Modulação da ativação de macrófagos por fatores secretados por adipócitos NIH3T3-L1 durante a infecção por Mycobacterium bovis BCG, IN VITRO

Abstract

Introduction: Considered a chronic and multifactorial disease, obesity is one of the main public health problems in the world. In addition to excess body fat, obesity is characterized by a lowgrade inflammatory state (metainflamation). Genetic, environmental factors and metabolic disorders have been proposed as triggers for inflammation that involves the signaling/interaction between adipocytes and macrophages. Factors secreted by adipocytes (FSA), are proposed as modulators of macrophagic profile (M1, M2) and are able to intensify the metabolic inflammation. Among them, there are cytokines, adipokines and extracellular vesicles (EV's). In addition, transcriptional factors it has an important role, PPARγ is a nuclear receptor activated by lipid ligands, with immunoregulatory properties in the macrophage function. PPARγ induces the lipid bodies (LB)s formation during intracellular pathogens infections, such as Mycobacterium bovis BCG. The LBs are dynamic organelles with functions in the lipid stock, synthesis of inflammatory mediators, cell signaling and function as sites for the survival of pathogens. The mechanisms involved in the signaling pathway activation and lipid metabolism by factors secreted by adipocytes and macrophages, as well as their impact on the course of infection by intracellular pathogens are not well known. Methods: LBs count was performed by fluorescence microscopy, cytokine assay by ELISA, and PPARy expression by Western Blot. Isolation of EVs was done by a series of centrifugations followed by characterization and quantification by flow cytometry, Zeta Size and Micro BCA. Results: BCG infection induce LBs formation, which was potentialized by FSA presence. Concomitantly, the expression of PPARy also was potentialized by FSA. Moreover, GW9662 inhibited the expression of this receptor during the infection. The production of TNF-α was negatively regulated, while IL-10 and KC were up modulated in macrophages infected and stimulated with FSA. Further, the adiponectin production was increased in control macrophages stimulated with FSA, an effect that was inhibited during the BCG infection. Furthermore, subpopulations of EV's, the microvesicles or the exosomes, positively modulate LBs biogenesis and macrophage activation, inducing TNF-α and IL-10 synthesis. In addition, the microvesicles were able to induce KC synthesis, which was negatively modulated in the infected groups, an effect that was not observed for macrophages stimulated with exosomes. Conclusion: Our results suggest that FSA have an important role on BCG-infected macrophages, attributed to the formation of LBs, PPARγ expression, synthesis of cytokines and adipokines, which can contribute to changes in the polarization state. However, future studies will still be carried out to elucidate the real cross-talk between adipocytes and macrophages in the face of infections by intracellular pathogens such as M. bovis BCG