ABSTRACT
Despite recent developments, there is a high medical need for new treatment options for chronic inflammatory dermatoses like allergic contact dermatitis (ACD) and psoriasis. Particularly, more predictive skin inflammation models are required to facilitate the process of drug discovery. Murine contact hypersensitivity (CHS) models adequately reflect ACD and are also used to characterize therapeutic approaches for psoriasis. Using the hapten 2,4-dinitrofluorobenzene (DNFB), we established new subacute and subchronic DNFB-induced CHS models in C57BL/6 mice, which more closely reflect the characteristics of chronic T-cell-dependent inflammatory dermatoses as pronounced keratinocyte proliferation, strong hypervascularization, immune cell infiltration and overexpression of T cell and inflammatory cytokines. For the subacute DNFB model, we demonstrated anti-inflammatory activity of the glucocorticoid, prednisolone, as well as of neutralization of TNFα, IL-12/IL-23 or IL-18. In the subchronic DNFB-induced CHS model, deficiency for MyD88 and IL-12/IL-35 p35 chain but not IL-12/IL-23 p40 chain led to decreased skin inflammation. Furthermore, as exemplified by the dose-dependently effective therapeutic prednisolone treatment, the subchronic model allows the continuous therapy of a pre-established stable contact dermatitis. Altogether, prolonged DNFB-induced mouse CHS models closely reflect ACD sensitive to glucocorticoids as standard therapy, reveal a more chronic skin inflammation and are responsive to cytokine antagonization.
Subject(s)
Dermatitis, Allergic Contact/etiology , Dinitrofluorobenzene/toxicity , Animals , Anti-Inflammatory Agents/therapeutic use , Chronic Disease , Dermatitis, Allergic Contact/drug therapy , Disease Models, Animal , Mice , Mice, Inbred C57BL , Mice, Nude , Prednisolone/therapeutic use , Psoriasis/etiologyABSTRACT
T-cell-mediated processes play an essential role in the pathogenesis of several inflammatory skin diseases such as atopic dermatitis, allergic contact dermatitis and psoriasis. The aim of this study was to investigate the role of the IL-2-inducible tyrosine kinase (Itk), an enzyme acting downstream of the T-cell receptor (TCR), in T-cell-dependent skin inflammation using three approaches. Itk knockout mice display significantly reduced inflammatory symptoms in mouse models of acute and subacute contact hypersensitivity (CHS) reactions. Systemic administration of a novel small molecule Itk inhibitor, Compound 44, created by chemical optimization of an initial high-throughput screening hit, inhibited Itk's activity with an IC50 in the nanomolar range. Compound 44 substantially reduced proinflammatory immune responses in vitro and in vivo after systemic administration in two acute CHS models. In addition, our data reveal that human Itk, comparable to its murine homologue, is expressed mainly in T cells and is increased in lesional skin from patients with atopic dermatitis and allergic contact dermatitis. Finally, silencing of Itk by RNA interference in primary human T cells efficiently blocks TCR-induced lymphokine secretion. In conclusion, Itk represents an interesting new target for the therapy of T-cell-mediated inflammatory skin diseases.
Subject(s)
Dermatitis/drug therapy , Protein Kinase Inhibitors/pharmacology , Protein-Tyrosine Kinases/antagonists & inhibitors , Animals , Base Sequence , Dermatitis/enzymology , Dermatitis/immunology , Dermatitis, Allergic Contact/drug therapy , Dermatitis, Allergic Contact/enzymology , Dermatitis, Allergic Contact/immunology , Dermatitis, Atopic/drug therapy , Dermatitis, Atopic/enzymology , Dermatitis, Atopic/immunology , Dinitrochlorobenzene/immunology , Dinitrochlorobenzene/toxicity , Disease Models, Animal , Drug Evaluation, Preclinical , Female , Gene Expression Profiling , Humans , In Vitro Techniques , Lymphoid Tissue/enzymology , Lymphoid Tissue/immunology , Mice , Mice, Knockout , Protein-Tyrosine Kinases/deficiency , Protein-Tyrosine Kinases/genetics , Psoriasis/drug therapy , Psoriasis/enzymology , Psoriasis/immunology , RNA Interference , RNA, Messenger/genetics , RNA, Messenger/metabolism , RNA, Small Interfering/genetics , Recombinant Fusion Proteins/antagonists & inhibitors , Recombinant Fusion Proteins/genetics , T-Lymphocytes/drug effects , T-Lymphocytes/enzymology , T-Lymphocytes/immunology , Up-RegulationABSTRACT
In Arabidopsis, the EMBYRONIC FLOWER2 (EMF2), VERNALISATION2 (VRN2) and FERTILISATION INDEPENDENT ENDOSPERM2 (FIS2) genes encode related Polycomb-group (Pc-G) proteins. Their homologues in animals act together with other Pc-G proteins as part of a multimeric complex, Polycomb Repressive Complex 2 (PRC2), which functions as a histone methyltransferase. Despite similarities between the fis2 mutant phenotype and those of some other plant Pc-G members, it has remained unclear how the FIS2/EMF2/VRN2 class Pc-G genes interact with the others. We have identified a weak emf2 allele that reveals a novel phenotype with striking similarity to that of severe mutations in another Pc-G gene, CURLY LEAF (CLF), suggesting that the two genes may act in a common pathway. Consistent with this, we demonstrate that EMF2 and CLF interact genetically and that this reflects interaction of their protein products through two conserved motifs, the VEFS domain and the C5 domain. We show that the full function of CLF is masked by partial redundancy with a closely related gene, SWINGER (SWN), so that null clf mutants have a much less severe phenotype than emf2 mutants. Analysis in yeast further indicates a potential for the CLF and SWN proteins to interact with the other VEFS domain proteins VRN2 and FIS2. The functions of individual Pc-G members may therefore be broader than single mutant phenotypes reveal. We suggest that plants have Pc-G protein complexes similar to the Polycomb Repressive Complex2 (PRC2) of animals, but the duplication and subsequent diversification of components has given rise to different complexes with partially discrete functions.
