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Background Montmorillonite is a type of nanoclay that originates from the clay fraction of the soil and is incorporated into polymers to form nanocomposites with enhanced mechanical strength, barrier, and flammability properties used for food packaging, automotive, and medical devices. However, with implementation in such consumer applications, the interaction of montmorillonite-based composites or derived byproducts with biological systems needs to be investigated. Methods Herein we examined the potential of Cloisite Na+ (pristine) and Cloisite 30B (organically modified montmorillonite nanoclay) and their thermally degraded byproducts’ to induce toxicity in model human lung epithelial cells. The experimental set-up mimicked biological exposure in manufacturing and disposal areas and employed cellular treatments with occupationally relevant doses of nanoclays previously characterized using spectroscopical and microscopical approaches. For nanoclay-cellular interactions and for cellular analyses respectively, biosensorial-based analytical platforms were used, with induced cellular changes being confirmed via live cell counts, viability assays, and cell imaging. Results Our analysis of byproducts’ chemical and physical properties revealed both structural and functional changes. Real-time high throughput analyses of exposed cellular systems confirmed that nanoclay induced significant toxic effects, with Cloisite 30B showing time-dependent decreases in live cell count and cellular viability relative to control and pristine nanoclay, respectively. Byproducts produced less toxic effects; all treatments caused alterations in the cell morphology upon exposure. Conclusions Our morphological, behavioral, and viability cellular changes show that nanoclays have the potential to produce toxic effects when used both in manufacturing or disposal environments. General significance The reported toxicological mechanisms prove the extensibility of a biosensorial-based platform for cellular behavior analysis upon treatment with a variety of nanomaterials.

Toxicity of engineered nanomaterials is associated with their inherent properties, both physical and chemical. Recent studies have shown that exposure to multi-walled carbon nanotubes (MWCNTs) promotes tumors and tumor-associated pathologies and lead to carcinogenesis in model in vivo systems. Here in we examined the potential of purified MWCNTs used at occupationally relevant exposure doses for particles not otherwise regulated to affect human lung epithelial cells. The uptake of the purified MWCNTs was evaluated using fluorescence activated cell sorting (FACS), while the effects on cell fate were assessed using 2- (4-iodophenyl) - 3- (4-nitrophenyl) - 5-(2, 4-disulfophenyl) -2H-tetrazolium salt colorimetric assay, cell cycle and nanoindentation. Our results showed that exposure to MWCNTs reduced cell metabolic activity and induced cell cycle arrest. Our analysis further emphasized that MWCNTs-induced cellular fate results from multiple types of interactions that could be analyzed by means of intracellular biomechanical changes and are pivotal in understanding the underlying MWCNTs-induced cell transformation.

Single-walled carbon nanotubes (SWCNTs) implementation in a variety of biomedical applications from bioimaging, to controlled drug delivery and cellular-directed alignment for muscle myofiber fabrication, has raised awareness of their potential toxicity. Nanotubes structural aspects which resemble asbestos, as well as their ability to induce cyto and genotoxicity upon interaction with biological systems by generating reactive oxygen species or inducing membrane damage, just to name a few, have led to focused efforts aimed to assess associated risks prior their user implementation. In this study, we employed a non-invasive and real-time electric cell impedance sensing (ECIS) platform to monitor behavior of lung epithelial cells upon exposure to a library of SWCNTs with user-defined physicochemical properties. Using the natural sensitivity of the cells, we evaluated SWCNT-induced cellular changes in relation to cell attachment, cell–cell interactions and cell viability respectively. Our methods have the potential to lead to the development of standardized assays for risk assessment of other nanomaterials as well as risk differentiation based on the nanomaterials surface chemistry, purity and agglomeration state.

The toxicity of engineered nanomaterials in biological systems depends on both the nanomaterial properties and the exposure duration. Herein we used a multi-tier strategy to investigate the relationship between user-characterized multi-walled carbon nanotubes (MWCNTs) exposure duration and their induced biochemical and biomechanical effects on model human lung epithelial cells (BEAS-2B). Our results showed that exposure to MWCNTs leads to time-dependent intracellular uptake and generation of reactive oxygen species (ROS), along with time-dependent gradual changes in cellular biomechanical properties. In particular, the amount of internalized MWCNTs followed a sigmoidal curve with the majority of the MWCNTs being internalized within 6h of exposure; further, the sigmoidal uptake correlated with the changes in the oxidative levels and cellular biomechanical properties respectively. Our study provides new insights into the time-dependent induced toxicity caused by exposure to occupationally relevant doses of MWCNTs and could potentially help establish bases for early risk assessments of other nanomaterials toxicological profiles.

Nanodiamonds (NDs) are an emerging class of engineered nanomaterials that hold great promise for the next generation of bionanotechnological products to be used for drug and gene delivery, or for bio-imaging and biosensing. Previous studies have shown that upon their cellular uptake, NDs exhibit high biocompatibility in various in vitro and in vivo set-ups. Herein we hypothesized that the increased NDs biocompatibility is a result of minimum membrane perturbations and their reduced ability to induce disruption or damage during cellular translocation. Using multi-scale combinatorial approaches that simulate ND-membrane interactions, we correlated NDs real-time cellular uptake and kinetics with the ND-induced membrane fluctuations to derive energy requirements for the uptake to occur. Our discrete and real-time analyses showed that the majority of NDs internalization occurs within 2 h of cellular exposure, however, with no effects on cellular viability, proliferation or cellular behavior. Furthermore, our simulation analyses using coarse-grained models identified key changes in the energy profile, membrane deformation and recovery time, all functions of the average ND or ND-based agglomerate size. Understanding the mechanisms responsible for ND-cell membrane interactions could possibly advance their implementation in various biomedical applications.

Digitoxin belongs to a naturally occurring class of cardiac glycosides (CG); digitoxin is clinically approved for heart failure and known for its anti-cancer effects against non-small lung cancer cells (NSCLC). However, concerns associated with its narrow therapeutic index and its concentration-dependent mechanism of action are rising. Thus, before digitoxin implementation in designing and developing safer and more effective CG-based anti-cancer therapies, its pharmacological and safety profiles need to be fully elucidated. In this research we used a combinatorial approach to evaluate the anti-cancer mechanisms of digitoxin in real-time. Our approach employed a non-invasive electric cell impedance sensing technique as a proxy to monitor NSCLC behavior post-exposure to toxic, therapeutic and sub-therapeutic concentrations of the drug. By developing structure–function combinatorial relations we showed that digitoxin targets cancer cells in a time and dose-dependant manner by activating pro-apoptotic and anti-proliferative signaling cascades that results in strengthening cellular adhesion and sequestration of key regulatory proliferation protein from the nucleus.

The unique properties of single walled carbon nanotubes (SWCNTs) make them viable candidates for versatile implementation in the next generation of biomedical devices for targeted delivery of chemotherapeutic agents or cellular-sensing probes. Such implementation requires user-tailored changes in SWCNT’s physicochemical characteristics to allow for efficient cellular integration while maintaining nanotubes’ functionality. However, isolated reports showed that user-tailoring could induce deleterious effects in exposed cells, from decrease in cellular proliferation, to changes in cellular adhesion, generation of reactive oxygen species or phenotypical variations, just to name a few. Before full implementation of SWCNTs is achieved, their toxicological profiles need to be mechanistically correlated with their physicochemical properties to determine how the induced cellular fate is related to the exposure conditions or samples’ characteristics. Our study provides a comprehensive analysis of the synergistic cyto- and genotoxic effects resulted from short-term exposure of human lung epithelial cells to pristine (as manufactured) and user-tailored SWCNTs, as a function of their physicochemical properties. Specifically, through a systematic approach we are correlating the nanotube uptake and nanotube-induced cellular changes to the sample’s physicochemical characteristics (e.g., metal impurities, length, agglomerate size, surface area, dispersion, and surface functionalization). By identifying changes in active hallmarks involved in cell-cell connections and maintaining epithelial layer integrity, we also determine the role that short-term exposure to SWCNTs plays in the overall cellular fate and cellular transformation. Lastly, we assess cellular structure-function relationships to identify non-apoptotic pathways induced by SWCNTs exposure that could however lead to changes in cellular behavior and cellular transformation. Our results show that the degree of cell transformation is a function of the physicochemical properties of the SWCNT, with the nanotube with higher length, higher metal content and larger agglomerate size reducing cell viability to a larger extent. Such changes in cell viability are also complemented by changes in cell structure, cycle and cell-cell interactions, all responsible for maintaining cell fate.