Mitochondrial Fission Promotes the Continued Clearance of Apoptotic Cells by Macrophages.

Clearance of apoptotic cells (ACs) by phagocytes (efferocytosis) prevents post-apoptotic necrosis and dampens inflammation. Defective efferocytosis drives important diseases, including atherosclerosis. For efficient efferocytosis, phagocytes must be able to internalize multiple ACs. We show here that uptake of multiple ACs by macrophages requires dynamin-related protein 1 (Drp1)-mediated mitochondrial fission, which is triggered by AC uptake. When mitochondrial fission is disabled, AC-induced increase in cytosolic calcium is blunted owing to mitochondrial calcium sequestration, and calcium-dependent phagosome formation around secondarily encountered ACs is impaired. These defects can be corrected by silencing the mitochondrial calcium uniporter (MCU). Mice lacking myeloid Drp1 showed defective efferocytosis and its pathologic consequences in the thymus after dexamethasone treatment and in advanced atherosclerotic lesions in fat-fed Ldlr-/- mice. Thus, mitochondrial fission in response to AC uptake is a critical process that enables macrophages to clear multiple ACs and to avoid the pathologic consequences of defective efferocytosis in vivo

Therapeutic Administration of Recombinant Paracoccin Confers Protection against Paracoccidioides brasiliensis Infection: Involvement of TLRs.

Paracoccin (PCN) is an N-acetylglucosamine-binding lectin from the human pathogenic fungus Paracoccidioides brasiliensis. Recombinant PCN (rPCN) induces a T helper (Th) 1 immune response when prophylactically administered to BALB/c mice, protecting them against subsequent challenge with P. brasiliensis. In this study, we investigated the therapeutic effect of rPCN in experimental paracoccidioidomycosis (PCM) and the mechanism accounting for its beneficial action. METHODOLOGY/PRINCIPAL FINDINGS: Four distinct regimens of rPCN administration were assayed to identify which was the most protective, relative to vehicle administration. In all rPCN-treated mice, pulmonary granulomas were less numerous and more compact. Moreover, fewer colony-forming units were recovered from the lungs of rPCN-treated mice. Although all therapeutic regimens of rPCN were protective, maximal efficacy was obtained with two subcutaneous injections of 0.5 microg rPCN at 3 and 10 days after infection. The rPCN treatment was also associated with higher pulmonary levels of IL-12, IFN-gamma, TNF-alpha, nitric oxide (NO), and IL-10, without IL-4 augmentation. Encouraged by the pulmonary cytokine profile of treated mice and by the fact that in vitro rPCN-stimulated macrophages released high levels of IL-12, we investigated the interaction of rPCN with Toll-like receptors (TLRs). Using a reporter assay in transfected HEK293T cells, we verified that rPCN activated TLR2 and TLR4. The activation occurred independently of TLR2 heterodimerization with TLR1 or TLR6 and did not require the presence of the CD14 or CD36 co-receptors. The interaction between rPCN and TLR2 depended on carbohydrate recognition because it was affected by mutation of the receptor\u27s N-glycosylation sites. The fourth TLR2 N-glycan was especially critical for the rPCN-TLR2 interaction. CONCLUSIONS/SIGNIFICANCE: Based on our results, we propose that PCN acts as a TLR agonist. PCN binds to N-glycans on TLRs, triggers regulated Th1 immunity, and exerts a therapeutic effect against P. brasiliensis infection

Growth curve, biochemical profile and phytochemical analyses in calli obtained from the procambium segments of Bacupari

Garcinia brasiliensis, popularly known as Bacupari, is native to the Amazon and commonly used in folk medicine for its therapeutic properties. This plant is rich in bioactive compounds like benzophenones. However, there are no works about the in vitro establishment and achievement of secondary metabolites in this plant. Thus, the aim of this work was to determine the growth curve and to perform the biochemical and phytochemical analyses in calli obtained from the procambium segments of Bacupari. The growth curve of calli followed a sigmoidal pattern, with four distinct phases (lag, exponential, linear, deceleration). Total soluble sugars were higher on the inoculation day and the reducing sugars on the 20 th day. Amino acids increased from the 60 th day up to the stabilization on the 120 th day. The protein content varied, but it seemed to be related to the amino acids metabolism. The phytochemical screening showed the presence of phenolic and flavonoid compounds in the calli and the HPLC analysis allowed the identification of Fukugetin, Guttiferone A and 7-epiclusianone