Bencheng Lin

Toxicity of TDCPP and TCEP on PC12 cell: Changes in CAMKII, GAP43, tubulin and NF-H gene and protein levels

TDCPP and TCEP are two major types of organophosphorus flame retardants (OPFRs) that are bioaccumulative and persistent in the environment. The toxicity effects of TDCPP and TCEP on PC12 cell are not well understood. In the present study, we investigated morphology, viability and apoptosis in cultured PC12 cells in response to TDCPP and TCEP. The mRNA and protein expression levels of CAMKII, GAP43, tubulin and NF-H were quantified in PC12 cells treated with varying concentrations of the two agents. Results indicate that, upon treatment with the two OPFRs, cell growth decreased, apoptosis increased, morphology was altered and significant changes were found in the gene and protein levels. Treatment with TDCPP caused a reduction in the levels of each of the six proteins studied and in the gene levels of GAP43, NF-H and the two tubulins, but it resulted in an increase in CAMKII gene levels. Treatment with TCEP resulted in similar changes in gene levels to TDCPP and led to decreases in the protein levels of GAP43 and the tubulins while increasing the CAMKII and NF-H protein levels. These results suggest that changes in the gene and protein levels of the regulatory proteins (CAMKII, GAP43) and the structural proteins (tubulin, NF-H) are due to different mechanisms of the toxins, and these proteins may be useful biomarkers for the cytotoxicity and neurotoxicity.

Neurotoxicity induced by zinc oxide nanoparticles: age-related differences and interaction.

This study mainly investigated the neurotoxicity induced by zinc oxide nanoparticle (ZnO NP) in different-aged mice and the interaction between age and ZnO NP exposure. Sixty adult and old male C57BL/6J mice were assigned to four groups based on a two-factor (age and ZnO NP exposure) design. Results showed that ZnO NPs (5.6 mg/kg, intraperitoneal) induced increased production of pro-inflammatory cytokines in the serum and the brain of mice. A synergistic reaction between aging and ZnO NP exposure occurred regarding serum interleukin 1 (IL-1) and interleukin 6 (IL-6). In the brain, increased oxidative stress level, impaired learning and memory abilities, and hippocampal pathological changes were identified, especially in old mice, following ZnO NP exposure. Then, a potential mechanism of cognitive impairment was examined. The contents of hippocampal cAMP response element binding protein (CREB), phosphorylated CREB, synapsin I, and cAMP were decreased in an age-dependent manner, and the most substantial decrease occurred in old mice treated with ZnO NPs. These findings demonstrated for the first time that aging and ZnO NP exposure synergistically influenced systemic inflammation, and indicated old individuals were more susceptible to ZnO NP-induced neurotoxicity. One of the mechanisms might due to the supression of cAMP/CREB signaling.

Studies of single-walled carbon nanotubes-induced hepatotoxicity by NMR-based metabonomics of rat blood plasma and liver extracts

The toxicological effects of single-walled carbon nanotubes (SWCNTs) were investigated after intratracheal instillation in male Wistar rats over a 15-day period using metabonomic analysis of 1H (nuclear magnetic resonance) NMR spectra of blood plasma and liver tissue extracts. Concurrent liver histopathology examinations and plasma clinical chemistry analyses were also performed. Significant changes were observed in clinical chemistry features, including alkaline phosphatase, total protein, and total cholesterol, and in liver pathology, suggesting that SWCNTs clearly have hepatotoxicity in the rat. 1H NMR spectra and pattern recognition analyses from nanomaterial-treated rats showed remarkable differences in the excretion of lactate, trimethylamine oxide, bilineurin, phosphocholine, amylaceum, and glycogen. Indications of amino acid metabolism impairment were supported by increased lactate concentrations and decreased alanine concentrations in plasma. The rise in plasma and liver tissue extract concentrations of choline and phosphocholine, together with decreased lipids and lipoproteins, after SWCNTs treatment indicated a disruption of membrane fluidity caused by lipid peroxidation. Energy, amino acid, and fat metabolism appeared to be affected by SWCNTs exposure. Clinical chemistry and metabonomic approaches clearly indicated liver injury, which might have been associated with an indirect mechanism involving nanomaterial-induced oxidative stress.

The combined toxicological effects of titanium dioxide nanoparticles and bisphenol A on zebrafish embryos

Environmental pollutants co-exist and exhibit interaction effects that are different from those associated with a single pollutant. As one of the more commonly manufactured nanomaterials, titanium dioxide nanoparticles (TiO2-NPs) are most likely to bind to other contaminants in water. In this paper, we aimed to study the combined toxicological effects of TiO2-NPs and bisphenol A (BPA) on organism. First, in vitro adsorption experiments were conducted to determine the adsorptive interaction between TiO2-NPs and BPA. Second, zebrafish embryo toxicity tests were performed to monitor for changes in the toxicological effects associated with the two chemicals. The study results demonstrated that adsorptive interactions exist between the two chemicals and increased toxicity effects which included an advanced toxicological effect time, decreased survival, increased morphological abnormalities, and delayed embryo hatching. Also, we suggest that the mode of combined action has a synergistic effect. Based on this, we postulate that concomitant exposure to TiO2-NPs and BPA increased BPA bioavailability and uptake into cells and organisms. Further studies are required to understand the mechanisms of interactions of this mixture.

Single-wall carbon nanotubes induce oxidative stress in rat aortic endothelial cells

Oxidative stress is a major factor contributing to endothelial cell damage. Single-wall carbon nanotubes (SWCNTs) have oxidative properties; however, the oxidative effects of SWCNTs on endothelial cells are not fully understood. In the present study, we investigated the effects of oxidative stress induced by SWCNTs on rat aortic endothelial cells (RAECs). Various markers of cellular damage were assessed, such as biochemical and ES immunity indexes, and DNA and protein damage. Our findings suggest that RAEC endured oxidative damage following SWCNT exposure. Specifically, after SWCNTs exposure, non-enzymatic antioxidant glutathione was activated prior to superoxide dismutase activation in order to defend against oxidative stress. Additionally, it was found that as SWCNT concentration increased, so did the stress protein, heme oxygenase-1 (HO-1), expression levels. These changes may induce RAEC damage, and result in many serious diseases.

Single-walled carbon nanotubes promote rat vascular adventitial fibroblasts to transform into myofibroblasts by SM22-α expression.

The aim of this study was to explore whether single-wall carbon nanotubes (SWCNTs) can be used as artery tissue-engineering materials by promoting vascular adventitial fibroblasts (VAFs) to transform into myofibroblasts (MFs) and to find the signal pathway involved in this process. VAFs were primary cultured and incubated with various doses of SWCNTs suspension (0, 0.8, 3.2, 12.5, 50, and 200 μg/mL). In the present study, we used three methods (MTT, WST-1, and WST-8) at the same time to detect the cell viability and immunofluorescence probe technology to investigate the effects of oxidative injury after VAFs incubated with SWCNTs. Immunocytochemical staining was used to detect SM22-α expression to confirm whether VAFs transformed into MFs. The protein levels were detected by western blotting. The results of immunocytochemical staining showed that SM22-α was expressed after incubation with 50 μg/mL SWCNTs for 96 hours, but with oxidative damage. The mRNA and protein levels of SM22-α, C-Jun N-terminal kinase, TGF-β1, and TGF-β receptor II in VAFs increased with the dose of SWCNTs. The expression of the p-Smad2/3 protein was upregulated while the Smad7 protein was significantly down-regulated. Smad4 was translocated to the nucleus to regulate SM22-α gene expression. In conclusion, SWCNTs promoted VAFs to transform into MFs with SM22-α expression by the C-Jun N-terminal kinase/Smads signal pathway at the early stage (48 hours) but weakened quickly. SWCNTs also promoted the transformation by the TGF-βl/Smads signal pathway at the advanced stage in a persistent manner. These results indicate that SWCNTs can possibly be used as artery tissue-engineering materials.

Biodistribution of single-walled carbon nanotubes in rats

Background: Carbon nanotube (CNT) mediated drug delivery systems have recently aroused a great deal of interest. Such delivery systems for drugs, proteins and genes have been preliminarily studied using cellular and animal models. For further study of the related biological behaviours of CNTs in vivo, a fast and convenient tracing method is particularly necessary. Methods: We adopted concentrated nitric acid/concentrated sulfuric acid oxidation combined with ultrasonication to treat SWCNTs, then detected and analyzed the samples before and after treatment by transmission electron microscopy (TEM), Fourier transform infrared spectrometry (FT-IR), energy dispersive X-ray spectrometry (EDX) and X-ray photoelectron spectroscopy (XPS). The iodogen oxidative method was used to synthesize iodinated single-walled carbon nanotubes, followed by intratracheal instillation of 125I-labelled SWCNTs to determine their distribution in rats. Results: SWCNTs form more hydroxyl and carboxyl functional groups with no change in their essential characteristics after treatment by violet acid oxidation combined with ultrasonication. 125I can easily form C–I covalent bonds to SWCNTs. The proportion of iodine-125 labelled SWCNTs is 46.14%, the radiochemical purity is 98.95%. In order of total radioactivity concentration in the main organs/tissues and body fluids for 125I-SWCNTs, it was shown that trachea > urine > stomach > small intestine > serum > bladder > blood vessel > kidney > liver > lung > adrenal > femoral head > spleen > testis > thymus > thyroid > heart > fat > muscle > brain. Conclusions: In this paper, we developed a generally adoptable tracing method for studying the biodistribution of SWCNTs (single walled carbon nanotubes) in vivo. SWCNTs could be labelled with radioactive 125I atoms. The 125I labelling method is reliable and effective and affords a quantitative analysis of CNTs accumulated in animal tissues. This result will provide an important reference for future research into the biomedical and pharmacological applications of SWCNTs.

Toxic effect comparison of three typical sterilization nanoparticles on oxidative stress and immune inflammation response in rats

Zinc oxide, titanium dioxide and silver nanoparticles are used as sterilization materials to enhance the performance of disinfectants. Here, the toxicological effects on the liver, spleen, thymus gland, immune function and inflammatory responses in rats induced by these nanoparticles were investigated after intratracheal instillation in male Wistar rats. Moreover, the relationships between the particle size, particle crystalline structure, chemical composition, chemical stability and toxicological effects of these typical nanoparticles in rats were explored. Exposure to nanoparticles increased the oxidative stress level in peripheral blood and the homogenates of the liver, spleen and thymus as well as disorders in regulating the cytokine network and blood cell count in the peripheral blood. Furthermore, the histopathological study revealed that pulmonary exposure to nanoparticles produced persistent, progressive liver inflammatory responses and cell necrosis, while no observable damage was found in the kidney, thymus gland or spleen tissue from the experimental groups. Our results demonstrate that oxidative stress might be important for inducing the toxic effects of these nanoparticles, and three nanoparticles can influence the immune function of rats. A comparative analysis of the toxic effects of nanomaterials demonstrated significant differences. Nano-ZnO induced the most significant toxicity, whereas nano-TiO2 induced the least. Particle composition and chemical stability probably played a primary role in the toxicological effects of different nanoparticles.

Comparative proteomic analysis of ovary for Chinese rare minnow (Gobiocypris rarus) exposed to chlorophenol chemicals.

UNLABELLED Pentachlorophenol (PCP) and 2,4,6-trichlorophenol (TCP) are suspected of disrupting the endocrine system and thus affecting human and wildlife reproduction, but the potential common mechanisms and biomarkers of chlorophenols (CPs) in the ovary are not fully elucidated. In the present study, the female rare minnow (Gobiocypris rarus) was exposed to PCP (0.5, 5.0, and 50 μg/L), TCP (1.0, 10, and 100 μg/L) and 17β-estradiol (as a positive control) for 28 days, and the matrix-assisted laser desorption/ionization (MALDI) tandem time-of-flight (TOF/TOF) mass spectrometry analysis was employed to investigate the alteration of protein expression in the ovary. After comparison of the protein profiles from treated and control groups, 22 protein spots were observed to be altered in abundance (>2-fold) from female treated groups, and 14 protein spots were identified successfully. These proteins were related to molecular response patterns, endocrine effects, metabolic pathways, and even the possible carcinogens in response to CP exposure. The seven differentially expressed mRNA encoding proteins were measured by quantitative real-time PCR (QRT-PCR) and histopathology was also measured. Our data demonstrate that alterations of multiple pathways may be associated with the toxic effects of CPs on ovaries. BIOLOGICAL SIGNIFICANCE Although numerous studies have shown the affection of the endocrine system with exposure to chlorophenols (CPs), there is little report on the alterations of protein expression in the ovaries from rare minnows following exposure to PCP or TCP. In the present study, a comparative proteomic approach using two dimensional gel electrophoresis and mass spectrometry (MALDI-TOF/TOF MS) has been developed to identify certain proteins in the ovaries of Chinese rare minnow, whose abundance changes during exposure to CPs. After comparison of the protein profiles from treated and control groups, 22 protein spots were observed to be altered in abundance (>2-fold) from female treated groups, and 14 protein spots were identified successfully. These proteins were related to molecular response patterns, endocrine effects, metabolic pathways, and even the possible carcinogens in response to CP exposure. Because the mechanism often involves changes in the expression of multiple proteins rather than a single protein, a global analysis of the protein alterations can result in valuable information to understand the CP action mechanism. All the above results demonstrate that the Vtg, SUMO, Lec-3 and PIMT protein are potential biomarkers and involved in the toxicity pathway of CP exposure in aquatic animals, which should be the primary focus of studies on the CP ovary toxicity mechanism in the future.

Respiratory exposure to single-walled carbon nanotubes induced changes in vascular homeostasis and the expression of peripheral blood related genes in a rat model

Epidemiological studies have demonstrated that nanometre particles in polluted air can increase the risk of CVD, which is dangerous to mankind. However, little is known regarding the indirect toxic effects on the cardiovascular system of respiratory tract exposure to nanometre particles. As a typical nanomaterial, SWCNTs have gained enormous popularity because of their unique properties. However, increasing attention has been paid to the potential pulmonary toxic effects of respiratory tract exposure to SWCNT than to the potential link of this exposure to cardiovascular disease risk. In this study, a rat intratracheal instillation model was used to evaluate the systemic and secondary effects of respiratory tract exposure to SWCNT, specifically changes in lung tissues, the circulatory system and vascular function. We found increased levels of inflammatory factors and interstitial inflammation in the lungs in this rat model. In addition, up-regulated levels of cytokines and an increase in white blood cells, platelets and fibrinogen were detected in the plasma. These changes were followed by increased blood viscosity in the high dose SWCNT exposure group. In addition, damage to the ultrastructure of the vascular intima in the rats was observed. Changes in coagulation and fibrinolysis activating factors were detected in the plasma. Lower expression of t-PA and higher expression of vWF were observed in the vascular intima of rats exposed to SWCNT at 10.5 and 17.5 mg per kg b.w. for 30 days and 60 days. After exposure to SWCNT for 60 days at 17.5 mg per kg b.w., decreased expression of t-PA gene and increased expression of TM, p38MAPK and PAI-1 genes were observed in the peripheral blood of the rats. Based on these results, we conclude that cardiovascular toxicity caused by respiratory tract exposure to SWCNTs may be induced by indirect effects on vascular homeostasis, which is different from previously reported direct cardiac effects of SWCNT. The present work established a correlation between pulmonary changes and CVD following pulmonary exposure to SWCNT. This study indicates a possible pathophysiological mechanism for CVD caused by pulmonary exposure to SWCNT. More importantly, these results supplement toxicological evaluation data for the risk of CVD caused by respiratory tract exposure to SWCNT.