Influence of silver doping on pro-inflammatory and pro-fibrogenic effects of nano-titanium dioxide in murine lung.

Silver is usually loaded on nano-titanium dioxide (TiO2 ) through photodeposition method to enhance visible-light catalytic functions for environment purification. However, little is known about how the toxicity changes after silver doping and how the physicochemical properties of loaded components affect nanocomposite toxicity. In this study, Ag-TiO2 with different sizes and contents of silver particles were obtained by controlling photodeposition time (PDT) and silver addition amount. Pro-inflammatory and pro-fibrogenic responses of these photocatalysts were evaluated in male C57BL/6J murine lung. As a result, silver was well assembled on TiO2 , promoting visible-light catalytic activity. Notably, the size of silver particles increased with PDT. Meanwhile, toxicity results showed that pure TiO2 (P25) mainly caused neutrophil infiltration, while 2 wt/wt% silver-loaded TiO2 recruited more types of inflammatory cells in the lung. Both of them caused the increase of proinflammatory cytokines while decreasing the anti-inflammatory cytokine in bronchoalveolar lavage fluid. However, 2 wt/wt% silver doping also accelerated the lung pro-fibrogenic response of photocatalysts in the subacute phase from evidence of collagen deposition and hydroxyproline concentrations. Mechanistically, the overactivation of TGFBR2 receptors in TGF-ß/smads pathways by silver-loaded TiO2 rather than pure TiO2 may be the reason why silver-loaded TiO2 can promote pro-fibrogenic effect response. Intriguingly, the increased toxicity caused by silver doping can be rescued by increasing the size of the loaded silver or decreasing the silver amount. These results may be important for the new understanding of the toxicity of TiO2 -based photocatalysts.

Ag/TiO2 nanohybrids induce fibrosis-related epithelial-mesenchymal transition in lung epithelial cells and the influences of silver content and silver particle size.

The controlled synthesis of silver nanoparticles (AgNPs) decorated TiO2 nanohybrids (Ag/TiO2) for photocatalysis has received considerable attention. These photocatalysts are widely used in environment and energy, resulting in human exposure through inhalation. Pure TiO2 is generally considered a low-toxic nanomaterial. However, little is known about the toxicity after AgNPs loading. In this study, silver-decorated TiO2 nanohybrids were controllably synthesized by the photodeposition method, and their toxic effects on murine lung and human lung epithelial cells were explored. As a result, silver loading significantly enhanced the effect of TiO2 photocatalyst on EMT in lung epithelial cells, potentially acting as a pro-fibrogenic effect in murine lung. Meanwhile, the increase in autophagy vacuoles, LC3-II marker, stub-RFP-sens-GFP-LC3 fluorescence assay, and LC3 turnover assay showed that silver loading also significantly increased autophagy flux. Furthermore, analysis of autophagy inhibition by 3-Methyladenine indicated that the promotion of EMT by silver loading was related to the increased autophagy flux. Intriguingly, the autophagy and EMT biological effects could be alleviated when the silver loading amount was reduced or silver particle size was increased, and the enhanced pro-fibrogenic effect was mitigated at the same time. This study supplemented safety information of Ag-decorated TiO2 nanohybrids and provided methods of controlled synthesis for reducing toxicity.

Progress on the toxicity of quantum dots to model organism-zebrafish.

In vivo toxicological studies are currently necessary to analyze the probable dangers of quantum dots (QDs) to the environment and human safety, due to the fast expansion of QDs in a range of applications. Because of its high fecundity, cost-effectiveness, well-defined developmental phases, and optical transparency, zebrafish has long been considered the "gold standard" for biosafety assessment of chemical substances and pollutants. In this review, the advantages of using zebrafish in QD toxicity assessment were explored. Then, the target organ toxicities such as developmental toxicity, immunotoxicity, cardiovascular toxicity, neurotoxicity, and hepatotoxicity were summarized. The hazardous effects of different QDs, including cadmium-containing QDs like CdTe, CdSe, and CdSe/ZnS, as well as cadmium-free QDs like graphene QDs (GQDs), graphene oxide QDs (GOQDs), and others, were emphasized and described in detail, as well as the underlying mechanisms of QDs generating these effects. Furthermore, general physicochemical parameters determining QD-induced toxicity in zebrafish were introduced, such as chemical composition and surface coating/modification. The limitations and special concerns of using zebrafish in QD toxicity studies were also mentioned. Finally, we predicted that the utilization of high-throughput screening assays and omics, such as transcriptome sequencing, proteomics, and metabolomics will be popular topic in nanotoxicology.

CdTe quantum dots trigger oxidative stress and endoplasmic reticulum stress-induced apoptosis and autophagy in rat Schwann cell line RSC96.

In the current study, the cytotoxicity and mechanisms of cadmium telluride quantum dots (CdTe QDs) on RSC96 cells were evaluated by exposing different doses of CdTe QDs for 24 h. Two types of cell death, including apoptosis and autophagy, as well as two important organelles, mitochondria and endoplasmic reticulum, were focused after CdTe QDs exposure. The results showed that CdTe QDs induced apoptosis in RSC96 cells in a concentration-dependent manner; promoted the accumulation of intracellular reactive oxygen species; decreased the mitochondrial membrane potential; caused the release of cytochrome c; and also increased the expression of Bcl-2 associated X protein, caspase-3, and cytochrome c proteins and decreased the expression of Bcl-2 protein. Further results also confirmed that CdTe QDs could be internalized by RSC96 cells, and the exposure and internalization of CdTe QDs could induce excessive endoplasmic reticulum stress in the cells, and the expression levels of binding immunoglobulin protein, C/EBP homologous protein, and caspase12 proteins were increased in a concentration-dependent manner. Moreover, autophagy-related proteins LC3II, Beclin1, and P62 all increased after CdTe QDs exposure, suggesting that CdTe QDs exposure both promoted autophagosome formation and inhibited autophagosome degradation, and that CdTe QDs affected the autophagic flow in RSC96 cells. In conclusion, CdTe QDs are able to cause apoptosis and autophagy in RSC96 cells through mitochondrial and endoplasmic reticulum stress pathways, and the possible neurotoxicity of CdTe QDs should be further investigated.

Protein corona mitigated the cytotoxicity of CdTe QDs to macrophages by targeting mitochondria.

Despite the potential of cadmium telluride quantum dots (CdTe QDs) in bioimaging and drug delivery, their toxic effects have been documented. It is known that the immunotoxicity of CdTe QDs targeting macrophages is one of their adverse effects, and the protein corona (PC) will affect the biological effects of QDs. In order to prove whether the PC-CdTe QDs complexes could alleviate the toxicity of CdTe QDs without weakening their luminescence, we investigated the impact of protein corona formed in fetal bovine serum (FBS) on the cytotoxicity of CdTe QDs to mitochondria. RAW264.7 cells were used as the model to compare the effects of CdTe QDs and PC-CdTe QDs complexes on the structure, function, quantity, morphology, and mitochondrial quality control of mitochondria. As result, the protein corona form in FBS alleviated the inhibition of CdTe QDs on mitochondrial activity, the damage to mitochondrial membrane, the increase of ROS, and the reduction of ATP content. Also, CdTe QDs increased the number of mitochondria in macrophages, while the complexes did not. In line with this, the morphology of mitochondrial network in macrophages which were exposed to CdTe QDs and PC-CdTe QDs complexes was different. CdTe QDs transformed the network into fragments, punctuations, and short rods, while PC-CdTe QDs complexes made the mitochondrial network highly branched, which was related to the imbalance of mitochondrial fission and fusion. Mechanically, CdTe QDs facilitated mitochondrial fission and inhibited mitochondrial fusion, while protein corona reversed the phenomenon caused by QDs. Besides mitochondrial dynamics, mitochondrial biogenesis and mitophagy were also affected. CdTe QDs increased the expression of mitochondrial biogenesis signaling molecules including PGC-1α, NRF-1 and TFAM, while PC-CdTe QDs complexes played the opposite role. With regard to mitophagy, they both showed promoting effect. In conclusion, the formation of protein corona alleviated the toxic effects of CdTe QDs on the mitochondria in macrophages and affected mitochondrial quality control. Under the premise of ensuring the fluorescence properties of CdTe QDs, these findings provided useful insight into reducing the toxicity of CdTe QDs from two perspectives: protein corona and mitochondria, and shared valuable information for the safe use of QDs.

Urban fine particulate matter causes cardiac hypertrophy through calcium-mediated mitochondrial bioenergetics dysfunction in mice hearts and human cardiomyocytes.

In recent years, the cardiovascular toxicity of urban fine particulate matter (PM2.5) has sparked significant alarm. Mitochondria produce 90% of ATP and make up 30% of the volume of cardiomyocytes. Thus knowledge of myocardial mitochondrial dysfunction due to PM2.5 exposure is essential for further cardiotoxic effects. Here, the mechanism of PM2.5-induced cardiac hypertrophy through calcium overload and mitochondrial dysfunction was investigated in vivo and in vitro. Male and female BALB/c mice were given 1.28, 5.5, and 11 mg PM2.5/kg bodyweight weekly through oropharyngeal inhalation for four weeks and were assigned to low, medium, and high dose groups, respectively. PM2.5-induced myocardial edema and cardiac hypertrophy were detected in the high-dose group. Mitochondria were scattered and ruptured with abnormal ultrastructural morphology. In vitro experiments on human cardiomyocyte AC16 showed that exposure to PM2.5 for 24 h caused opened mitochondrial permeability transition pore --leading to excessive calcium production, decreased mitochondrial membrane potential, weakened mitochondrial respiratory metabolism capacity, and decreased ATP production. Nevertheless, the administration of calcium chelator ameliorated the mitochondrial damage in the PM2.5-treated group. Our in vivo and in vitro results confirmed that calcium overload under PM2.5 exposure triggered mTOR/AKT/GSK-3ß activation, leading to mitochondrial bioenergetics dysfunction and cardiac hypertrophy.

The apoptosis induced by CdTe quantum dots through the mitochondrial pathway in dorsal root ganglion cell line ND7/23.

Recently, the use of CdTe quantum dots in the field of biomedicine, such as biological imaging, biosensors, cell markers, and drug carriers, is increasing due to their special physical and chemical properties. However, their biosafety assessment lags far behind their rapid application. In this study, we observed that CdTe quantum dots with certain exposed doses and time decreased the cell viability and increased the apoptosis rates in ND7/23 cells. In general, CdTe quantum dots exposure could promote the accumulation of reactive oxygen species (ROS) in cells and decrease the mitochondrial membrane potential, which led to pathological changes and subcellular organelle damages. We hypothesized that the mitochondrial pathway could be involved in CdTe quantum dots-induced apoptosis. The results suggested that CdTe quantum dots exposure increased the expression levels of three mitochondrial pathway markers, for example, caspase-3, cytochrome c, and Bax while decreased Bcl-2 protein expression, following with cytochrome c falling out of the inner membrane of mitochondrial and releasing into the cytoplasm. The application of caspase-3 protein inhibitor Ac-DEVD-CHO could decrease apoptosis rates in ND7/23 cells. The results, taken together, demonstrated that CdTe quantum dots could induce apoptosis of ND7/23 cells through the mitochondrial pathway. Our findings provide a novel insight for researchers to explore CdTe quantum dots' toxic mechanisms to reduce their adverse effects.

NADPH oxidases regulate endothelial inflammatory injury induced by PM2.5 via AKT/eNOS/NO axis.

Fine particulate matter (PM2.5 )-induced detrimental cardiovascular effects have been widely concerned, especially for endothelial cells, which is the first barrier of the cardiovascular system. Among potential mechanisms involved, reactive oxidative species take up a crucial part. However, source of oxidative stress and its relationship with inflammatory response have been rarely studied in PM2.5 -induced endothelial injury. Here, as a key oxidase that catalyzes redox reactions, NADPH oxidase (NOX) was investigated. Human umbilical vein endothelial cells (EA.hy926) were exposed to Standard Reference Material 1648a of urban PM2.5 for 24 h, which resulted in NOX-sourced oxidative stress, endothelial dysfunction, and inflammation induction. These are manifested by the up-regulation of NOX, increase of superoxide anion and hydrogen peroxide, elevated endothelin-1 (ET-1) and asymmetric dimethylarginine (ADMA) level, reduced nitric oxide (NO) production, and down-regulation of phosphorylation of endothelial NO synthase (eNOS) with increased levels of inducible NO synthase, as well as the imbalance between tissue-type plasminogen activator (tPA) and plasminogen activator inhibitor 1 (PAI-1), and changes in the levels of pro-inflammatory and anti-inflammatory factors. However, administration of NOX1/4 inhibitor GKT137831 alleviated PM2.5 -induced elevated endothelial dysfunction biomarkers (NO, ET-1, ADMA, iNOS, and tPA/PAI-1), inflammatory factors (IL-1ß, IL-10, and IL-18), and adhesion molecules (ICAM-1, VCAM-1, and P-selectin) and also passivated NOX-dependent AKT and eNOS phosphorylation that involved in endothelial activation. In summary, PM2.5 -induced NOX up-regulation is the source of ROS in EA.hy926, which activated AKT/eNOS/NO signal response leading to endothelial dysfunction and inflammatory damage in EA.hy926 cells.

CdTe and CdTe@ZnS quantum dots induce IL-1ß-mediated inflammation and pyroptosis in microglia.

CdTe quantum dots (QDs) are still widely considered as excellent fluorescent probes because of their far more superior optical performance and fluorescence efficiency than non­cadmium QDs. Thus, it is important to find ways to control their toxicity. In this study, CdTe QDs and CdTe@ZnS QDs both could cause IL-1ß-mediated inflammation following with pyroptosis in BV2 cells, but the toxic effects caused by CdTe@ZnS QDs was weaker than CdTe QDs, which demonstrated the partial protection of ZnS shell. When investigating the molecular mechanisms of QDs causing the inflammatory injury, the findings suggested that cadmium-containing QDs exposure activated NF-κB that participated in the NLRP3 inflammasome priming and pro-IL-1ß expression. After that, QDs-induced excessive ROS generation triggered the NLRP3 inflammasome activation and resulted in active caspase-1 to process pro-IL-1ß into mature IL-1ß release and inflammatory cell death, i.e. pyroptosis. Fortunately, the inhibitions of caspase-1, NF-κB and ROS or knocking down of NLRP3 all effectively attenuated the increases in the IL-1ß secretion and cell death caused by QDs in BV2 cells. This study provided two methods to alleviate the toxicity of cadmium-containing QDs, in which one is to encapsulate bare-core QDs with a shell and the other is to inhibit their toxic pathways. Since the latter way is more effective than the former one, it is significant to evaluate QDs through a mechanism-based risk assessment to identify controllable toxic targets.

A metabolomics study: CdTe/ZnS quantum dots induce polarization in mice microglia.

In this study, a metabolomic analysis was used to reveal the neurotoxicity of the CdTe/ZnS QDs via microglia polarization. A gas chromatography-mass spectrometer (GC-MS) was applied to uncover the metabonomic changes in microglia (BV-2 cell line) after exposure to 1.25 µM CdTe/ZnS QDs. 11 annotated metabolic pathways (KEGG database) were significantly changed in all exposed groups (3 h, 6 h, 12 h), 3 of them were related to glucose metabolism. The results of the Seahorse XFe96 Analyzer indicated that the CdTe/ZnS QDs increased the glycolysis level of microglia by 86% and inhibited the aerobic respiration level by 54% in a non-hypoxic environment. In vivo study, 3 h after the injection of CdTe/ZnS QDs (2.5 mM) through the tail vein in mice, the concentration of the CdTe/ZnS QDs in hippocampus reached the peak (1.25 µM). The polarization level of microglia (Iba-1 immunofluorescence) increased 2.7 times. In vitro study, the levels of the extracellular TNF-α, IL-1ß and NO of BV-2 cells were all increased significantly after a 6 h or 12 h exposure. According to the results of the Cell Counting Kit-8, after a 6 h or 12 h exposure to the CdTe/ZnS QDs, the exposed microglia could significantly decrease the number of neurons (HT-22 cell line). This study proved that CdTe/ZnS QDs could polarize microglia in the brain and cause secondary inflammatory damage to neurons. There are potential risks in the application of the CdTe/ZnS QDs in brain tissue imaging.

Toxicological study of metal and metal oxide nanoparticles in zebrafish.

Metal and metal oxide nanoparticles have been widely used in catalytic, electronic and biomedical fields. It is necessary to investigate their toxicity and potential hazards to human and aquatic ecosystems. Zebrafish (Danio rerio), as a promising animal model, has been increasingly utilized to assess the toxicity of nanoparticles. Zebrafish has numerous characteristics for toxicity evaluation, such as short life cycle and high fecundity. This review describes the advantages of using zebrafish in the toxicity assessment of metal and metal oxide nanoparticles. Then we focus on the toxic effects, particularly the acute toxicity and the chronic ones, induced by nanoparticles in zebrafish. Target organ toxicities are also mentioned, including immunotoxicity, developmental toxicity, neurotoxicity, reproductive toxicity, cardiovascular toxicity and hepatotoxicity. The toxic effects of selected metal nanoparticles, including Au, Ag, Cu, and metal oxide nanoparticles such as TiO2 , Al2 O3 , CuO, NiO and ZnO, as well as the underlying mechanisms of nanoparticles causing these effects, are also highlighted and described in detail. Furthermore, we introduce the general factors that affect nanoparticle-induced toxicity in zebrafish. The drawbacks and advantages of using the zebrafish model in nanotoxicity studies are also argued. Finally, we suggest that the application of zebrafish to assess chronic toxicity of metal and metal oxide nanoparticles and the joint toxicity of metal and metal oxide nanoparticles and other pollutants could be hot topics in nanotoxicology.

The role of NLRP3 inflammasome activation in the neuroinflammatory responses to Ag2Se quantum dots in microglia.

Silver selenide quantum dots (Ag2Se QDs) provide bright prospects for the application of QDs in the field of biomedicine because they contain low-toxic compounds and show great advantages in the imaging of deep tissues and tiny vascular structures. However, the biosafety of these novel QDs has not been thoroughly evaluated, especially in one main target for toxicity-the central nervous system (CNS). Our previous studies have suggested severe inflammatory responses to cadmium-containing QDs in the hippocampus, which gives us a hint regarding the risk assessment of Ag2Se QDs. In this study, microglial activation followed by enhanced levels of pro-inflammatory cytokines was observed in the hippocampus of mice intravenously injected with Ag2Se QDs. When using the microglial BV2 cells to investigate the underlying mechanisms, we found that the NLRP3 inflammasome activation was involved in the IL-1ß-mediated inflammation induced by Ag2Se QDs. On the one hand, Ag2Se QD-activated NF-κB participated in the NLRP3 inflammasome priming and assembly as well as the pro-IL-1ß upregulation. On the other hand, Ag2Se QD-induced ROS generation, particularly mtROS, triggered the NLRP3 inflammasome activation and resulted in active caspase-1 to process pro-IL-1ß into mature IL-1ß release. These findings not only indicated that it is important to evaluate the biosafety of novel QDs, even those containing low-toxic compounds, but also provided an unbiased and mechanism-based risk assessment of similar nanoparticles.

Identification of an amphioxus intelectin homolog that preferably agglutinates gram-positive over gram-negative bacteria likely due to different binding capacity to LPS and PGN.

Intelectin is a recently described galactofuranose-binding lectin that plays a role in innate immunity in vertebrates. Little is known about intelectin in invertebrates, including amphioxus, the transitional form between vertebrates and invertebrates. We cloned an amphioxus intelectin homolog, AmphiITLN-like, coding 302 amino acids with a conserved fibrinogen-related domain (FReD) in the N-terminus and an Intelectin domain in the C-terminus. In situ hybridization in adult amphioxus showed that AmphiITLN-like transcripts were highly expressed in the digestive tract and the skin. Quantitative real-time PCR revealed that AmphiITLN-like is significantly up-regulated in response to Staphylococcus aureus challenge, but only modestly to Escherichia coli. In addition, recombinant AmphiITLN-like expressed in E. coli agglutinates Gram-negative and Gram-positive bacteria to different degrees in a calcium dependent manner. Recombinant AmphiITLN-like could bind lipopolysaccharide (LPS) and peptidoglycan (PGN), the major cell wall components of Gram-negative and Gram-positive bacteria, respectively, with a higher affinity to PGN. Our work identified and characterized for the first time an amphioxus intelectin homolog, and provided insight into the evolution and function of the intelectin family.