Discovery of High Abundances of Aster-Like Nanoparticles in Pelagic Environments: Characterization and Dynamics.

This study reports the discovery of Aster-Like Nanoparticles (ALNs) in pelagic environments. ALNs are pleomorphic, with three dominant morphotypes which do not fit into any previously defined environmental entities [i.e., ultramicro-prokaryotes, controversed nanobes, and non-living particles (biomimetic mineralo-organic particles, natural nanoparticles or viruses)] of similar size. Elemental composition and selected-area electron diffraction patterns suggested that the organic nature of ALNs may prevail over the possibility of crystal structures. Likewise, recorded changes in ALN numbers in the absence of cells are at odds with an affiliation to until now described viral particles. ALN abundances showed marked seasonal dynamics in the lakewater, with maximal values (up to 9.0 ± 0.5 × 107 particles·mL-1) reaching eight times those obtained for prokaryotes, and representing up to about 40% of the abundances of virus-like particles. We conclude that (i) aquatic ecosystems are reservoirs of novel, abundant, and dynamic aster-like nanoparticles, (ii) not all virus-like particles observed in aquatic systems are necessarily viruses, and (iii) there may be several types of other ultra-small particles in natural waters that are currently unknown but potentially ecologically important.

Mycoplasmas are no exception to extracellular vesicles release: Revisiting old concepts.

Release of extracellular vesicles (EV) by Gram-negative and positive bacteria is being frequently reported. EV are nano-sized, membrane-derived, non-self-replicating, spherical structures shed into the extracellular environment that could play a role in bacteria-host interactions. Evidence of EV production in bacteria belonging to the class Mollicutes, which are wall-less, is mainly restricted to the genus Acholeplasma and is scanty for the Mycoplasma genus that comprises major human and animal pathogens. Here EV release by six Mycoplasma (sub)species of clinical importance was investigated. EV were obtained under nutritional stress conditions, purified by ultracentrifugation and observed by electron microscopy. The membrane proteins of EV from three different species were further identified by mass spectrometry as a preliminary approach to determining their potential role in host-pathogen interactions. EV were shown to be released by all six (sub)species although their quantities and sizes (30-220 nm) were very variable. EV purification was complicated by the minute size of viable mycoplasmal cells. The proteins of EV-membranes from three (sub)species included major components of host-pathogen interactions, suggesting that EV could contribute to make the host-pathogen interplay more complex. The process behind EV release has yet to be deciphered, although several observations demonstrated their active release from the plasma membrane of living cells. This work shed new light on old concepts of "elementary bodies" and "not-cell bound antigens".

A shear-induced network of aligned wormlike micelles in a sugar-based molecular gel. From gelation to biocompatibility assays.

A new low molecular weight hydrogelator with a saccharide (lactobionic) polar head linked by azide-alkyne click chemistry was prepared in three steps. It was obtained in high purity without chromatography, by phase separation and ultrafiltration of the aqueous gel. Gelation was not obtained reproducibly by conventional heating-cooling cycles and instead was obtained by shearing the aqueous solutions, from 2 wt% to 0.25 wt%. This method of preparation favored the formation of a quite unusual network of interconnected large but thin 2D-sheets (7nm-thick) formed by the association side-by-side of long and aligned 7nm diameter wormlike micelles. It was responsible for the reproducible gelation at the macroscopic scale. A second network made of helical fibres with a 10-13nm diameter, more or less intertwined was also formed but was scarcely able to sustain a macroscopic gel on its own. The gels were analysed by TEM (Transmission Electronic Microscopy), cryo-TEM and SAXS (Small Angle X-ray Scattering). Molecular modelling was also used to highlight the possible conformations the hydrogelator can take. The gels displayed a weak and reversible transition near 20°C, close to room temperature, ascribed to the wormlike micelles 2D-sheets network. Heating over 30°C led to the loss of the gel macroscopic integrity, but gel fragments were still observed in suspension. A second transition near 50°C, ascribed to the network of helical fibres, finally dissolved completely these fragments. The gels showed thixotropic behaviour, recovering slowly their initial elastic modulus, in few hours, after injection through a needle. Stable gels were tested as scaffold for neural cell line culture, showing a reduced biocompatibility. This new gelator is a clear illustration of how controlling the pathway was critical for gel formation and how a new kind of self-assembly was obtained by shearing.

Ultrastructural analysis of healthy synovial fluids in three mammalian species.

A better knowledge of synovial fluid (SF) ultrastructure is required to further understand normal joint lubrication and metabolism. The aim of the present study was to elucidate SF structural features in healthy joints from three mammalian species of different size compared with features in biomimetic SF. High-resolution structural analysis was performed using transmission electron microscopy (TEM) and scanning electron microscopy (SEM) and environmental SEM/wet scanning transmission electron microscopy mode complemented by TEM and SEM cryogenic methods. Laser-scanning confocal microscopy (LCM) was used to locate the main components of SF with respect to its ultrastructural organization. The present study showed that the ultrastructure of healthy SF is built from a network of vesicles with a size range from 100 to a few hundred nanometers. A multilayered organization of the vesicle membranes was observed with a thickness of about 5 nm. LCM study of biological SF compared with synthetic SF showed that the microvesicles consist of a lipid-based membrane enveloping a glycoprotein gel. Thus, healthy SF has a discontinuous ultrastructure based on a complex network of microvesicles. This finding offers novel perspectives for the diagnosis and treatment of synovial joint diseases.

Internalization pathways into cancer cells of gadolinium-based radiosensitizing nanoparticles.

Over the last few decades, nanoparticles have been studied in theranostic field with the objective of exhibiting a long circulation time through the body coupled to major accumulation in tumor tissues, rapid elimination, therapeutic potential and contrast properties. In this context, we developed sub-5 nm gadolinium-based nanoparticles that possess in vitro efficient radiosensitizing effects at moderate concentration when incubated with head and neck squamous cell carcinoma cells (SQ20B). Two main cellular internalization mechanisms were evidenced and quantified: passive diffusion and macropinocytosis. Whereas the amount of particles internalized by passive diffusion is not sufficient to induce in vitro a significant radiosensitizing effect, the cellular uptake by macropinocytosis leads to a successful radiotherapy in a limited range of particles incubation concentration. Macropinocytosis processes in two steps: formation of agglomerates at vicinity of the cell followed by their collect via the lamellipodia (i.e. the "arms") of the cell. The first step is strongly dependent on the physicochemical characteristics of the particles, especially their zeta potential that determines the size of the agglomerates and their distance from the cell. These results should permit to control the quantity of particles internalized in the cell cytoplasm, promising ambitious opportunities towards a particle-assisted radiotherapy using lower radiation doses.