Common fragrance chemicals activate dendritic cells in coculture with keratinocytes.

BACKGROUND: Fragrances are important contact allergens; however, investigation of their skin sensitization potency has been challenging in new approach methods (NAMs). Many fragrance chemicals are susceptible to autoxidation or can be metabolized by enzymes constitutively expressed in skin keratinocytes. Strong sensitizers can be formed in both of these processes. Further, keratinocytes can modulate the dendritic cell (DC) activation and maturation potential, a key event in the acquisition of contact allergy. OBJECTIVES: To evaluate the 2D coculture model consisting of keratinocytes and DCs using different weak to moderate sensitizing fragrance chemicals. Further, to investigate fragrances and related oxidation products in the in vitro model and compare to in vivo data. METHODS: Chemicals were tested in the coculture activation test (COCAT), consisting of HaCaT keratinocytes and THP-1 cells. THP-1 cell surface expression of costimulatory and adhesion molecules (CD86 and CD54) collected after 24 h incubation with the chemicals was analysed using flow cytometry. RESULTS: Twenty-four molecules were tested positive, three were negative (n = 27). Four pairs were evaluated, with aldehydes showing a 6- to 13-fold stronger responses compared to their corresponding alcohols. CONCLUSIONS: Results provide insight into the activation of DC in their natural environment of keratinocytes. α,ß-Unsaturated alcohols were classified as weaker sensitizers compared to their corresponding aldehydes. In sum, testing of fragrances retrieved results in good agreement with in vivo data.

The HaCaT/THP-1 Cocultured Activation Test (COCAT) for skin sensitization: a study of intra-lab reproducibility and predictivity.

The Cocultured Activation Test (COCAT) consists of cocultured HaCaT (human keratinocyte cell line) and THP-1 cells (surrogate of antigen presenting cells). Individually, these cell lines are used to address key event 2 and 3 of the skin sensitization Adverse Outcome Pathway (AOP). Their exposure in coculture was found to have the potential to increase their response to sensitizing chemicals, enable the detection of pro-haptens and support the identification of skin sensitization potency. The present study was undertaken to assess the predictive capacity of COCAT to both skin sensitization hazard and potency and to assess the intra-laboratory reproducibility of COCAT based on the blind testing of chemicals. Results showed a reproducibility between runs of 80 % for 15 coded chemicals. 100 % sensitivity (9/9), 75 % specificity (3/4) and 92.3 % accuracy (12/13) was found for skin sensitization hazard prediction, while the tests of two chemicals were inconclusive. Including additional chemicals tested during the optimization phase in addition to the blind tested chemicals, the skin sensitization UN GHS sub-categories were correctly predicted for 85.7 % (12/14) Sub-category 1A chemicals, 83.3 % (10/12) Sub-category 1B chemicals and 92.3 % (12/13) 'No Category' chemicals, resulting in an overall accuracy of 87.4 % (34/39). The present study shows the COCAT to be a promising method for the identification of skin sensitization hazard and potency sub-categorization according to the UN GHS classification.