Publikationsdatenbank

Investigation of the sensitisation potential of various textile dyes using a biphasic mice local lymph node assay (LLNA) and an in vitro loose-fit coculture-based sensitisation assay (LCSA) (2010)

Art

Hochschulschrift

Autor

Ahuja, Varun (WE 14)

Quelle

Berlin: Mensch und Buch Verlag, 2010 — V, 115 Seiten

ISBN: 978-3-86664-789-3

Sprache

Englisch

Verweise

URL (Volltext): https://refubium.fu-berlin.de/handle/fub188/7692

Kontakt

Institut für Pharmakologie und Toxikologie

Koserstr. 20
14195 Berlin
+49 30 838 53221
pharmakologie@vetmed.fu-berlin.de

Abstract / Zusammenfassung

Contact dermatitis is one of the most common skin diseases, with a great socio-economic impact. Disperse dyes, which are suitable for dyeing synthetic fibres, are responsible for the great majority of allergic contact dermatitis (ACD) cases to textile dyes. The aim of the present study was to investigate the sensitising potential of various disperse dyes using a biphasic protocol of the local lymph node assay (LLNA) and an in vitro loose-fit coculture based sensitisation assay (LCSA). In the biphasic LLNA, mice were shaved over a surface of approximately 2 cm2 on their backs, and treated using a “sensitisation-challenge protocol”. The shaved surface was treated once daily on days 1 to 3 with 50 l of the test solution. Animals remained untreated on days 4 to 14. On days 15 to 17, mice were treated with 25 l of the test solution on the dorsum of both ears. Mice were sacrificed on day 19 with deep CO2 anaesthesia, the lymph nodes were prepared and various end points, such as ear thickness, ear punch weight, lymph node weight, lymph node cell count, and the proportion of various lymphocyte subpopulations were determined. The results of treated mice were compared to those of the control group treated with the vehicle alone. The LCSA involves single layer of human non- differentiating keratinocytes and of allogenic floating monocytes which are cocultured in serum-free medium in the presence of a cytokine cocktail. The coculture develops into a system consisting of activated keratinocytes and dendritic cell-related cells. The half-maximal increase in CD86 expression on the dendritic cell-like cells (EC50 values) was used to compare the sensitising potential of tested substances. Our results from the LLNA experiments showed that almost all of the tested textile dyes caused a significant increase in lymph node cell count and lymph node weight except Disperse orange 3. Disperse blue 124 and Disperse blue 106 showed the most pronounced increase in cell count and lymph node weight already at low concentrations of 0.003%, followed by Disperse orange 37, Disperse Red 1, Disperse Blue 35 and Disperse Blue 1. Disperse yellow 3 showed the least increase in cell count and lymph node weight parameters among all the textile dyes tested. Based on these results, the disperse dyes could be arranged in four groups on the basis of their sensitising potency in the following decreasing order (in parenthesis: lowest concentration causing a significant increase in lymph node cell number): group 1, strong: Disperse blue 124 and Disperse blue 106 (0.003%), group 2, moderate: Disperse red 1 and Disperse blue 1 (3%), group 3, weak: Disperse orange 37 and Disperse blue 35 (10%) and group 4, very weak: Disperse yellow 3 and Disperse orange 3 (increase at 30% or no increase at 30%). We also observed an increase in ear thickness and ear- punch weight in most of the concentrations tested for various textile dyes. Many allergens cause an increase in ear parameters also, but the relative effects on lymph node parameters are more significant as compared to the ear parameters. We observed a decrease in CD4+ and CD8+ cells and an increase in CD19+, CD45+ and CD45+/1A+ cells in most of the cases, which is characteristic for allergens. The CD4+/CD69+ cells increased in only few experiments mainly with Disperse blue 124 and Disperse blue 106. In the LCSA experiments, the textile dyes can be arranged on the basis of EC50 values in the following decreasing order of their sensitisation potency: Disperse blue 124 > Disperse yellow 3 > Disperse orange 37 > Disperse blue 106 > Disperse red 1 > Disperse blue 35 > Disperse orange 3. The results of the LCSA experiments are mostly comparable with the LLNA experiments. In conclusion, this study shows that the biphasic LLNA protocol was proficient enough to study the sensitisation potential of tested textile dyes. The in vitro loose-fit-coculture assay also gave comparable results to LLNA, and was found to be robust enough to investigate the sensitisation potential of test substances.