Local cortisol activation is involved in EGF-induced immunosuppression.

The major effects of the epidermal growth factor receptor (EGFR) signalling pathway on keratinocytes are cell proliferation, cell differentiation, and wound healing. In addition to these effects, an immunosuppressive effect of EGFR signalling has been reported. However, the precise mechanism of immunosuppression by EGFR signalling is not well understood. In this study, we clarified the involvement of increased local cortisol activation in EGFR signalling-induced immunosuppression in keratinocytes. EGF treatment up-regulated the expression of 11ß-hydroxysteroid dehydrogenase 1 (11ß-HSD1) and supernatant cortisol levels in a dose-dependent manner in keratinocytes. 11ß-HSD1 is an enzyme that catalyses the conversion of cellular hormonally inactive cortisone into active cortisol. qRT-PCR and ELISA assays indicated that EGF significantly decreased tumour necrosis factor α (TNF- α)-induced interleukin-6 (IL-6) expression in keratinocytes. Similarly, 11ß-HSD1 overexpression significantly decreased TNF-α-induced IL-6 expression. We evaluated the role of 11ß-HSD1 in immunosuppression through EGFR signalling. Blockade of 11ß-HSD1 via 11ß-HSD1 inhibitor reversed both the expression and production of TNF-α-induced IL-6, which was decreased by EGF in keratinocytes. Therefore, increased local cortisol activation by 11ß-HSD1 is involved in EGFR signalling-induced immunosuppression in keratinocytes. Finally, we evaluated whether EGFR inhibition by cetuximab affects the expression of 11ß-HSD1. We found that 0.1 µg cetuximab decreased 11ß-HSD1 transcript levels in keratinocytes. The changes in 11ß-HSD1 were more apparent in TNF-α-treated cells. As 11ß-HSD1 expression in keratinocytes is associated with inflammation and cell proliferation, this mechanism may be associated with adverse skin reactions observed in patients treated with EGFR inhibitors.

Local Glucocorticoid Activation by 11ß-Hydroxysteroid Dehydrogenase 1 in Keratinocytes: The Role in Hapten-Induced Dermatitis.

Over the past decade, extra-adrenal cortisol production was reported in various tissues. The enzyme that catalyzes the conversion of hormonally inactive cortisone into active cortisol in cells is 11ß-hydroxysteroid dehydrogenase 1 (11ß-HSD1). We recently reported that 11ß-HSD1 is also expressed in keratinocytes and regulates inflammation and keratinocyte proliferation. To investigate the function of 11ß-HSD1 in keratinocytes during inflammation in vivo, we created keratinocyte-specific 11ß-HSD1 knockout (K5-Hsd11b1-KO) mice and analyzed the inflammatory response in models of hapten-induced contact irritant dermatitis. K5-Hsd11b1-KO mice showed enhanced ear swelling in low-dose oxazolone-, 2,4,6-trinitro-1-chlorobenzene (TNCB)-, and 2,4-dinitrofluorobenzene-induced irritant dermatitis associated with increased inflammatory cell infiltration. Topical application of corticosterone dose dependently suppressed TNCB-induced ear swelling and cytokine expression. Similarly in mouse keratinocytes in vitro, corticosterone dose dependently suppressed 2,4,6-trinitrobenzenesulfonic acid-induced IL-1α and IL-1ß expression. The effect of 11-dehydrocorticosterone was attenuated in TNCB-induced irritant dermatitis in K5-Hsd11b1-KO mice compared with wild-type mice. In human samples, 11ß-HSD1 expression was decreased in epidermis of psoriasis vulgaris compared with healthy skin. Taken together, these data suggest that corticosterone activation by 11ß-HSD1 in keratinocytes suppresses hapten-induced irritant dermatitis through suppression of expression of cytokines, such as IL-1α and IL-1ß, in keratinocytes.

A vitamin D analog inhibits Th2 cytokine- and TGFß -induced periostin production in fibroblasts: a potential role for vitamin D in skin sclerosis.

Scleroderma is an autoimmune disease characterized by extracellular matrix deposition and inflammation. Topical vitamin D analogs have been reported as effective treatments for scleroderma. We previously reported that a matricellular protein, periostin (POSTN), contributes to pathogenesis of scleroderma as POSTN knockout mice were resistant to bleomycin (BLM)-induced scleroderma. We investigated whether a vitamin D analog affects the expression of POSTN in dermal fibroblasts and in a BLM-induced scleroderma model. The vitamin D analog, maxacalcitol (22-oxacalcitriol [OCT]), was applied to dermal fibroblasts and POSTN expression was measured. The effect of OCT on Th2 cytokine- and TGFß-induced POTSN and Collagen 1 α 1 (Col1A1) expression was also assessed. In vivo, OCT was administered to BLM-induced scleroderma model and outcomes were determined by dermal thickness, collagen density and POSTN expression. Treatment with OCT significantly decreased POSTN expression in dermal fibroblasts. Th2 cytokine- and TGFß-induced expression of POSTN and Col1A1 was also suppressed by OCT. In vivo, OCT administration decreased the density of collagen bundles and POSTN expression in a BLM-induced scleroderma model. In addition to the previously reported immunosuppressive effect, the vitamin D analog OCT might be effective to treat scleroderma, in part through inhibition of Th2 cytokine- and TGFß-induced POSTN expression.

Th2 cytokines enhance TrkA expression, upregulate proliferation, and downregulate differentiation of keratinocytes.

BACKGROUND: Nerve growth factor (NGF), a neurotrophin that plays a critical role in developmental neurobiology, is released by proliferating keratinocytes and induces proliferation. OBJECTIVE: The aim of this study was to investigate the role of tyrosine kinase receptor A (TrkA), a high-affinity receptor of NGF, in human keratinocytes. METHODS: Expression of TrkA and NGF in skin diseases was investigated by immunohistochemistry. Expression of TrkA in cells was examined by Western blotting and RT-PCR. Cell proliferation was assessed by BrdU assay. RESULTS: We first determined the expression of TrkA and NGF in skin samples from patients with atopic dermatitis, prurigo nodularis, psoriasis vulgaris, and seborrheic keratosis. TrkA was only expressed in proliferating basal cells, and its expression was enhanced in atopic dermatitis samples. NGF expression was enhanced in atopic dermatitis and prurigo nodularis samples and in some samples from seborrheic keratosis patients. Investigation of the role of TrkA in vitro using normal human epidermal keratinocytes (NHEK) revealed that TrkA was significantly enhanced by the T helper type 2 (Th2) cytokines interleukin (IL)-4 and IL-13 but not by other inflammatory cytokines, such as IL-1ß, tumor necrosis factor α, interferon γ, or epidermal growth factor. On the other hand, expression of NGF was not altered by Th2 cytokines. Notably, inhibition of TrkA significantly reversed the effects of IL-4 on proliferation and differentiation. Furthermore, overexpression of TrkA enhanced proliferation of NHEK. These results indicate that IL-4-induced TrkA expression in keratinocytes modulates proliferation and differentiation of these cells. CONCLUSION: Increased TrkA expression in keratinocytes in atopic dermatitis may contribute to the observed epidermal hyperproliferation in these patients.

The predominant drug-specific T-cell population may switch from cytotoxic T cells to regulatory T cells during the course of anticonvulsant-induced hypersensitivity.

BACKGROUND: Delayed hypersensitivity is responsible for severe cutaneous adverse drug reactions (cADRs), especially in Stevens-Johnson syndrome (SJS), toxic epidermal necrolysis, and drug-induced hypersensitivity syndrome (DIHS) (also known as drug rash with eosinophilia and systemic symptoms [DRESS] syndrome). The drug-induced lymphocyte stimulation test (DLST), or lymphocyte transformation test (LTT), is used to identify the culprit drug in severe cADR cases. OBJECTIVE: The aim of this study was to examine the immune reactions in cADR patients through the identification of the drug-specific proliferating cells by flow cytometric DLST (FCM-DLST). METHODS: The peripheral blood mononuclear cells of 16 anticonvulsant-induced cADR patients were investigated by conventional DLST and a FCM-DLST protocol in which CFSE dilution and BrdU incorporation were combined. FCM-DLST allowed for the identification of the drug-specific proliferating cells in six cases. Three of these cases were DIHS cases, whereas there was one case of SJS, one case of maculopapular rash (MP), and one case of erythema multiforme (EM) among the six cases. RESULTS: In FCM-DLST, drug-specific proliferating T cells were detected as CFSE(low) BrdU(high) cells. These cells corresponded to the cells incorporating (3)H-thymidine in conventional DLST. Although CD4(+) T-cell proliferation dominated the observed proliferation in most of the cases (in the recovery stage of the three DIHS cases, the MP case, and the EM case), drug-specific CD8(+) cytotoxic T lymphocytes (CTLs) were detected, especially in the acute stages of the SJS case and one of the DIHS cases. There was a dramatic switch in the predominant drug-specific proliferating T-cell population in the course of one of the cases of DIHS in which CD8(+) CTLs were predominant initially, whereas CD4(+) T cells were predominant later. Moreover, drug-specific CD4(+) CD25(+) Foxp3(+) regulatory T cells (Tregs) proliferated during the recovery stage in one DIHS case. CONCLUSIONS: FCM-DLST revealed that the cell proliferation detected by conventional DLST is a heterogeneous proliferation of both CD8(+) CTLs and CD4(+) T cells that likely includes Tregs. However, the number of cADR cases in this study was limited, which limits the conclusions that can be drawn from it.