Integrated high-throughput drug screening microfluidic system for comprehensive ocular toxicity assessment.

Traditional experimental methodologies suffer from a few limitations in the toxicological evaluation of the preservatives added to eye drops. In this study, we overcame these limitations by using a microfluidic device. We developed a microfluidic system featuring a gradient concentration generator for preservative dosage control with microvalves and micropumps, automatically regulated by a programmable Arduino board. This system facilitated the simultaneous toxicological evaluation of human corneal epithelial cells against eight different concentrations of preservatives, allowing for quadruplicate experiments in a single run. In our study, the IC50 values for healthy eyes and those affected with dry eyes syndrome showed an approximately twofold difference. This variation is likely attributable to the duration for which the preservative remained in contact with corneal cells before being washed off by the medium, suggesting the significance of exposure time in the cytotoxic effect of preservatives. Our microfluidic system, automated by Arduino, simulated healthy and dry eye environments to study benzalkonium chloride toxicity and revealed significant differences in cell viability, with IC50 values of 0.0033% for healthy eyes and 0.0017% for dry eyes. In summary, we implemented the pinch-to-zoom feature of an electronic tablet in our microfluidic system, offering innovative alternatives for eye research.

Proteomic analysis using isobaric tags for relative and absolute quantification technology reveals mechanisms of toxic effects of tris (1,3-dichloro-2-propyl) phosphate on RAW264.7 macrophage cells.

Tris (1,3-dichloro-2-propyl) phosphate (TDCIPP) is one of the most commonly used organophosphorus flame retardants. Immuno-toxicity induced by TDCIPP is becoming of increasing concern. However, effects of TDCIPP on immune cells and mechanisms resulting in those effects are poorly understood. In this study, it was determined, for the first time, by use of isobaric tags for relative and absolute quantification (iTRAQ) based proteomic techniques expression of global proteins in RAW264.7 cells exposed to 10 µM TDCIPP. A total of 180 significantly differentially expressed proteins (DEPs) were identified. Of these, 127 were up-regulated and 53 were down-regulated. The DEPs associated with toxic effects of TDCIPP were then screened by use of Gene Ontology and the Kyoto Encyclopedia of Genes and Genomes for enrichment analysis. Results showed that these DEPs were involved in a number of pathways including apoptosis, DNA damage, cell cycle arrest, immune-toxicity, and signaling pathways, such as the Toll-like receptor, PPAR and p53 signaling pathways. The complex regulatory relationships between different DEPs, which might play an important role in cell death were also observed in the form of a protein-protein interaction network. Meanwhile, mitochondrial membrane potential (MMP) in RAW264.7 cells after TDCIPP treatment was also analyzed, the collapse of the MMP was speculated to play an important role in TDCIPP induced apoptosis. Moreover, some of the important regulator proteins discovered in this study, such as Chk1, Aurora A, would provide novel insight into the molecular mechanisms involved in toxic responses to TDCIPP.

Establishment of an in vitro method of rabbit embryo toxicity with toxicokinetics study.

This report introduces a novel method, rabbit whole embryo culture (WEC) combined with toxicokinetics (TK), for toxicity testing. Rodent WEC has been extensively used for in vitro screening of developmental toxicity. To improve the reliability of in vitro data, it is important to consider TK and species specificity. To test the utility and effectiveness of this method, we investigated the toxic effect of thalidomide on rabbit embryos and its behavior in test systems both in vitro and in vivo under the same experimental condition. The data showed that thalidomide induced embryo malformations such as embryonic brain hypoplasia, short limb buds, and declined embryonic growth both in vitro and in vivo. The toxic effect increased with the increasing exposure of the embryo to thalidomide. In addition, we observed similar toxic effects and exposure-effect relationships in vivo and in vitro. Therefore, we preliminarily conclude that this new method can effectively predict and explain thalidomide toxicity. Furthermore, we investigated the behavior of test compounds in the WEC system for the first time, and this method is expected to be an important technique for in vitro toxicity study after extensive verification.

Dynamic microfluidic bioreactor-Hip simulator (DMBH) system for implant toxicity monitoring.

The generation of degradation products (DPs) like ions and organo-metallic particles from corroding metallic implants is an important healthcare concern. These DPs generate local and systemic toxicity. The impact on local toxicity is well documented, however, little is known about systemic toxicity. This is mainly due to the limited scope of the current microtiter plate-based (static) toxicity assay techniques. These methods do not mimic the systemic (dynamic) conditions. In this study, it is hypothesized that DPs incubated with cells in static conditions might provide improper systemic toxicity results, as there is no movement mimicking the blood circulation around cells. This study reports the development of a three-chambered prototype microfluidic system connected to the operational hip implant simulator to test the cellular response induced by the DPs. This setup is called a dynamic microfluidic bioreactor-hip simulator system. We hypothesize that a dynamic microfluidic system will provide a realistic toxicology response induced by DPs than a static cell culture plate. To prove the hypothesis, Neuro2a (N2a) cells were used as representative cells to study systemic neurotoxicity by the implant DPs. The microfluidic bioreactor system was validated by comparing the cell toxicity against the traditional static system and using COMSOL modeling for media flow with DPs. The hip implant simulator used in this study was a state-of-the-art sliding hip simulator developed in our lab. The results suggested that static toxicity was significantly more compared to dynamic microfluidic-based toxicity. The newly developed DMBH system tested for in situ systemic toxicity on N2a cells and demonstrated very minimum toxicity level (5.23%) compared to static systems (31.16%). Thus, the new DMBH system is an efficient tool for in situ implant metal systemic toxicity testing.

Evaluation of Technology-Enabled Monitoring of Patient-Reported Outcomes to Detect and Treat Toxic Effects Linked to Immune Checkpoint Inhibitors.

Importance: Immune checkpoint inhibitors can produce distinct toxic effects that require prompt recognition and timely management. Objective: To develop a technology-enabled, dynamically adaptive protocol that can provide the accurate information needed to inform specific remedies for immune toxic effects in patients treated with immune checkpoint inhibitors. Design, Setting, and Participants: An open-label cohort study was conducted at a single tertiary referral center from September 6, 2019, to September 3, 2020. The median follow-up duration was 63 (interquartile range, 35.5-122) days. Fifty patients with genitourinary cancers treated with immune checkpoint inhibitors were enrolled. Interventions: A fit-for-purpose electronic platform was developed to enable active patient and care team participation. A smartphone application downloaded onto patients' personal mobile devices prompted them to report their symptoms at least 3 times per week. The set of symptoms and associated queries were paired with alert thresholds for symptoms requiring clinical action. Main Outcomes and Measures: The primary end point of this interim analysis was feasibility, as measured by patient and care team adherence, and lack of increase in care team staffing. Operating characteristics were estimated for each symptom alert and used to dynamically adapt the alert thresholds to ensure sensitivity while reducing unnecessary alerts. Results: Of the 50 patients enrolled, 47 had at least 1 follow-up visit and were included in the analysis. Median age was 65 years (range, 37-86), 39 patients (83%) were men, and 39 patients (83%) had metastatic cancer, with the most common being urothelial cell carcinoma and renal cell carcinoma (22 [47%] patients each). After initial onboarding, no further care team training or additional care team staffing was required. Patients had a median study adherence rate of 74% (interquartile range, 60%-86%) and 73% of automated alerts were reviewed within 3 days by the clinic team. Symptoms with the highest positive predictive value for adverse events requiring acute intervention included dizziness (21%), nausea/vomiting (26%), and shortness of breath (14%). The symptoms most likely to result in unnecessary alerts were arthralgia and myalgia, fatigue, and cough. Conclusions and Relevance: The findings of this cohort study suggest an acceptable and fiscally sound method can be developed to create a dynamic learning system to detect and manage immune-related toxic effects.

Drug Evaluation Based on a Multi-Channel Cell Chip with a Horizontal Co-Culture.

We developed a multi-channel cell chip containing a three-dimensional (3D) scaffold for horizontal co-culture and drug toxicity screening in multi-organ culture (human glioblastoma, cervical cancer, normal liver cells, and normal lung cells). The polydimethylsiloxane (PDMS) multi-channel cell chip (PMCCC) was based on fused deposition modeling (FDM) technology. The architecture of the PMCCC was an open-type cell chip and did not require a pump or syringe. We investigated cell proliferation and cytotoxicity by conducting 3-(4,5-dimethylthiazol-2-yl)-2,5-dphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays and analysis of oleanolic acid (OA)-treated multi-channel cell chips. The results of the MTT and LDH assays showed that OA treatment in the multi-channel cell chip of four cell lines enhanced chemoresistance of cells compared with that in the 2D culture. Furthermore, we demonstrated the feasibility of the application of our multi-channel cell chip in various analysis methods through Annexin V-fluorescein isothiocyanate/propidium iodide staining, which is not used for conventional cell chips. Taken together, the results demonstrated that the PMCCC may be used as a new 3D platform because it enables simultaneous drug screening in multiple cells by single point injection and allows analysis of various biological processes.

Assessing how in vitro assay types predict in vivo toxicology data.

In vivo animal bioassays are increasingly being supplemented with in vitro assays to serve as the new standard for chemical toxicity tests. Despite this shift, investigators face challenges related to increased reliance on in vitro data. The aim of this study was to deploy a streamlined method to assess the ability of in vitro data to predict similar results as in vivo data by correlating chemical toxicity rankings obtained using Benchmark Doses and Benchmark Dose Lower Limits (BMD(L)s) derived from in vivo and in vitro assays. In vitro and in vivo assay characteristics were assessed for their impact on the predictive ability of in vitro data. Minimum best-fit BMD(L)s were calculated for chemicals using Environmental Protection Agency's (EPA's) Benchmark Dose Software (BMDS). Forty-one chemicals met the inclusion criteria of this study. Relative chemical toxicity rankings were assessed through Kappa statistics, Pearson correlations, and/or Ordinary Least Squares (OLS) regressions. Results illustrated likely ability of in vitro data to predict similar results as short-term in vivo data. Further, rankings derived from in vitro cytotoxicity assays, unlike stress response assays, significantly correlated with rankings derived from short-term in vivo assays. These results support the use of in vitro data as a prioritization tool within toxicity testing.

A novel smartphone-based electrochemical cell sensor for evaluating the toxicity of heavy metal ions Cd2+, Hg2+, and Pb2+ in rice.

A novel smartphone-based electrochemical cell sensor was developed to evaluate the toxicity of heavy metal ions, such as cadmium (Cd2+), lead (Pb2+), and mercury (Hg2+) ions on Hep G2 cells. The cell sensor was fabricated with reduced graphene oxide (RGO)/molybdenum sulfide (MoS2) composites to greatly improve the biological adaptability and amplify the electrochemical signals. Differential pulse voltammetry (DPV) was employed to measure the electrical signals induced by the toxicity of heavy metal ions. The results showed that Cd2+, Hg2+, and Pb2+ significantly reduced the viability of Hep G2 cells in a dose-dependent manner. The IC50 values obtained by this method were 49.83, 36.94, and 733.90 µM, respectively. A synergistic effect was observed between Cd2+ and Pb2+ and between Hg2+ and Pb2+, and an antagonistic effect was observed between Cd2+ and Hg2+, and an antagonistic effect at low doses and an additive effect at high doses were found in the ternary mixtures of Cd2+, Hg2+, and Pb2+. These electrochemical results were confirmed via MTT assay, SEM and TEM observation, and flow cytometry. Therefore, this new electrochemical cell sensor provided a more convenient, sensitive, and flexible toxicity assessment strategy than traditional cytotoxicity assessment methods.

Development and validation of dual-cardiotoxicity evaluation method based on analysis of field potential and contractile force of human iPSC-derived cardiomyocytes / multielectrode assay platform.

A recent in vitro cardiovascular safety pharmacology test uses cardiomyocytes derived from human induced pluripotent stem cells (hiPSCs) to overcome the limitations of the classical test systems, such as species differences and local channel analysis. The Comprehensive in vitro Proarrhythmia Assay (CiPA) is a new proarrhythmia screening paradigm proposed by a CiPA steering expert group, which essentially requires iPSCs derived cardiomyocyte-based electrophysiological evaluation technology. Moreover, the measurement of the contractile force is also emerging as an important parameter to recapitulate non-proarrhythmic cardiotoxicity. Therefore, we constructed an multielectrode assay (MEA) evaluation method that can measure the electrophysiological changes with 6 reference drugs in hiPSC-derived cardiomyocytes. Subsequently, it was confirmed that the electrophysiological were changed in accordance with the mechanism of action of the drugs. Furthermore, based on the multi-probe impedance, we confirmed the decrease in contractile force due to treatment with drugs, and developed a platform to evaluate cardiotoxicity according to drugs along with field potential changes. Our excitation-contraction coupling cardiotoxicity assessment is considered to be more supportive in cardiac safety studies on pharmacologic sensitivity by complementing each assessment parameter.

Improving reliability and reducing costs of cardiotoxicity assessments using laser-induced cell poration on microelectrode arrays.

Drug-induced cardiotoxicity is a major barrier to drug development and a main cause of withdrawal of marketed drugs. Drugs can strongly alter the spontaneous functioning of the heart by interacting with the cardiac membrane ion channels. If these effects only surface during in vivo preclinical tests, clinical trials or worse after commercialization, the societal and economic burden will be significant and seriously hinder the efficient drug development process. Hence, cardiac safety pharmacology requires in vitro electrophysiological screening assays of all drug candidates to predict cardiotoxic effects before clinical trials. In the past 10 years, microelectrode array (MEA) technology began to be considered a valuable approach in pharmaceutical applications. However, an effective tool for high-throughput intracellular measurements, compatible with pharmaceutical standards, is not yet available. Here, we propose laser-induced optoacoustic poration combined with CMOS-MEA technology as a reliable and effective platform to detect cardiotoxicity. This approach enables the acquisition of high-quality action potential recordings from large numbers of cardiomyocytes within the same culture well, providing reliable data using single-well MEA devices and single cardiac syncytia per each drug. Thus, this technology could be applied in drug safety screening platforms reducing times and costs of cardiotoxicity assessments, while simultaneously improving the data reliability.

Fish early life stage toxicity prediction from acute daphnid toxicity and quantum chemistry.

One step towards reduced animal testing is the use of in silico screening methods to predict toxicity of chemicals, which requires high-quality data to develop models that are reliable and clearly interpretable. We compiled a large data set of fish early life stage no observed effect concentration endpoints (FELS NOEC) based on published data sources and internal studies, containing data for 338 molecules. Furthermore, we developed a new quantitative structure-activity-activity relationship (QSAAR) model to inform estimation of this endpoint using a combination of dimensionality reduction, regularization, and domain knowledge. In particular, we made use of a sparse partial least squares algorithm (sPLS) to select relevant variables from a huge number of molecular descriptors ranging from topological to quantum chemical properties. The final QSAAR model is of low complexity, consisting of 2 latent variables based on 8 molecular descriptors and experimental Daphnia magna acute data (EC50, 48 h). We provide a mechanistic interpretation of each model parameter. The model performs well, with a coefficient of determination r 2 of 0.723 on the training set (cross-validated q 2 = 0.686) and comparable predictivity on a test data set of chemically related molecules with experimental Daphnia magna data (r 2 test = 0.687, RMSE = 0.793 log units).

Induced pluripotent stem cell-derived vascular networks to screen nano-bio interactions.

The vascular bioactivity/safety of nanomaterials is typically evaluated by animal testing, which is of low throughput and does not account for biological differences between animals and humans such as ageing, metabolism and disease profiles. The development of personalized human in vitro platforms to evaluate the interaction of nanomaterials with the vascular system would be important for both therapeutic and regenerative medicine. A library of 30 nanoparticle (NP) formulations, in use in imaging, antimicrobial and pharmaceutical applications, was evaluated in a reporter zebrafish model of vasculogenesis and then tested in personalized humanized models composed of human-induced pluripotent stem cell (hiPSC)-derived endothelial cells (ECs) with "young" and "aged" phenotypes in 3 vascular network formats: 2D (in polystyrene dish), 3D (in Matrigel) and in a blood vessel on a chip. As a proof of concept, vascular toxicity was used as the main readout. The results show that the toxicity profile of NPs to hiPSC-ECs was dependent on the "age" of the endothelial cells and vascular network format. hiPSC-ECs were less susceptible to the cytotoxicity effect of NPs when cultured in flow than in static conditions, the protective effect being mediated, at least in part, by glycocalyx. Overall, the results presented here highlight the relevance of in vitro hiPSC-derived vascular systems to screen vascular nanomaterial interactions.

Characterizing the reproducibility in using a liver microphysiological system for assaying drug toxicity, metabolism, and accumulation.

Liver microphysiological systems (MPSs) are promising models for predicting hepatic drug effects. Yet, after a decade since their introduction, MPSs are not routinely used in drug development due to lack of criteria for ensuring reproducibility of results. We characterized the feasibility of a liver MPS to yield reproducible outcomes of experiments assaying drug toxicity, metabolism, and intracellular accumulation. The ability of the liver MPS to reproduce hepatotoxic effects was assessed using trovafloxacin, which increased lactate dehydrogenase (LDH) release and reduced cytochrome P450 3A4 (CYP3A4) activity. These observations were made in two test sites and with different batches of Kupffer cells. Upon culturing equivalent hepatocytes in the MPS, spheroids, and sandwich cultures, differences between culture formats were detected in CYP3A4 activity and albumin production. Cells in all culture formats exhibited different sensitivities to hepatotoxicant exposure. Hepatocytes in the MPS were more functionally stable than those of other culture platforms, as CYP3A4 activity and albumin secretion remained prominent for greater than 18 days in culture, whereas functional decline occurred earlier in spheroids (12 days) and sandwich cultures (7 days). The MPS was also demonstrated to be suitable for metabolism studies, where CYP3A4 activity, troglitazone metabolites, diclofenac clearance, and intracellular accumulation of chloroquine were quantified. To ensure reproducibility between studies with the MPS, the combined use of LDH and CYP3A4 assays were implemented as quality control metrics. Overall results indicated that the liver MPS can be used reproducibly in general drug evaluation applications. Study outcomes led to general considerations and recommendations for using liver MPSs. Study Highlights WHAT IS THE CURRENT KNOWLEDGE ON THE TOPIC? Microphysiological systems (MPSs) have been designed to recreate organ- or tissue-specific characteristics of extracellular microenvironments that enhance the physiological relevance of cells in culture. Liver MPSs enable long-lasting and stable culture of hepatic cells by culturing them in three-dimensions and exposing them to fluid flow. WHAT QUESTION DID THIS STUDY ADDRESS? What is the functional performance relative to other cell culture platforms and the reproducibility of a liver MPS for assessing drug development and evaluation questions, such as toxicity, metabolism, and pharmacokinetics? WHAT DOES THIS STUDY ADD TO OUR KNOWLEDGE? The liver MPS systematically detected the toxicity of trovafloxacin. When compared with spheroids and sandwich cultures, this system had a more stable function and different sensitivity to troglitazone, tamoxifen, and digoxin. Quantifying phase II metabolism of troglitazone and intracellular accumulation of chloroquine demonstrated the potential use of the liver MPS for studying drug metabolism and pharmacokinetics. Quality control criteria for assessing chip function were key for reliably using the liver MPS. HOW MIGHT THIS CHANGE CLINICAL PHARMACOLOGY OR TRANSLATIONAL SCIENCE? Due to its functional robustness and physiological relevance (3D culture, cells expose to fluid flow and co-culture of different cell types), the liver MPS can, in a reproducible manner: (i) detect inflammatory-induced drug toxicity, as demonstrated with trovafloxacin, (ii) detect the toxicity of other drugs, such as troglitazone, tamoxifen, and digoxin, with different effects than those detected in spheroids and sandwich cultures, (iii) enable studies of hepatic function that rely on prolonged cellular activity, and (iv) detect phase II metabolites and drug accumulation to potentially support the interpretation of clinical data. The integration of MPSs in drug development will be facilitated by careful evaluation of performance and reproducibility as performed in this study.

A Scaffold Free 3D Bioprinted Cartilage Model for In Vitro Toxicology.

Bioprinting has emerged as a promising method for precise spatiotemporal patterning of biological materials such as living cells, genetic materials, and proteins, which are sensitive to any other fabrication techniques. Bioprinting allows the generation of tissue constructs and models that closely mimic the anatomical and physiological attributes of a chosen tissue. In vitro toxicology assays can greatly benefit from bioprinting as drugs can be screened with higher efficiencies in a significantly reduced period. This protocol describes a method for fabricating bioprinted cartilage constructs which can be used for in vitro toxicology studies employing a scalable "tissue strand" bioprinting modality.

Evaluate the comparability of two automated liquid handling systems for clinical toxicology assays.

Automated liquid handling (ALH) platforms are increasingly implemented in clinical laboratories to improve analytical reproducibility and replace manual handling during sample analysis. In clinical toxicology laboratories, ALH platforms are primarily utilized to perform sample preparation and extraction prior to subsequent analysis by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). When performing analysis with complex human biological matrices, verifying the performance characteristics of ALH platforms is required to ensure the assays' accuracy and reproducibility. Here, we evaluated and compared the analytical performances of Perkin Elmer JANUS® and Tecan Fluent® ALH systems in parallel, based on their performance in two toxicology assays designed to identify and quantify various opiates, semisynthetic opiates, and their metabolites. The comparability of the instrument platforms was evaluated by comparing assay analytical measuring range, total analytical imprecisions, and patient samples measurement when the ALH platforms are incorporated as part of the clinical assay's workflow. We have shown that both ALH platforms meet quality and performance criteria suitable for clinical toxicology assays. Nevertheless, the two platforms exhibit biases when measuring unknown patient samples. Such variations in their analytical performances may cause discrepancies when comparing results obtained from two different ALH platforms. In conclusion, it is important to consider how variations in ALH platform performances can affect patient results interpretation when implementing them in clinical laboratories.

Whole body electronic cigarette exposure system for efficient evaluation of diverse inhalation conditions and products.

Objectives: To develop and test a new system for whole body exposure of small animals to support investigation of the biological effects of aerosol generated by electronic cigarette (e-cig) products under diverse inhalation conditions with improved control and monitoring of the e-cig vape exposure and nicotine delivered to the animal's systemic circulation. Methods: A computer-controlled design, with built-in sensors for real time monitoring of O2, CO2, relative humidity, and temperature within the exposure chambers and port for measuring total particulate matter (TPM) was developed, constructed and tested. This design accommodates a variety of commercial vaping devices, offers software flexibility to adjust exposure protocols to mimic different users' puffing patterns, enables variable nicotine delivery to the animal's systemic circulation; minimizes travel time and alterations of aerosol quality or quantity by delivering aerosol directly to the exposure chamber, offers local or remote operation of up to six distinct exposure chambers from a single control unit, and can simultaneously test different exposure conditions or products in diverse animal groups, which reduces inter-run variability, saves time, and increases productivity. Results: The time course pattern of TPM concentration during different phases of the exposure cycle was measured. With increased puffing duration or number of exposure cycles, higher TPM exposure and plasma cotinine levels were observed with plasma cotinine levels in the range reported in light or heavy smokers. Conclusion: Overall, this novel, versatile, and durable exposure system facilitates high-throughput evaluation of the relative safety and potential toxicity of a variety of e-cig devices and liquids.

Metallic nanoparticle production and exposure/deposition system for toxicological research applications using zebrafish.

Metallic nanoparticles (NPs) have been accepted for various applications ranging from cosmetics to medicine. However, no method has been established in the scientific community that is capable of analyzing various metals, sizes, and levels of exposures without the concern of background chemical contaminations. We present here a system utilizing soft-landing ion mobility (SLIM) exposures of laser ablated metallic clusters capable of operating pressures of reduced vacuum (1 Torr) up to ambient (760 Torr) in the presence of a buffer gas. Clusters experience kinetic energies of less than 1 eV upon exiting the SLIM, allowing for the exposure of NPs to take place in a passive manner. While there is no mass-selection of cluster sizes in this work, it does show for the first time the creation and soft-landing of nanoclusters at ambient pressures. Factors such as area coverage and percentage distribution were studied, as well as the different effects that varying surfaces may cause in the agglomeration of the clusters. Furthermore, the system was successfully used to study the effects of silver nanoparticle exposure and determine the specific organs the NPs accumulate in using zebrafish (Danio rerio) as a model organism. This method provides a novel way to synthesize NPs and expose biological organisms for various toxicological analysis.

Organic Electrochemical Transistors for Real-Time Monitoring of In Vitro Silver Nanoparticle Toxicity.

Nanomaterials are being widely used in medical applications and consumer products such as cosmetics, fabrics, and food packaging, although their impact on health and the environment is yet to be understood. Strategies enabling reliable and reproducible safety assessment of nanomaterials are needed because predicting their toxic effects is challenging as there is no simple correlation between their properties and the interaction with living systems. Here, the real-time monitoring of toxic effects induced by nanoparticles on cells using organic electrochemical transistors (OECTs) is reported. Noteworthy, OECTs are able to assess the coating-dependent toxicity of nanoparticles on both barrier and non-barrier tissue cells and, moreover, to monitor the cell health status as a function of exposure time, allowing useful insight on the interaction processes between nanomaterials and cells. These results demonstrate that OECTs are effective devices for real-time cell monitoring and in vitro assessment of nanomaterial toxicity.

In quest of effect directed analysis in the smart laboratory: Automated system for flow-through evaluation of membranotropic effects of emerging contaminants.

The rate-determining step of the human exposome workflow is the acquisition of physiologically relevant data (e.g., effect directed analysis), which can be performed retrospectively or with ad hoc experiments. In this contribution, an automated system is proposed for evaluating potential interaction mechanisms of xenobiotics across cell membranes, the so-called membranotropic effects, using liposomes as a mimicry of biological membranes, and fluorescent membrane probes. The smart fluidic method features real-time acquisition of fluorescence readouts, data processing and feedback in a fully unsupervised mode. As a proof of concept applicability, the behavior of newly synthesized cholesterol-laden biomimetic liposomes, and the in-vitro potential toxicant action of bisphenol A and diclofenac as model of emerging contaminants on cell membrane surrogates were investigated in a flow-through format. Unattended operation resulted in excellent intermediate precision (<1.5%) and unveiled that diclofenac affected the liposomal bilayer order very slightly, regardless of the cholesterol concentration, because it accumulates at a superficial level, while the membranotropic effect of bisphenol A was more pronounced at low concentration levels of cholesterol because at increased levels, the membrane reduces its permeability.

A non-contact impedimetric biosensing system for classification of toxins associated with cytotoxicity testing.

We report on a novel impedance spectroscopy measurement and data analysis technique for cytotoxicity testing. The technique combines non-contact measurement with real-time impedance data analysis based on the toxin dose dependency of the outputs, making it suitable for high throughput screening. A multi-electrode array was designed and fabricated such that a standard well plate could be positioned above the electrodes, negating the requirement for bespoke culture wells with integrated electrodes. For cytotoxicity testing, endothelial cells, type ECV304, within the wells were exposed to various concentrations of 3 toxins, dimethyl sulphoxide, cadmium chloride and saponin, which exhibit different modes of action on cells. Impedance spectra were recorded every 30 min over a 24 h period. From the spectra 'toxin maps' were produced which presented the correlation between impedance output and dose of toxin versus frequency and time. The results demonstrated characteristic toxin maps for each toxin and significantly differences between the three toxins studied. Using complementary measurement methods, we showed that these differences in toxin maps related to morphological and physiological changes in the cells due to the differing mode of action of each toxin.