Inflammasome gene expression is associated with immunopathology in human localized cutaneous leishmaniasis.

Localized cutaneous leishmaniasis (LCL) can ultimately progress to chronic ulcerated lesions with strong local inflammatory reactions. The functional role of certain inflammasomes in mediating inflammation caused by Leishmania braziliensis needs to be addressed. By combining PCR-array, quantitative real-time PCR and immunohistochemical analysis, we identified inflammasome genes, such as IL-1ß, NLRP3, NLRP1, NLRC5, AIM2 and P2RX7, that were upregulated in LCL patients. Temporal gene expression studies showed that the early phase of LCL displayed increased NLRP3 and reduced AIM2 and NLRP1 expression, while the late stages showed increased AIM2 and NLRP1 and lower NLRP3 expression. Our findings also showed that AIM2, NLRP1, and P2RX7 promoted susceptibility to experimental L. braziliensis infection. These results highlight the importance of inflammasome machinery in human LCL and suggest that inflammasome machinery plays a role in the acute and chronic phases of the disease.

Biological predictors shared by dementia and bullous pemphigoid patients point out a cross-antigenicity between BP180/BP230 brain and skin isoforms.

Bullous pemphigoid (BP) following dementia diagnosis has been reported in the elderly. Skin and brain tissues express BP180 and BP230 isoforms. Dementia has been associated with rs6265 (Val66Met) polymorphism of the brain-derived neurotrophic factor (BDNF) gene and low serum BDNF. Here we investigated a possible cross-antigenicity between BP180/BP230 brain and skin isoforms. We assessed antibodies against BP180/BP230 and BDNF levels by ELISA and BDNF Val66Met SNP by PCR in three groups: 50 BP patients, 50 patients with dementia, and 50 elderly controls. Heatmap hierarchical clustering and data mining decision tree were used to analyze the patients' demographic and laboratorial data as predictors of dementia-BP association. Sixteen percent of BP patients with the lowest serological BDNF presented dementia-BP clinical association. Anti-BP180/230 positivity was unexpected observed among dementia patients (10%, 10%) and controls (14%, 1%). Indirect immunofluorescence using healthy human skin showed a BP pattern in two of 10 samples containing antibodies against BP180/BP230 obtained from dementia group but not in the control samples. Neither allelic nor genotypic BDNF Val66Met SNP was associated with dementia or with BP (associated or not with clinical manifestation of dementia). Heatmap analysis was able to differentiate the three studied groups and confirmed the ELISA results. The comprehensive data mining analysis revealed that BP patients and dementia patients shared biological predictors that justified the dementia-BP association. Autoantibodies against the BP180/BP230 brain isoforms produced by dementia patients could cross-react with the BP180/BP230 skin isoforms, which could justify cases of dementia preceding the BP disease.

Anti-Desmoglein 1 and 3 Autoantibody Levels in Endemic Pemphigus Foliaceus and Pemphigus Vulgaris from Brazil.

BACKGROUND: Pemphigus is a group of autoimmune blistering diseases of which the major forms are pemphigus foliaceus (PF) and vulgaris (PV). In Brazil, PF occurs in an endemic form also known as fogo selvagem. The main autoantibody in PF is against desmoglein 1 (DSG1), while in PV the main antibody is anti-desmoglein 3 (DSG3), but often anti-DSG1 is also present. The aim of the present study was to analyze the levels of anti-DSG1 and antiDSG3 autoantibodies in Brazilian PF and PV patients, considering different stages of the disease for PF patients and comparing these levels to those of healthy individuals living in and outside the endemic regions. METHODS: Levels of anti-DSG1 and anti-DSG3 were measured in the sera of Brazilian PF (n = 68) and PV (n = 20) patients as well as in clinically healthy (control) individuals (n = 48) by Enzyme Linked Immunosorbent Assay (ELISA). Comparisons were made using Kruskal-Wallis and Mann-Whitney tests. RESULTS: As expected, anti-DSG1 was more prevalent among PF patients (84% against 43% in PV), while antiDSG3 was more prevalent in PV patients (50% against 4% in PF). Levels of anti-DSG1 in PF patients in remission differed from those in patients undergoing active disease (p = 0.003), and patients in long-term remission (more than two years without presenting new lesions) were similar to control individuals living in the endemic region and surrounding area (p = 0.09). Moreover, patients with a more severe form of the disease had higher levels of anti-DSG1 (at least 134 U/mL, mean of 233 U/mL) than patients with a less severe form (fewer lesions) (mean of 193 U/mL, including two negative individuals). CONCLUSIONS: Despite the importance of these antibodies for diagnosis and management purposes, their presence in a healthy individual and in patients under remission indicates that caution should be taken when using anti-DSG for diagnosis, especially in endemic areas.

Genome-wide gene expression profiling reveals unsuspected molecular alterations in pemphigus foliaceus.

Pemphigus foliaceus (PF) is a complex autoimmune disease characterized by bullous skin lesions and the presence of antibodies against desmoglein 1. In this study we sought to contribute to a better understanding of the molecular processes in endemic PF, as the identification of factors that participate in the pathogenesis is a prerequisite for understanding its biological basis and may lead to novel therapeutic interventions. CD4+ T lymphocytes are central to the development of the disease. Therefore, we compared genome-wide gene expression profiles of peripheral CD4+ T cells of various PF patient subgroups with each other and with that of healthy individuals. The patient sample was subdivided into three groups: untreated patients with the generalized form of the disease, patients submitted to immunosuppressive treatment, and patients with the localized form of the disease. Comparisons between different subgroups resulted in 135, 54 and 64 genes differentially expressed. These genes are mainly related to lymphocyte adhesion and migration, apoptosis, cellular proliferation, cytotoxicity and antigen presentation. Several of these genes were differentially expressed when comparing lesional and uninvolved skin from the same patient. The chromosomal regions 19q13 and 12p13 concentrate differentially expressed genes and are candidate regions for PF susceptibility genes and disease markers. Our results reveal genes involved in disease severity, potential therapeutic targets and previously unsuspected processes involved in the pathogenesis. Besides, this study adds original information that will contribute to the understanding of PF's pathogenesis and of the still poorly defined in vivo functions of most of these genes.

Characterization of regulatory T cell (Treg) function in patients infected with Leishmania braziliensis.

Th1 immune responses are crucial for eliminating Leishmania parasites. However, despite strong Th1 responses, cutaneous leishmaniasis (CL) patients infected with Leishmania braziliensis develop the disease, while milder Th1 responses are found in sub-clinical (SC) infections. Therefore, CL patients may experience impaired regulatory T cell (Treg) function, causing excessive Th1 responses and tissue damage. To address this hypothesis, we characterized the function of circulating Tregs in L. braziliensis infected CL patients and compared them to Tregs from uninfected controls (UC) and SC subjects. The frequency of circulating Tregs was similar in CL patients, UC and SC subjects. Moreover, CL patients Tregs suppressed lymphocyte proliferation and PBMC pro-inflammatory cytokine production more efficiently than UC Tregs, and also produced higher levels of IL-10 than UC and SC Tregs. Furthermore, PBMC and mononuclear cells from lesions of CL patients responded normally to Treg-induced suppression. Therefore, the lesion development in CL patients infected with L. braziliensis is not associated with impairment in Treg function or failure of cells to respond to immunomodulation. Rather, the increased Treg activation in CL patients may impair parasite elimination, resulting in establishment of chronic infection. Thus, immunological strategies that interfere with this response may improve leishmaniasis treatment.

Chemokines and chemokine receptors coordinate the inflammatory immune response in human cutaneous leishmaniasis.

Cutaneous leishmaniasis (CL) includes different clinical manifestations displaying diverse intensities of dermal inflammatory infiltrate. Diffuse CL (DCL) cases are hyporesponsive, and lesions show very few lymphocytes and a predominance of macrophages. In contrast, localized CL (LCL) cases are responsive to leishmanial antigen, and lesions exhibit granulocytes and mononuclear cell infiltration in the early phases, changing to a pattern with numerous lymphocytes and macrophages later in the lesion. Therefore, different chemokines may affect the predominance of cell infiltration in distinct clinical manifestations. In lesions from LCL patients, we examined by flow cytometry the presence of different chemokines and their receptors in T cells, and we verified a higher expression of CXCR3 in the early stages of LCL (less than 30 days of infection) and a higher expression of CCR4 in the late stages of disease (more than 60 days of infection). We also observed a higher frequency of T cells producing IL-10 in the late stage of LCL. Using immunohistochemistry, we observed a higher expression of CCL7, CCL17 in lesions from late LCL, as well as CCR4 suggesting a preferential recruitment of regulatory T cells in the late LCL. Comparing lesions from LCL and DCL patients, we observed a higher frequency of CCL7 in DCL lesions. These results point out the importance of the chemokines, defining the different types of cells recruited to the site of the infection, which could be related to the outcome of infection as well as the clinical form observed.

Color image processing and content-based image retrieval techniques for the analysis of dermatological lesions.

This paper presents color image processing methods for the analysis of dermatological images in the context of a content-based image retrieval (CBIR) system. Tests were conducted on the classification of tissue components in skin lesions, in terms of necrotic tissue, fibrin, granulation, and mixed composition. The images were classified based on color components by an expert dermatologist following a black-yellow-red model. Indexing and retrieval of images were performed based on texture information obtained from the red, green, blue, hue, and saturation components of the color images. The performance of the CBIR system was measured in terms of precision and recall. Initial results demonstrate the potential of the proposed methods with the best precision result of 70% obtained for the characterization of mixed tissue composition.

CD4+CD25+ T cells in skin lesions of patients with cutaneous leishmaniasis exhibit phenotypic and functional characteristics of natural regulatory T cells.

Endogenous regulatory T (Treg) cells are involved in the control of infections, including Leishmania infection in mice. Leishmania viannia braziliensis is the main etiologic agent of cutaneous leishmaniasis (CL) in Brazil, and it is also responsible for the more severe mucocutaneous form. Here, we investigated the possible involvement of Treg cells in the control of the immune response in human skin lesions caused by L. viannia braziliensis infection. We show that functional Treg cells can be found in skin lesions of patients with CL. These cells express phenotypic markers of Treg cells--such as CD25, cytotoxic T lymphocyte-associated antigen 4, Foxp3, and glucocorticoid-induced tumor necrosis factor receptor--and are able to produce large amounts of interleukin-10 and transforming growth factor- beta . Furthermore, CD4+CD25+ T cells derived from the skin lesions of 4 of 6 patients with CL significantly suppressed in vitro the phytohemagglutinin-induced proliferative T cell responses of allogeneic peripheral-blood mononuclear cells (PBMCs) from healthy control subjects at a ratio of 1 Treg cell to 10 allogeneic PBMCs. These findings suggest that functional Treg cells accumulate at sites of Leishmania infection in humans and possibly contribute to the local control of effector T cell functions.