Skin is a target of allergic reactions to aromatic amine hair dye precursors, such as p-phenylenediamine (PPD). As conversion of PPD on or in the skin is expected to be required for the induction of allergic contact dermatitis, we analyzed the role of oxidation and N-acetylation as major transformation steps. PPD and its oxidative and N-acetylated derivatives were tested for their sensitizing potential in vitro using a dendritic cell (DC) activation assay and in vivo using the local lymph node assay (LLNA). PPD did not induce relevant DC activation but induced a positive LLNA response. In contrast, DC activation was obtained when PPD was chemically pre-oxidized or after air oxygen exposure. Under both conditions, the potent sensitizing PPD oxidation product Bandrowski's base was identified along with other di- and trimeric species, indicating that PPD oxidation products provide an effective immune stimulation (danger signal). In contrast mono- and diacetylated PPD did not induce DC activation or a positive LLNA response. We conclude that dermal N-acetylation of PPD competes with the formation of oxidized PPD whereas skin exposure conditions allowing auto-oxidation, as in the LLNA, provide an effective danger signal necessary to induce skin sensitization to PPD.

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Development of in vitro models to identify sensitizing chemicals receives public interest since animal testing should be avoided whenever possible. In this article we analyze two essential properties of sensitizing chemicals: skin penetration and dendritic cell (DC) activation. Activation of immature DC derived from peripheral blood monocytes was evaluated by flow cytometric analysis of CD86 positive cells and quantitative measurement of interleukin-1beta and aquaporin P3 gene expression. The sensitizer 2,4,6-trinitrobenzenesulfonic acid induced a concentration-dependent response for all parameters, whereas the irritant sodium lauryl sulfate did not. When two related aromatic amines, p-toluylenediamine (PTD) and hydroxyethyl-p-phenylenediamine (HE-PPD) were tested, both induced substantial DC activation indicating their potential sensitizing properties. These findings contrasted with in vivo results: in murine local lymph node assays (LLNA) PTD, but not HE-PPD, was sensitizing using acetone/aqua/olive oil as vehicle. Skin penetration measurement revealed that this was due to bioavailability differences. On retesting HE-PPD in the LLNA using the penetration enhancer dimethylsulfoxide as vehicle, it induced a specific response. We conclude that in vitro analysis of DC activation capability of the two selected chemicals demonstrates that prediction of skin sensitization potential is possible provided that skin penetration data indicate sufficient bioavailability of the test compound.

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We constructed balancer-chromosomes for the large autosomes ofDrosophila hydei and screened more than 16000 chromosomes for male sterile mutations in order to dissect spermatogenesis genetically. 365 mutants on the X chromosome and the autosomes 2, 3, and 4 were recovered and analysed cytologically in squash preparations under phase-contrast optics. The majority of the mutations allows a rather advanced differentiation of the spermatozoa. At the light-microscopical level, it is possible to classify these mutations with respect to individualization, coiling or motility of the mutant spermatozoa. In contrast, a small number of mutants exhibits conspicuous, pleiotropic phenotypes. Gonial divisions, the shaping of the spermatocyte nucleus and male meiotic divisions are controlled by X chromosomal or autosomal genes which can mutate to male sterile alleles. A number of nonallelic 3 chromosome male sterile mutations interfere with the unfolding of the Y chromosomal lampbrush loops. Other autosomal male sterile mutations modify the morphology of these lampbrush loops. Another group of mutations inhibits the formation of the nebenkern while the development of the spermatid nucleus and the flagellum can proceed. Such male sterile mutations can decouple the development of nucleus, protein body, nebenkern, and flagellum of the spermatid. Thus, we can describe spermatogenesis inDrosophila as the coordinate execution of the individual developmental programs of the different components of the spermatozoon.