Virulence factors of Paracoccidioides brasiliensis as therapeutic targets: a review.

Paracoccidiodomycosis (PCM) is a systemic mycosis caused by the fungus Paracoccidioides brasiliensis and Paracoccidioides lutzii. The disease requires long and complicated treatment. The aim of this review is to address the fungal virulence factors that could be the target of the development of new drugs for PCM treatment. Virulence factors favoring the process of fungal infection and pathogenicity are considered as a microbial attribute associated with host susceptibility. P. brasiliensis has some known virulence factors which are 43 kDa glycoprotein (gp 43) which is an important fungal antigen, 70 kDa glycoprotein (gp 70), the carbohydrates constituting the fungal cell wall α-1,3, glucan and ß-1,3-glucan, cell adhesion molecules and the presence of melanin pigments. The discovery and development of drugs that interact with these factors, such as inhibitors of ß-1,3-glucan, reduced synthesis of gp 43, inhibitors of melanin production, is of great importance for the treatment of PCM. The study of virulence factors favors the understanding of pathogen-host relationships, aiming to evaluate the possibility of developing new therapeutic targets and mechanisms that these molecules play in the infectious process, favoring the design of a more specific treatment for this disease.

Targeting the Homoserine Dehydrogenase of Paracoccidioides Species for Treatment of Systemic Fungal Infections.

This work evaluated new potential inhibitors of the enzyme homoserine dehydrogenase (HSD) of Paracoccidioides brasiliensis, one of the etiological agents of paracoccidioidomycosis. The tertiary structure of the protein bonded to the analogue NAD, and l-homoserine was modeled by homology. The model with the best output was subjected to gradient minimization, redocking, and molecular dynamics simulation. Virtual screening simulations with 187,841 molecules purchasable from the Zinc database were performed. After the screenings, 14 molecules were selected and analyzed by the use of absorption, distribution, metabolism, excretion, and toxicity criteria, resulting in four compounds for in vitro assays. The molecules HS1 and HS2 were promising, exhibiting MICs of 64 and 32 µg · ml-1, respectively, for the Pb18 isolate of P. brasilensis, 64 µg · ml-1 for two isolates of P. lutzii, and also synergy with itraconazole. The application of these molecules to human-pathogenic fungi confirmed that the HSD enzyme may be used as a target for the development of drugs with specific action against paracoccidioidomycosis; moreover, these compounds may serve as leads in the design of new antifungals.

Role of omega-3 polyunsaturated fatty acids in the production of prostaglandin E2 and nitric oxide during experimental murine paracoccidioidomycosis.

There has recently been increased interest in the potential health effects of omega-3 polyunsaturated fatty acids on the immune system. Paracoccidioidomycosis is the most important endemic mycosis in Latin America. Macrophages have a fundamental role and act as first line of organism defense. The purpose of this study was to analyze the effect of n-3 fatty acids on the production of PGE2 and NO by mice infected with Pb18 and fed a diet enriched with LNA for 8 weeks. To study the effect of omega-3 fatty acids on macrophage activity during experimental paracoccidioidomycosis, mice were infected with Pb18 and fed a diet supplemented with LNA. PGE2 in the serum of animals was analyzed and NO in the supernatants of macrophages cultured and challenged in vitro with Pb18 was measured. Omega-3 fatty acids seemed to decrease the production of PGE2 in vivo in the infected group fed an LNA-supplemented diet during the 4th and 8th weeks of the experiment. At the same time, we observed an increase in synthesis of NO by peritoneal macrophages in this group. Omega-3 fatty acids thus appear to have an immunomodulatory effect in paracoccidioidomycosis.

Effect of low intensity helium-neon (HeNe) laser irradiation on experimental paracoccidioidomycotic wound healing dynamics.

The effect of HeNe laser on the extracellular matrix deposition, chemokine expression and angiogenesis in experimental paracoccidioidomycotic lesions was investigated. At days 7, 8 and 9 postinfection the wound of each animal was treated with a 632.8 nm HeNe laser at a dose of 3 J cm(-2). At day 10 postinfection, the wounds were examined by using histologic and immunohistochemical methods. Results revealed that laser-treated lesions were lesser extensive than untreated ones, and composed mainly by macrophages and lymphocytes. High IL-1beta expression was shown in the untreated group whereas in laser-treated animals the expression was scarce. On the other hand, the expression of CXCL-10 was found to be reduced in untreated animals and quite intensive and well distributed in the laser-treated ones. Also, untreated lesions presented vascular endothelial growth factor (VEGF) in a small area near the center of the lesion and high immunoreactivity for hypoxia-inducible factor-1 (HIF-1), whereas laser-treated lesions expressed VEGF surrounding blood vessels and little immunoreactivity for HIF-1. Laser-treated lesions presented much more reticular fibers and collagen deposition when compared with the untreated lesion. Our results show that laser was efficient in minimizing the local effects observed in paracoccidioidomycosis and can be an efficient tool in the treatment of this infection, accelerating the healing process.

Effect of low intensity Helium-Neon (HeNe) laser irradiation on experimental paracoccidioidomycotic wound healing dynamics

The effect of HeNe laser on the extracellular matrix deposition, chemokine expression and angiogenesis in experimental paracoccidioidomycotic lesions was investigated. At days 7, 8 and 9 postinfection the wound of each animal was treated with a 632.8 nm HeNe laser at a dose of 3 J cm(-2). At day 10 postinfection, the wounds were examined by using histologic and immunohistochemical methods. Results revealed that laser-treated lesions were lesser extensive than untreated ones, and composed mainly by macrophages and lymphocytes. High IL-1beta expression was shown in the untreated group whereas in laser-treated animals the expression was scarce. On the other hand, the expression of CXCL-10 was found to be reduced in untreated animals and quite intensive and well distributed in the laser-treated ones. Also, untreated lesions presented vascular endothelial growth factor (VEGF) in a small area near the center of the lesion and high immunoreactivity for hypoxia-inducible factor-1 (HIF-1), whereas laser-treated lesions expressed VEGF surrounding blood vessels and little immunoreactivity for HIF-1. Laser-treated lesions presented much more reticular fibers and collagen deposition when compared with the untreated lesion. Our results show that laser was efficient in minimizing the local effects observed in paracoccidioidomycosis and can be an efficient tool in the treatment of this infection, accelerating the healing process.

Therapeutic administration of KM+ lectin protects mice against Paracoccidioides brasiliensis infection via interleukin-12 production in a toll-like receptor 2-dependent mechanism.

KM(+) is a mannose-binding lectin from Artocarpus integrifolia that induces interleukin (IL)-12 production by macrophages and protective T helper 1 immune response against Leishmania major infection. In this study, we performed experiments to evaluate the therapeutic activity of jackfruit KM(+) (jfKM(+)) and its recombinant counterpart (rKM(+)) in experimental paracoccidioidomycosis. To this end, jfKM(+) or rKM(+) was administered to BALB/c mice 10 days after infection with Paracoccidiodes brasiliensis. Thirty days postinfection, lungs from the KM(+)-treated mice contained significantly fewer colony-forming units and little to no organized granulomas compared to the controls. In addition, lung homogenates from the KM(+)-treated mice presented higher levels of nitric oxide, IL-12, interferon-gamma, and tumor necrosis factor-alpha, whereas higher levels of IL-4 and IL-10 were detected in the control group. With mice deficient in IL-12, Toll-like receptor (TLR) 2, TLR4, or TLR adaptor molecule MyD88, we demonstrated that KM(+) led to protection against P. brasiliensis infection through IL-12 production, which was dependent on TLR2. These results demonstrated a beneficial effect of KM(+) on the severity of P. brasiliensis infection and may expand its potential use as a novel immunotherapeutic molecule.

Oxidative stress response in Paracoccidioides brasiliensis: assessing catalase and cytochrome c peroxidase.

Paracoccidioides brasiliensis is a dimorphic fungus that infects humans and establishes infection in the yeast form. We are interested in the mechanisms this fungus uses to evade the human immune system, and in its survival strategies within infected host cells. Reactive oxygen species play an important role in host defence, but are detoxified by pathogen-derived antioxidant enzymes to prevent oxidative damage. The transcriptional and post-transcriptional regulation of P. brasiliensis catalase and cytochrome-c peroxidase (CCP) antioxidant enzymes upon culture treatment with hydrogen peroxide (H(2)O(2)) is described. High H(2)O(2) concentrations (up to 100 mm) still permitted 70-100% survival of exponential and stationary phase yeast cells, though stationary phase cells were consistently more resistant. P. brasiliensis has both cytosolic and peroxisomal catalase isoenzymes and a single cytochrome-c peroxidase. High-dose treatments with H(2)O(2) led to an early increase in total catalase and CCP enzymatic activities, indicative of post-transcriptional regulation. The expression levels of the catalase genes increased three to fourfold when the cells were treated with 50 mm H(2)O(2) for 40 or 50 min. Lipid peroxidation, as assessed by the thiobarbituric acid method, was relatively low upon treatment with H(2)O(2), which was consistent with our results demonstrating that P. brasiliensis has a powerful antioxidant defence system enabling it to survive H(2)O(2)-mediated stress.