Immunohistochemical demonstration of TGF-ß and decorin in paracoccidioidal granulomas

Abstract

Different patterns of granulomas have been observed in 6-to 8-weekold mice after ip inoculation with 5 x 10 6 yeast cells of Paracoccidioides brasiliensis. Transforming growth factor-ß (TGF-ß) is a cytokine that has been shown to participate in fibrosis and granuloma formation; its activities seem to be modulated by the small proteoglycan decorin. In the present study, TGF-ß and decorin expression in epiploon granulomas was assessed by immunohistochemistry in susceptible (B10.A) and resistant (A/J) mice after 15, 30, 120 and 150 days of P. brasiliensis ip infection. The epiploon was collected, fixed in Methacarn solution and embedded in paraffin, and 5-µm thick sections were used for immunohistochemical analysis employing the streptavidin-biotin-peroxidase technique. The former mouse strain developed fatal disease with many disseminated lesions increasing in size and number during the infection and the latter developed mild disease with the presence of encapsulated granulomas. In the epiploon, TGF-ß was present on macrophages, giant cells, lymphocytes and fibroblasts, and absent on neutrophils. It was also detected in areas of fibrosis and necrosis, as well as disperse in amorphous extracellular matrix, mostly in resistant mice. Decorin was present circumscribing macrophages and giant cells containing fungi, but absent on these cells. In both mouse strains, decorin was found at the periphery of the lesions, and markedly in milky spot granulomas. In resistant mice, positivity was found around fibrotic and necrotic areas of encapsulated and residual lesions containing lysed fungi. Decorin was found associated with thick fibers around encapsulated lesions. In susceptible mice, the size and number of lesions increased with the progression of the disease and were correlated with the weaker expression of decorin. We suggest an association of decorin with the fibrogenic process observed in paracoccidioidal granulomas.

TGF-β1 induces transendothelial migration of the pathogenic fungus Sporothrix schenckii by a paracellular route involving extracellular matrix proteins

“…Studies performed with mice suggested a role for TGF-b1 in determining a protective response against Candida albicans, as demonstrated by the enhancement of TGF-b1 production by resistant infected animals (Spaccapelo et al, 1995). However, the local production of this cytokine was demonstrated in liver granulomas of human candidiasis patients (Letterio et al, 2001) and in omental granulomas of mice infected with Paracoccidioides brasiliensis (Nishikaku & Burger, 2003).…”

“…It has been suggested that the enhanced expression of TGFb1 in fungi-infected granuloma (Spaccapelo et al, 1995;Letterio et al, 2001;Nishikaku & Burger, 2003) could favour residence inside host cells by inhibiting immune responses, as already described for other pathogens (Reed, 1999). However, the present work reinforces another possible contribution of this cytokine, by stimulating the association of a pathogenic fungus, such as S. schenckii, with host tissue stroma and, thus, providing additional clues that implicate FN-and LM-recognizing cytoadhesins as virulence factors for this pathogenic fungus. )…”

Immunolocalization of IFN-gamma in the lesions of resistant and susceptible mice to Paracoccidioides brasiliensis infection

“…In our experimental model of P. brasiliensis infection, we could detect distinct patterns of ECM components using immunohistochemical reactions (Xidieh et al, 1999;Nishikaku & Burger, 2003a, b), the presence of some cytokines (Nishikaku & Burger, 2003a;Nishikaku et al, 2008) and of proteolytic enzymes (Nishikaku et al, 2009a) at the lesions of infected mouse strains.…”

“…The peripancreatic/perisplenic omentum, the target organ of ip P. brasiliensis infection, (Xidieh et al, 1999;Nishikaku & Burger, 2003c) was collected and fixed in Methacarn solution (60% methanol, 30% chloroform, and 10% acetic acid) for 3-4 h in a shaker at 4°C. Tissues were embedded in paraffin, and 5 µm sections were used for histologic and immunohistochemical procedures according to Nishikaku & Burger (2003a).…”

“…The immunohistochemical reactions were done according to the protocol described previously (Nishikaku & Burger, 2003a;Nishikaku et al, 2008). In brief, slides with deparaffinized tissue sections were incubated overnight at 4°C with anti-mouse IFN-c mAb (hybridoma XMG 1.2, dilution in PBS -0.3% Tween 20).…”