Abstract
Histoplasma capsulatum var. capsulatum is a dimorphic fungus that causes histoplasmosis, a human systemic mycosis with worldwide distribution. In the present work, we demonstrate that H. capsulatum yeasts are able to induce cytokine secretion by the human lung epithelial cell line A549 in integrin- and Src-family kinase (SFK)-dependent manners. This conclusion is supported by small interfering RNA (siRNA) directed to α3 and α5 integrins, and PP2, an inhibitor of SFK activation. siRNA and PP2 reduced IL-6 and IL-8 secretion in H. capsulatum-infected A549 cell cultures. In addition, α3 and α5 integrins from A549 cells were capable of associating with H. capsulatum yeasts, and this fungus promotes recruitment of these integrins and SFKs to A549 cell membrane rafts. Corroborating this finding, membrane raft disruption with the cholesterol-chelator methyl-β-cyclodextrin reduced the levels of integrins and SFKs in these cell membrane domains. Finally, pretreatment of A549 cells with the cholesterol-binding compound, and also a membrane raft disruptor, filipin, significantly reduced IL-6 and IL-8 levels in A549-H.capsulatum cultures. Taken together, these results indicate that H. capsulatum yeasts induce secretion of IL-6 and IL-8 in human lung epithelial cells by interacting with α3 and α5 integrins, recruiting these integrins to membrane rafts, and promoting SFK activation.
Highlights
Histoplasmosis is a human systemic mycosis caused by the fungal pathogen Histoplasma capsulatum
To verify whether H. capsulatum yeasts induce secretion of inflammatory cytokines by epithelial cells, the human lung epithelial cell line A549 was incubated with H. capsulatum yeasts for different periods of time (5-24 h), and levels of IL-6, IL-8, and IL-10 in these culture supernatants were determined by ELISA
By MTT assay, we verified that cell viability was unaffected when A549 cells were incubated with H. capsulatum yeasts for 5–24 h (Supplementary Table 1)
Summary
Histoplasmosis is a human systemic mycosis caused by the fungal pathogen Histoplasma capsulatum. This fungus presents two varieties: H. capsulatum var. Duboisii that are etiological agents of the classical and the African histoplasmoses, respectively. Classical histoplasmosis is widely distributed in the Americas. In the United States, highly endemic areas include the Mississippi and Ohio River valleys. This mycosis occurs in countries of Central and South America, and in Brazil, outbreaks of histoplasmosis have been reported after exposure to H. capsulatum fragments (Martins et al, 2003; Guimarães et al, 2006; Oliveira et al, 2006; Rocha-Silva et al, 2014).
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