Δ-FeOOH as Support for Immobilization Peroxidase: Optimization via a Chemometric Approach.

Owing to their high surface area, stability, and functional groups on the surface, iron oxide hydroxide nanoparticles have attracted attention as enzymatic support. In this work, a chemometric approach was performed, aiming at the optimization of the horseradish peroxidase (HRP) immobilization process on Δ-FeOOH nanoparticles (NPs). The enzyme/NPs ratio (X1), pH (X2), temperature (X3), and time (X4) were the independent variables analyzed, and immobilized enzyme activity was the response variable (Y). The effects of the factors were studied using a factorial design at two levels (-1 and 1). The biocatalyst obtained was evaluated for the ferulic acid (FA) removal, a pollutant model. The materials were characterized by X-ray powder diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), and scanning electron microscopy (SEM). The SEM images indicated changes in material morphology. The independent variables X1 (-0.57), X2 (0.71), and X4 (0.42) presented the significance effects estimate. The variable combinations resulted in two significance effects estimates, X1*X2 (-0.57) and X2*X4 (0.39). The immobilized HRP by optimized conditions (X1 = 1/63 (enzyme/NPs ratio, X2 = pH 8, X4 = 60 °C, and 30 min) showed high efficiency for FA oxidation (82%).

Efficient uptake of mannosylated proteins by a human Schwann cell line.

Complex carbohydrate structures are essential molecules of infectious microbes and host cells, and are involved in cell signaling associated with inflammatory and immune responses. The uptake of mannose-tailed glycans is usually carried out by macrophages, dendritic cells (DCs), and other professional phagocytes to trigger MHC class I- and MHC class II-restricted antigen presentation, and to promote T cell effector responses. Since Schwann cells (SCs) have been proposed as immunocompetent cells, we investigated whether a human cell line (ST88-14 cells) could bind mannosylated ligands in a specific manner. The saturation of uptake of mannosylated molecules by ST88-14 cells and the internalization and distribution pathway of these ligands were tested by cytometry and confocal plus electron microscopy, respectively. This uptake showed a dose-dependent increase, the saturation point being reached at high concentrations of mannosyl residues/240 mM mannose. Merging of man/BSA-FITC and S100 labeling showed their partial, but, significant colocalization. Ultrastructural analysis of ST88-14 cells after incubation with HRP-colloidal gold, without or with subsequent chasing at 37C, showed an initial location on the cell surface and temperature- and time-dependent internalization of the probe. Our findings suggest an efficient mannosylated ligand uptake system through putative lectin(s) that may be operational in inflammatory and immune responses.

A cardiomyocyte mannose receptor system is involved in Trypanosoma cruzi invasion and is down-modulated after infection.

Mannosyl binding sites were detected "in vitro" on cardiomyocytes (CM) surface using horseradish peroxidase (HRP) as the ligand. Binding assays revealed a specific recognition system, which was time- and concentration-dependent. The binding required physiological pH and was inhibited by EDTA and trypsin treatments. HRP binding was reduced by pre-incubations with low concentrations of D-mannose. Ultrastructural analysis of the endocytic process was followed using HRP coupled to colloidal gold particles (HRP-Au). The tracer was found within caveolae characterizing early steps of the receptor-mediated endocytosis. The addition of 10 mM D-mannose to the interaction medium blocked Trypanosoma cruzi uptake by CM. The labeling of CM with a subsaturating concentration of HRP-Au before their infection showed, by ultrastructural studies, that its association with trypomastigote forms occurred frequently near to HRP-gold particles that could also be seen to comprise the parasitophorous vacuole. After infection of CM with T. cruzi, a considerable reduction on HRP binding was noticed. Binding was almost completely restored by treating the infected cultures with the trypanocidal drug Nifurtimox. Our "in vitro" findings suggest that cardiomyocyte's mannose receptors localized at the sarcolemma mediates T. cruzi recognition and can be down-modulated by parasite infection.