Rab7 controls lipid droplet-phagosome association during mycobacterial infection.

Lipid droplets (LDs) are organelles that have multiple roles in inflammatory and infectious diseases. LD act as essential platforms for immunometabolic regulation, including as sites for lipid storage and metabolism, inflammatory lipid mediator production, and signaling pathway compartmentalization. Accumulating evidence indicates that intracellular pathogens may exploit host LDs as source of nutrients and as part of their strategy to promote immune evasion. Notably, numerous studies have demonstrated the interaction between LDs and pathogen-containing phagosomes. However, the mechanism involved in this phenomenon remains elusive. Here, we observed LDs and PLIN2 surrounding M. bovis BCG-containing phagosomes, which included observations of a bacillus cell surrounded by lipid content inside a phagosome and LAM from mycobacteria co-localizing with LDs; these results were suggestive of exchange of contents between these compartments. By using beads coated with M.tb lipids, we demonstrated that LD-phagosome associations are regulated through the mycobacterial cell wall components LAM and PIM. In addition, we demonstrated that Rab7 and RILP, but not Rab5, localizes to LDs of infected macrophages and observed the presence of Rab7 at the site of interaction with an infected phagosome. Moreover, treatment of macrophages with the Rab7 inhibitor CID1067700 significantly inhibited the association between LDs and LAM-coated beads. Altogether, our data demonstrate that LD-phagosome interactions are controlled by mycobacterial cell wall components and Rab7, which enables the exchange of contents between LDs and phagosomes and may represent a fundamental aspect of bacterial pathogenesis and immune evasion.

Microglia/Astrocytes-Glioblastoma Crosstalk: Crucial Molecular Mechanisms and Microenvironmental Factors.

In recent years, the functions of glial cells, namely, astrocytes and microglia, have gained prominence in several diseases of the central nervous system, especially in glioblastoma (GB), the most malignant primary brain tumor that leads to poor clinical outcomes. Studies showed that microglial cells or astrocytes play a critical role in promoting GB growth. Based on the recent findings, the complex network of the interaction between microglial/astrocytes cells and GB may constitute a potential therapeutic target to overcome tumor malignancy. In the present review, we summarize the most important mechanisms and functions of the molecular factors involved in the microglia or astrocytes-GB interactions, which is particularly the alterations that occur in the cell's extracellular matrix and the cytoskeleton. We overview the cytokines, chemokines, neurotrophic, morphogenic, metabolic factors, and non-coding RNAs actions crucial to these interactions. We have also discussed the most recent studies regarding the mechanisms of transportation and communication between microglial/astrocytes - GB cells, namely through the ABC transporters or by extracellular vesicles. Lastly, we highlight the therapeutic challenges and improvements regarding the crosstalk between these glial cells and GB.

Lipid Body Organelles within the Parasite Trypanosoma cruzi: A Role for Intracellular Arachidonic Acid Metabolism.

Most eukaryotic cells contain varying amounts of cytosolic lipidic inclusions termed lipid bodies (LBs) or lipid droplets (LDs). In mammalian cells, such as macrophages, these lipid-rich organelles are formed in response to host-pathogen interaction during infectious diseases and are sites for biosynthesis of arachidonic acid (AA)-derived inflammatory mediators (eicosanoids). Less clear are the functions of LBs in pathogenic lower eukaryotes. In this study, we demonstrated that LBs, visualized by light microscopy with different probes and transmission electron microscopy (TEM), are produced in trypomastigote forms of the parasite Trypanosoma cruzi, the causal agent of Chagas' disease, after both host interaction and exogenous AA stimulation. Quantitative TEM revealed that LBs from amastigotes, the intracellular forms of the parasite, growing in vivo have increased size and electron-density compared to LBs from amastigotes living in vitro. AA-stimulated trypomastigotes released high amounts of prostaglandin E2 (PGE2) and showed PGE2 synthase expression. Raman spectroscopy demonstrated increased unsaturated lipid content and AA incorporation in stimulated parasites. Moreover, both Raman and MALDI mass spectroscopy revealed increased AA content in LBs purified from AA-stimulated parasites compared to LBs from unstimulated group. By using a specific technique for eicosanoid detection, we immunolocalized PGE2 within LBs from AA-stimulated trypomastigotes. Altogether, our findings demonstrate that LBs from the parasite Trypanosoma cruzi are not just lipid storage inclusions but dynamic organelles, able to respond to host interaction and inflammatory events and involved in the AA metabolism. Acting as sources of PGE2, a potent immunomodulatory lipid mediator that inhibits many aspects of innate and adaptive immunity, newly-formed parasite LBs may be implicated with the pathogen survival in its host.

Differential TLR2 downstream signaling regulates lipid metabolism and cytokine production triggered by Mycobacterium bovis BCG infection.

The nuclear receptor PPARγ acts as a key modulator of lipid metabolism, inflammation and pathogenesis in BCG-infected macrophages. However, the molecular mechanisms involved in PPARγ expression and functions during infection are not completely understood. Here, we investigate signaling pathways triggered by TLR2, the involvement of co-receptors and lipid rafts in the mechanism of PPARγ expression, lipid body formation and cytokine synthesis in macrophages during BCG infection. BCG induces NF-κB activation and increased PPARγ expression in a TLR2-dependent manner. Furthermore, BCG-triggered increase of lipid body biogenesis was inhibited by the PPARγ antagonist GW9662, but not by the NF-κB inhibitor JSH-23. In contrast, KC/CXCL1 production was largely dependent on NF-κB but not on PPARγ. BCG infection induced increased expression of CD36 in macrophages in vitro. Moreover, CD36 co-immunoprecipitates with TLR2 in BCG-infected macrophages, suggesting its interaction with TLR2 in BCG signaling. Pretreatment with CD36 neutralizing antibodies significantly inhibited PPARγ expression, lipid body formation and PGE2 production induced by BCG. Involvement of CD36 in lipid body formation was further confirmed by decreased BCG-induced lipid body formation in CD36 deficient macrophages. Similarly, CD14 and CD11b/CD18 blockage also inhibited BCG-induced lipid body formation, whereas TNF-α synthesis was not affected. Disruption of rafts recapitulates the latter result, inhibiting lipid body formation, but not TNF-α synthesis in BCG-infected macrophages. In conclusion, our results suggest that CD36-TLR2 cooperation and signaling compartmentalization within rafts, divert host response signaling through PPARγ-dependent and NF-κB-independent pathways, leading to increased macrophage lipid accumulation and down-modulation of macrophage response.

Host cell lipid bodies triggered by Trypanosoma cruzi infection and enhanced by the uptake of apoptotic cells are associated with prostaglandin E2 generation and increased parasite growth.

Lipid bodies (lipid droplets) are lipid-rich organelles with functions in cell metabolism and signaling. Here, we investigate the mechanisms of Trypanosoma cruzi-induced lipid body formation and their contributions to host-parasite interplay. We demonstrate that T. cruzi-induced lipid body formation in macrophages occurs in a Toll-like receptor 2-dependent mechanism and is potentiated by apoptotic cell uptake. Lipid body biogenesis and prostaglandin E2 (PGE2) production triggered by apoptotic cell uptake was largely dependent of α(v)ß3 and transforming growth factor-ß signaling. T. cruzi-induced lipid bodies act as sites of increased PGE synthesis. Inhibition of lipid body biogenesis by the fatty acid synthase inhibitor C75 reversed the effects of apoptotic cells on lipid body formation, eicosanoid synthesis, and parasite replication. Our findings indicate that lipid bodies are highly regulated organelles during T. cruzi infection with roles in lipid mediator generation by macrophages and are potentially involved in T. cruzi-triggered escape mechanisms.

Neutrophils recruited to the site of Mycobacterium bovis BCG infection undergo apoptosis and modulate lipid body biogenesis and prostaglandin E production by macrophages.

Neutrophil influx to sites of mycobacterial infections is one of the first events of tuberculosis pathogenesis. However, the role of early neutrophil recruitment in mycobacterial infection is not completely understood. We investigated the rate of neutrophil apoptosis and the role of macrophage uptake of apoptotic neutrophils in a pleural tuberculosis model induced by BCG. Recruited neutrophils were shown to phagocyte BCG and a large number of neutrophils undergo apoptosis within 24 h. Notably, the great majority of apoptotic neutrophils were infected by BCG. Increased lipid body (lipid droplets) formation, accompanied by prostaglandin E(2) (PGE(2)) and TGF-beta1 synthesis, occurred in parallel to macrophage uptake of apoptotic cells. Lipid body and PGE(2) formation was observed after macrophage exposure to apoptotic, but not necrotic or live neutrophils. Blockage of BCG-induced lipid body formation significantly inhibited PGE(2) synthesis. Pre-treatment with the pan-caspase inhibitor zVAD inhibited BCG-induced neutrophil apoptosis and lipid body formation, indicating a role for apoptotic neutrophils in macrophage lipid body biogenesis in infected mice. In conclusion, BCG infection induced activation and apoptosis of infected neutrophils at the inflammatory site. The uptake of apoptotic neutrophils by macrophages leads to TGF-beta1 generation and PGE(2)-derived lipid body formation, and may have modulator roles in mycobacterial pathogenesis.

Toll-like receptor-2-mediated C-C chemokine receptor 3 and eotaxin-driven eosinophil influx induced by Mycobacterium bovis BCG pleurisy.

An acute and persistent eosinophil infiltration is observed during Mycobacterium bovis BCG pleural infection in mice. Eosinophil accumulation, lipid body formation, and eotaxin production were significantly reduced in BCG-infected Toll-like receptor-2 (TLR2)-deficient mice compared to wild-type mice. Neutralization of eotaxin or CCR3 drastically inhibited BCG-induced eosinophil accumulation and lipid body formation, indicating that BCG-induced eosinophil recruitment and activation is largely dependent of TLR2-mediated eotaxin generation.