Title : Evaluation of the immunomodulatory role of butyrate in macrophages infected with paracoccidioides brasiliensis
Abstract:
Introduction: The microbiota, composed of bacteria, archaea, fungi, and viruses, constantly interacts with the host in a symbiotic and dynamic relationship, performing essential functions for the maintenance of homeostasis. Among its functions is the fermentation of dietary fibers, which generates key nutrients and produces bioactive metabolites such as short-chain fatty acids (SCFAs). These SCFAs, which include acetate, propionate, and butyrate, play relevant roles in both metabolic regulation and immune response modulation. Among them, butyrate stands out for exerting immunomodulatory effects, acting on G protein-coupled receptors (GPCRs) and inhibiting epigenetic enzymes such as histone deacetylases. This dual action enables potential adjustments in inflammatory pathways, resulting in a balance between pro- and anti-inflammatory responses. Experimental evidence suggests that butyrate enhances the microbicidal activity of macrophages and may inhibit the growth of pathogenic fungi, raising interest in its use as a potential therapeutic adjuvant.
The purpose of this study was to investigate the effect of butyrate on the innate immune response against the pathogenic fungus Paracoccidioides brasiliensis, focusing on both cytokine production and the fungicidal capacity of macrophages. Methodology: The fungal isolate Pb18 was cultured for 7 days at 37 °C in semi-solid BHI-dextrose medium to obtain viable cells. RAW264.7 macrophages and bone marrow-derived macrophages (BMDMs) were then infected and treated with butyrate (10–20 mM). Lipopolysaccharide (LPS, 1 μg/ml) was used as a positive control for inflammatory activation. After 24 and 48 hours of incubation, cytokines IL-1β, TNF-α, and IL-10 were quantified by ELISA. In parallel, the fungicidal activity of macrophages was assessed through the quantification of colony-forming units (CFUs). To investigate the contribution of phagolysosomal acidification to the microbicidal effect, Bafilomycin A1 (0.5 μM) was added prior to cell infection. Results: Butyrate significantly reduced TNF-α levels in macrophages stimulated with LPS as well as in those infected with P. brasiliensis, confirming its anti-inflammatory effect. In BMDMs, butyrate treatment decreased IL-10 and increased IL-1β, suggesting inflammasome activation. Moreover, butyrate enhanced macrophage fungicidal capacity, but this effect was reversed in the presence of Bafilomycin, indicating that intracellular acidification is a central mechanism for the fungicidal activity induced by this SCFA. Conclusion: These findings highlight the immunomodulatory potential of butyrate in macrophages infected with P. brasiliensis, characterized by the reduction of TNF-α and IL-10 levels and the increase of IL-1β. The observed decrease in fungal viability reinforces its role in stimulating acidification-dependent microbicidal capacity. Thus, butyrate emerges as a possible adjuvant strategy for the treatment of paracoccidioidomycosis.
