MilliporeSigma
  • Home
  • Search Results
  • rPbPga1 from Paracoccidioides brasiliensis Activates Mast Cells and Macrophages via NFkB.

rPbPga1 from Paracoccidioides brasiliensis Activates Mast Cells and Macrophages via NFkB.

PLoS neglected tropical diseases (2015-09-01)
Clarissa Xavier Resende Valim, Elaine Zayas Marcelino da Silva, Mariana Aprigio Assis, Fabricio Freitas Fernandes, Paulo Sergio Rodrigues Coelho, Constance Oliver, Maria Célia Jamur
ABSTRACT

The fungus Paracoccidioides brasiliensis is the leading etiological agent of paracoccidioidomycosis (PCM), a systemic granulomatous disease that typically affects the lungs. Cell wall components of P. brasiliensis interact with host cells and influence the pathogenesis of PCM. In yeast, many glycosylphosphatidylinositol (GPI)-anchored proteins are important in the initial contact with the host, mediating host-yeast interactions that culminate with the disease. PbPga1 is a GPI anchored protein located on the surface of the yeast P. brasiliensis that is recognized by sera from PCM patients. Endogenous PbPga1 was localized to the surface of P. brasiliensis yeast cells in the lungs of infected mice using a polyclonal anti-rPbPga1 antibody. Furthermore, macrophages stained with anti-CD38 were associated with P. brasiliensis 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-α, IL-4 and NO. Mast cells, identified by toluidine blue staining, were also associated with P. brasiliensis containing granulomas. Co-culture of P. Brasiliensis 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 P. brasiliensis yeast cells, but not by rPbPga1, as determined by the release of beta-hexosaminidase. However, 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. 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 P. brasiliensis.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Methanol, NMR reference standard
Sigma-Aldrich
Methanol, HPLC Plus, ≥99.9%, poly-coated bottles
Sigma-Aldrich
Osmium tetroxide, Sealed ampule.
Sigma-Aldrich
Trypan Blue, Dye content 60 %
Sigma-Aldrich
Methanol, anhydrous, 99.8%
Sigma-Aldrich
Methanol, suitable for HPLC, gradient grade, 99.93%
Sigma-Aldrich
Hydrogen peroxide solution, contains potassium stannate as inhibitor, 30-32 wt. % in water, semiconductor grade, 99.999% trace metals basis
Sigma-Aldrich
2,2,2-Tribromoethanol, 97%
Sigma-Aldrich
Hydrogen peroxide solution, 34.5-36.5%
Sigma-Aldrich
Hydrogen peroxide solution, 30 % (w/w) in H2O, contains stabilizer
Sigma-Aldrich
Trypan Blue, powder, BioReagent, suitable for cell culture
Sigma-Aldrich
Os EnCat® 40, extent of labeling: 0.3 mmol/g Os loading
Sigma-Aldrich
3,3′-Diaminobenzidine, ≥97% (HPLC)
Sigma-Aldrich
Hematoxylin, certified by the Biological Stain Commission
Supelco
Hematoxylin
Sigma-Aldrich
Trypan Blue solution, 0.4%, liquid, sterile-filtered, suitable for cell culture
Sigma-Aldrich
TNP, ≥95% (HPLC)
Sigma-Aldrich
Methanol-12C, 99.95 atom % 12C
Sigma-Aldrich
o-Xylene, anhydrous, 97%
Sigma-Aldrich
3,3′-Diaminobenzidine, 98%
Sigma-Aldrich
Osmium tetroxide solution, suitable for electron microscopy, 4% in H2O
Sigma-Aldrich
Osmium tetroxide solution, suitable for electron microscopy, 2% in H2O
Supelco
Methanol solution, contains 0.10 % (v/v) formic acid, UHPLC, suitable for mass spectrometry (MS), ≥99.5%
Supelco
Methanol solution, NMR reference standard, 4% in methanol-d4 (99.8 atom % D), NMR tube size 5 mm × 8 in.
Sigma-Aldrich
Methanol solution, NMR reference standard, 4% in methanol-d4 (99.8 atom % D), NMR tube size 3 mm × 8 in.