Fluorescence and electron microscopy methods for exploring antimicrobial peptides mode(s) of action.

Due to the increasing resistance of microbial pathogens to the available drugs, the identification of new antimicrobial agents with a new mechanism of action is urgently needed. In this context, cationic antimicrobial peptides (AMPs) are considered promising candidates. Although there is evidence that, in contrast to conventional antibiotics, microbial membranes are the principal target of a large number of AMPs, thus making it difficult for the pathogen to acquire resistance, their mode(s) of action is not yet completely clear.

Morphological analysis of the interaction of charged surfactant vesicles (SVs) with human cultured cells.

We analyzed the binding and fusogenic properties of surfactant vesicles (SVs), composed of ionic and nonionic surfactants and cholesterol, with the surface of different human lymphoid cells. The influence of charge on SVs-cell interaction was evaluated by monitoring the presence of fluorescent sodium calcein artificially entrapped in the vesicles using optical fluorescence microscopy and laser scanning confocal microscopy. Our results clearly indicate that only negatively charged vesicles bind and fuse with the plasma membrane of human lymphoid cells, and the number of SVs bound to the cell surface was variable among the positive cells.

A novel mechanism of CD4 down-modulation induced by monosialoganglioside GM3. Involvement of serine phosphorylation and protein kinase c delta translocation.

In this report the molecular mechanism(s) involved in the rapid and selective endocytosis of cell surface glycoprotein CD4 induced by exogenous monosialoganglioside GM3 in human peripheral blood lymphocytes have been investigated. Inhibition of the GM3-induced CD4 down-modulation was observed in the presence of specific protein kinase C (PKC) inhibitors. Scanning confocal microscopy revealed the translocation and clustering on the cell surface of PKC isozymes delta and theta (more evidently than alpha and beta) after GM3 treatment, suggesting the involvement of these isozymes in the ganglioside-induced CD4 down-modulation.

Colocalization and complex formation between prosaposin and monosialoganglioside GM3 in neural cells.

Prosaposin, the precursor of saposins A, B, C, and D, was recently identified as a neurotrophic factor in vitro as well as in vivo. Its neurotrophic activity has been localized to a linear 12-amino acid sequence located in the NH2-terminal portion of the saposin C domain. In this study, we show the colocalization of prosaposin and ganglioside GM3 on NS20Y cell plasma membrane by scanning confocal microscopy.

Carbohydrate binding activity in human spermatozoa: localization, specificity, and involvement in sperm-egg fusion.

Sperm carbohydrate binding activity is involved in gamete recognition. We identified a human sperm protein extracted under reducing conditions, and with a molecular mass of 65 kDa on sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), and which binds D-mannose coupled to albumin (DMA) in presence of cations and a neutral pH. Epifluorescence microscopy showed that fluorescein-DMA binds to dead or permeabilized sperm heads.