Leishmania infantum Exoproducts Inhibit Human Invariant NKT Cell Expansion and Activation.

Leishmania infantum is one of the major parasite species associated with visceral leishmaniasis, a severe form of the disease that can become lethal if untreated. This obligate intracellular parasite has developed diverse strategies to escape the host immune response, such as exoproducts (Exo) carrying a wide range of molecules, including parasite virulence factors, which are potentially implicated in early stages of infection. Herein, we report that L. infantum Exo and its two fractions composed of extracellular vesicles (EVs) and vesicle-depleted-exoproducts (VDEs) inhibit human peripheral blood invariant natural killer T (iNKT) cell expansion in response to their specific ligand, the glycolipid α-GalactosylCeramide (α-GalCer), as well as their capacity to promptly produce IL-4 and IFNγ. Using plate-bound CD1d and α-GalCer, we found that Exo, EV, and VDE fractions reduced iNKT cell activation in a dose-dependent manner, suggesting that they prevented α-GalCer presentation by CD1d molecules. This direct effect on CD1d was confirmed by the observation that CD1d:α-GalCer complex formation was impaired in the presence of Exo, EV, and VDE fractions. Furthermore, lipid extracts from the three compounds mimicked the inhibition of iNKT cell activation. These lipid components of L. infantum exoproducts, including EV and VDE fractions, might compete for CD1-binding sites, thus blocking iNKT cell activation. Overall, our results provide evidence for a novel strategy through which L. infantum can evade immune responses of mammalian host cells by preventing iNKT lymphocytes from recognizing glycolipids in a TCR-dependent manner.

Accelerated solvent extraction method for the quantification of polycyclic aromatic hydrocarbons in cocoa beans by gas chromatography-mass spectrometry.

An accelerated solvent extraction (ASE) procedure for use with gas chromatography-mass spectrometry (GC-MS) was optimized for the determination of eight polycyclic aromatic hydrocarbons (PAHs) in cocoa beans. Plackett-Burman and rotatable central composite design (RCCD) indicated that three variables affected the recoveries of PAHs during the extraction and purification steps: agitation time in the second liquid-liquid partition, weight of silica gel in the column, and volume of hexane for PAH elution from the column. After obtaining the optimal conditions, a single laboratory method validation was performed. Linearity was demonstrated for benzo[a]pyrene in the concentration range from 0.5 to 8.0mgkg-1 of sample, corresponding to 1.25-20.0µgkg-1 of cocoa on a fat basis. For the other analytes, linearity was observed from 0.75 to 8.0µgkg-1 of sample (1.88-20.0µgkg-1 of cocoa on a fat basis). Significant matrix effects were found for chrysene and benzo[b]fluoranthene. The precision of the method was verified with relative standard deviations (RSDs) ranging from 2.57 to 14.13% and from 4.36 to 19.77% under repeatability and intermediate precision conditions, respectively. The average recoveries of the eight PAHs ranged from 74.99 to 109.73%. These parameters, limits and measurement uncertainties met the performance criteria established by European Union regulations, except for the theoretical limit of detection for chrysene. The method was applied to the analysis of samples of Brazilian cocoa beans, and only one sample was found to have a PAH content above the maximum limit defined by the European Union legislation. This optimized and validated method is intended to be used as part of the official Brazilian monitoring programs investigating contaminants and residues in food.

Innate-like and conventional T cell populations from hemodialyzed and kidney transplanted patients are equally compromised.

Clinicians are well aware of existing pharmacologically-induced immune deficient status in kidney-transplanted patients that will favor their susceptibility to bacterial or viral infections. Previous studies indicated that advanced Stage 4-5 Chronic Kidney Disease might also be regarded as an immune deficiency-like status as well, even though the mechanisms are not fully understood. Here, we analyzed the ex vivo frequency and the functional properties of both conventional and innate-like T (ILT) lymphocyte subsets in the peripheral blood of 35 patients on hemodialysis, 29 kidney transplanted patients and 38 healthy donors. We found that peripheral blood cell count of ILT cells, as iNKT (invariant Natural Killer T) and MAIT (mucosal-associated invariant T), were significantly decreased in hemodialyzed patients compared to healthy controls. This deficiency was also observed regarding conventional T cells, including the IL-17-producing CD4(+) Th17 cells. Pertaining to regulatory T cells, we also noticed major modifications in the global frequency of CD4(+)CD25(+)Foxp3(+) T lymphocytes, including the resting suppressive CD45RA(+)Foxp3lo and activated suppressive CD45RA-Foxp3hi T cell subpopulations. We found no significant differences between the immune status of hemodialyzed and kidney-transplanted subjects. In conclusion, we demonstrated that both ILT and conventional T cell numbers are equally impaired in hemodialyzed and kidney-transplanted patients.