After 2 h of incubation with rPbPga1 or LPS there was a significant increase in luciferase activity in Raw 264.7 Luc cells which was increased at 3 h (Fig 3A). polyclonal anti-rPbPga1 antibody. Furthermore, macrophages stained with anti-CD38 were associated with containing granulomas. Additionally, rPbPga1 activated the transcription factor NFkB in the macrophage cell line Raw 264.7 Luc cells, containing the luciferase gene downstream of the NFkB promoter. After 24 h of incubation with rPbPga1, alveolar macrophages from BALB/c mice were stimulated to release TNF-, IL-4 and NO. Mast cells, identified by toluidine blue staining, were also associated with containing granulomas. Co-culture of yeast cells with RBL-2H3 mast cells induced morphological changes on the surface of the mast cells. Furthermore, RBL-2H3 mast cells were degranulated by yeast cells, but not by rPbPga1, as determined by the release of beta-hexosaminidase. However, Naspm trihydrochloride RBL-2H3 cells activated by rPbPga1 released the inflammatory interleukin IL-6 and also activated the transcription factor NFkB in GFP-reporter mast cells. The transcription factor NFAT was not activated when the mast cells were incubated with rPbPga1. Conclusions/Significance The results indicate that PbPga1 may act as a modulator protein in PCM pathogenesis and serve as a useful target for additional studies on the pathogenesis of is thought to infect the host through the respiratory tract. Cell wall components of interact with host cells producing granulomas, thus influencing the pathogenesis of PCM. PbPga1 is an granulomas. Furthermore, recombinant PbPga1 was able to activate both alveolar macrophages and mast cells via the transcription factor NFkB to release inflammatory mediators. The results of this study indicate that the surface antigen, PbPga1, may play an important role in PCM pathogenesis by activating macrophages and mast cells. Additionally, PbPga1 may be a target for new Naspm trihydrochloride strategies for detecting and treating PCM. Introduction The fungus is the etiological agent of paracoccidioidomycosis (PCM), the most prevalent systemic mycosis in Latin America [1C3], and is considered the major cause of death from systemic mycosis in Brazil [4]. is a thermodimorphic fungus that at room temperature grows as long, thin, multicellular hyphae which produce infectious propagules in the form of asexual conidia. After inhalation of the mycelium into the lungs, it switches to the pathogenic yeast form at body temperature [5C9]. Within the lungs the yeast is initially sequestered in granulomas which controls the spread of the fungus to other organs [10]. The host response to infection is dependent on the interaction between the fungi and host immune cells present in the lung. Macrophages and mast cells are among the cells that participate in the host response to fungal infection. Macrophages are activated by yeast and present fungicidal activity and [6, 11]. During the early stages of infection, fungal dissemination is limited by the activation of macrophages which produce high levels of TNF- [12] and nitric oxide (NO) [13]. Mast cells are considered sentinel cells of the innate immune system. They reside in the connective tissue at the interface between the Naspm trihydrochloride environment and the host and are encountered in the skin as well Naspm trihydrochloride as in the respiratory and gastrointestinal tracts. They function in the host response against many pathogens, such as viruses, bacteria and parasites. However, little is known about their reaction to fungal infections [14C16]. Mast cells can also be activated through FcRI (high affinity IgE receptor) or other cell surface receptors such as PRRs (Pattern Recognition Receptors) to participate in the innate immune response. The presence of large amounts of immunoglobulin E in the blood of PCM patients provides evidence that mast cells can participate in the acquired immune response to [17]. Mast cell activation by pathogens culminates in the release of Rabbit polyclonal to PELI1 interleukins and other mediators that contribute to the recruitment, differentiation and activation of immature monocytes and macrophages as well as leading to granuloma formation [18, 19]. The interaction between the host and the pathogenic fungi occurs by contact of the host cells with the fungal cell wall or its components. Thus, the cell wall of pathogenic fungi plays a major role in the pathogenesis of the fungus. The cell wall of many ascomycetes consists of a network of polysaccharides in which many proteins are covalently linked to the cell wall [20, 21]. In transcriptome identified GPI-anchored proteins that play an important role in the virulence of pathogenic fungi. However, the function of most of the proteins that were identified remains unknown [2]. Two GPI-anchored proteins, phospholipase B1 (PLB1) and the glycoprotein Dfg5P, have been functionally analyzed in [32, 33]. Inhibition of PLB1 leads.