Emiliano Antiga

Expression of cytokines, chemokines and other effector molecules in two prototypic autoinflammatory skin diseases, pyoderma gangrenosum and sweet's syndrome

Pyoderma gangrenosum (PG) and Sweets syndrome (SS) are two inflammatory skin diseases presenting with painful ulcers and erythematous plaques, respectively; both disorders have a debilitating clinical behaviour and PG is potentially life‐threatening. Recently, PG and SS have been included among the autoinflammatory diseases, which are characterized by recurrent episodes of sterile inflammation, without circulating autoantibodies and autoreactive T cells. However, an autoinflammatory pattern clearly supporting this inclusion has never been demonstrated. We studied 16 patients with PG, six with SS and six controls, evaluating, using a sandwich‐based protein antibody array method, the expression profile of inflammatory effector molecules in PG, SS and normal skin. The expressions of interleukin (IL)‐1 beta and its receptor I were significantly higher in PG (P = 0·0001 for both) and SS (P = 0·004–0·040) than in controls. In PG, chemokines such as IL‐8 (P = 0·0001), chemokine (C‐X‐C motif) ligand (CXCL) 1/2/3 (P = 0·002), CXCL 16 (P = 0·003) and regulated upon activation normal T cell expressed and secreted (RANTES) (P = 0·005) were over‐expressed. In SS, IL‐8 (P = 0·018), CXCL 1/2/3 (P = 0·006) and CXCL 16 (P = 0·036) but not RANTES were over‐expressed, suggesting that chemokine‐mediated signals are lower than in PG. Fas/Fas ligand and CD40/CD40 ligand systems were over‐expressed in PG (P = 0·0001 for Fas, P = 0·009 for Fas ligand, P = 0·012 for CD40, P = 0·0001 for CD40 ligand), contributing to tissue damage and inflammation, while their role seems to be less significant in SS. Over‐expression of cytokines/chemokines and molecules amplifying the inflammatory network supports the view that PG and SS are autoinflammatory diseases. The differences in expression profile of inflammatory effectors between these two disorders may explain the stronger local aggressiveness in PG than SS.

Guidelines for the diagnosis and treatment of dermatitis herpetiformis

Dermatitis herpetiformis is a rare disease that should be considered the cutaneous expression of a gluten‐sensitive enteropathy indistinguishable from celiac disease. Dermatitis herpetiformis is often misdiagnosed and to date no guidelines for the management of dermatitis herpetiformis have been published in Literature. The present guidelines have been prepared for dermatologists by the Group for Cutaneous Immunopathology of the Italian Society of Dermatology and Venereology. They reflect the best data available at the time of preparation and the clinical experience of the authors and the members of the Italian Group for Cutaneous Immunopathology. The diagnosis of dermatitis herpetiformis is established clinically, histologically, immunopathologically and serologically. A gluten‐free diet (GFD) is the treatment of choice for patients with dermatitis herpetiformis. Dapsone and/or other drugs should be used during the period until the GFD is effective. In conclusion, the present guidelines provide evidence‐based guidance for the diagnosis and treatment of dermatitis herpetiformis.

Regulatory T cells in the skin lesions and blood of patients with systemic sclerosis and morphoea

Background  Systemic sclerosis (SSc) and morphoea are connective tissue diseases characterized by fibrosis of the skin. Although to date their pathogenesis has not been clearly defined, it is thought that autoimmunity may play a role in the development of the skin lesions observed in both these diseases. As regulatory T cells (Tregs) play a key role in the modulation of immune responses, it has recently been suggested that Treg impairment may lead to the development of autoimmune diseases.

Cutaneous lupus erythematosus: First multicenter database analysis of 1002 patients from the European Society of Cutaneous Lupus Erythematosus (EUSCLE)

In this prospective, cross-sectional, multicenter study, we assessed clinical and laboratory characteristics from patients with cutaneous lupus erythematosus (CLE) using the Core Set Questionnaire of the European Society of Cutaneous Lupus Erythematosus (EUSCLE). 1002 (768 females, 234 males) patients with different subtypes of CLE, such as acute CLE (ACLE, 304 patients), subacute CLE (SCLE, 236 patients), chronic CLE (CCLE, 397 patients), and intermittent CLE (ICLE, 65 patients), from 13 European countries were collected and statistically analyzed by an SPSS database. The main outcome measures included gender, age at onset of disease, LE-specific and LE-nonspecific skin lesions, photosensitivity, laboratory features, and the criteria of the American College of Rheumatology (ACR) for the classification of systemic lupus erythematosus. The mean age at onset of disease was 43.0±15.7 years and differed significantly between the CLE subtypes. In 347 (34.6%) of the 1002 patients, two or more CLE subtypes were diagnosed during the course of the disease and 453 (45.2%) presented with LE-nonspecific manifestations. Drug-induced CLE and Sjögrens Syndrome had the highest prevalence in SCLE patients (13.1% and 14.0%, respectively). Photosensitivity was significantly more frequent in patients with ACLE, SCLE, and ICLE compared with those with CCLE. The detection of antinuclear antibodies such as anti-Ro/SSA and anti-La/SSB antibodies revealed further significant differences between the CLE subtypes. In summary, the EUSCLE Core Set Questionnaire and its database facilitate the analysis of clinical and laboratory features in a high number of patients with CLE and will contribute to standardized assessment and monitoring of the disease in Europe.

Chronic Idiopathic and Chronic Autoimmune Urticaria: Clinical and Immunopathological Features of 68 Subjects

Skin tests with autologous serum elicit an immediate wheal-and-flare response in about 30-50% of chronic idiopathic urticaria subjects, suggesting that an autoimmune mechanism might be involved in the pathogenesis of this disease. The aim of the present work, involving 68 subjects with chronic idiopathic urticaria, was to distinguish between the serum-positive and serum-negative cases and highlight the clinical differences between the two groups on the basis of the Breneman scale score. We also tried to correlate the finding of a positive response to the autologous serum skin test with other autoimmune diatheses or fully developed autoimmune disorders. Our results did not demonstrate any significant differences between the two groups with regard to mean age, sex distribution, angioedema and mucosal/cutaneous atopy. However, all subjects with positive autologous serum skin test presented more severe clinical features than serum-negative subjects. We found no differences between the two groups in the incidence of autoimmune disease.

The effects of tacrolimus ointment on regulatory T lymphocytes in atopic dermatitis.

Only very recently studies were conducted in order to evaluate the impact of regulatory T (Treg) cells in the pathophysiology of atopic dermatitis (AD). Nine adult patients with moderate-to-severe AD in riacutization period of a chronic disease were given tacrolimus ointment, while seven hydrocortisone butyrate ointment, that served as controls. We performed lesional-skin biopsies before and after treatment, that were stained immunohistochemically with monoclonal antibodies to CD4, CD25, forkhead/winged helix transcription factor (FoxP3), interleukin (IL)-10 and transforming growth factor (TGF)-β. CD4+ cells were significantly reduced in post-treatment series. Tacrolimus treatment achieved a significant reduction of CD25+ cells. FoxP3+ cells were present in untreated AD lesions. Both treatments did not significantly modify the number of FoxP3+ cells. The number of IL-10+ cells increased in post-treatment series. Tacrolimus enhanced the production of TGF-β, while hydrocortisone did not.Restoration of TGF-β-producing Treg cells may represent another important pharmacodynamic effect of tacrolimus on AD.

The comparative effects of tacrolimus and hydrocortisone in adult atopic dermatitis: an immunohistochemical study

Background  While many studies have demonstrated the efficacy and safety of tacrolimus ointment in the treatment of atopic dermatitis (AD), only a few have investigated the effects of tacrolimus on inflammatory cells and their cytokine gene expression in patients with AD.

The Treg/Th17 cell ratio is reduced in the skin lesions of patients with pyoderma gangrenosum

DEAR EDITOR, Pyoderma gangrenosum (PG) is a rare, inflammatory skin disease that, together with other conditions such as Sweet syndrome (SS), is included within the group of neutrophilic dermatoses. Although its pathogenesis remains poorly understood, the treatments with the best clinical evidence are tumour necrosis factor inhibitors, high-dose systemic corticosteroids and ciclosporin, suggesting the pivotal role of inflammatory pathways in the development of PG. Interestingly, recent studies have highlighted the role of T helper (Th)17 cells in neutrophilic dermatoses, and an increase of interleukin (IL)-17 and IL-23 expression was found in PG lesions. Together with Th17 cells, regulatory T cells (Tregs) play a major role in human disease. Accordingly, recent reports suggest that controlling the balance between Tregs and Th17 cells may be a promising therapeutic strategy for inflammatory diseases. However, no data are present in the literature about Tregs in PG. In this study, we investigated the proportions of Tregs and Th17 cells in the skin of 15 patients with PG (seven male, eight female; age range 27–69 years), on neither immunosuppressive treatment nor topical steroids for at least 4 weeks prior to entering the study. Their clinical findings are presented in Table S1 (see Supporting Information). As a control, skin samples from five patients with SS (two male, three female; age range 31–64 years) and six healthy subjects (HS) (three male, three female; age range 28–59 years) were collected. The trial was approved by the local ethics committee and conducted according to the Declaration of Helsinki. All of the patients and controls provided written informed consent. Treg and Th17 markers were analysed by immunohistochemistry using the monoclonal antibodies anti-CD4 (1 : 20; Dako, Copenhagen, Denmark), anti-CD25 (1 : 25; Histo-Line Laboratories, Milan, Italy), anti-CD161 (1 : 80; AbD Serotec, Oxford, U.K.), antiforkhead box protein P3 (anti-FOXP3) (1 : 80; Abcam, Cambridge, U.K.), anti-IL-10 (1 : 300; Dako), anti-IL-17 (1 : 1000; Abcam), anti-RORct (1 : 2000; R&D Systems, Minneapolis, MN, U.S.A.) and antitransforming growth factor (anti-TGF)-b1 (1 : 2000; Abcam), as described previously. The results of quantitative analysis of these markers are described in Table S2 (see Supporting Information). The stained cells were counted in three consecutive microscopic fields (4009). Furthermore, FOXP3/CD4, TGF-b/CD4, IL-10/CD4 and RORct/CD4 cell ratios were calculated. The results were analysed with the Mann– Whitney U-test and were considered significant with a P-value < 0 05. In PG and SS, CD4 and CD25 cells were located in the whole dermis, with some cells scattered in the epidermis (Fig. 1a,b,d,e). The number of CD4 cells in PG was significantly higher than in SS (P < 0 001), while no differences were found for CD25 cells. By contrast, CD4 and CD25 cells were significantly less represented in HS than in the other two groups (P < 0 001) (Fig. 2a). Some FOXP3 cells were detected within the superficial dermis of patients with PG (Fig. 1g); their number was higher than in HS (P = 0 004) but lower than in SS (P < 0 001) (Fig. 2a). Interestingly, the FOXP3/CD4 cell ratio was significantly lower in PG than in SS (P < 0 001) and HS (P < 0 001) (Fig. 2a). Some IL-10 cells were found in the superficial dermis of patients with PG (Fig. 1j); their number was significantly lower than in SS (P < 0 001) and higher than in HS (P < 0 001). Moreover, the IL-10/CD4 cell ratio was reduced in PG vs. SS and HS (P < 0 001 and P = 0 03, respectively) (Fig. 2a). TGF-b staining was diffusely distributed within the superficial and medium dermis in PG and SS. Moreover, some TGF-b cells could be detected in the same areas (Fig. 1m,n). Their number was similar in both PG and SS. By contrast, the TGF-b/CD4 cell ratio was significantly reduced in PG (P = 0 01). Moreover, although HS showed a lower number of TGF-b cells than PG (P < 0 001) and SS (P < 0 001), their TGF-b/CD4 cell ratio was significantly higher (HS vs. PG, P = 0 001; HS vs. SS, P < 0 001) (Fig. 2a). Regarding Th17 markers, RORct cells were distributed in the upper dermis in PG (Fig. 1p); their number was similar to that found in SS. By contrast, the RORct/CD4 ratio was significantly lower in PG than in SS (P = 0 008) (Fig. 2a). As expected, no RORct expression was found in HS. The numbers of both CD161 and IL-17 cells, which were distributed predominantly in the superficial and medium dermis (Fig. 1s, t,v,w), were similar in PG and SS, while no CD161 nor IL-17 expression was detected in HS (Fig. 2a). Finally, in order to quantify the balance between Tregs and Th17 cells, we calculated the ratio between FOXP3 and RORct, which was significantly lower in PG than in SS (P < 0 001) (Fig. 2b).

Regulatory T cells in skin lesions and blood of patients with bullous pemphigoid

Although regulatory T cells (Tregs) are affected in several autoimmune skin diseases, only two studies have been performed in patients with bullous pemphigoid (BP) with contrasting results.

Serum levels of the Th1 promoter IL-12 and the Th2 chemokine TARC are elevated in erythema multiforme and Stevens-Johnson syndrome/toxic epidermal necrolysis and correlate with soluble Fas ligand expression. An immunoenzymatic study from the Italian Group of Immunopathology.

Background: No data exist as to Th2 chemokines in erythema multiforme (EM) and Stevens-Johnson syndrome (SJS)/toxic epidermal necrolysis (TEN). Objective: To evaluate thymus- and activation-regulated chemokine (TARC), macrophage-derived chemokine (MDC) and regulated upon activation, normal T-lymphocyte-expressed and secreted chemokine (RANTES) expression in EM and SJS/TEN and to correlate with the serum levels of the Th1 promoter interleukin (IL)-12 and soluble Fas ligand (sFasL). Materials and Methods: IL-12, sFasL, TARC, MDC and RANTES expression were analyzed by ELISA techniques in 31 untreated EM (n = 24) or SJS/TEN (n = 7) patients and in 28 healthy donors (HD). Results: EM and SJS/TEN exhibited significantly higher levels of TARC, IL-12 and sFasL with respect to HD. TARC upregulation paralleled both the IL-12 (p = 0.0225) and sFasL increase (p = 0.0194). Conclusions: Our results support a role of TARC in the pathophysiology of EM/SJS/TEN and confirm the coexistence of a Th2 response in addition to the predominant Th1 profile.