The in vitro skin sensitization test; human cell line activation test (h-CLAT) using THP-1 cells

Sakaguchi, Hitoshi; Ashikaga, Takao; Kosaka, Nanae; Sono, Sakiko; Nishiyama, Naohiro; Itagaki, Hiroshi

doi:10.1016/j.toxlet.2007.05.257

Cited by 13 publications

(5 citation statements)

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“…After the early observation that CD86 is upregulated in dendritic cells upon treatment with sensitizers (Ashikaga et al 2002;Manome et al 1999), a large body of evidence has accumulated that this is a predictive marker in different cell lines and primary cells (Reuter et al 2011;Sakaguchi et al 2007). Next to CD86, also upregulation of CD54, CD80, CD83, among other markers, were evaluated, but overall, less consistent results were obtained (Azam et al 2006;Ryan et al 2004b), with best predictive value obtained for CD54 next to CD86 (Azam et al 2006).…”

Section: Expression Of Cell Surface Markersmentioning

confidence: 99%

Reporter cell lines for skin sensitization testing

Natsch

Emter

2015

Arch Toxicol

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Skin sensitization has been described as an adverse outcome pathway (AOP), comprising a number of molecular events leading to the final adverse effect. In a new paradigm of toxicology, attempts are made to collect information using single mechanistic tests addressing different targets along such an AOP and to then integrate this information to arrive at a final toxicological prediction. This proposal is strongly influenced by the availability of methods for high-throughput screening of cellular events. Reporter cell lines are a particularly useful tool in such screening paradigms, as they can deliver highly reproducible and easily measureable results, and they can be designed to quantify induction or suppression at the transcription level of very specific molecular targets within cells. The first cell-based assay for skin sensitization, which has recently received ECVAM and OECD endorsement, is the reporter cell assay KeratinoSens™, reflecting activation of the Nrf2 pathway, and other assays measuring the Nrf2 pathway are under development or validation. An alternative approach (THP-G8) was recently developed based on activation of the Interleukin-8 gene. Here, we review these assays, their role in the AOP, their mechanistic interrelationships, their use for hazard and risk assessment, and their application in integrated testing strategies. At the same time, this study reviews (1) other cellular markers for sensitizers, and the potential to develop new reporter gene assays providing additional, non-redundant information, and (2) it presents approaches and new experimental data on attempts to further improve the predictivity of the existing assay.

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Section: Expression Of Cell Surface Markersmentioning

confidence: 99%

Reporter cell lines for skin sensitization testing

Natsch

Emter

2015

Arch Toxicol

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“…Cosmetics Europe (The European Cosmetics Industry Trade Association) member companies have developed three in vitro tests which include the Direct Peptide Reactivity Assay (DPRA), 13 the myeloid U937 skin sensitization test (MUSST), 14 and the human cell line activation test (h-CLAT). 15 The Procter & Gamble, Wella-Cosmital Laboratories has also developed an in vitro test that is based on human peripheral blood monocyte-derived dendritic cells (MoDCs). 16 Recently, the European consortium, SENS-IT-IV, has developed an in vitro test that uses a KC cell line (NCTC 2544) and it is based on the selective induction of IL-18 in the KCs by contact allergens.…”

Section: Introductionmentioning

confidence: 99%

In vitro micro-physiological immune-competent model of the human skin

2016

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Skin allergy, in particular, allergic contact dermatitis and irritant contact dermatitis, are common occupational and environmental health problems affecting the quality of life of a significant proportion of the world population. Since all new ingredients to be incorporated into a product are potential skin allergens, it is essential that these ingredients be first tested for their allergenic potential. However, despite the considerable effort using animal models to understand the underlying mechanism of skin sensitization, to date, the molecular and cellular responses due to skin contact with sensitizers are still not fully understood. To replace animal testing and to improve the prediction of skin sensitization, significant attention has been directed to the use of reconstructed organotypic in vitro models of human skin. Here we describe a miniaturized immune competent in vitro model of human skin based on 3D co-culture of immortalized human keratinocytes (HaCaT) as a model of the epidermis barrier and human leukemic monocyte lymphoma cell line (U937) as a model of human dendritic cells. The biological model was fitted in a microfluidic-based cell culture system that provides a dynamic cellular environment that mimics the in vivo environment of skin. The dynamic perfusion of culture media significantly improved the tight junction formation as evidenced by measuring higher values of TEER compared to static culture. This setting also maintained the high viability of cells over extended periods of time up to 17 days. The perfusion-based culture also allows growth of the cells at the air-liquid interface by exposing the apical side of the cells to air while providing the cell nutrients through a basolateral fluidic compartment. The microsystem has been evaluated to investigate the effect of the chemical and physical (UV irradiation) stimulation on the skin barrier (i.e. the TJ integrity). Three-tiered culture differential stimulation allowed the investigation of the role of the keratinocyte layer as a protection barrier to chemical/biological hazards.

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“…Whereas the LLNA has officially been validated only for classification and labeling, and thus hazard prediction, it is also used by both academia and industry for potency prediction, because a significant correlation between the EC3 values and the in vivo sensitization capacity in humans was established . A number of in vitro and in chemico assays for detecting skin sensitizers has been proposed in recent years, and great progress has been made for hazard identification with several assays giving a predictive accuracy of 85%. − Additionally, for several assays, a certain correlation with potency was noted, , but a close prediction of the LLNA EC3 result currently is not possible based on global models. This is partly due to the fact that specific classes of compounds have higher or lower relative activities in certain in vitro assays and thus are over- or underpredicted, respectively, if the prediction is based on correlations derived from large databases of structurally very diverse molecules.…”

Section: Introductionmentioning

confidence: 99%

Structure–Activity Relationship between the in Vivo Skin Sensitizing Potency of Analogues of Phenyl Glycidyl Ether and the Induction of Nrf2-Dependent Luciferase Activity in the KeratinoSens in Vitro Assay

Delaine

Niklasson

Emter

et al. 2011

Chem. Res. Toxicol.

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Because of regulatory constraints and ethical considerations, research on alternatives to animal testing to predict the skin sensitization potential of novel chemicals has become a high priority. Ideally, these alternatives should not only predict the hazard of novel chemicals but also rate the potency of skin sensitizers. Currently, no alternative method gives reliable potency estimations for a wide range of chemicals in differing structural classes. Performing potency estimations within specific structural classes has thus been proposed. Detailed structure-activity studies for the in vivo sensitization capacity of a series of analogues of phenyl glycidyl ether (PGE) were recently published. These studies are part of an investigation regarding the allergenic activity of epoxy-resin monomers. Here we report data on the same chemicals in the KeratinoSens in vitro assay, which is based on a stable transgenic keratinocyte cell line with a luciferase gene under the control of an antioxidant response element. A strong correlation between the EC3 values in the local lymph node assay (LLNA) and both the luciferase-inducing concentrations and the cytotoxicity in the cell-based assay was established for six analogues of PGE. This correlation allowed the potency in the LLNA of two novel structurally closely related derivatives to be predicted by read-across with errors of 1.4- and 2.6-fold. However, the LLNA EC3 values of two structurally different bifunctional monomers were overpredicted on the basis of this data set, indicating that accurate potency estimation by read-across based on in vitro data might be restricted to a relatively narrow applicability domain.

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The in vitro skin sensitization test; human cell line activation test (h-CLAT) using THP-1 cells

Cited by 13 publications

References 0 publications

Reporter cell lines for skin sensitization testing

Reporter cell lines for skin sensitization testing

In vitro micro-physiological immune-competent model of the human skin

Structure–Activity Relationship between the in Vivo Skin Sensitizing Potency of Analogues of Phenyl Glycidyl Ether and the Induction of Nrf2-Dependent Luciferase Activity in the KeratinoSens in Vitro Assay

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