The acute pulmonary toxicity in mice induced by multiwall carbon nanotubes,benzene, and their combination

Carbon nanotubes (CNTs) have been synthesized and produced on large scale for their wide application. They have high absorption ability to organic contaminants (such as benzene) and can form CNTs‐benzene combination with benzene. In this article, the acute pulmonary toxicity, induced by multiwall carbon nanotubes (MWCNTs), benzene, and their combination, was studied by administrating the three test materials into mice lungs via intratracheal instillation. The biochemical parameters in bronchoalveolar lavage fluid (BALF) and pathological lesions in lungs were used as endpoints to evaluate the pulmonary toxicity of the three test materials at 3‐day and 7‐day postexposure, respectively. After the mice were intratracheally instilled with MWCNTs, benzene and MWCNTs‐benzene combination at doses of 6.67 mg/kg, 2.67 mg/kg, and 9.34 mg/kg (containing 6.67 mg/kg MWCNTs and 2.67 mg/kg benzene), the total protein, alkaline phosphatase (ALP), acid phosphatase (ACP), and lactate dehydrogenase (LDH) in BALF and pathological lesions in lungs were examined. At 3‐day postexposure, MWCNTs induced obvious pulmonary toxicity and benzene only induced slight pulmonary toxicity, whereas their combination induced very severe pulmonary toxicity. At 7‐day postexposure, MWCNTs and benzene did not induce pulmonary toxicity individually, whereas their combination still induced severe pulmonary toxicity. These data indicated that, at the instilled doses in this experiment, the MWCNTs can alone induce acute pulmonary toxicity in mice and the benzene does not induce pulmonary toxicity, but the pulmonary toxicity of MWCNTs is enhanced after they form MWCNTs‐benzene combination with low dose of benzene. The enhanced pulmonary toxicity may be due to the change of MWCNTs aggregation ability after benzene is adsorbed on them. © 2009 Wiley Periodicals, Inc. Environ Toxicol 25: 409–417, 2010.     

Comparison of the effects of multiwall carbon nanotubes on the epithelial cells and macrophages

Abstract

Effects of two kinds of multiwall carbon nanotubes (MWCNTs) on cells were examined. The effects of MWNT-7, which has been reported to be carcinogenic, and MWCNT-B, whose toxicity is unclear, were examined in both epithelial cells and macrophages. Human lung carcinoma A549 cells were used as representative epithelial cells and differentiated human monocyte THP-1 cells, as well as rat pulmonary macrophages NR8383, were employed to examine possible harmful effects of the MWCNTs. The MWCNTs induced the production of chemokines such as interleukin-8 (IL-8). MWCNTs were found to more strongly affect macrophages than epithelial cells. In addition, the toxicity was more pronounced in the MWNT-7 exposed cells than in those exposed to MWCNT-B. Cytochalasin D and amiloride treatment of differentiated THP-1 cells reduced cell-associated MWCNTs and IL-8 induction. To confirm these cellular influences in vivo, intratracheal administration of each type of MWCNT was performed by pharyngeal aspiration in the mouse lung. Analysis of bronchoalveolar lavage fluid (BALF) showed increase of inflammatory monocyte in MWNT-7 exposed animals at 1week after. In addition, neutrophils in the BALF were also significantly increased MWNT-7 exposed animals at 1 week and 1 month after. Aspiration of MWNT-7 caused formation of granulomas in the lung. Formation of the granulomas was not observed in the case of MWCNT-B. These results suggest that cellular uptake of the MWCNTs by phagocytosis and chemokine induction is important aspects of their toxicity.     

二氯乙酸钠联合顺铂诱导人肺腺癌A549细胞株凋亡的实验研究

目的研究二氯乙酸钠(Sodium dichloroacetate,DCA-Na)、顺铂(DDP)联合对人肺腺癌A549细胞株增殖和凋亡的影响及其作用机制。方法用MTT法检测DCA-Na、DDP应用对A549细胞增殖抑制作用的影响;流式细胞仪(Annexin V-FITC/PI法)检测DCA-Na、DDP单药及两药联合作用于A549细胞凋亡率的变化;用分光光度法检测DCA-Na作用于A549细胞后半胱氨酸天门冬氨酸蛋白酶(Caspase)-3、-8、-9蛋白的活性。结果DCA-Na、DDP单药组均对A549细胞增殖有抑制作用,且呈明显的剂量-时间依赖性。0.4、2μg/mL的DDP与37.5、75、150μg/mL的DCA-Na联合作用A549细胞24、48、72 h后的抑制率显著高于同浓度DDP单药组(P<0.05),且2μg/mL的DDP与75μg/mL的DCA联合在48 h表现为协同作用。流式细胞仪检测显示,A549细胞联合组凋亡率显著高于各单药组(P<0.05)。DCA-Na作用于A549细胞后,分别在12、24、48、72 h测得Caspase-3、-8、-9蛋白活性显著高于对照组(P<0.05)。结论 DCA-Na对人肺腺癌A549细胞的增殖有抑制作用,并且随着药物浓度增加、作用时间延长,其抑制作用也增加(呈剂量和时间依赖性)。一定浓度范围的DCA-Na和DDP联合作用于人肺腺癌A549细胞能够产生协同作用。DCA-Na可诱导人肺腺癌A549细胞凋亡,且与DDP联合后其诱导凋亡作用更为显著。     

Toxicological assessments of an ethanol extract complex of Descurainia sophia and Peucedanum praeruptorum: Subacute oral toxicity and genotoxicity studies

An ethanol extract complex of Descurainia sophia seeds and Peucedanum praeruptorum roots, called BP10A, has antitumor potential against colorectal cancer. In the present study, we evaluated the 28-day oral toxicity and the genotoxicity of BP10A. The subacute toxicity test was done through oral administration to mice. ICR mice (n = 10) received daily oral BP10A doses of 0, 500, 1000 and 2000 mg/kg for 28 consecutive days. During administration, general clinical signs, food consumption, organ weights, and hematologic, biochemical and histopathological parameters in male and female mice were assessed. No significant adverse effects up to the highest dose (2000 mg/kg) were found. The genotoxicity was evaluated using a battery of tests, including an in vitro bacterial reverse mutation (Ames) test, an in vivo micronucleus test using bone marrow cells in ICR mice and a chromosomal aberration test using CHL/IU cells. BP10A did not show any genotoxic signs in the Ames (up to 5000 μg/plate), micronucleus (up to 5000 mg/kg) and the chromosomal aberration tests (550-1750 μg/mL). Therefore, BP10A was considered safe based on the subacute toxicity and genotoxicity results, indicating that it is a useful pharmaceutical material with no adverse toxicity.     

An integrative cellular metabolomic study reveals downregulated tricarboxylic acid cycle and potential biomarkers induced by tetrabromobisphenol A in human lung A549 cells

Tetrabromobisphenol A (TBBPA) is extensively utilized as a brominated flame retardant in numerous chemical products. As an environmental contaminant, the potential human toxicity of TBBPA has been attracting increasing attention. Nonetheless, the exact underlying mechanisms of toxicological effects caused by TBBPA remain uncertain. In this study, we investigated the potential mechanisms of TBBPA toxicity in vitro in the A549 cell line, one of the widely used type II pulmonary epithelial cell models in toxicology research. Cell viability was determined after treatment with varying concentrations of TBBPA. Liquid chromatography–mass spectrometry (LC–MS) metabolomics and metabolic flux approaches were utilized to evaluate metabolite and tricarboxylic acid (TCA) cycle oxidative flux changes. Our findings demonstrated that TBBPA significantly reduced the viability of cells and attenuated mitochondrial respiration in A549 cells. Additionally, LC–MS data showed significant reductions in TCA cycle metabolites including citrate, malate, fumarate, and alpha-ketoglutarate in 50 μM TBBPA-treated A549 cells. Metabolic flux analysis indicated reduced oxidative capacity in mitochondrial metabolism following TBBPA exposure. Moreover, diverse metabolic pathways, particularly alanine, aspartate, and glutamate metabolism and the TCA cycle, were found to be dysregulated. In total, 12 metabolites were significantly changed (p < .05) in response to 50 μM TBBPA exposure. Our results provide potential biomarkers of TBBPA toxicity in A549 cells and help elucidate the molecular mechanisms of pulmonary toxicity induced by TBBPA exposure.     

Thymosin alpha‐1; a natural peptide inhibits cellular proliferation,cell migration,the level of reactive oxygen species and promotes the activity of antioxidant enzymes in human lung epithelial adenocarcinoma cell line (A549)

This research was conducted to investigate the biochemical effects of thymosin alpha‐1 using human lung cancer cells (A549). The A549 cells were treated with different concentrations of Thα1 for 24 h and the growth, inhibition of cells was determined. Thα1 revealed anti‐proliferative effect at 24 and 48 μg/ml after 24 h. Furthermore, it indicated antioxidant properties by significantly enhancing the activity of catalase (12 μg/ml), superoxide dismutase (6 and 12 μg/ml), and glutathione peroxidase (3, 6 and 12 μg/ml) and reducing the production of cellular ROS. Our results showed that Thα1 inhibits the migration of A549 cells in a concentration‐dependent manner after 24 and 48 h. Moreover, the effect of Thα1 on apoptosis was investigated by Hoechst 33342 staining and cell cycle analysis. Results demonstrated no significant effect on the induction of apoptosis in A549 cells. In conclusion, our results showed the antioxidant properties of Thα1 on A549 cancer cells.     

Evaluation of cytotoxic,genotoxic and inflammatory response in human alveolar and bronchial epithelial cells exposed to titanium dioxide nanoparticles

The toxicity of titanium dioxide nanoparticles (TiO₂‐NPs), used in several applications, seems to be influenced by their specific physicochemical characteristics. Cyto‐genotoxic and inflammatory effects induced by a mixture of 79% anatase/21% rutile TiO₂‐NPs were investigated in human alveolar (A549) and bronchial (BEAS‐2B) cells exposed to 1–40 µg ml^–1 30 min, 2 and 24 h to assess potential pulmonary toxicity. The specific physicochemical properties such as crystallinity, NP size and shape, agglomerate size, surface charge and specific surface area (SSA) were analysed. Cytotoxic effects were studied by evaluating cell viability using the WST1 assay and membrane damage using LDH analysis. Direct/oxidative DNA damage was assessed by the Fpg‐comet assay and the inflammatory potential was evaluated as interleukin (IL)‐6, IL‐8 and tumour necrosis factor (TNF)‐α release by enzyme‐linked immunosorbant assay (ELISA). In A549 cells no significant viability reduction and moderate membrane damage, only at the highest concentration, were detected, whereas BEAS‐2B cells showed a significant viability reduction and early membrane damage starting from 10 µg ml^–1. Direct/oxidative DNA damage at 40 µg ml^–1 and increased IL‐6 release at 5 µg ml^–1 were found only in A549 cells after 2 h. The secretion of pro‐inflammatory cytokine IL‐6, involved in the early acute inflammatory response, and oxidative DNA damage indicate the promotion of early and transient oxidative‐inflammatory effects of tested TiO₂‐NPs on human alveolar cells. The findings show a higher susceptibility of normal bronchial cells to cytotoxic effects and higher responsiveness of transformed alveolar cells to genotoxic, oxidative and early inflammatory effects induced by tested TiO₂‐NPs. This different cell behaviour after TiO₂‐NPs exposure suggests the use of both cell lines and multiple end‐points to elucidate NP toxicity on the respiratory system. Copyright © 2014 John Wiley & Sons, Ltd.     

In-vivo laxative and toxicological evaluation and in-vitro antitumour effects of Mitracarpus frigidus aerial parts

Objectives To evaluate the in‐vitro antitumour properties, and the in‐vivo laxative and toxicological effects of the methanolic extract of the aerial parts of Mitracarpus frigidus (MFM). Methods The in‐vitro antitumour activity of MFM was evaluated against three human tumour cell lines: Jurkat, HL60 and MCF‐7. The laxative activity and the effect of MFM on intestinal motility were evaluated in rats at the doses of 100, 300 and 1000 mg/kg. Acute oral toxicity was performed at 10, 100, 1000 and 2000 mg/kg and subchronic toxicity was evaluated at 100, 300 and 1000 mg/kg of MFM during a 42‐day period. After subchronic administration of MFM the biochemical, haematological and histopathological parameters were analysed. Also, the total content of anthraquinones was determined. Key findings MFM was cytotoxic only against HL60 and Jurkat cells with 89 and 83% growth inhibition, respectively. The laxative activity of MFM was similar to bisacodyl. Regarding the effect on intestinal motility, MFM showed a significant increase in the pathway of charcoal compared with the group treated with saline. Furthermore, MFM showed no in‐vivo toxicity at the doses tested. Free and anthraquinone C‐ and O‐glycosides were detected in MFM. Conclusions MFM showed significant antitumour activity for leukaemic cells. Moreover, it presented laxative potential and no in‐vivo toxicity.     

1,8‐Cineole induces relaxation in rat and guinea‐pig airway smooth muscle

Objectives 1,8‐Cineole is a monoterpene with anti‐inflammatory, vascular and intestinal smooth muscle relaxant activity. We have evaluated the potential bronchodilatatory activity of this compound. Methods 1,8‐Cineole was tested against carbachol, histamine, K⁺ 80 mM and ovalbumin‐induced bronchial contractions in Wistar rat or guinea‐pig tissues. Some of the guinea‐pigs had been previously sensitized with an intramuscular injection of 5% (w/v) ovalbumin/saline solution. Control animals received 0.3 ml saline. In separate experimental groups the response to 1,8‐cineole (1–30 mg/kg), phenoterol (0.05–5 mg/kg) or vehicle (0.3% Tween in saline) was studied. Key findings 1,8‐Cineole decreased, in vivo, rat bronchial resistance with similar efficacy as phenoterol (66.7 ± 3.2% vs 72.1 ± 5.3%). On the other hand, the maximal relaxant response to 1,8‐cineole in carbachol‐precontracted rat tracheas was 85.5 ± 5.7% (IC50 = 408.9 (328–5196) μg/ml) compared with 80.2 ± 4.8% (IC50 = 5.1 (4.3–6.1) μg/ml) with phenoterol. The addition of 1,8‐cineole to guinea‐pig tracheal rings tonically contracted with K⁺ 80 mM induced a concentration‐related relaxation. The maximal relaxation elicited by 1,8‐cineole was 113.6 ± 11.7% (IC50 127.0 (115.9–139.2) μg/ml) compared with 129.7 ± 14.6% (IC50 0.13 (0.12–0.14) μg/ml) achieved after phenoterol administration. In addition, the incubation of tracheal rings with 1,8‐cineole (100, 300 or 1000 μg/ml), 15 min before inducing phasic contractions with K⁺ 80 mM, decreased the maximal amplitude of the contraction by 31.6 ± 4.6, 75.7 ± 2.7 and 92.2 ± 1.5%, respectively. In another set of experiments, neither the maximal response nor the IC50 for the 1,8‐cineole‐induced relaxation were different between normal and ovalbumin‐sensitized tissues. Moreover, the relaxation of bronchial rings contracted after exposure to 1 μg/ml ovalbumin occurred at a faster rate in rings pre‐incubated with 1,8‐cineole when compared with rings pre‐incubated with vehicle only (Tween 0.3%). Therefore, in the first minute after the antigen challenge, the tracheal tissue relaxed after the peak contraction by 6.5, 21.4 (P < 0.05 vs control) and 66.9% (P < 0.05 vs control) in the presence of 100, 300 or 1000 μg/ml 1,8‐cineole, respectively. Conclusions 1,8‐Cineole relaxed rat and guinea‐pig (nonsensitized and ovalbumin‐sensitized) airway smooth muscle by a nonspecific mechanism.     

2,3,7,8-Tetrachlorodibenzo-p-dioxin: Acute oral toxicity in hamsters

The acute oral toxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was evaluated in hamsters. Male golden Syrian hamsters received 0 (corn oil only), 300, 600, 1000, 3000, or 6000 μg TCDD/kg body wt by single-dose gavage. To facilitate dosage, TCDD was fed as a suspension in acetone/corn oil (1:9). The 55-day single-dose oral LD50 was 5051 μg/kg (3876–18,487 μg/kg, 95% confidence interval). Deaths occurred from Day 9 through Day 43 following single-dose administration. Initially, no hamsters exhibited adverse effects related to dosage with TCDD, but 4–5 weeks following dosage, hamsters of the 1000, 3000, and 6000 μg/kg dose groups began to develop unkempt hair coats. A dose-related depression of mean body weight when compared to control weights was apparent in hamsters of the 1000, 3000, and 6000 μg/kg dose groups by 3 weeks posttreatment. Gross necropsy revealed the primary target organs affected in the hamster to be the same as in other laboratory species, namely, liver and thymus; this was accompanied by a slight decrease in adipose tissue reserves. No other significant effects were noted upon necropsy. Based on these results, the hamster appears to be much less sensitive than other laboratory species to the acute oral toxicity of TCDD.     

Pulmonary response of mice to a sequential exposure of side-stream cigarette smoke and multi-walled carbon nanotubes

Abstract

Due to their unique properties, nano-sized carbon materials are predicted to have numerous applications in industry, but significant evidence exists to suggest their potential to cause toxicity. To determine if pre-exposure to side-stream cigarette smoke (SSCS) influences their toxicity, we examined the pulmonary response of smoke-exposed mice to multi-walled carbon nanotubes (MWCNT). Female A/J mice were exposed to SSCS in a whole body exposure chamber at approximately 40?mg/m³ for 4 weeks (6?h/d, 5?d/wk) and challenged with a single dose of MWCNT (40?µg) by the pharyngeal aspiration technique. A total of four groups were compared: air/phosphate buffered saline (PBS)-control, SSCS/PBS, air/MWCNT, and SSCS/MWCNT. At days 1 and 3 post-MWCNT treatment, lung tissues and bronchoalveolar lavage fluid (BALF) were collected and analyzed. In comparison with controls, significantly higher levels of total BAL cells were obtained from mice exposed to SSCS and MWCNT alone or combination. Influx of polymorphonuclear leukocytes (PMN) into BALF greatly increased in MWCNT alone and SSCS/MWCNT groups at both days 1 and 3 compared with controls. However, pre-exposure to SSCS significantly suppressed PMN response to MWCNT on day 1 but not day 3. Total BALF protein, lactate dehydrogenase, and mucin were significantly elevated in MWCNT and SSCS/MWCNT groups but not in the SSCS group, except mucin at day 3, when compared with controls. These results demonstrate that MWCNT markedly increases pulmonary toxicity in mice and SSCS pre-exposure plays a minor role in modulating MWCNT-induced lung toxicity at the concentrations and time points selected in the present study.     

Alleviating effects of reduced graphene oxide against lead-induced cytotoxicity and oxidative stress in human alveolar epithelial (A549) cells

Broad application of reduced graphene oxide (rGO) and ubiquitous lead (Pb) pollution may increase the possibility of combined exposure of humans. Information on the combined effects of rGO and Pb in human cells is scarce. This work was designed to explore the potential effects of rGO on Pb-induced toxicity in human alveolar epithelial (A549) cells. Prepared rGO was polycrystalline in nature. The formation of a few layers of visible creases and silky morphology due to high aspect ratio was confirmed. Low level (25 μg/mL) of rGO was not toxic to A549 cells. However, Pb exposure (25 μg/mL) induced cell viability reduction, lactate dehydrogenase enzyme leakage with rounded morphology in A549 cells. Remarkably, Pb-induced cytotoxicity was significantly mitigated by rGO co-exposure. Pb-induced mitochondrial membrane potential loss, cell cycle arrest and higher activity of caspase-3 and -9 enzymes were also alleviated by rGO co-exposure. Moreover, we observed that Pb exposure causes generation of pro-oxidants (e.g., reactive oxygen species, hydrogen peroxide and lipid peroxidation) and antioxidant depletion (e.g., glutathione and antioxidant enzymes). In addition, the effects of Pb on pro-oxidant and antioxidant markers were significantly reverted by GO co-exposure. Inductively coupled plasma-mass spectrometry suggested that due to the adsorption of Pb on rGO sheets, accessibility of Pb ions for A549 cells was limited. Hence, rGO reduced the toxicity of Pb in A549 cells. This research warrants further study to work on detailed underlying mechanisms of the mitigating effects of rGO against Pb-induced toxicity on a molecular level.     

TGF‐β1 mediated MAPK signaling pathway promotes collagen formation induced by Nano NiO in A549 cells

Nickel oxide nanoparticles (Nano NiO) could induce pulmonary fibrosis, however, the mechanisms are still unknown. The aim of the present study was to explore the roles of transforming growth factor‐β1 (TGF‐β1), mitogen‐activated protein kinase (MAPK) pathway and MMPs/TIMPs balance in Nano NiO‐induced pulmonary fibrosis. For that purpose, we first established Nano NiO‐induced human lung adenocarcinoma epithelial cells (A549 cells) model of collagen excessive formation through detecting the levels of hydroxyproline (Hyp) and type I collagen (Col‐I). Then the protein levels of TGF‐β1, MAPKs, and MMPs/TIMPs were assessed by Western blot. The results showed that Nano NiO resulted in the increased contents of Hyp, Col‐I, and TGF‐β1, the MAPK pathway activation and MMPs/TIMPs imbalance with a dose‐dependent manner. In addition, to investigate whether TGF‐β1 mediated MAPK signaling pathway, A549 cells were treated by 100 μg/mL Nano NiO combined with TGF‐β1, p38 MAPK, and ERK1/2 inhibitors (10 μM SB431542, 10 μM SB203580, and 10 μM U0126), respectively. We found that MAPK signal pathway was suppressed by TGF‐β1 inhibitor. Meanwhile, the increased contents of Hyp and Col‐I, and MMPs/TIMPs imbalance were alleviated by the p38 MAPK and ERK1/2 inhibitors, respectively. These findings indicated that the MAPK pathway and MMPs/TIMPs imbalance were involved in collagen excessive formation induced by Nano NiO.     

Synthesis and Anticonvulsant Activity of N‐(2‐Hydroxy‐ethyl)amide Derivatives

A series novel of N‐(2‐hydroxyethyl)amide derivatives was synthesized and screened for their anticonvulsant activities by the maximal electroshock (MES) test, and their neurotoxicity was evaluated by the rotarod test (Tox). The maximal electroshock test showed that N‐(2‐hydroxyethyl)decanamide 1g , N‐(2‐hydroxyethyl)palmitamide 1l , and N‐(2‐hydroxyeth‐yl)stearamide 1n were found to show a better anticonvulsant activity and also had lower toxicity than the marked anti‐epileptic drug valproate. In the anti‐MES potency test, these compounds exhibited median effective doses (ED₅₀) of 22.0, 23.3, 20.5 mg/kg, respectively, and median toxicity doses (TD₅₀) of 599.8, >1000, >1000 mg/kg, respectively, resulting in a protective index (PI) of 27.5, >42.9, >48.8, respectively. This is a much better protective index than that of the marked anti‐epileptic drug valproate (PI = 1.6). To further investigate the effects of the anticonvulsant activity in several different models, compounds 1g , 1l , and 1n were tested having evoked convulsions with chemical substances, including pentylenetetrazloe, isoniazide, 3‐mercaptopropionic acid, bicuculline, thiosemicarbazide, and strychnine.     

Pulmonary toxicity of polyvinyl chloride particles after a single intratracheal instillation in rats. Time course and comparison with silica

Our previous in vitro studies indicated that emulsion polyvinyl chloride (PVC) particles (PVC-E3), with a mean diameter of 2 microm, exhibited a moderate toxicity in different pulmonary cell cultures. The in vitro cytotoxicity and pro-inflammatory potential of PVC-E3 particles were reduced when the additives had been "washed off" (PVC-W3), indicating that PVC-particle associated toxicity is probably related to the residual additives. In the present study, male Wistar rats (230 +/- 18 g) received a single intratracheal instillation of vehicle, crystalline silica particles [Min-U-Sil, 10 mg/kg body weight (BW)], PVC-E3 (10 or 50 mg/kg BW), or PVC-W3 (10 or 50 mg/kg BW). After 2, 7, 28, or 90 days, the rats were sacrificed (n = 6) and pulmonary injury and inflammation were determined by measuring lung weight, lactate dehydrogenase (LDH) activity and protein concentrations in bronchoalveolar lavage fluid (BALF), differential BALF cell count, and histopathology. Silica exposure resulted in pulmonary inflammation and damage at all time points with a progressive deterioration. Exposure to high concentrations of PVC particles caused pulmonary inflammation and damage, which were similar to the silica-exposed group at 2 days, but at 90 days, most parameters had returned to the control level, except for minor histopathological lesions. PVC-E3 did not induce more damage than PVC-W3. Two days after exposure, PVC-W3 caused less neutrophil but more eosinophil influx than PVC-E3. Although the pulmonary toxicity of both PVC-E3 and PVC-W3 appeared limited, this in vivo study has not confirmed the conclusion from the in vitro toxicity tests that removal of residual additives reduces the toxicity of PVC-E3 particles.     

Microcystin‐LR induces a wide variety of biochemical changes in the A549 human non‐small cell lung cancer cell line: Roles for protein phosphatase 2A and its substrates

Our previous studies have described the toxic effects of microcystin‐LR (MC‐LR) in various normal cell lines and human hepatoma SMMC‐7721 cells, but the specific effects of MC‐LR in other types of cancer cells with respect to protein phosphatase 2A (PP2A) have not been fully elaborated. A549 human lung adenocarcinoma cells have been identified to express organic anion‐transporting polypeptides (OATP) involved in cellular uptake of MC‐LR, and thus probably make an appropriate in vitro model to assess MC‐LR's cytotoxicity. Hence, in our present study, A549 cells were treated with various concentrations of MC‐LR for 24 h. The presence of MC‐LR in A549 cells was confirmed, and PP2A activity, PP2A substrates, cytoskeleton, apoptosis, and proliferation were subsequently explored. The results showed that 5–10 μM MC‐LR inhibited PP2A activity significantly but 0.5–1 μM MC‐LR did not change PP2A activity dramatically. The inhibition could result from the hyperphosphorylation of PP2A/C at Tyr307, an elevation in the total PP2A/C expression and the dissociation of α4/PP2A/C complexes. Moreover, MC‐LR led to rearrangements of filamentous actin and microtubules, which might be correlated with the hyperphosphorylation of Ezrin, VASP and HSP27 due to PP2A inhibition and mitogen‐activated protein kinase (MAPK) activation. However, exposure to MC‐LR for 24 h failed to trigger either apoptosis or proliferation, which might be related to PP2A‐inhibition‐induced hyperphosphorylation of Bcl‐2 and Bad and the activation status of Akt. In conclusion, our data indicated that MC‐LR induced extensive molecular and cellular alterations in A549 cells through a PP2A‐centered pathway, which differed in some respects from our previous study in SMMC‐7721 cells. To our knowledge, this is the first report comprehensively demonstrating the effects of MC‐LR in A549 cells, and our findings provide insights into the mechanism of MC‐LR toxicity in cancer cells. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1065–1078, 2017.     

Comparative study on the acute pulmonary toxicity induced by 3 and 20nm TiO(2) primary particles in mice

The acute pulmonary toxicity induced by 3-nm TiO₂ primary particles was preliminary investigated after they were intratracheally instilled at doses of 0.4, 4 and 40 mg/kg into lungs of mice. The biochemical parameters in bronchoalveolar lavage fluid (BALF) and pathological examination were used as endpoints to assess their pulmonary toxicity at 3-day postexposure. As such, the pulmonary toxicity assessment of 20-nm TiO₂ primary particles was performed using the same method. It was found that the 3-nm TiO₂ primary particles induced no pulmonary toxicity at dose of 0.4 mg/kg, moderate toxicity at 4 mg/kg and lung overload at 40 mg/kg, and this kind of particles did not produce more pulmonary toxicity than the 20-nm ones at any instilled doses. As regards physicochemical characteristics of the two TiO₂ particles, their pH values in medium, other than particle size, surface area and aggregation, may play important role in affecting their pulmonary toxicity.     

Perfluorooctane sulfonate induces apoptosis in lung cancer A549 cells through reactive oxygen species‐mediated mitochondrion‐dependent pathway

Perfluorooctane sulfonate (PFOS) is a widespread environmental contaminant that is detected in the lung of mammals. The mechanisms underlying PFOS‐induced lung cytotoxicity remain unclear. The main purpose of this study was to evaluate the cytotoxic effects of PFOS on human lung cancer A549 cells and its possible molecular mechanism. A549 cells were treated with PFOS (0, 25, 50, 100 and 200 μm ) and the cellular apoptosis, mitochondrial membrane potential as well as intracellular reactive oxygen species were determined. In this study, PFOS induced a dose‐dependent increase in A549 cell toxicity via an apoptosis pathway as characterized by increased percentage of sub‐G1, activation of caspase‐3 and ?9, and increased ratio of Bax/bcl‐2 mRNA expression. In addition, there was obvious oxidative stress, represented by decreased glutathione level, increased malondialdehyde level and superoxide dismutase activity. N‐Acetylcysteine, as an antioxidant that is a direct reactive oxygen species scavenger, can effectively block PFOS‐induced reactive oxygen species generation, mitochondrial membrane potential loss and cell apoptosis. These data indicate that PFOS induces apoptosis in A549 cells through a reactive oxygen species‐mediated mitochondrial dysfunction pathway mechanism. Copyright © 2012 John Wiley & Sons, Ltd.