enzymes (approximately 20%). These results strongly indicate that in Cd-stressed seedlings, pretreatment with HRW induces the accumulation of H₂ (biosynthesized or permeated), which further stimulates the biosynthesis of NO through the NR pathway. Finally, H₂ and NO together enhance the antioxidant capabilities of seedlings in response to Cd toxicity.

Fomesafen, a long-lived protoporphyrinogen-oxidase inhibitor, specially developed for post-emergence control of broad-leaf weeds, is used widely in soybean fields in northern China (Dayan and Duke, 2010). The impact of fomesafen on microbial communities in rhizosphere soils, however, is unknown. In this study we examined fomesafen degradation as well as its effects in the rhizosphere of soybean plants grown in a greenhouse. Fomesafen had shorter half-life in rhizosphere soil than previously reported for bulk soil from the same location (87 vs 120 days). The enzyme activity of soil extracts and the microbial community composition of 16S rRNA genes (16S) amplified from soil DNA were also investigated. Although not immediately apparent, both the high (37.5 mg kg⁻¹) and low (18.75 mg kg⁻¹) doses of fomesafen significantly decreased urease and invertase activities in the rhizosphere soil from days 30 and 45 respectively until the end of the experiment (90 days). Analysis of 16S amplicons demonstrated that fomesafen had a dose dependent effect, decreasing alpha diversity and altering beta diversity. Significant phylum level decreases were observed in five of the ten phyla that were most abundant in the control. Proteobacteria was the only phylum whose relative abundance increased in the presence of fomesafen, driven by increases in the genera Methylophilacaea, Dyella, and Sphingomonas. The functional implications of changes in 16S abundance as predicted using PICRUSt suggested that fomesafen enriched for enzymes involved in xenobiotic metabolism and detoxification (cytochrome P450s and glutathione metabolism). Our data suggest that, despite being degraded more rapidly in the rhizosphere than in bulk soil, fomesafen had long-lasting functional impacts on the soil microbial community.

Hydroquinone (HQ), one of the main metabolites of benzene, is a well-known human leukemogen. However, the specific mechanism of how benzene or HQ contributes to the development of leukemia is unknown. In a previous study, we demonstrated the upregulation of DNA methyltransferase (DNMT) expression in HQ-induced malignant transformed TK6 (HQ-TK6) cells. Here, we investigated whether a regulatory loop between the long noncoding RNA FAS-AS1 and DNMT3b exists in HQ-TK6 cells and benzene-exposed workers. We found that the expression of FAS-AS1 was downregulated in HQ-TK6 cells and workers exposed to benzene longer than 1.5 years via histone acetylation, and FAS-AS1 expression was negatively correlated with the time of benzene exposure. Restoration of FAS-AS1 in HQ-TK6 cells promoted apoptosis and inhibited tumorigenicity in female nude mice. Interestingly, treatment with a DNMT inhibitor (5-aza-2-deoxycytidine), histone deacetylase inhibitor (trichostatin A), or DNMT3b knockout led to increased FAS-AS1 through increased H3K27ac protein expression in HQ-TK6 cells, and DNMT3b knockout decreased H3K27ac and DNMT3b enrichment to the FAS-AS1 promoter region, which suggested that DNMT3b and/or histone acetylation involve FAS-AS1 expression. Importantly, restoration of FAS-AS1 resulted in reduced expression of DNMT3b and SIRT1 and increased expression of FAS in both HQ-TK6 cells and xenograft tissues. Moreover, the average DNMT3b expression in 17 paired workers exposed to benzene within 1.5 years was decreased, but that of the remaining 103 paired workers with longer exposure times was increased. Conversely, DNMT3b was negatively correlated with FAS-AS1 expression. Both FAS-AS1 and DNMT3b influenced the enrichment of H3K27ac in the FAS promoter region by regulating the expression of SIRT1, consequently upregulating FAS expression. Taken together, these observations demonstrate crosstalk between FAS-AS1 and DNMT3b via a mutual inhibition loop and indicate a new mechanism by which FAS-AS1 regulates the expression of FAS in benzene-related carcinogenesis.

In ecological risk assessment, acute to chronic ratio (ACR) uncertainty factors are routinely applied to acute mortality benchmarks to estimate chronic toxicity thresholds. To investigate variability of aquatic ACRs, we first compiled and compared 56 and 150 pairs of acute and subchronic/chronic growth/reproductive toxicity data for fishes (Pimephales promelas (53), Danio rerio (2), and Oryzias latipes (1)) and the crustacean Daphnia magna, respectively, for 172 chemicals with different modes of action (MOA). We found that there were only significant relationships between P. promelas acute median lethal concentrations and growth lowest-observed effect concentrations for class 1 (nonpolar narcosis) chemicals, though significant relationships were demonstrated for D. magna to all Verhaar et al. MOA classes (Class 1: nonpolar narcosis, Class 2: polar narcosis, Class 3: reactive chemicals, and Class 4: AChE inhibitors and estrogenics). Probabilistic ecological hazard assessment using chemical toxicity distributions was subsequently employed for each MOA class to estimate acute and chronic thresholds, respectively, to identify MOA and species specific ecological thresholds of toxicological concern. Finally, novel MOA and species specific ACRs using both chemical toxicity distribution comparison and individual ACR probability distribution approaches were identified using representative MOA and chemical categories. Our data-driven approaches and newly identified ACR values represent robust alternatives to application of default ACR values, and can also support future research and risk assessment and management activities for other chemical classes when toxicity information is limited for chemicals with specific MOAs within invertebrates and fish.

Fine particulate matter (PM₂.₅) is an essential risk factor of chronic obstructive pulmonary disease (COPD). Recent studies showed weak association between PM₂.₅ and COPD incidence, but smokers who exposed to higher PM₂.₅ concentration had more opportunity to gain COPD. Cigarette smoking is the most important risk factor of COPD. Thus, we hypothesized: the role of PM₂.₅ played on cigarette-inflamed airways was more significant than normal airways. The study firstly established an animal model of C57BL/6J mice with cigarette smoke exposure and PM₂.₅ orotracheal administration. After calculating pathological scores, mean linear intercept and mean alveolar area, we found PM₂.₅ aggravated pathological injury of cigarette-inflamed lungs, but the injury on normal lungs was not significant. Meanwhile, inflammatory factors as T-bet, IFN-γ and IL-1α were tested using qRT-PCR and ELISA. The results showed PM₂.₅ aggravated inflammation of cigarette-inflamed lungs, but the effect on normal lungs was not significant. The most important pathogenesis of COPD is abnormal apoptosis in airway epithelium, due to oxidative stress following long-term exposure to cigarette smoke. Then, apoptotic responses were detected in lungs. TUNEL analysis demonstrated that PM₂.₅ promoted DNA fragmentation of cigarette-inflamed lungs, but the effect on normal lungs was not significant. Western-blot and immunohistochemistry showed caspase activated significantly in PM₂.₅-cigarette smoke exposed lungs and activated caspase 3 located mainly on bronchial epithelium. Next, human bronchial epithelial cells were cultured treated with cigarette smoke solution (CSS) with or without PM₂.₅. Z-VAD-FMK, a pan-caspase inhibitor, was used to suppress the activation of caspases. After analyzing cell viability, DNA fragmentation, mitochondrial activities and caspase activities, the results clarified that PM₂.₅ aggravated apoptosis in cigarette-inflamed bronchial epithelial cells and the responses could be suppressed by Z-VAD-FMK. Our results gave a new idea about the mechanism of PM₂.₅ on COPD and inferred cigarette-inflamed airways were more vulnerable to PM₂.₅ than normal airways.

Boscalid as one of the most widely used succinate dehydrogenase inhibitor (SDHI) fungicides has been frequently detected in both freshwater and estuarine environments. Its acute toxic effects on zebrafish and freshwater algae have been reported in our previous studies. To further investigate its chronic toxic effects to aquatic organisms, adult zebrafish were exposed for 28 days to a series of environmentally relevant boscalid concentrations in this study. Growth indicators and histopathology were determined in this study. Results indicated that boscalid inhibited the growth of zebrafish and induced damage in the kidneys and liver. Carbohydrate and lipid metabolism as the key pathways of energy metabolism in growth of zebrafish were also investigated. Results showed boscalid caused an increase in the activity of hexokinase (HK), the content of glycogen, glucose-6-phosphatase (G6Pase), and insulin (INS) in liver and a decrease in blood glucose content and succinate dehydrogenase (SDH) activity. Boscalid reduced the total content of triacylglyceride (TG) and cholesterol (TC) and the activity of fatty acid synthase (FAS) and acetyl coenzyme A carboxylase (ACC) in the liver. Correspondingly, expression of the genes related to carbohydrate and lipid metabolism in liver and intestine was affected by boscalid, especially in the significant upregulation of G6Pase and pparα and downregulation of SGLT-1 and AMY. Results suggested that boscalid could affect carbohydrate metabolism of adult zebrafish via regulation of gluconeogenesis and glycolysis at 0.1 mg/L. Moreover, boscalid might induce an increase in β-oxidation and a decrease in lipid synthesis at 0.01 mg/L. In conclusion, our study identified that carbohydrate and lipid metabolism are the possible biological pathways that mediate boscalid-induced developmental effects.

The effects of Pb pollution on cattle and sheep raised in an ancient mining area were studied through the use of blood Pb (PbB) levels and δ-aminolevulinic acid dehydratase (δ-ALAD) activity. Lead levels in livestock blood from the mining area (n=110) were significantly elevated when compared to the controls (n=79). In 91.4% of cattle (n=58) and 13.5% of sheep (n=52) sampled in the mining area, PbB levels corresponded to subclinical exposure (6–35μg/dl). Two young cattle (<2 years) from the mining area (n=5) had PbB levels indicative of clinical poisoning (>35μg/dl). Elevated PbB was also accompanied by δ-ALAD activity inhibition in blood, which confirms that measurable effects of Pb poisoning were taking place. Observed PbB levels suggest that a potential risk to human consumers of beef from the Pb polluted areas may also exist, as has been shown previously for game meat from the same mining area.

Time-dependent changes in enzymatic and non-enzymatic antioxidants, and lipid peroxidation were investigated in roots of rice (Oryza sativa) grown hydroponically with Cd, with or without pretreatment of salicylic acid (SA). Exposure to 50 μM Cd significantly decreased root growth, and activities of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD), but increased the concentrations of H2O2, malondialdehyde (MDA), ascorbic acid (AsA), glutathione (GSH) and non-protein thiols (NPT). However, pretreatment with 10 μM SA enhanced the activities of antioxidant enzymes and the concentrations of non-enzymatic antioxidants, but lowered the concentrations of H2O2 and MDA in the Cd-stressed rice compared with the Cd treatment alone. Pretreatment with SA alleviated the Cd-induced inhibition of root growth. The results showed that pretreatment with SA enhanced the antioxidant defense activities in Cd-stressed rice, thus alleviating Cd-induced oxidative damage and enhancing Cd tolerance. The possible mechanism of SA-induced H2O2 signaling in mediating Cd tolerance was discussed. Pretreatment with SA enhanced the antioxidant defense activities in Cd-stressed rice, thus alleviating Cd-induced oxidative damage and enhancing Cd tolerance.

Wheat is one of several cereals that is capable of accumulating higher amounts of Cd in plant tissues. It is important to understand the Cd²⁺ transport processes in roots that result in excess Cd accumulation. Traditional destructive technologies have limited capabilities in analyzing root samples due to methodological limitations, and sometimes may result in false conclusions. The mechanisms of Cd²⁺ uptake into the roots of wheat seedlings (Triticum aestivum L.) were investigated by assessing the impact of various inhibitors and channel blockers on Cd accumulation as well as the real-time net Cd²⁺ flux at roots with the non-destructive scanning ion-selective electrode technique. The P-type ATPase inhibitor Na3VO4 (500 μM) had little effect on Cd uptake (p < 0.05) and the kinetics of transport in the root of wheat, suggesting that Cd²⁺ uptake into wheat root cells is not directly dependent on H⁺ gradients. While, the uncoupler 2,4-dinitrophenol significantly limited Cd²⁺ uptake (p < 0.05) and transport kinetics in the root of wheat, suggesting the existence of metabolic mediation in the Cd²⁺ uptake process by wheat. The Cd content at the whole-plant level in wheat was significantly (p < 0.05) decreased upon pretreatment with the Ca²⁺ channel blockers La³⁺ or Gd³⁺ and Verapamil, but not in case of pretreatment with the K⁺ channel blocker tetraethylammonium (TEA). In addition, the inhibitors of the Ca²⁺ channel, as well as high concentrations of Ca²⁺, reduced the real-time net Cd²⁺ fluxes at the root surface in SIET experiments. These results indicate that Cd²⁺ moves across the plasma lemma of the wheat root via Ca²⁺ channels. In addition, our results suggested a role for protein synthesis in mediating Cd²⁺ uptake and transport by wheat.