Marcel B. M. Teunissen

In vitro and in situ expression of IL-23 by keratinocytes in healthy skin and psoriasis lesions: enhanced expression in psoriatic skin

Keratinocytes contribute to cutaneous immune responses through the expression of cytokines. We investigated whether human keratinocytes can express IL-23, a newly defined IFN-γ-inducing cytokine composed of a unique p19 subunit and a p40 subunit shared with IL-12. Cultured keratinocytes from normal and lesional psoriatic skin were found to express constitutively mRNA for both subunits of IL-23. Low but significant levels of the heterodimeric IL-23 protein could be detected in cell lysates and supernatants from stimulated keratinocytes by immunoblotting and ELISA. Functional analysis showed that these low levels of keratinocyte-derived IL-23 were sufficient to enhance the IFN-γ production by memory T cells. Immunostaining of skin sections confirmed expression of both subunits of IL-23 by keratinocytes in situ and also revealed expression of this cytokine in the dermal compartment. IL-23 expression was significantly higher in psoriatic lesional skin, compared with normal and psoriatic nonlesional skin. The immunostained preparations of cultured cells and IL-23 levels in culture supernatants did not show any difference between normal and psoriatic keratinocytes indicating no intrinsic aberration of IL-23 expression in keratinocytes from psoriatic skin. Double staining of cytospin preparations demonstrated that IL-23 p19 is also expressed by epidermal Langerhans cells, dermal dendritic cells, and macrophages. Psoriasis is a chronic inflammatory skin disease mediated by IFN-γ-expressing type 1 memory T cells. As IL-23 is important to activate memory T cells to produce IFN-γ, its augmented expression of IL-23 by keratinocytes and cutaneous APC may contribute to the perpetuation of the inflammation process in this disease.

Overrepresentation of IL-17A and IL-22 Producing CD8 T Cells in Lesional Skin Suggests Their Involvement in the Pathogenesis of Psoriasis

Background Although recent studies indicate a crucial role for IL-17A and IL-22 producing T cells in the pathogenesis of psoriasis, limited information is available on their frequency and heterogeneity and their distribution in skin in situ. Methodology/Principal Findings By spectral imaging analysis of double-stained skin sections we demonstrated that IL-17 was mainly expressed by mast cells and neutrophils and IL-22 by macrophages and dendritic cells. Only an occasional IL-17pos, but no IL-22pos T cell could be detected in psoriatic skin, whereas neither of these cytokines was expressed by T cells in normal skin. However, examination of in vitro-activated T cells by flow cytometry revealed that substantial percentages of skin-derived CD4 and CD8 T cells were able to produce IL-17A alone or together with IL-22 (i.e. Th17 and Tc17, respectively) or to produce IL-22 in absence of IL-17A and IFN-γ (i.e. Th22 and Tc22, respectively). Remarkably, a significant proportional rise in Tc17 and Tc22 cells, but not in Th17 and Th22 cells, was found in T cells isolated from psoriatic versus normal skin. Interestingly, we found IL-22 single-producers in many skin-derived IL-17Apos CD4 and CD8 T cell clones, suggesting that in vivo IL-22 single-producers may arise from IL-17Apos T cells as well. Conclusions/Significance The increased presence of Tc17 and Tc22 cells in lesional psoriatic skin suggests that these types of CD8 T cells play a significant role in the pathogenesis of psoriasis. As part of the skin-derived IL-17Apos CD4 and CD8 T clones developed into IL-22 single-producers, this demonstrates plasticity in their cytokine production profile and suggests a developmental relationship between Th17 and Th22 cells and between Tc17 and Tc22 cells.

Psoriasis: dysregulation of innate immunity.

The current understanding of the function of natural killer (NK) T cells in innate immunity and their potential to control acquired specific immunity, as well as the remarkable efficacy of antitumour necrosis factor‐α biological treatments in psoriasis, forces us to refine the current T‐cell hypothesis of psoriasis pathogenesis, and to give credit to the role of innate immunity. Psoriasis might be envisioned to be a genetically determined triggered state of otherwise dormant innate immunity. This aggravated state of innate immunity is represented by the activity of NK T cells, dendritic cells, neutrophils and keratinocytes, leading to the recruitment and activation of preferentially type 1 T cells, possibly in an antigen‐independent way. Keratinocytes in psoriasis then are sensitive to the effects of T‐cell activation and cytokine production, interferon (IFN)‐γ, by responding with psoriasiform hyperplasia. The chronic inflammation of psoriatic lesions suggests that this might be due to a deficiency in downregulation processes (e.g. a defect in the regulatory T‐cell repertoire) and/or the persistence of an unknown trigger resulting in an exaggerated innate immune response.

Cutting Edge: Loss of TLR2, TLR4, and TLR5 on Langerhans Cells Abolishes Bacterial Recognition

It is unknown whether closely related epidermal dendritic cells, Langerhans cells (LCs), and dermal dendritic cells (DDCs) have unique functions. In this study, we show that human DDCs have a broad TLR expression profile, whereas human LCs have a selective impaired expression of cell surface TLR2, TLR4, and TLR5, all involved in bacterial recognition. This distinct TLR expression profile is acquired during the TGF-β1-driven development of LCs in vitro. Consequently, and in contrast to DDCs, LCs weakly respond to bacterial TLR2, TLR4, and TLR5 ligands in terms of cytokine production and maturation, as well as to whole Gram-positive and Gram-negative bacteria, whereas their responsiveness to viral TLR ligands and viruses is fully active and comparable to DDCs. Unresponsiveness of LCs to bacteria may be a mechanism that contributes to tolerance to bacterial commensals that colonize the skin.

Interleukin-17 in inflammatory skin disorders

Purpose of reviewRecently, a novel and unique subset of interleukin (IL)-17-producing CD4+ T helper (Th17) cells, distinct from Th1 and Th2 cells, was discovered. The question is addressed as to what extent inflammatory skin diseases are associated with the actions of this newly discovered Th17 cell subset. Recent findingsTh17 cells are involved in protection against bacterial pathogens. In addition, it is now clear that Th17 cells may also be crucial in the pathogenesis of various chronic inflammatory diseases that were formerly categorized as Th1-mediated disorders. SummaryIn this review, we summarize the current knowledge of IL-17 and Th17 cells and discuss the possible role of IL-17 in the pathology of psoriasis, contact hypersensitivity and atopic dermatitis. Whereas IL-17 may play an important role in the pathogenesis of psoriasis and contact hypersensitivity, its role in atopic dermatitis is still unclear.

Composition of Innate Lymphoid Cell Subsets in the Human Skin: Enrichment of NCR+ ILC3 in Lesional Skin and Blood of Psoriasis Patients

Innate lymphoid cells (ILCs) are increasingly appreciated as important regulators of tissue homeostasis and inflammation. However, their role in human skin remains obscure. We found that healthy peripheral blood CD117(+) ILC3, lacking the natural cytotoxicity receptor (NCR) NKp44 (NCR(-) ILC3), CD117(-)NCR(-)CRTH2(-)CD161(+) ILC1, and CRTH2(+) ILC2, express the skin-homing receptor cutaneous lymphocyte antigen (CLA). NCR(+) ILC3 were scarce in peripheral blood. Consistently, we identified in normal skin ILC2 and NCR(-) ILC3, a small proportion of CD161(+) ILC1, and hardly any NCR(+) ILC3, whereas NCR(+) ILC3 were present in cultured dermal explants. The skin ILC2 and NCR(+) ILC3 subsets produced IL-13 and IL-22, respectively, upon cytokine stimulation. Remarkably, dermal NCR(-) ILC3 converted to NCR(+) ILC3 upon culture in IL-1β plus IL-23, cytokines known to be involved in psoriatic inflammation. In line with this observation, significantly increased proportions of NCR(+) ILC3 were present in lesional skin and peripheral blood of psoriasis patients as compared with skin and blood of healthy individuals, respectively, whereas the proportions of ILC2 and CD161(+) ILC1 remained unchanged. NCR(+) ILC3 from skin and blood of psoriasis patients produced IL-22, which is regarded as a key driver of epidermal thickening, suggesting that NCR(+) ILC3 may participate in psoriasis pathology.

Deactivation of endothelium and reduction in angiogenesis in psoriatic skin and synovium by low dose infliximab therapy in combination with stable methotrexate therapy: a prospective single-centre study

Psoriasis and psoriatic arthritis are inflammatory diseases that respond well to anti-tumour necrosis factor-α therapy. To evaluate the effects of anti-tumour necrosis factor-α treatment on expression of adhesion molecules and angiogenesis in psoriatic lesional skin and synovial tissue, we performed a prospective single-centre study with infliximab therapy combined with stable methotrexate therapy. Eleven patients with both active psoriasis and psoriatic arthritis received infusions of infliximab (3 mg/kg) at baseline, and at weeks 2, 6, 14 and 22 in an open-label study. In addition, patients continued to receive stable methotrexate therapy in dosages ranging from 5 to 20 mg/week. Clinical assessments, including Psoriasis Area and Severity Index (PASI) and Disease Activity Score (DAS), were performed at baseline and every 2 weeks afterward. In addition, skin biopsies from a target psoriatic plaque and synovial tissue biopsies from a target joint were taken before treatment and at week 4. Immunohistochemical analysis was performed to detect the number of blood vessels, the expression of adhesion molecules and the presence of vascular growth factors. Stained sections were evaluated by digital image analysis. At week 16, the mean PASI was reduced from 12.3 ± 2.4 at baseline to 1.8 ± 0.4 (P ≤ 0.02). The mean DAS was reduced from 6.0 ± 0.5 to 3.6 ± 0.6 (P ≤ 0.02). We found some fluctuations in DAS response as compared with the change in PASI, with the latter exhibiting a steady decrease over time. After 4 weeks the cell infiltrate was reduced in both skin and synovium. There was a significant reduction in the number of blood vessels in dermis and synovium at week 4. A significant reduction in the expression of αvβ3 integrin, a marker of neovascularization, was also found in both skin and synovium at week 4. In addition, a significant reduction in the expression of adhesion molecules was observed in both skin and synovium at week 4. We also observed a trend toward reduced expression of vascular endothelial growth factor in both skin and synovium. In conclusion, low-dose infliximab treatment leads to decreased neoangiogenesis and deactivation of the endothelium, resulting in decreased cell infiltration and clinical improvement in psoriasis and psoriatic arthritis.

Clinical improvement in chronic plaque-type psoriasis lesions after narrow-band UVB therapy is accompanied by a decrease in the expression of IFN-gamma inducers -- IL-12, IL-18 and IL-23.

Abstract:  Type‐1 cytokine‐producing T cells are important in the pathogenesis of psoriasis vulgaris, for which efficient therapy is provided by means of narrow‐band ultraviolet‐B (NB‐UVB). The expression of the type‐1 cytokine interferon‐γ (IFN‐γ) is regulated by interleukin‐12 (IL‐12), IL‐15, IL‐18 and IL‐23; however, not much is known about the effect of this therapy on the levels of these cytokines in lesional psoriatic skin in situ. In this study, we investigated the effects of NB‐UVB therapy on the expression of IFN‐γ‐inducing cytokines. Ten patients with chronic plaque‐type psoriasis selected to be treated with NB‐UVB therapy were recruited for these experiments and the expression of cytokines IL‐12, IL‐15, IL‐18, IL‐23 and IFN‐γ in lesional psoriatic skin before, during and after therapy was determined with the help of immunohistochemistry. Double staining was performed in order to determine the cell types expressing these cytokines. The decrease in the psoriasis area and severity index was accompanied by a significant decrease in the expression of IFN‐γ, and concomitantly, significant reduction of IFN‐γinducers – IL‐12, IL‐18 and IL‐23. Thus, we concluded that the decrease of IFN‐γ expression in psoriasis lesions after NB‐UVB therapy could be a result of diminished expression of IL‐12, IL‐18 and IL‐23 in lesional skin. Therapies targeting these three cytokines should, therefore, be considered in the treatment of psoriasis.

Immunohistochemical Analysis of Regulatory T Cell Markers FOXP3 and GITR on CD4+CD25+ T Cells in Normal Skin and Inflammatory Dermatoses

Regulatory T cells (Treg) are a subset of T lymphocytes that play a central role in immunologic tolerance and in the termination of immune responses. The identification of these cells in normal and inflammatory conditions may contribute to a better understanding of underlying pathology. We investigated the expression of FOXP3 and GITR in normal skin and in a panel of different inflammatory dermatoses. Immunohistochemical double stainings in skin tissue sections revealed that FOXP3 and GITR were almost exclusively present on T cells that express both CD4 and CD25. Further, immunohistochemical double staining, as well as fluorescence-activated cell sorter analysis, on peripheral blood T cells showed that most FOXP3+ cells expressed GITR and vice versa, whereas a minority were single-positive for these markers. The mean frequency of FOXP3+ T cells in spongiotic dermatitis, psoriasis, and lichen planus was in the same range (25-29%), but the frequency of these cells in leishmaniasis appeared to be lower (∼15%), although this was not statistically significant. The mean frequency of GITR+ T cells was fairly similar in all conditions studied (14-20%). Normal human skin also contained FOXP3+ and GITR+ cells in the same frequency range as in diseased skin, but the absolute numbers were, of course, much lower. In conclusion, frequencies of FOXP3+ and GITR+ T cells were similar in all inflammatory skin diseases studied and normal skin, despite the well-known differences among the inflammatory conditions under investigation.

Increased frequencies of IL‐31‐producing T cells are found in chronic atopic dermatitis skin

Abstract:  Interleukin (IL)‐31 has been associated with pruritus, a characteristic feature of atopic dermatitis (AD). Local T cell responses may be responsible for the increased level of IL‐31 mRNA observed in AD. We investigated the frequency of IL‐31‐producing T cells in AD lesions, as well as their cytokine profile. T cells were isolated from chronic AD lesions, autologous blood and healthy donor skin. Intracellular expression of IL‐31, IFN‐γ, IL‐13, IL‐17 and IL‐22 was measured using flow cytometry. T cells from AD lesions contained significantly higher percentages of IL‐31‐producing T cells compared to autologous blood and donor skin. Many IL‐31‐producing T cells co‐produced IL‐13 and to lesser extent IL‐22, but rarely IFN‐γ or IL‐17. A substantial part of the IL‐31‐producing T cells did not co‐produce any of the other cytokines and could therefore not be linked to any of the known functionally different T cell subsets. The T cell infiltrates were also relatively enriched for Th2/Tc2 and Th22/Tc22 cells, while frequencies of Th1/Tc1 and Th17 cells were decreased. This is the first report describing the detection of IL‐31 at protein level in skin‐infiltrating T cells. We show here that T cells in chronic AD skin produce IL‐31 and that AD lesions contain increased levels of these IL‐31‐producing T cells. This suggests that a substantial part of previously reported increased IL‐31 mRNA levels in AD skin is T cell derived and that these cells may be involved in the pathogenesis of AD.