A chitin-like component on sclerotic cells of Fonsecaea pedrosoi inhibits Dectin-1-mediated murine Th17 development by masking ß-glucans.

Fonsecaea pedrosoi (F. pedrosoi), a major agent of chromoblastomycosis, has been shown to be recognized primarily by C-type lectin receptors (CLRs) in a murine model of chromoblastomycosis. Specifically, the ß-glucan receptor, Dectin-1, mediates Th17 development and consequent recruitment of neutrophils, and is evidenced to have the capacity to bind to saprophytic hyphae of F. pedrosoi in vitro. However, when embedded in tissue, most etiological agents of chromoblastomycosis including F. pedrosoi will transform into the sclerotic cells, which are linked to the greatest survival of melanized fungi in tissue. In this study, using immunocompetent and athymic (nu/nu) murine models infected subcutaneously or intraperitoneally with F. pedrosoi, we demonstrated that T lymphocytes play an active role in the resolution of localized footpad infection, and there existed a significantly decreased expression of Th17-defining transcription factor Rorγt and inefficient recruitment of neutrophils in chronically infected spleen where the inoculated mycelium of F. pedrosoi transformed into the sclerotic cells. We also found that Dectin-1-expressing histocytes and neutrophils participated in the enclosure of transformed sclerotic cells in the infectious foci. Furthermore, we induced the formation of sclerotic cells in vitro, and evidenced a significantly decreased binding capacity of human or murine-derived Dectin-1 to the induced sclerotic cells in comparison with the saprophytic mycelial forms. Our analysis of ß-glucans-masking components revealed that it is a chitin-like component, but not the mannose moiety on the sclerotic cells, that interferes with the binding of ß-glucans by human or murine Dectin-1. Notably, we demonstrated that although Dectin-1 contributed to the development of IL-17A-producing CD3+CD4+ murine splenocytes upon in vitro-stimulation by saprophytic F. pedrosoi, the masking effect of chitin components partly inhibited Dectin-1-mediated Th17 development upon in vitro-stimulation by induced sclerotic cells. Therefore, these findings extend our understanding of the chronicity of chromoblastomycosis.

Fonsecaea pedrosoi infection induces differential modulation of costimulatory molecules and cytokines in monocytes from patients with severe and mild forms of chromoblastomycosis.

The host defense mechanism in chromoblastomycosis has not been thoroughly investigated. It has been suggested that cell-mediated immunity in patients with long-standing chromoblastomycosis is somehow impaired. As a result, these individuals became unable to develop an efficient immune reaction. Many studies have shown that monocyte-derived macrophages exhibit critical activities in immunity to microorganisms. Moreover, the ability of cells from the monocytic lineage to process and present antigens, to produce cytokines, and to provide costimulatory signals confirms their pivotal role in the initiation of specific immune responses. In the present study, it was observed that monocytes from patients with a severe form of disease had a higher production of IL-10 and a lower expression of HLA-DR and costimulatory molecules when stimulated with specific antigen or LPS. Immune modulation with recombinant IL-12 or anti-IL-10 can restore the antigen-specific Th1-type immune response in chromoblastomycosis patients by up-regulating HLA-DR and costimulatory molecules in monocytes. Therefore, our data show that monocytes from patients with different clinical forms of chromoblastomycosis present distinct phenotypic and functional profiles. This observation suggests possible mechanisms that control the T cell response and influence their role in the development of pathology.

Cryptic "chromo-fibroma".

Chromoblastomycosis is an uncommon chronic fungal infection capable of presenting in a variety of clinical guises. Herein, we present the histopathological features of an unusual dermal response engendered by this organism, consisting of dermal effacement by a spindle cell proliferation arranged in sweeping fascicles.

Phagocytosis, production of nitric oxide and pro-inflammatory cytokines by macrophages in the presence of dematiaceous [correction of dematiaceus] fungi that cause chromoblastomycosis.

Chromoblastomycosis is characterized by the slow development of polymorphic skin lesions (nodules, verrucas, tumores, plaques and scar tissue). Inside the host, infectious propagules adhere to epithelial cells and differentiate into sclerotic forms, which effectively resist destruction by host effector cells and allow onset of chronic disease. A cellular immune response against fungi is essential to control infection. Amongst the cells of the immune system, macrophages play the most important role in controlling fungal growth. In this study, we show that the fungicidal characteristic of macrophages is dependent on the fungal species that causes chromoblastomycosis. We began by observing that the phagocytic index was higher for Fonsecaea pedrosoi and Rhinocladiella aquaspersa compared with that of other fungi. Complement-mediated phagocytosis was more important for Phialophora verrucosa and R. aquaspersa and was inhibited by mannan when F. pedrosoi and R. aquaspersa conidia were phagocytosed by macrophages. We showed that macrophages killed significantly only R. aquaspersa. We also found that the phagocytosis of fungi has functional consequences for macrophages as phagocytosis resulted in down-modulation of MHC-II and CD80 expression as well as in the inhibition of the basal liberation of NO. However, the inhibition of the basal liberation of NO nor the down-modulation of MHC and co-stimulatory molecules were observed in the presence of R. aquaspersa.

Monitoring of extracellular matrix metabolism and cross-linking in tissue, serum and urine of patients with chromoblastomycosis, a chronic skin fibrosis.

BACKGROUND: Chromoblastomycosis is a fungal disease leading to a granulomatous reaction associated with dermal fibrosis. METHODS: In an attempt to elucidate the mechanisms leading to improvement in the cutaneous lesions after treatment with terbinafine, a new antifungal drug, we analysed collagen content and cross-linking before and at the end of the treatment. The turnover of extracellular matrix was monitored for 1 year by following up serum and urinary metabolites. RESULTS: The serum levels of type III collagen and its N-terminal propeptide were correlated with the lesion size (P < 0.035) after 4 and 12 months of treatment respectively. After 4 months of treatment, urinary pyridinoline was higher (P = 0.04) in patients whose lesion size was reduced by more than 50% and serum hyaluronan was lower in patients who had lesions active for less than 5 years (P < 0.05). The treatment increased pyridinoline and pentosidine cross-links in the lesions but significantly reduced the collagen content (P = 0.05). CONCLUSION: This is the first demonstration that, in addition to its fungicidal activity, terbinafine acts in vivo as an antifibrotic drug.

Immunohistochemical study of type I collagen turn-over and of matrix metalloproteinases in chromoblastomycosis before and after treatment by terbinafine.

The distribution of type I collagen, the major component of human dermis, was characterized by immunohistochemistry in skin lesions of chromoblastomycosis, a chronic cutaneous mycosis, before and after a specific antifungal treatment with terbinafine to study the changes induced in the lesions by the treatment. Newly synthesized type I collagen was studied with an antibody directed against the aminoterminal propeptide of the molecule (PINP), whereas mature, cross-linked type I collagen was detected with an antibody against the carboxyterminal telopeptide of type I collagen (ICTP). The isopeptide N epsilon gamma-glutamyl lysine (N epsilon gamma GL), synthesized by transglutaminase and able to cross-link several components of the extracellular matrix, has also been investigated with two monoclonal antibodies to determine if it is involved in the stabilisation of the fibrotic cutaneous lesions. The degradative process involved in the remodelling has also been assessed by immunohistochemistry with anti-metalloproteinase (MMP-1 and MMP-9) and anti-tissue inhibitor (TIMP-1) antibodies. All tissue macrophages stained for CD68 and MMP-9, but not for MMP-1, while the polymorphonuclear neutrophils had an elastase and a weak MMP-9 phenotype. The fibroblasts of fibrotic areas stained constantly for N epsilon gamma GL and PINP. The immunostaining of extracellular matrix for ICTP and N epsilon gamma GL, and the number of PINP-positive fibroblasts, decreased significantly after one year of antifungal treatment. Terbinafine treatment decreases the synthesis of type I collagen and leads to a partial reversal of the cutaneous fibrotic lesions, independently of the cure of the fungal infection.

Expression of heat shock protein 27 in chromomycosis.

We report on a 58-year-old woman with long-lasting (36 years) chromomycosis on the foot and secondary self-inoculation from foot to hand 4 years ago. Mycological classification was performed after culture on Sabouraud glucose agar. We used haematoxylin and eosin and Giemsa staining and an antibody to heat shock protein (HSP) 27 (Stress Gen, Clone G3.1) on paraffin-embedded and cryostat specimens of chromomycosis. The mycological culture revealed the fungus Fonsecaea pedosoi. Histopathology revealed dermal fibrosis with persistent fungi (Medlar bodies), numerous mast cells and pseudoepitheliomatous hyperplasia. Immunohistochemically, HSP 27 was positively identified in F. pedrosoi. Moreover, in differentiating keratinocytes in the pseudoepitheliomatous lesions of chromomycosis, HSP 27 was increasingly expressed from basal layers to stratum spinosum in the epidermis but not in keratinocytes directly bordering Medlar bodies. In chromomycosis, HSP 27 is expressed, in accordance with its role as a marker of differentiation and proliferation, in keratinocytes and also in F. pedrosoi. It remains unknown if these results might explain the therapeutic efficacy of hyperthermic treatment.

Collagen cross-linking by pyridinoline occurs in non-reversible skin fibrosis.

The extent of the covalent cross-linking of collagen molecules by pyridinoline was measured in skin lesions from patients with chromomycosis, a chronic fungal infection leading to an extensive and chronic dermal fibrosis. These data were compared to those collected from patients with a localized cutaneous leishmaniasis, an acute inflammatory process leading to an extensive and reversible remodelling of the extracellular matrix. The amount of the mature cross-linking amino acid pyridinoline increased in chromomycosis patients when compared to controls and was significantly higher than in leishmaniasis patients. These data confirm and extend our previous studies on liver fibrosis showing that a high level of pyridinoline is associated to irreversible fibrotic lesions. They also suggest that an increase in the mature collagen cross-linking by pyridinoline in the course of fibrosis is not restricted to liver, but might be a general feature of irreversible and chronic fibrosis.