Adaptation of a skin sensitization assay to a chemically defined culture. (June 2019)

• Record Type: Journal Article
• Authors: Marigliani, Bianca; Silva, Josemar Vinícius Maiworm Abreu; Balottin, Luciene Bottentuit López; Silva, Karina Ribeiro; Baptista, Leandra Santos; Campos, Claudia Barbosa Ladeira de; Augusto, Elisabeth de Fatima Pires
• Is Part Of: Toxicology in vitro. Volume 57(2019)
• Page Start: 145
• Page End: 153
• Publication Date: 2019-06
• Subjects: Toxicity tests -- Animal use alternatives -- Culture media, serum-free -- In vitro techniques -- Dermatitis, allergic contact; UN GHS United Nations Globally Harmonized System of Classification and Labelling of Chemicals -- LLNA Local Lymph Node Assay -- AOP Adverse outcome pathway -- OECD Organization for Economic Co-operation and Development -- h-CLAT Human-Cell Line Activation Test -- FBS Fetal bovine serum -- GLC d-Glucose -- LAC l-Lactate -- GLN l-Glutamine -- GLU l-Glutamic acid -- DT Doubling time -- CV75 Chemical concentration giving 75% cell viability -- LA Lactic acid -- DNCB 2, 4-Dinitrochlorobenzene -- NI nickel sulfate; Toxicity testing -- In vitro -- Periodicals; Toxicology -- Periodicals; 615.9
• DOI: 10.1016/j.tiv.2018.12.004
• Languages: English
• ISSNs: 0887-2333

Abstract

Abstract: There are currently three in vitro methods adopted by the Organization for the Economic Co-operation and Development for testing chemicals based on the third key event of the skin sensitization adverse outcome pathway, the activation of dendritic cells. All of them use culture medium supplemented with fetal bovine serum (FBS), which brings technical disadvantages and animal welfare concerns. The objective of this study was to analyze the possibility of eliminating the use of FBS in the human Cell Line Activation Test (h-CLAT). After successful implementation of the h-CLAT using THP-1 cells cultured in FBS-containing medium, several attempts to adapt THP-1 cells to four different serum-free media were made. The best results were obtained with gradual adaptation to RPMI-1640 medium with HL-1™ Supplement and to X-VIVO™ 10. Adapted cells were cryopreserved and submitted to the reactivity check. After being approved, they were used in dose finding and proficiency assays. Despite minor adjustments in the original protocol, it was possible to correctly predict the sensitizing potential of the ten proficiency substances using THP-1 cells adapted to X-VIVO™ 10, which indicates that it is possible to eliminate the use of FBS in the h-CLAT, using a chemically defined medium.