Ruth Pendlington

Xenobiotic metabolism in human skin and 3D human skin reconstructs: A review

In this review, we discuss and compare studies of xenobiotic metabolism in both human skin and 3D human skin reconstructs. In comparison to the liver, the skin is a less studied organ in terms of characterising metabolic capability. While the skin forms the major protective barrier to environmental chemical exposure, it is also a potential target organ for adverse health effects. Occupational, accidental or intended-use exposure to toxic chemicals could result in acute or delayed injury to the skin (e.g. inflammation, allergy, cancer). Skin metabolism may play a role in the manifestation or amelioration of adverse effects via the topical route. Today, we have robust testing strategies to assess the potential for local skin toxicity of chemical exposure. Such methods (e.g. the local lymph node assay for assessing skin sensitisation; skin painting carcinogenicity studies) incorporate skin metabolism implicitly in the in vivo model system used. In light of recent European legislation (i.e. 7(th) Amendment to the Cosmetics Directive and Registration Evaluation and Authorisation of existing Chemicals (REACH)), non-animal approaches will be required to reduce and replace animal experiments for chemical risk assessment. It is expected that new models and approaches will need to account for skin metabolism explicitly, as the mechanisms of adverse effects in the skin are deconvoluted. 3D skin models have been proposed as a tool to use in new in vitro alternative approaches. In order to be able to use 3D skin models in this context, we need to understand their metabolic competency in relation to xenobiotic biotransformation and whether functional activity is representative of that seen in human skin.

Paraben Transport and Metabolism in the Biomimetic Artificial Membrane Permeability Assay (BAMPA) and 3-Day and 21-Day Caco-2 Cell Systems

Noncellular and cellular in vitro models for predicting intestinal absorption were used to investigate the transport and metabolism of parabens. The biomimetic artificial membrane permeability assay (BAMPA) membrane was constructed by impregnating a lipid solution on a hydrophobic filter. Caco-2 cells at passage numbers 65 to 80 were cultured in either the accelerated 3-day Biocoat™ system or the standard 21-day Transwell™ cell culture system. Paraben transport across the BAMPA system showed a parabolic relationship. The lowest log P (p-hydroxybenzoic acid) and highest log P compounds (heptyl and octyl parabens) had apparent permeabilities (Papp) less than 1.0 × 10-6 cm/s and Papp was maximal at approximately 8.5 × 10-6cm/s for the intermediate log P (ethylparaben) compound. With the Biocoat™, a similar parabolic relationship was found. In the 21-day Caco-2 cells, the parabens were metabolized by esterases at to p-hydroxybenzoic acid. In conclusion, the in vitro models added complementary insight into the absorption process, such as the transport route, intrinsic permeability, and extent of metabolism of the parabens. This study indicated that presystemic metabolism of orally ingested parabens to the p-hydroxybenzoic acid in the intestine may limit systemic exposure to alkyl-paraben esters in vivo.

Determining Epidermal Disposition Kinetics for Use in an Integrated Nonanimal Approach to Skin Sensitization Risk Assessment

Development of risk assessment methods for skin sensitization in the absence of toxicological data generated in animals represents a major scientific and technical challenge. The first step in human skin sensitization induction is the transport of sensitizer from the applied dose on the skin surface to the epidermis, where innate immune activation occurs. Building on the previous development of a time course in vitro human skin permeation assay, new kinetic data for 10 sensitizers and 2 nonsensitizers are reported. Multicompartmental modeling has been applied to analyze the data and determine candidate dose parameters for use in integrated risk assessment methods: the area under the curve (AUC) and maximum concentration (C(max)) in the epidermis. A model with two skin compartments, representing the stratum corneum and viable skin (epidermis and dermis), was chosen following a formal model selection process. Estimates of the uncertainty, as well as average values of the epidermal disposition kinetics parameters, were made by fitting to the time course skin permeation data from individual skin donors. A potential reduced time course method is proposed based on two time points at 4 and 24 h, which gives results close to those from the full time course for the current data sets. The time course data presented in this work have been provided as a resource for development of predictive in silico skin permeation models.

DEVELOPMENT OF A MODIFIED IN VITRO SKIN ABSORPTION METHOD TO STUDY THE EPIDERMAL/DERMAL DISPOSITION OF A CONTACT ALLERGEN IN HUMAN SKIN

In vitro skin absorption methods exist in Organisation for Economic Co-operation and Development (OECD) guideline form (No. 428) and are used to estimate the degree of systemic penetration of chemicals through skin. More detailed kinetics of permeation through skin compartments are not described well by existing methods. This study was designed to assess the practical feasibility of generating compartmental (stratum corneum/epidermal/dermal) disposition and kinetic data of topically applied chemicals. For chemically induced effects initiated in the skin (e.g., skin allergy), the delivery of tissue concentrations of chemical will impact the incidence and severity of biological effect. Explicit data on the kinetics of chemical disposition in skin have not traditionally been needed for skin allergy risk assessment: current in vivo assays embody delivery implicitly. Under the 7th Amendment to the European Cosmetics Directive, in vivo assays (such as the local lymph node assay for skin sensitization) will not be permitted to assess cosmetic ingredients. New in vitro and in silico alternative approaches and ways of predicting risk of adverse effects in humans need to be developed, and new methods such as that described here provide a way of estimating delivered concentrations and the effect of formulation changes on that delivery. As we continue to deconstruct the contributing factors of skin allergy in humans, it will be useful to have methods available that can measure skin tissue compartment exposure levels delivered from different exposure use scenarios. Here we provide such a method. The method could also be used to generate useful data for developing in silico kinetic models of compartmental skin delivery and for refining data for skin delivery in relation to the evaluation of systemic toxicity.

In Vitro Skin Absorption of the Topically Applied Metal Ion-Chelating Agent Diethylenetriaminepenta-acetic Acid

Diethylenetriaminepenta-acetic acid (DTPA) is a metal ion-chelating agent that has antimicrobial properties and potential therapeutic properties against metal-induced toxicities such as nickel allergy. In this study, the absorption properties of DTPA applied topically to rat skin are investigated in vitro, using a flow-through diffusion skin absorption model. [14C]DTPA was applied in solution in 60% ethanol (pH 6). Overall skin penetration into receptor fluid resulting from a topical dose of 0.13 mg/cm2 DTPA for 24 h was low at 1.27%. The local tissue distribution of DTPA was investigated using microautoradiography, and effects on the tissue were assessed by histology. Diethylenetriaminepenta-acetic acid was primarily associated with the stratum corneum and upper layers of the skin; minimal levels were observed in the dermis.