To reduce variance between different experiments, thickness ratios were calculated

To reduce variance between different experiments, thickness ratios were calculated. from healthy controls and psoriasis patients were cultured alone or co-cultured with activated memory CD4+ T cells. Besides IL-1, IL-17A induced direct expression of IL-19 and IL-24 in skin fibroblasts and keratinocytes. Importantly, intrinsic higher expression of IL-19 in psoriatic Chlorocresol skin fibroblasts was observed in comparison to healthy skin fibroblasts. Neutralization of IL-17A in the human skin fibroblast-T cell co-culture system significantly suppressed IL-19 and IL-24 expression. Together, our data show that IL-17A-induced IL-19 and IL-24 expression in skin stromal cells contribute to keratinocyte proliferation. IL-19 expression is usually normalized in patients with psoriasis treated with anti-IL-17 therapy is not obvious. Previously, our group established a psoriasis-like skin inflammation model in mice using topical application of imiquimod (IMQ) (16). This model successfully re-captures most critical features of acute plaque formation in psoriasis such as keratinocyte hyper proliferation, acanthosis and parakeratosis (16). Like in human psoriasis, enhanced activity of the IL-23/IL-17 pathway was also involved in the IMQ-induced psoriasis mouse model (16). However, in contrast to the chronic natural course in human psoriasis, this mouse model does not develop into a chronic state of psoriasis, because of stabilization and even improvement of skin inflammation after 5 to 6 days. Interestingly, a clinical study in psoriasis patients showed that, non-lesional skin treated with IMQ in the beginning developed typical features of psoriasis such as acanthosis and parakeratosis (17). Nevertheless, both clinical and histological features subsided thereafter and in this human model of IMQ-induced psoriasis, the induced lesions showed spontaneous improvement after 5 to 6 days. This improvement was accompanied by significantly lower expression of IL-17A and with a higher expression of IL-10 (17). This suggests that upregulation of IL-10 is usually involved in the spontaneous improvement of psoriasis symptoms after 5 to 6 days in murine IMQ model and probably explains the spontaneous improvement observed in the IMQ mouse model. Therefore, we used an anti-IL-10 antibody to investigate whether we could achieve enhanced expression of IL-17 in the IMQ-induced psoriasis mouse model and the accompanying visible psoriatic symptoms beyond day 5. assays with human skin fibroblasts from patients with psoriasis and healthy skin were performed to evaluate the direct induction of IL-19 by IL-17. In addition, an ex lover vivo human psoriasis skin co-culture system was used to examine the effects of biologics targeting IL-17A on IL-19 expression. Material and Methods IMQ-Induced Psoriasis Mouse Model BALB/c mice (8-11 week-old) received daily topical application of 62.5mg 5% Aldara (3M Pharmaceuticals) on their shaved back skin. Control mice (n=6, pooled from two impartial experiments) were treated with a thin layer of petrolatum (Fagron). Daily evaluation of the local psoriasis area and severity index (PASI) has been explained previously (16). Every other day, 20 mg/kg body weight of anti-IL-10 or isotype control antibody (n=10 each, pooled from two impartial experiments) was intraperitoneally (i.p.) Chlorocresol injected, or 5 mg/kg body weight of dexamethasone (n=7, pooled from two impartial experiments) was subcutaneously (s.c.) injected as an anti-inflammatory platinum standard. Five and ten days after IMQ Rabbit Polyclonal to Cytochrome P450 46A1 induction, mice were sacrificed for analysis. Food and water were provided ad libitum, and mice were kept under specific pathogen-free conditions. All experiments were approved by the Erasmus MC Dutch Animal Ethics Committee (DEC). Histology and Immunohistochemistry After sacrifice, skin biopsies were taken and snap-frozen in TissueTek (Bayer). Sections were cut with a Leica cryostat. Gr-1 antibody (clone RB6-8C5) and Ki-67 antibody (Dako, A0047) were utilized for IHC staining. Subsequent steps were performed as explained earlier (16). Images were analyzed with LAX V4.12 program (Leica microsystems) or NDP view2 (Hamamatsu photonics). To measure epidermal thickness, the average of four measurements was used as the representative thickness per sample. To reduce variance between different experiments, thickness ratios were calculated. Specifically, each skin thickness was divided by the mean skin thickness of the isotype group from that experiment, and thereby mean values of thickness for isotype groups were usually set at one. Flow Cytometry Back skin (ca. 1 cm2) was digested in Chlorocresol 50 g/mL Liberase (Roche) at 4C immediately and then at 37C for 1 hour to create a single cell suspension and cells were stained with the following antibodies: CD45-BV785 (Biolegend, clone 104), CD11b-eF450 (eBioscience, clone M1/70), Ly6C-APC-Cy7 (BD pharmingen, clone.