Key Role of Choline Head Groups in Large Unilamellar Phospholipid Vesicles for the Interaction with and Rupture by Silica Nanoparticles

For highly abundant silica nanomaterials, detrimental effects on proteins and phospholipids are postulated as critical molecular initiating events that involve hydrogen-bonding, hydrophobic, and/or hydrophilic interactions. Here, large unilamellar vesicles with various well-defined phospholipid compositions are used as biomimetic models to recapitulate membranolysis, a process known to be induced by silica nanoparticles in human cells. Differential analysis of the dominant phospholipids determined in membranes of alveolar lung epithelial cells demonstrates that the quaternary ammonium head groups of phosphatidylcholine and sphingomyelin play a critical and dose-dependent role in vesicle binding and rupture by amorphous colloidal silica nanoparticles. Surface modification by either protein adsorption or by covalent coupling of carboxyl groups suppresses the disintegration of these lipid vesicles, as well as membranolysis in human A549 lung epithelial cells by the silica nanoparticles. Furthermore, molecular modeling suggests a preferential affinity of silanol groups for choline head groups, which is also modulated by the pH value. Biomimetic lipid vesicles can thus be used to better understand specific phospholipid–nanoparticle interactions at the molecular level to support the rational design of safe advanced materials.     

Amorphous Silica Nanoparticles Promote Monocyte Adhesion to Human Endothelial Cells: Size‐Dependent Effect

There is evidence that nanoparticles can induce endothelial dysfunction. Here, the effect of monodisperse amorphous silica nanoparticles (SiO₂‐NPs) of different diameters on endothelial cells function is examined. Human endothelial cell line (EA.hy926) or primary human pulmonary artery endothelial cells (hPAEC) are seeded in inserts introduced or not above triple cell co‐cultures (pneumocytes, macrophages, and mast cells). Endothelial cells are incubated with SiO₂‐NPs at non‐cytotoxic concentrations for 12 h. A significant increase (up to 2‐fold) in human monocytes adhesion to endothelial cells is observed for 18 and 54 nm particles. Exposure to SiO₂‐NPs induces protein expression of adhesion molecules (ICAM‐1 and VCAM‐1) as well as significant up‐regulation in mRNA expression of ICAM‐1 in both endothelial cell types. Experiments performed with fluorescent‐labelled monodisperse amorphous SiO₂‐NPs of similar size evidence nanoparticle uptake into the cytoplasm of endothelial cells. It is concluded that exposure of human endothelial cells to amorphous silica nanoparticles enhances their adhesive properties. This process is modified by the size of the nanoparticle and the presence of other co‐cultured cells.     

Cytotoxicity of carboxyl carbon nanotubes on human embryonic lung fibroblast cells and its mechanism

The wide use in various fields and the great potentials in biomedical applications of carbon nanotubes highlight the need to study their toxicity and biocompatibility for recent years. This work aimed to investigate the cytotoxicity of carbon nanotubes on human embryonic lung fibroblast cells and their inter-related affecting factors. Three carboxyl modified carbon nanotubes, short carboxyl single-walled carbon nanotubes (SWCNTs-COOH), short carboxyl double-walled carbon nanotubes (DWCNTs-COOH) and short carboxyl multi-walled carbon nanotubes (MWCNTs-COOH) were chosen as subjects for the evaluation of carbon nanotubes cytotoxity. Different concentrations of carboxyl carbon nanotubes were incubated with human embryonic lung fibroblast (HELF) cells for 48?h, respectively, and the electron microscopy was used to observe the cell growth and morphology. The results showed that MWCNTs-COOH, which had a better dispersion in water was much more cytotoxic than the other two carbon nanotubes. From Cell Counting Kit-8 assay and acridine orange staining, MWCNTs-COOH could inhibit the cell growth and induce cell apoptosis with a dose–effect relationship and oxidative stress may be one of the mechanisms.     

Cationic silica nanoparticles as gene carriers: synthesis, characterization and transfection efficiency in vitro and in vivo

The potential of cationic SiO2 nanoparticles was investigated for in vivo gene transfer in this study. Cationic SiO2 nanoparticles with surface modification were generated using amino-hexyl-amino-propyltri-methoxysilane (AHAPS). The zeta potential of the nanoparticles at pH = 7.4 varied from -31.4 mV (unmodified particles; 10 nm) to +9.6 mV (modified by AHAPS). Complete immobilization of DNA at the nanoparticle surface was achieved at a particle ratio of 80 (w/w nanoparticle/DNA ratio). The surface modified nanoparticle had a size of 42 nm with a distribution from 10-100 nm. The ability of these particles to transfect pCMVbeta reporter gene was tested in Cos-1 cells, and optimum results were obtained in the presence of FCS and chloroquine at a particle ratio of 80. These nanoparticles were tested for their ability to transfer genes in vivo in the mouse lung, and a two-times increase in the expression levels was found with silica particles in comparison to EGFP alone. Very low or no cell toxicity was observed, suggesting silica nanoparticles as potential alternatives for gene transfection.     

Mechanisms of Antibacterial Activity of MgO: Non‐ROS Mediated Toxicity of MgO Nanoparticles Towards Escherichia coli

The toxicity of metal oxide nanomaterials and their antimicrobial activity is attracting increasing attention. Among these materials, MgO is particularly interesting as a low cost, environmentally‐friendly material. The toxicity of MgO, similar to other metal oxide nanomaterials, is commonly attributed to the production of reactive oxygen species (ROS). We investigated the toxicity of three different MgO nanoparticle samples, and clearly demonstrated robust toxicity towards Escherichia coli bacterial cells in the absence of ROS production for two MgO nanoparticle samples. Proteomics data also clearly demonstrate the absence of oxidative stress and indicate that the primary mechanism of cell death is related to the cell membrane damage, which does not appear to be due to lipid peroxidation.     

Shape‐Dependent Cytotoxicity and Proinflammatory Response of Poly(3,4‐ethylenedioxythiophene) Nanomaterials

Poly(3,4‐ethylenedioxythiophene) (PEDT) is recognized as one of the most promising conducting polymers for future applications in the fields of electronics, optics, energy storage/conversion, and biomedical science. The toxicity of PEDT could be considered to affect the potential for its widespread application. Herein, the cytotoxicity and proinflammatory response of PEDT nanomaterials of three different shapes toward human lung fibroblast (IMR90) and mouse alveolar macrophage (J774A.1) cells are investigated. The shape‐dependent toxicity of the PEDT nanomaterials is evaluated by examining cell morphological change, cytotoxicity, apoptosis/necrosis, oxidative stress, and immune response. The cytotoxicity and apoptosis of the nanomaterials increase with their decreasing aspect ratio in both cell lines. The formation of reactive oxygen species in cells treated with PEDT nanomaterials is dependent on the shape and concentration of the nanomaterial. Proinflammatory cytokines, such as interleukin‐1, interleukin‐6, and tumor necrosis factor α from macrophages, are induced by PEDT nanomaterial‐treated cells.     

Preferential killing of cancer cells and activated human T cells using ZnO nanoparticles

Nanoparticles are increasingly being recognized for their potential utility in biological applications including nanomedicine. Here we examine the response of normal human cells to ZnO nanoparticles under different signaling environments and compare it to the response of cancerous cells. ZnO nanoparticles exhibit a strong preferential ability to kill cancerous T cells ( approximately 28-35x) compared to normal cells. Interestingly, the activation state of the cell contributes toward nanoparticle toxicity, as resting T cells display a relative resistance while cells stimulated through the T cell receptor and CD28 costimulatory pathway show greater toxicity in direct relation to the level of activation. Mechanisms of toxicity appear to involve the generation of reactive oxygen species, with cancerous T cells producing higher inducible levels than normal T cells. In addition, nanoparticles were found to induce apoptosis and the inhibition of reactive oxygen species was found to be protective against nanoparticle induced cell death. The novel findings of cell selective toxicity, towards potential disease causing cells, indicate a potential utility of ZnO nanoparticles in the treatment of cancer and/or autoimmunity.     

Endosomal Escape of Polymer‐Coated Silica Nanoparticles in Endothelial Cells

Current investigations into hazardous nanoparticles (i.e., nanotoxicology) aim to understand the working mechanisms that drive toxicity. This understanding has been used to predict the biological impact of the nanocarriers as a function of their synthesis, material composition, and physicochemical characteristics. It is particularly critical to characterize the events that immediately follow cell stress resulting from nanoparticle internalization. While reactive oxygen species and activation of autophagy are universally recognized as mechanisms of nanotoxicity, the progression of these phenomena during cell recovery has yet to be comprehensively evaluated. Herein, primary human endothelial cells are exposed to controlled concentrations of polymer‐functionalized silica nanoparticles to induce lysosomal damage and achieve cytosolic delivery. In this model, the recovery of cell functions lost following endosomal escape is primarily represented by changes in cell distribution and the subsequent partitioning of particles into dividing cells. Furthermore, multilamellar bodies are found to accumulate around the particles, demonstrating progressive endosomal escape. This work provides a set of biological parameters that can be used to assess cell stress related to nanoparticle exposure and the subsequent recovery of cell processes as a function of endosomal escape.     

Analysis of stress responsive genes induced by single-walled carbon nanotubes in BJ Foreskin cells

Nanotechnology is finding its use as a potential technology in consumer products, defense, electronics, and medical applications by exploiting the properties of nanomaterials. Single-walled carbon nanotubes are novel forms of these nanomaterials with potential for large applications. However, the toxicity studies on this material are not explored in detail and therefore limiting its use. It has been earlier reported that single-walled carbon nanotubes induces oxidative stress and also dictates activation of specific signaling pathway in keratinocytes. The present study explores the effect of single-walled carbon nanotubes on stress genes in human BJ Foreskin cells. The results show induction of oxidative stress in BJ Foreskin cells by single-walled carbon nanotubes and increase in stress responsive genes. The genes included inducible genes like HMOX1, HMOX2, and Cyp1B1. In addition we validated increase for four genes by SWCNT, namely ATM, CCNC, DNAJB4, and GADD45A by RT-PCR. Moreover results of the altered stress related genes have been discussed and that partially explains some of the toxic responses induced by single-walled carbon nanotubes.     

Surface modifications of ZnO nanoparticles and their cytotoxicity

ZnO is a well-known UV absorber. At small particle sizes its absorption efficiency is substantially increased and this property, combined with transparency to visible light, has attracted growing interest in its applications in personal care products such as sunscreens. However, some recent studies suggest that ZnO nanoparticles could induce considerable toxicity to certain cells and microorganisms. Aiming to reduce cytotoxicity of ZnO nanoparticles without impairing their unique properties, this paper examines the influence of surface modifications to ZnO nanoparticles using coatings such as silica (SiO2) and Poly methyl Acrylic Acid (PMAA). It was found that both PMAA and SiO2 coatings were physically attached to the ZnO surface and their presence did not weaken UV absorption of the original nanoparticles. Uncoated ZnO and SiO2-coated ZnO exhibited similar cytotoxicity to human lymphoblastoid cells, while PMAA-coated ZnO nanoparticles had little adverse effect except at high concentrations. The type of coating was also shown to affect the generation of Reactive Oxygen Species (ROS). The correlation between cell viability and ROS level led to conclusions that enhanced oxidative stress could be one of the reasons for cytotoxicity of ZnO nanoparticles.     

On-chip evaluation of shear stress effect on cytotoxicity of mesoporous silica nanoparticles

In this work, nanotoxicity in the bloodstream was modeled, and the cytotoxicity of sub-50 nm mesoporous silica nanoparticles to human endothelial cells was investigated under microfluidic flow conditions. Compared to traditional in vitro cytotoxicity assays performed under static conditions, unmodified mesoporous silica nanoparticles show higher and shear stress-dependent toxicity to endothelial cells under flow conditions. Interestingly, even under flow conditions, highly organo-modified mesoporous silica nanoparticles show no significant toxicity to endothelial cells. This paper clearly demonstrates that shear stress is an important factor to be considered in in vitro nanotoxicology assessments and provides a simple device for pursuing this consideration.     

SO(2) inhalation modulates the expression of pro-inflammatory and pro-apoptotic genes in rat heart and lung

SO(2) is a common air pollutant, and human exposure to SO(2) has become increasingly widespread due to the combustion of fossil fuels. The epidemiological studies have linked SO(2) exposure not only with many respiratory responses, but also with cardiovascular diseases. Also, its possible toxicity has been implicated by determining oxidative stress, DNA damage and membrane channel alteration in rat heart and lung. However, its detailed mechanisms remain unclear. In the present study, rats were treated with 7, 14 and 28 mg/m(3) SO(2) for 6h/day for 7 days, and the mRNA levels of TNF-α, IL-1β, iNOS, ICAM-1, Bax and Bcl-2 and subsequent insults were determined in the heart and lung. The results indicate that SO(2) inhalation markedly elevated TNF-α and IL-1β mRNA levels and secretions, enhanced iNOS and ICAM-1 mRNA levels and the ratio of Bax/Bcl-2 in a concentration-dependent manner, and induced occurrence of apoptosis. This suggests that SO(2) inhalation induced an inflammatory response and subsequent insults via modulating pro-inflammatory and pro-apoptotic genes in the heart and lung, which contributed to the increased risk of respiratory and cardiovascular diseases.     

The Janus faces of nanoparticles

There is an paradox apparent in the fact that nanoparticles have potential use in nanomedicine for imaging and therapy, whereas combustion-derived NP are thought to be responsible for adverse health effects of air pollution. The nanotechnology industry is in the process of producing a number of new nanoparticles which are as-yet unquantified with regard to both hazard and potential for human exposure. The toxicology of combustion-derived nanoparticles is developing and there is now considerable understanding of how they might drive both adverse lung and cardiovascular effects, including the importance of small size, large relative surface area and oxidative stress. Medicinal nanoparticles are being developed and tested on a case-by-case basis using testing protocols from biomaterials and drug safety and with regard to risk-benefit. There are considerable differences in physical and chemical properties and biodegradability between medicinal nanoparticles and the industrial and combustion-derived nanoparticles studied by particle toxicologists and we would anticipate that the bulk of medicinal NP types will be of low toxicity. However, to resolve the nanoparticle paradox there is a need to advance understanding of the characteristics that control acute and chronic toxicity, translocation, biodegradation and elimination of all of the types of particles likely to gain access to the human body. Much would be gained in this area by collaboration between particle toxicologists and nanopharmacologists.     

In vitro safety toxicology data for evaluation of gold nanoparticles-chronic cytotoxicity, genotoxicity and uptake

Safety and toxic effects of nanoparticles are still largely unexplored due to the multiple aspects that influence their behaviour toward biological systems. Here, we focus the attention on 12 nm spherical gold nanoparticle coated or not with hyaluronic acid compared to its precursor counterpart salt. Results ranging from the effects of a 10-days exposure in an in vitro model with BALB/c 3T3 fibroblast cells show how 12 nm spherical gold nanoparticles are internalized from 3T3 cells by endo-lysosomal pathway by an indirect measurement technique; and how gold nanoparticles, though not being a severe cytotoxicant, induce DNA damage probably through an indirect mechanism due to oxidative stress. While coating them with hyaluronic acid reduces gold nanoparticles cytotoxicity and slows their cell internalization. These results will be of great interest to medicine, since they indicate that gold nanoparticles (with or without coating) are suitable for therapeutic applications due to their tunable cell uptake and low toxicity.     

In vitro and in vivo toxicity of CdTe nanoparticles

Cadmium telluride (CdTe) nanoparticles exhibit strong and stable fluorescence that is attractive for many applications such as biological probing and solid state lighting. The evaluation of nanoparticle toxicity is important for realizing these practical applications. However, no systematic studies of CdTe nanoparticle toxicity have been reported. We investigated and compared the size- and concentration-dependent cytotoxicity of CdTe nanoparticles in human hepatoma HepG2 cells using the MTT assay. CdTe nanoparticles elicited cytotoxicity in a concentration- and size-dependent manner, with smaller-sized particles exhibiting somewhat higher potency. Lesser cytotoxicity of partially purified CdTe-Red particles (following methanol precipitation and resuspension) suggested that free cadmium ions may contribute to cytotoxicity. We also evaluated the acute toxicity of CdTe-Red particles following intravenous exposure in male rats (2 micromol/kg). Few signs of functional toxicity or clinical (urinary or blood) changes were noted. Interestingly, motor activity was transiently reduced (2 hours after treatment) and then significantly increased at a later timepoint (24 hours after dosing). These studies provide a framework for further characterizing the in vitro and in vivo toxic potential of different types of CdTe nanoparticles and suggest that the nervous system may be targeted by these nanoparticles under some conditions.     

Single-walled carbon nanotubes induces oxidative stress in rat lung epithelial cells

Single-walled carbon nanotubes (SWCNT) show unique properties find applications in micro devices; electronics to biological systems specially drug delivery and gene therapy. However the manufacture and extensive use of nanotubes raises concern about its safe use and human health. Very few studies have been carried out on toxicity of carbon nanotubes in experimental animals and humans, thus resulted in limiting their use. The extensive toxicological studies using in vitro and in vivo models are necessary and are required to establish safe manufacturing guidelines and also the use of SWCNT. These studies also help the chemists to prepare derivative of SWCNT with less or no toxicity. The present study was undertaken to determine the toxicity exhibited by SWCNT in rat lung epithelial cells as a model system. Lung epithelial cells (LE cells) were cultured with or without SWCNT and reactive oxygen species (ROS) produced were measured by change in fluorescence using dichloro fluorescein (DCF). The results show increased ROS on exposure to SWCNT in a dose and time dependent manner. The decrease in glutathione content suggested the depletion and loss of protective mechanism against ROS in SWCNT treated cells. Use of rotenone, the inhibitor of mitochondrial function have no effect on ROS levels suggested that mitochondria is not involved in SWCNT induced ROS production. Studies carried out on the effect of SWCNT on superoxide dismutase (SOD-1 and SOD-2) levels in LE cells, indicates that these enzyme levels decreased by 24 hours. The increased ROS induced by SWCNT on LE cells decreased by treating the cells with 1 mM of glutathione, N-Acetyl Cysteine, and Vitamin C. These results further prove that SWCNT induces oxidative stress in LE cells and shows loss of antioxidants.     

Comparative cytotoxicity evaluation of different size gold nanoparticles in human dermal fibroblasts

The aim of this work was to compare the effects of three commercially available gold nanoparticles (AuNPs) of different sizes (30, 50 and 90 nm) on the viability of normal human dermal fibroblasts (NHDF). In addition, we evaluated protective effect of N-Acetyl-L-cysteine (NAC), total glutathione content (GSH/GSSG), superoxide dismutase (SOD) activity and reactive oxygen species (ROS) production to investigate if oxidative stress was involved in the cytotoxic response of these AuNPs. Although AuNP-induced cytotoxicity was dose and time dependent, nanoparticle size slightly influenced the cytotoxic response of AuNPs assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and lactate dehydrogenase. Regarding oxidative parameters, NAC produced no significant protection of NHDF cells against treatment with any of the three AuNPs. Independently on nanoparticle size, GSH/GSSG content was drastically depleted after 24 h of incubation with the three AuNPs (less than 15% in all cases), while no statistically significant changes on SOD activity were reported (～90% of activity). The three AuNPs also caused a notable increase in the ROS production of NHDF cells. In conclusion, our data suggest that AuNP-induced cytotoxicity in NHDF is mediated by oxidative stress and it is independent of nanoparticle size.     

Programmed Nanoparticle‐Loaded Nanoparticles for Deep‐Penetrating 3D Cancer Therapy

Tumors are 3D, composed of cellular agglomerations and blood vessels. Therapies involving nanoparticles utilize specific accumulations due to the leaky vascular structures. However, systemically injected nanoparticles are mostly uptaken by cells located on the surfaces of cancer tissues, lacking deep penetration into the core cancer regions. Herein, an unprecedented strategy, described as injecting “nanoparticle‐loaded nanoparticles” to address the long‐lasting problem is reported for effective surface‐to‐core drug delivery in entire 3D tumors. The “nanoparticle‐loaded nanoparticle” is a silica nanoparticle (≈150 nm) with well‐developed, interconnected channels (diameter of ≈30 nm), in which small gold nanoparticles (AuNPs) (≈15 nm) with programmable DNA are located. The nanoparticle (AuNPs)‐loaded nanoparticles (silica): (1) can accumulate in tumors through leaky vascular structures by protecting the inner therapeutic AuNPs during blood circulation, and then (2) allow diffusion of the AuNPs for penetration into the entire surface‐to‐core tumor tissues, and finally (3) release a drug triggered by cancer‐characteristic pH gradients. The hierarchical “nanoparticle‐loaded nanoparticle” can be a rational design for cancer therapies because the outer large nanoparticles are effective in blood circulation and in protection of the therapeutic nanoparticles inside, allowing the loaded small nanoparticles to penetrate deeply into 3D tumors with anticancer drugs.     

Influence of shape,adhension and simulated lung mechanics on amorphous silica nanoparticle toxicity

Prevailing theories suggest that acicular, or fiber-like, particles induce enhanced toxicity over isotropic material through hindrance of phagocyte-mediated clearance mechanisms and through the aggravation of proximal cells via mechanical interactions. Currently, the degree to which either of these mechanisms operates is not well understood. To gain a more fundamental understanding of acicular particle toxicity, we have synthesized submicron and nanoscale amorphous silica spheres and rods as model materials for shape-driven toxicological experimentation. To accentuate contributions from mechanical damage in vitro, exposure studies were performed in the presence and absence of simulated lung mechanics. To promote and mitigate cell–particle contact-mediated mechanical interactions, the adhesion of the particles to the cell membrane was respectively modified by the physisorption of fibronectin and chemisorption of the polyethylene glycol to the silica particle surface. Lactic acid dehydrogense (LDH) and interleukin (IL)-8 release were used as endpoints for cytotoxicity and inflammation, respectively. The results indicate that particle exposures in the presence of physiological stretch induce increased LDH release and IL-8 expression regardless of shape. Moreover, it is evident that shape-induced aggregation may play a significant role in mitigating particle clearance pathways.