TY - JOUR AB - This manuscript discusses the challenges of applying New Approach Methodologies (NAMs) for safe by design and regulatory risk assessment of advanced nanomaterials (AdNMs). The authors propose a framework for Next Generation Risk Assessment of AdNMs involving NAMs that is aligned to the conventional risk assessment paradigm. This framework is exposure-driven, endpoint-specific, makes best use of pre-existing information, and can be implemented in tiers of increasing specificity and complexity of the adopted NAMs. The tiered structure of the approach, which effectively combines the use of existing data with targeted testing will allow safety to be assessed cost-effectively and as far as possible with an even more limited use of vertebrates. The regulatory readiness of state-of-the-art emerging NAMs is assessed in terms of Transparency, Reliability, Accessibility, Applicability, Relevance and Completeness, and their relevance for AdNMs are discussed in relation to each step of the risk assessment paradigm along with providing perspectives for future developments in the respective scientific and regulatory areas. AU - Hristozov, D.* AU - Badetti, E.* AU - Bigini, P.* AU - Brunelli, A.* AU - Dekkers, S.* AU - Diomede, L.* AU - Doak, S.H.* AU - Fransman, W.* AU - Gajewicz-Skretna, A.* AU - Giubilato, E.* AU - Gómez-Cuadrado, L.* AU - Grafström, R.C.* AU - Gutleb, A.C.* AU - Halappanavar, S.* AU - Hischier, R.* AU - Hunt, N.* AU - Katsumiti, A.* AU - Kermanizadeh, A.* AU - Marcomini, A.* AU - Moschini, E.* AU - Oomen, A.G.* AU - Pizzol, L.* AU - Rumbo, C.* AU - Schmid, O. AU - Shandilya, N.* AU - Stone, V.* AU - Stoycheva, S.* AU - Stöger, T. AU - Merino, B.S.* AU - Tran, L.* AU - Tsiliki, G.* AU - Vogel, U.B.* AU - Wohlleben, W.* AU - Zabeo, A.* C1 - 71312 C2 - 56068 CY - Radarweg 29, 1043 Nx Amsterdam, Netherlands TI - Next generation risk assessment approaches for advanced nanomaterials: current status and future perspectives. JO - NanoImpact VL - 35 PB - Elsevier PY - 2024 SN - 2452-0748 ER - TY - JOUR AB - BACKGROUND: The establishment of reliable and robust in vitro models for hazard assessment, a prerequisite for moving away from animal testing, requires the evaluation of model transferability and reproducibility. Lung models that can be exposed via the air, by means of an air-liquid interface (ALI) are promising in vitro models for evaluating the safety of nanomaterials (NMs) after inhalation exposure. We performed an inter-laboratory comparison study to evaluate the transferability and reproducibility of a lung model consisting of the human bronchial cell line Calu-3 as a monoculture and, to increase the physiologic relevance of the model, also as a co-culture with macrophages (either derived from the THP-1 monocyte cell line or from human blood monocytes). The lung model was exposed to NMs using the VITROCELL® Cloud12 system at physiologically relevant dose levels. RESULTS: Overall, the results of the 7 participating laboratories are quite similar. After exposing Calu-3 alone and Calu-3 co-cultures with macrophages, no effects of lipopolysaccharide (LPS), quartz (DQ12) or titanium dioxide (TiO2) NM-105 particles on the cell viability and barrier integrity were detected. LPS exposure induced moderate cytokine release in the Calu-3 monoculture, albeit not statistically significant in most labs. In the co-culture models, most laboratories showed that LPS can significantly induce cytokine release (IL-6, IL-8 and TNF-α). The exposure to quartz and TiO2 particles did not induce a statistically significant increase in cytokine release in both cell models probably due to our relatively low deposited doses, which were inspired by in vivo dose levels. The intra- and inter-laboratory comparison study indicated acceptable interlaboratory variation for cell viability/toxicity (WST-1, LDH) and transepithelial electrical resistance, and relatively high inter-laboratory variation for cytokine production. CONCLUSION: The transferability and reproducibility of a lung co-culture model and its exposure to aerosolized particles at the ALI were evaluated and recommendations were provided for performing inter-laboratory comparison studies. Although the results are promising, optimizations of the lung model (including more sensitive read-outs) and/or selection of higher deposited doses are needed to enhance its predictive value before it may be taken further towards a possible OECD guideline. AU - Braakhuis, H.M.* AU - Gremmer, E.R.* AU - Bannuscher, A.* AU - Drasler, B.* AU - Keshavan, S.* AU - Rothen-Rutishauser, B.* AU - Birk, B.* AU - Verlohner, A.* AU - Landsiedel, R.* AU - Meldrum, K.* AU - Doak, S.H.* AU - Clift, M.J.D.* AU - Erdem, J.S.* AU - Foss, O.A.H.* AU - Zienolddiny-Narui, S.* AU - Serchi, T.* AU - Moschini, E.* AU - Weber, P.* AU - Burla, S.* AU - Kumar, P. AU - Schmid, O. AU - Zwart, E.* AU - Vermeulen, J.P.* AU - Vandebriel, R.J.* C1 - 67823 C2 - 54301 CY - Radarweg 29, 1043 Nx Amsterdam, Netherlands TI - Transferability and reproducibility of exposed air-liquid interface co-culture lung models. JO - NanoImpact VL - 31 PB - Elsevier PY - 2023 SN - 2452-0748 ER - TY - JOUR AB - Air-liquid interface (ALI) lung cell models cultured on permeable transwell inserts are increasingly used for respiratory hazard assessment requiring controlled aerosolization and deposition of any material on ALI cells. The approach presented herein aimed to assess the transwell insert-delivered dose of aerosolized materials using the VITROCELL® Cloud12 system, a commercially available aerosol-cell exposure system. An inter-laboratory comparison study was conducted with seven European partners having different levels of experience with the VITROCELL® Cloud12. A standard operating procedure (SOP) was developed and applied by all partners for aerosolized delivery of materials, i.e., a water-soluble molecular substance (fluorescence-spiked salt) and two poorly soluble particles, crystalline silica quartz (DQ12) and titanium dioxide nanoparticles (TiO2 NM-105). The material dose delivered to transwell inserts was quantified with spectrofluorometry (fluorescein) and with the quartz crystal microbalance (QCM) integrated in the VITROCELL® Cloud12 system. The shape and agglomeration state of the deposited particles were confirmed with transmission electron microscopy (TEM). Inter-laboratory comparison of the device-specific performance was conducted in two steps, first for molecular substances (fluorescein-spiked salt), and then for particles. Device- and/or handling-specific differences in aerosol deposition of VITROCELL® Cloud12 systems were characterized in terms of the so-called deposition factor (DF), which allows for prediction of the transwell insert-deposited particle dose from the particle concentration in the aerosolized suspension. Albeit DF varied between the different labs from 0.39 to 0.87 (mean (coefficient of variation (CV)): 0.64 (28%)), the QCM of each VITROCELL® Cloud 12 system accurately measured the respective transwell insert-deposited dose. Aerosolized delivery of DQ12 and TiO2 NM-105 particles showed good linearity (R2 > 0.95) between particle concentration of the aerosolized suspension and QCM-determined insert-delivered particle dose. The VITROCELL® Cloud 12 performance for DQ12 particles was identical to that for fluorescein-spiked salt, i.e., the ratio of measured and salt-predicted dose was 1.0 (29%). On the other hand, a ca. 2-fold reduced dose was observed for TiO2 NM-105 (0.54 (41%)), which was likely due to partial retention of TiO2 NM-105 agglomerates in the vibrating mesh nebulizer of the VITROCELL® Cloud12. This inter-laboratory comparison demonstrates that the QCM integrated in the VITROCELL® Cloud 12 is a reliable tool for dosimetry, which accounts for potential variations of the transwell insert-delivered dose due to device-, handling- and/or material-specific effects. With the detailed protocol presented herein, all seven partner laboratories were able to demonstrate dose-controlled aerosolization of material suspensions using the VITROCELL® Cloud12 exposure system at dose levels relevant for observing in vitro hazard responses. This is an important step towards regulatory approved implementation of ALI lung cell cultures for in vitro hazard assessment of aerosolized materials. AU - Bannuscher, A.* AU - Schmid, O. AU - Drasler, B.* AU - Rohrbasser, A.* AU - Braakhuis, H.M.* AU - Meldrum, K.* AU - Zwart, E.P.* AU - Gremmer, E.R.* AU - Birk, B.* AU - Rissel, M.* AU - Moschini, E.* AU - Evans, S.J.* AU - Kumar, P. AU - Orak, S. AU - Doryab, A. AU - Erdem, J.S.* AU - Serchi, T.* AU - Vandebriel, R.J.* AU - Cassee, F.R.* AU - Doak, S.H.* AU - Petri-Fink, A.* AU - Zienolddiny, S.* AU - Clift, M.J.D.* AU - Rothen-Rutishauser, B.* C1 - 66869 C2 - 53337 CY - Radarweg 29, 1043 Nx Amsterdam, Netherlands TI - An inter-laboratory effort to harmonize the cell-delivered in vitro dose of aerosolized materials. JO - NanoImpact VL - 28 PB - Elsevier PY - 2022 SN - 2452-0748 ER - TY - JOUR AB - The widespread integration of engineered nanomaterials into consumer and industrial products creates new challenges and requires innovative approaches in terms of design, testing, reliability, and safety of nanotechnology. The aim of this review article is to give an overview of different product groups in which nanomaterials are present and outline their safety aspects for consumers. Here, release of nanomaterials and related analytical challenges and solutions as well as toxicological considerations, such as dose-metrics, are discussed. Additionally, the utilization of engineered nanomaterials as pharmaceuticals or nutraceuticals to deliver and release cargo molecules is covered. Furthermore, critical pathways for human exposure to nanomaterials, namely inhalation and ingestion, are discussed in the context of risk assessment. Analysis of NMs in food, innovative medicine or food contact materials is discussed. Specific focus is on the presence and release of nanomaterials, including whether nanomaterials can migrate from polymer nanocomposites used in food contact materials. With regard to the toxicology and toxicokinetics of nanomaterials, aspects of dose metrics of inhalation toxicity as well as ingestion toxicology and comparison between in vitro and in vivo conclusions are considered. The definition of dose descriptors to be applied in toxicological testing is emphasized. In relation to potential exposure from different products, opportunities arising from the use of advanced analytical techniques in more unique scenarios such as release of nanomaterials from medical devices such as orthopedic implants are addressed. Alongside higher product performance and complexity, further challenges regarding material characterization and safety, as well as acceptance by the general public are expected. AU - Tschiche, H.R.* AU - Bierkandt, F.S.* AU - Creutzenberg, O.* AU - Fessard, V.* AU - Franz, R.* AU - Greiner, R.* AU - Gruber-Traub, C.* AU - Haas, K.H.* AU - Haase, A.* AU - Hartwig, A.* AU - Hesse, B.D.* AU - Hund-Rinke, K.* AU - Iden, P.* AU - Kromer, C.* AU - Löschner, K.* AU - Mutz, D.* AU - Rakow, A.* AU - Rasmussen, K.* AU - Rauscher, H.* AU - Richter, H.* AU - Schoon, J.* AU - Schmid, O. AU - Som, C.* AU - Spindler, L.M.* AU - Tovar, G.E.M.* AU - Westerhoff, P.* AU - Wohlleben, W.* AU - Luch, A.* AU - Laux, P.* C1 - 65999 C2 - 53032 TI - Analytical and toxicological aspects of nanomaterials in different product groups: Challenges and opportunities. JO - NanoImpact VL - 28 PY - 2022 SN - 2452-0748 ER - TY - JOUR AB - Global transcriptomic responses in lungs of mice exposed for 24 h to individual multi-walled carbon nanotubes (MWCNTs, ten different types), nano titanium dioxide (nano TiO 2, nine different types) and one type of carbon black (CB) nanomaterials (NMs) were investigated using toxicogenomics tools to determine if gene or pathway dose-response modelling can be used to rank the potential of NMs to induce in vivo acute lung inflammation. In each study, female adult C57BL/6 mouse lungs were intratracheally exposed once with 18, 54 or 162 μg/mouse doses of individual NMs and control mice were exposed to vehicle only. A high dose of 486 μg/mouse was used in only one study that involved nanoTiO 2 . The pathway perturbations associated with NM features and the underlying toxicity mechanisms were identified using bioinformatics tools. Bench Mark Dose (BMD) response analysis was conducted to derive transcriptional BMD estimates for each differentially expressed gene and the associated pathways in NM-treated lungs compared to vehicle-treated controls. The resulting BMDs were used to rank the potency of different NMs to induce perturbations in pathways that mark the occurrence of acute lung inflammation in mice. The transcriptional BMDs were compared with the BMDs of an apical endpoint derived for the lung neutrophil influx in bronchoalveolar lavage fluid from the same mice, a commonly measured pro-inflammatory endpoint in vivo. The results showed that similar gene and pathway responses were induced by the NM variants investigated. Among the MWCNT types, NM-401 and Mitsui-7, the two long and rigid fibres exhibited the most potency and, the short and tangled NRCWE-026 exhibited the least potency compared to the other nine varieties. Although mixed rutile/anatase nano TiO 2 showed higher potency compared to other nano TiO 2 , owing to the large confidence intervals, a clear distinction in potency could not be made. CB behaved similarly to the less potent MWCNTs. The transcriptomic BMDs were largely comparable to the neutrophil influx BMDs, with mass-based BMDs showing better correlation for all approaches. The mass-based BMDs were more conservative compared to the surface area based BMDs. Although the NMs investigated did not show large differences in their responses that would allow a clear ranking, the concept described demonstrates that quantitative pathway responses can be used to rank the potency of NMs to induce in vivo lung inflammation. AU - Halappanavar, S.* AU - Rahman, L.* AU - Nikota, J.* AU - Poulsen, S.S.* AU - Ding, Y. AU - Jackson, P.* AU - Wallin, H.* AU - Schmid, O. AU - Vogel, U.* AU - Williams, A.* C1 - 56020 C2 - 46737 TI - Ranking of nanomaterial potency to induce pathway perturbations associated with lung responses. JO - NanoImpact VL - 14 PY - 2019 SN - 2452-0748 ER - TY - JOUR AB - Nanoparticle in vitro toxicity studies often report contradictory results with one main reason being insufficient material characterization. In particular the characterization of nanoparticles in biological media remains challenging. Our aim was to provide robust protocols for two of the most commonly applied techniques for particle sizing, i.e. dynamic light scattering (DLS) and differential centrifugal sedimentation (DCS) that should be readily applicable also for users not specialized in nanoparticle physico-chemical characterization. A large number of participants (40, although not all participated in all rounds) were recruited for a series of inter-laboratory comparison (ILC) studies covering many different instrument types, commercial and custom-built, as another possible source of variation. ILCs were organized in a consecutive manner starting with dispersions in water employing well-characterized near-spherical silica nanoparticles (nominal 19 nm and 100 nm diameter) and two types of functionalized spherical polystyrene nanoparticles (nominal 50 nm diameter). At first each laboratory used their in-house established procedures. In particular for the 19 nm silica particles, the reproducibility of the methods was unacceptably high (reported results were between 10 nm and 50 nm). When comparing the results of the first ILC round it was observed that the DCS methods performed significantly worse than the DLS methods, thus emphasizing the need for standard operating procedures (SOPs). SOPs have been developed by four expert laboratories but were tested for robustness by a larger number of independent users in a second ILC (11 for DLS and 4 for DCS). In a similar approach another SOP for complex biological fluids, i.e. cell culture medium containing serum was developed, again confirmed via an ILC with 8 participating laboratories. Our study confirms that well-established and fit-for-purpose SOPs are indispensable for obtaining reliable and comparable particle size data. Our results also show that these SOPs must be optimized with respect to the intended measurement system (e.g. particle size technique, type of dispersant) and that they must be sufficiently detailed (e.g. avoiding ambiguity regarding measurand definition, etc.). SOPs may be developed by a small number of expert laboratories but for their widespread applicability they need to be verified by a larger number of laboratories. AU - Langevin, D.* AU - Lozano, O.* AU - Salvati, A.* AU - Kestens, V.* AU - Monopoli, M.* AU - Raspaud, E.* AU - Mariot, S.* AU - Salonen, A.* AU - Thomas, S.* AU - Driessen, M.* AU - Haase, A.* AU - Nelissen, I.* AU - Smisdom, N.* AU - Pompa, P.P.* AU - Maiorano, G.* AU - Puntes, V.* AU - Puchowicz, D.* AU - Stępnik, M.* AU - Suárez, G.* AU - Riediker, M.* AU - Benetti, F.* AU - Mičetić, I.* AU - Venturini, M.* AU - Kreyling, W.G. AU - van der Zande, M.* AU - Bouwmeester, H.* AU - Milani, S.* AU - Rädler, J.O.* AU - Mülhopt, S.* AU - Lynch, I.* AU - Dawson, K.A.* C1 - 52679 C2 - 44098 SP - 97-107 TI - Inter-laboratory comparison of nanoparticle size measurements using dynamic light scattering and differential centrifugal sedimentation. JO - NanoImpact VL - 10 PY - 2018 SN - 2452-0748 ER -