MatTek scientists attended and presented posters at WC11, the virtual 11th World Congress on Animal Alternatives. See what they’ve been working on and download copies of their posters below.
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Poster Presentations:
Abstract #: 775
SUBACUTE 28-DAY RESPIRATORY TOXICITY ASSAY USING AN IN VITRO HUMAN AIRWAY MODEL
Markus J, Jackson GR, Debatis M, Klausner M, Hayden P, Letasiova S, Landry T, Ayehunie S
MatTek, Ashland, MA, USA and MatTek, Bratislava, Slovakia
Corresponding author’s e-mail: jmarkus@mattek.com
Determining the subacute (28-day) respiratory toxicity potential is an important component of establishing safety profile of chemicals and consumer products. Here we describe our efforts to develop an alternative, non-animal method for determining subacute respiratory toxicity using the EpiAirway™ in vitro human airway model. The first stage consisted of determination of acute toxicity. The EpiAirway tissues were exposed for three hours to four concentrations of test chemicals via apical application using either
aqueous or corn oil vehicles. After rinsing, the tissues were incubated for additional 21 hours in standard cultivation conditions. Barrier integrity (determined by measuring transepithelial electrical resistance (TEER)) and viability (MTT assay) were determined
and an IC75 concentration (concentration required to reduce the endpoint value to 75% of vehicle exposed controls) was determined. Based on the determined acute IC75 value, EpiAirway tissues were exposed to additional serial dilutions of the test chemicals, using
the IC75 as the baseline dose. Every Monday, Wednesday, and Friday the tissues were apically exposed for 3 hours and subsequently washed. TEER was measured before each application. Experiments continued for at least 30 days to determine no-observed-adverse-effect level (NOAEL) doses. Rank ordering of NOAEL levels obtained for 8 chemicals was as follows: formaldehyde << butyl amine < oxalic acid << vinyl acetate < morpholine < methyl methacrylate << dimethylacetamide < ethanol. These results indicate that in vitro airway tissue models using TEER as a convenient non-destructive endpoint are a promising alternative to animal tests for the assessment of subacute 28-day respiratory toxicity and NOAELs. With further in vivo correlation and validation, this test may be a useful non-animal alternative for determining safe human subacute exposure levels for inhaled chemicals.
Abstract #: 799
IN VITRO SKIN IRRITATION PROTOCOL FOR THE MEDICAL DEVICES EXTRACTS USING EPIDERM MODEL
Silvia Letasiova1, Jan Markus1, Bridget Breyfogle2, Michael A. Bachelor2, Helena Kandarova3
1 MatTek, Bratislava, Slovakia
2 MatTek, Ashland, MA, USA
3 Centre of Experimental Medicine SAS, Slovak Academy of Sciences
Corresponding author’s e-mail: sletasiova@mattek.com
Safety evaluation of medical devices is a complex process and evaluation of skin irritation potential is an indispensable part of this process. The rabbit skin irritation test developed by Draize has been successfully replaced by the reconstructed human epidermis protocol (RhE) (OECD TG 439). However, this protocol is optimized for neat chemicals and medical device (MD) extracts are dilute solutions with low irritation potential. To reflect the requirements of the ISO 10993 directive, an optimized protocol using known irritants and spiked polymers was developed in 2013 (1). After successful transfer and standardization, a validation scheme was prepared. All 17 laboratories trained in the protocol worldwide produced results with almost 100% agreement with predictions for selected references (2). In the follow-up approach, several medical devices benchmark materials (5 irritants and 2 vehicles) were evaluated in the controlled human patch testing (4 h and 18 h) and in EpiDerm in vitro skin irritation protocol. Results were then compared to existing rabbit skin irritation test data. Based on the preliminary studies an international round-robin validation study was conducted in 2016 to confirm the ability of the RhE models to correctly predict the intracutaneous irritation of extracts from medical devices (4 irritants and 3 non-irritant materials). Blinded polymer samples were extracted with sesame oil and saline according to ISO 10993-12 (3). The protocol employing EpiDerm tissues was able to correctly predict virtually all of the irritant polymers in the saline, sesame oil as well as in both solvent extracts. Our results confirmed the ability of in vitro approach using RhE tissue models to detect the presence of skin irritants at low concentrations in dilute medical device polymer extracts (3). The use of the reconstructed tissue models, as replacements for the rabbit intra-cutaneous test, is currently being implemented into the ISO 10993 standards used to evaluate medical device biocompatibility.
Keywords: EpiDerm; in vitro skin irritation protocol; medical devices extracts; validation; ISO10993:23
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Abstract #: 1104
ALT4EI: DETERMINATION OF EYE IRRITATING POTENTIAL OF 59 CHEMICALS USING EPIOCULAR™ TIME-TO-TOXICITY NEAT AND DILUTION PROTOCOLS
Lenka Hudecova1, Silvia Letasiova1, Helena Kandarova2, Els Adriaens3, Sandra Verstraelen4, An R. Van Rompay4
1 MatTek, Bratislava, Slovakia
2 Centre of Experimental Medicine SAS, Slovak Academy of Sciences, Dubravska cesta 9,
Bratislava, Slovakia
3 Adriaens Consulting BVBA, Bellemdorpweg 95, 9981 Aalter, Belgium
4 VITO NV, UNI Health, Boeretang 200, 2400 Mol, Belgium
Corresponding author’s e-mail: sletasiova@mattek.com
Determination of acute eye irritation potential is part of international regulatory requirements for the testing of chemicals. The objective of ALT4EI (ALTernatives for Eye Irritation) project was to confirm the testing strategy developed in CON4EI (CONsortium for in vitro Eye Irritation testing strategy)project. These projects focused on the development of tiered testing strategies for eye irritation assessment for all drivers of classification and evaluation of whether the test methods can discriminate chemicals not requiring classification for serious eye damage/eye irritancy (No Category) from chemicals requiring classification and labeling for Category 1 (Cat 1) and Category 2 (Cat 2). In the CON4EI project, a new testing strategy for the EpiOcular time-to-toxicity was developed, the sensitivity for predicting GHS Cat 1 and GHS Cat 2 chemicals was 73% and 64%, respectively, and the very high specificity of 97% was maintained. None of the Cat 1 chemicals was underpredicted as GHS No Category. Plus the goal of the ALT4EI project was to fill the remaining data gaps and strengthen the data set. A new set of 59 chemicals (41 liquids:(un)diluted, and 18 solids) was tested using the reconstructed human cornea-like epithelium, EpiOcular, in two EpiOcular time-to-toxicity tests (neat and dilution). The set of chemicals contained 32 chemicals not requiring classification (No Cat) and 27 chemicals requiring classification (16 Cat 2 and 11 Cat 1). The chemicals were tested blinded in two independent runs by MatTek. In this study, a testing strategy to achieve optimal prediction for all three classes developed in the CON4EI project (combining the most predictive time-points of both protocols and which tests liquids and solids separately) was used. Using the CON4EI testing strategy, we were able to identify correctly 63,6 % of the Cat 1 chemicals,56,6 % of the Cat 2, and 76,6 % of No Cat chemicals. Reproducibility between both runs was 88,7 %. The combination of the EpiOcular time-to-toxicity neat and dilution protocols seems to be promising in integrated testing strategy (ITS) for eye irritation assessment.
Keywords: ALT4EI; EpiOcular; ocular irritation assay; in vitro; testing strategy
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IN VITRO MODELING OF GASTROINTESTINAL EXPOSURE AND RESPONSE TO ENGINEERED NANOMATERIALS
Puskar Marek2, Markus Jan2, Llanos Pierre1, Klausner Mitchell1, Armento Alex1, Kaluzhny Yulia1, and Ayehunie Seyoum1
1 MatTek, Ashland MA, USA
2 MatTek, Bratislava, Slovakia
There is a growing number of applications for various engineered nanoparticles (ENP), however, their impact on human health and various organs is poorly understood. In this study we utilized a 3D reconstructed human intestinal microtissues, EpiIntestinal tissue model, to develop an in vitro system for assessment of toxicological profiles of ingested nanomaterials. The tissues were exposed to various concentrations of three types of nanoparticles: copper (II) oxide (CuO) (50 nm in size), zinc oxide (ZnO, 35-45 nm in size), and titanium oxide (TiO2, 40 nm in size). Sonicated nanoparticles were resuspended in 40 μl of buffer and applied apically onto the tissues for 4 hours. Following application, the tissues were washed and incubated for additional 24 hours in standard medium. Subsequently, the barrier integrity (TEER) and viability (MTT) were determined for each tissue. In addition, medium was collected to determine levels of selected pro-inflammatory cytokines released following the nanoparticle exposure. Using IC15 (concentration of nanoparticles that reduces barrier function or tissue viability by 15%) as a cut-off, we observed a dose-response reduction of barrier integrity and tissue viability for CuO and ZnO. On the other hand, the titanium oxide did not induce toxic effects even at the highest concentration. Similar observation was detected through the cytokine production – we have seen a dose-dependent increase of interleukin 8 (IL-8) in tissues exposed to CuO and ZnO. Overall, we have shown that the TEER measurement is a very reproducible and more sensitive endpoint than MTT. In conclusion, these results suggest that reconstructed small intestine tissues might become a sensitive tool not only for determining the toxicity of ingested nanoparticles but also for further studying the interactions of nanoparticles with the host gastrointestinal system.
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