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  • 1.
    Andres, M I
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Forsby, A
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Walum, E
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Polygodial-induced noradrenaline release in human neuroblastoma SH-SY5Y cells.1997Inngår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 11, nr 5, s. 509-11Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Polygodial is a natural sesquiterpene which exhibits pronounced pungency and a powerful antifeedant activity. At low concentrations, which do not alter general cell membrane permeability, polygodial increases the intracellular concentration of free calcium ([Ca(2+)](i)). Sensory neurotransmission depends on noradrenaline (NA) release, and vesicular exocytosis, in turn, is dependent on an increase in [Ca(2+)](i). The nociceptive response induced by polygodial could therefore be directly linked to intracellular calcium levels. Consequently, the objective of this work was to investigate the effect of polygodial on NA release. The human neuroblastoma cell line SH-SY5Y was selected as an in vitro model for sensory neurones. Semiconfluent cells were preloaded with tritiated NA ([(3)H]NA). After 3 min exposure of polygodial to the cells, released and unreleased radioactivity were measured. Polygodial induced a significant [(3)H]NA release at concentrations between 0.1 and 0.5 mug/ml with a maximum effect at 0.2 mug/ml (40% increased release of [(3)H]NA as compared with unstimulated control cells). No polygodial-induced transmitter release was seen at 3.5 and 5 mug/ml. For comparison, carbachol (1 rim) increased [(3)H]NA release by 10% and the KCl-induced (100 mm) [(3)H]NA release increased by 8% as compared with unstimulated SH-SY5Y cells. In conclusion polygodial, at the concentrations 0.1-0.5 mug/ml (equal to 0.4-2 mum), induces NA release which is dependent on polygodial-induced increase in [Ca(2+)](i).

  • 2.
    Attoff, Kristina
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Kertika, Dimitra
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Lundqvist, Jessica
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Oredsson, S.
    Forsby, Anna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. Stockholm Univ, Dept Neurochem, S-10691 Stockholm, Sweden.
    Acrylamide affects proliferation and differentiation of the neural progenitor cell line C17.2 and the neuroblastoma cell line SH-SY5Y2016Inngår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 35, s. 100-111Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Acrylamide is a well-known neurotoxic compound and people get exposed to the compound by food consumption and environmental pollutants. Since acrylamide crosses the placenta barrier, the fetus is also being exposed resulting in a risk for developmental neurotoxicity. In this study, the neural progenitor cell line C17.2 and the neuroblastoma cell line SH-SY5Y were used to study proliferation and differentiation as alerting indicators for developmental neurotoxicity. For both cell lines, acrylamide reduced the number of viable cells by reducing proliferation and inducing cell death in undifferentiated cells. Acrylamide concentrations starting at 10 fM attenuated the differentiation process in SH-SY5Y cells by sustaining cell proliferation and neurite outgrowth was reduced at concentrations from 10 pM. Acrylamide significantly reduced the number of neurons starting at 1 mu M and altered the ratio between the different phenotypes in differentiating C17.2 cell cultures. Ten micromolar of acrylamide also reduced the expression of the neuronal and astrocyte biomarkers. Although the neurotoxic concentrations in the femtomolar range seem to be specific for the SH-SY5Y cell line, the fact that micromolar concentrations of acrylamide seem to attenuate the differentiation process in both cell lines raises the interest to further investigations on the possible developmental neurotoxicity of acrylamide.

  • 3. Dejongh, J
    et al.
    Forsby, A
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Houston, J B
    Beckman, M
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Combes, R
    Blaauboer, B J
    An Integrated Approach to the Prediction of Systemic Toxicity using Computer-based Biokinetic Models and Biological In vitro Test Methods: Overview of a Prevalidation Study Based on the ECITTS Project.1999Inngår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 13, nr 4-5, s. 549-54Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Chemical toxicity was estimated by integrating in vitro study results with physiologically-based biokinetic models for eight neurotoxic compounds (benzene, toluene, lindane, acrylamide, parathion/oxon, caffeine, diazepam and phenytoin). In vitro studies on general and specific neurotoxicity were performed and biotransformation and tissue-blood distribution studies were used in modelling the biokinetic behaviour of the compounds. Subsequently, neurotoxicity was estimated from the integrated in vitro and kinetic studies. These results were compared with in vivo data from the literature on minimal neurotoxicity for these compounds, such as lowest-observed-effect levels (LOELs). The discrepancy between estimated and experimental LOELs ranged from 2- to 10-fold. LOEL estimates for compounds with a relatively low toxicity were more accurate than for compounds with a relatively high toxicity. LOELs for the most active compounds could only be established after consideration of additional in vitro results from the literature. The present study has generated encouraging results on the risk assessment of chemicals from in vitro studies and computer simulations and has identified some key directions for future research.

  • 4.
    Folch, Jaume
    et al.
    Unitat de Bioquimica, Facultat de Medicina i Ciències de la Salut, Universitat Rovira i Virgili, Tarragona, Spain.
    Alvira, Daniel
    Unitat de Farmacologia i Farmacognòsia and Institut de Biomedicina (IBUB), Facultat de Farmàcia, Universitat de Barcelona, Nucli Universitari de Pedralbes, Barcelona, Spain.
    López-Querol, Marta
    Unitat de Farmacologia, Facultat de Medicina i Ciències de la Salut, Universitat Rovira i Virgili, Tarragona, Spain.
    Tajes, Marta
    Unitat de Farmacologia i Farmacognòsia and Institut de Biomedicina (IBUB), Facultat de Farmàcia, Universitat de Barcelona, Nucli Universitari de Pedralbes, Barcelona, Spain.
    Sureda, Francesc X
    Unitat de Farmacologia, Facultat de Medicina i Ciències de la Salut, Universitat Rovira i Virgili, Tarragona, Spain.
    Forsby, Anna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Rimbau, Víctor
    Unitat de Farmacologia i Farmacognòsia, Facultat de Farmàcia, Universitat de Barcelona, Nucli Universitari de Pedralbes, Barcelona, Spain.
    Camins, Antoni
    Unitat de Farmacologia i Farmacognòsia and Institut de Biomedicina (IBUB), Facultat de Farmàcia, Universitat de Barcelona, Nucli Universitari de Pedralbes, Barcelona, Spain.
    Pallàs, Mercè
    Unitat de Farmacologia i Farmacognòsia and Institut de Biomedicina (IBUB), Facultat de Farmàcia, Universitat de Barcelona, Nucli Universitari de Pedralbes, Barcelona, Spain.
    Evaluation of transcriptional activity of caspase-3 gene as a marker of acute neurotoxicity in rat cerebellar granular cells2010Inngår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 24, nr 2, s. 465-471Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Caspase-3 is a key protein involved in the classical apoptosis mechanism in neurons, as in many other cells types. In the present research, we describe the transcriptional activity of caspase-3 gene as a marker of acute toxicity in a primary culture model of rat cerebellar granule neurons (CGNs). CGNs were incubated for 16h in complete medium containing the chemicals at three concentrations (10, 100 μM and 1 mM). A total of 48 different compounds were tested. Gene transcriptional activity was determined by low-density array assays, and by single Taqman caspase-3 assays. Results from the PCR arrays showed that the caspase-3 gene was up-regulated when CGNs were exposed to neurotoxic chemicals. Significative correlations were found between the transcriptional activity of caspase-3 and the activity of some other genes related to apoptosis, cell-cycle and ROS detoxification. In our experiments, acute exposure of CGNs to well-documented pro-apoptotic xenobiotics modulated significantly caspase-3 gene expression, whereas chemicals not related to apoptosis did not modify caspase-3 gene expression. In conclusion, acute exposure of CGNs to neurotoxic compounds modulates the transcriptional activity of genes involved in the classical apoptotic pathway, oxidative stress and cell-cycle control. Transcriptional activity of caspase-3 correlates significantly with these changes and it could be a good indicator of acute neurotoxicity.

  • 5. Forreryd, Andy
    et al.
    Norinder, Ulf
    Stockholms universitet, Samhällsvetenskapliga fakulteten, Institutionen för data- och systemvetenskap. Karolinska Institutet, Sweden.
    Lindberg, Tim
    Lindstedt, Malin
    Predicting skin sensitizers with confidence - Using conformal prediction to determine applicability domain of GARD2018Inngår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 48, s. 179-187Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    GARD - Genomic Allergen Rapid Detection is a cell based alternative to animal testing for identification of skin sensitizers. The assay is based on a biomarker signature comprising 200 genes measured in an in vitro model of dendritic cells following chemical stimulations, and consistently reports predictive performances similar to 90% for classification of external test sets. Within the field of in vitro skin sensitization testing, definition of applicability domain is often neglected by test developers, and assays are often considered applicable across the entire chemical space. This study complements previous assessments of model performance with an estimate of confidence in individual classifications, as well as a statistically valid determination of the applicability domain for the GARD assay. Conformal prediction was implemented into current GARD protocols, and a large external test dataset (n = 70) was classified at a confidence level of 85%, to generate a valid model with a balanced accuracy of 88%, with none of the tested chemical reactivity domains identified as outside the applicability domain of the assay. In conclusion, results presented in this study complement previously reported predictive performances of GARD with a statistically valid assessment of uncertainty in each individual prediction, thus allowing for classification of skin sensitizers with confidence.

  • 6. Hamm, Jon
    et al.
    Sullivan, Kristie
    Clippinger, Amy J.
    Strickland, Judy
    Bell, Shannon
    Bhhatarai, Barun
    Blaauboer, Bas
    Casey, Warren
    Dorman, David
    Forsby, Anna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi. Swedish Toxicology Sciences Research Center (Swetox), Sweden.
    Garcia-Reyero, Natàlia
    Gehen, Sean
    Graepel, Rabea
    Hotchkiss, Jon
    Lowit, Anna
    Matheson, Joanna
    Reaves, Elissa
    Scarano, Louis
    Sprankle, Catherine
    Tunkel, Jay
    Wilson, Dan
    Xia, Menghang
    Zhu, Hao
    Allen, David
    Alternative approaches for identifying acute systemic toxicity: Moving from research to regulatory testing2017Inngår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 41, s. 245-259Artikkel, forskningsoversikt (Fagfellevurdert)
    Abstract [en]

    Acute systemic toxicity testing provides the basis for hazard labeling and risk management of chemicals. A number of international efforts have been directed at identifying non-animal alternatives for in vivo acute systemic toxicity tests. A September 2015 workshop, Alternative Approaches for Identifying Acute Systemic Toxicity: Moving from Research to Regulatory Testing, reviewed the state-of-the-science of non-animal alternatives for this testing and explored ways to facilitate implementation of alternatives. Workshop attendees included representatives from international regulatory agencies, academia, nongovernmental organizations, and industry. Resources identified as necessary for meaningful progress in implementing alternatives included compiling and making available high-quality reference data, training on use and interpretation of in vitro and in silico approaches, and global harmonization of testing requirements. Attendees particularly noted the need to characterize variability in reference data to evaluate new approaches. They also noted the importance of understanding the mechanisms of acute toxicity, which could be facilitated by the development of adverse outcome pathways. Workshop breakout groups explored different approaches to reducing or replacing animal use for acute toxicity testing, with each group crafting a roadmap and strategy to accomplish near-term progress. The workshop steering committee has organized efforts to implement the recommendations of the workshop participants.

  • 7.
    Kotova, Natalia
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Hebert, N.
    Härnwall, Eva-Lena
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Vare, Daniel
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Mazurier, C.
    Douay, L.
    Jenssen, Dag
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för molekylär biovetenskap, Wenner-Grens institut.
    Grawe, J.
    A novel micronucleus in vitro assay utilizing human hematopoietic stem cells2015Inngår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 29, nr 7, s. 1897-1905Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The induction of micronucleated reticulocytes in the bone marrow is a sensitive indicator of chromosomal damage. Therefore, the micronucleus assay in rodents is widely used in genotoxicity and carcinogenicity testing. A test system based on cultured human primary cells could potentially provide better prediction compared to animal tests, increasing patient safety while also implementing the 3Rs principle, i.e. replace, reduce and refine. Hereby, we describe the development of an in vitro micronucleus assay based on animal-free ex vivo culture of human red blood cells from hematopoietic stem cells. To validate the method, five clastogens with direct action, three clastogens requiring metabolic activation, four aneugenic and three non-genotoxic compounds have been tested. Also, different metabolic systems have been applied. Flow cytometry was used for detection and enumeration of micronuclei. Altogether, the results were in agreement with the published data and indicated that a sensitive and cost effective in vitro assay to assess genotoxicity with a potential to high-throughput screening has been developed.

  • 8.
    Lindegren, H.
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Mogren, H.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    EL Andaloussi-Lilja, J.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Lundqvist, J.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Forsby, A.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Anionic linear aliphatic surfactants activate TRPV1: a possible endpoint for estimation of detergent induced eye nociception?2009Inngår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 23, nr 8, s. 1472-1476Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The transient receptor potential vanilloid type 1 (TRPV1) has been reported as one of the key components in the pain pathway. Activation of the receptor causes a Ca2+ influx in sensory C-fibres with secondary effects leading to neurogenic inflammation in the surrounding tissue. We have earlier reported specific activation of TRPV1 by surfactant-containing hygiene products. We have continued this project by investigating activation of the TRPV1 by shampoo and soap ingredients in low concentrations measured as intracellular Ca2+ influxes in stably TRPV1-expressing neuroblastoma SH-SY5Y cells. As a TRPV1 specific control, the TRPV1 antagonist capsazepine was used. The response was quantified as the product induced Ca2+ influx during 2 min in relation to the maximum response induced by the TRPV1 agonist capsaicin. The results show that anionic alkyl linear surfactant ingredients such as sodium lauryl sulphate, sodium laureth sulphate, ammonium lauryl sulphate, sodium C12-15 pareth sulphate and N-lauroylsarcosine concentration-dependently induced Ca2+ influx that could be addressed to TRPV1. The cationic surfactants benzalkonium chloride and cetylpyridinium chloride induced a Ca2+ influx that was not TRPV1 mediated as well as the zwitterionic surfactant cocamidopropyl betaine, the non-linear anionic surfactant sodium deoxycholate and the non-ionic surfactant Triton-X. These results reveal a new mechanistic pathway for surfactant-induced nociception.

  • 9.
    Lundqvist, Jessica
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    EL Andaloussi-Lilja, Johanna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Svensson, Christina
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Gustafsson Dorfh, Helena
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Forsby, Anna
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Optimisation of culture conditions for differentiation of C17.2 neural stem cells to be used for in vitro toxicity tests2013Inngår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 27, nr 5, s. 1565-1569Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Here we present a multipotent neuronal progenitor cell line for toxicity testing as an alternative to primary cultures of mixed cell types from brain tissue. The v-myc immortalised C17.2 cell line, originally cloned from mouse cerebellar neural stem cells, were maintained as monolayer in cell culture dishes in DMEM supplemented with fetal calf serum, horse serum and antibiotics. Different media and exposure scenarios were used to induce differentiation. The optimal condition which generated mixed cultures of neurons and astrocytes included serum-free DMEM:F12 medium with N2 supplements, brain-derived neurotrophic factor and nerve growth factor. The medium was changed every 3rd or 4th day to fresh N2 medium with supplements. After 7days, the culture contained two different morphological cell types, assumed to be neurons and glia cells. The presence of astrocytes and neurons in the culture was confirmed by RT-PCR and Western blot analyses, indicating increased mRNA and protein levels of the specific biomarkers glial fibrillary acidic protein (GFAP) and βIII-tubulin, respectively. Concomitantly, the expression of the neural progenitor cell marker nestin was down-regulated.

  • 10.
    Nordin-Andersson, M
    et al.
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Forsby, A
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Heldring, N
    Stockholms universitet, Naturvetenskapliga fakulteten, Institutionen för neurokemi.
    Dejongh, J
    Kjellstrand, P
    Walum, E
    Neurite degeneration in differentiated human neuroblastoma cells.1998Inngår i: Toxicology in Vitro, ISSN 0887-2333, E-ISSN 1879-3177, Vol. 12, nr 5, s. 557-60Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    We have studied neurite degeneration in differentiated human neuroblastoma (SH-SY5Y) cells. The axonopathy-inducing potency in vitro of caffeine, diazepam, methylmercury chloride (MeHg), triethyltin chloride (TET) and acrylamide (ACR) was elucidated. After 72 hours of exposure the neurite degeneration was determined (by morphological quantification) as well as the total protein content (general cytotoxicity). The concentrations that caused 20% reduction of number of neurites (ND(20)) for ACR (250+/-36 mum) and TET (0.097+/-0.03 mum) was significantly lower, 63% and 35%, respectively (P</=0.005), as compared to corresponding inhibition of general cytotoxicity (IC(20)). The effects of TET on the neurites may be related to the disturbance in Ca(2+)-signalling, and thus a secondary event. The ND(20)s for caffeine and diazepam, which are compounds without a known neurite degenerative potency in vivo, were higher as compared with the IC(20). For MeHg which is an extremely cyto- and neurotoxic compound the ND(20) was not statistically different from the IC(20), indicating that degeneration of the neurites is not a primary effect. This study indicates that the SH-SY5Y-neurite degeneration model is useful for the identification of axonopathy-inducing substances.

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