Jalal Bacharouche

Immunomodulation with Self-Crosslinked Polyelectrolyte Multilayer-Based Coatings

This study aims to design an optimal polyelectrolyte multilayer film of poly-l-lysine (PLL) and hyaluronic acid (HA) as an anti-inflammatory cytokine release system in order to decrease the implant failure due to any immune reactions. The chemical modification of the HA with aldehyde moieties allows self-cross-linking of the film and an improvement in the mechanical properties of the film. The cross-linking of the film and the release of immunomodulatory cytokine (IL-4) stimulate the differentiation of primary human monocytes seeded on the films into pro-healing macrophages phenotype. This induces the production of anti-inflammatory cytokines (IL1-RA and CCL18) and the decrease of pro-inflammatory cytokines secreted (IL-12, TNF-α, and IL-1β). Moreover, we demonstrate that cross-linking PLL/HA film using HA-aldehyde is already effective by itself to limit inflammatory processes. Finally, this functionalized self-cross-linked PLL/HA-aldehyde films constitutes an innovative and efficient candidate for immunomodulation of any kind of implants of various architecture and properties.

In situ and real time investigation of the evolution of a Pseudomonas fluorescens nascent biofilm in the presence of an antimicrobial peptide.

Against the increase of bacterial resistance to traditional antibiotics, antimicrobial peptides (AMP) are considered as promising alternatives. Bacterial biofilms are more resistant to antibiotics that their planktonic counterpart. The purpose of this study was to investigate the action of an AMP against a nascent bacterial biofilm. The activity of dermaseptin S4 derivative S4(1-16)M4Ka against 6 h-old Pseudomonas fluorescens biofilms was assessed by using a combination of Attenuated Total Reflectance-Fourier Transform InfraRed (ATR-FTIR) spectroscopy in situ and in real time, fluorescence microscopy using the Baclight™ kit, and Atomic Force Microscopy (AFM, imaging and force spectroscopy). After exposure to the peptide at three concentrations, different dramatic and fast changes over time were observed in the ATR-FTIR fingerprints reflecting a concentration-dependent action of the AMP. The ATR-FTIR spectra revealed major biochemical and physiological changes, adsorption/accumulation of the AMP on the bacteria, loss of membrane lipids, bacterial detachment, bacterial regrowth, or inhibition of biofilm growth. AFM allowed estimating at the nanoscale the effect of the AMP on the nanomechanical properties of the sessile bacteria. The bacterial membrane elasticity data measured by force spectroscopy were consistent with ATR-FTIR spectra, and they allowed suggesting a mechanism of action of this AMP on sessile P. fluorescens. The combination of these three techniques is a powerful tool for in situ and in real time monitoring the activity of AMPs against bacteria in a biofilm.

Auxiliary Biomembranes as a Directional Delivery System To Control Biological Events in Cell-Laden Tissue-Engineering Scaffolds

Delivery of growth factors is an indispensable part of tissue engineering. Here, we describe a detachable membrane-based release system composed of extracellular matrix components that can be attached to hydrogels to achieve directional release of bioactive molecules. This way, the release of cytokines/growth factors can be started at a desired point of tissue maturation or directly in vivo. As a model, we develop thin films of an interpenetrating network of double-cross-linked gelatin and hyaluronic acid derivatives. The use of the auxiliary release system with vascular endothelial growth factor results in extensive sprouting by encapsulated vascular endothelial cells. The presence of the release system with interleukin-4 results in clustering of encapsulated macrophages with a significant decrease in M1 macrophages (proinflammatory). This system can be used in conjunction with three-dimensional structures as an auxiliary system to control artificial tissue maturation and growth.

On the Infectivity of Bacteriophages in Polyelectrolyte Multilayer Films: Inhibition or Preservation of Their Bacteriolytic Activity?

Antibiotic resistance in bacterial cells has motivated the scientific community to design new and efficient (bio)materials with targeted bacteriostatic and/or bactericide properties. In this work, a series of polyelectrolyte multilayer films differing in terms of polycation-polyanion combinations are constructed according to the layer-by-layer deposition method. Their capacities to host T4 and φx174 phage particles and maintain their infectivity and bacteriolytic activity are thoroughly examined. It is found that the macroscopic physicochemical properties of the films, which includes film thickness, swelling ratio, or mechanical stiffness (as derived by atomic force microscopy and spectroscopy measurements), do not predominantly control the selectivity of the films for hosting infective phages. Instead, it is evidenced that the intimate electrostatic interactions locally operational between the loaded phages and the polycationic and polyanionic PEM components may lead to phage activity reduction and preservation/enhancement, respectively. It is argued that the underlying mechanism involves the screening of the phage capsid receptors (operational in cell recognition/infection processes) because of the formation of either polymer-phage hetero-assemblies or polymer coating surrounding the bioactive phage surface.

Low Biotinyl Glycogen: A Model for Single-Molecule Force Analysis of Branched Biological Macromolecules

Characterization of polysaccharides in their native-like forms is of great importance for their subsequent application in the pharmaceuticals and biomedicine fields. Commercial glycogen molecules immobilized on a modified gold surface were characterized using a combination of Single Molecule Force and InfraRed spectroscopies. For this, low biotinylated glycogen was synthetized and subsequently immobilized on a Bovine Serum Albumin/Streptavidin-coated gold modified surface. The presence of glycogen was ascertained by vibrational spectra and confirmed based on the specific interaction of concanavalin A functionalized AFM tips with -glucose residues. Theoretical values of the particle size of this glycogen fitted well with the structural observations. Conformational properties were deduced from the force curves using the FJC model. The studied glycogen showed a compact slightly ramified structure with molecular elongations up to 1000 nm. The investigation of other conformational properties showed similarities with bacterial glycogen, however some differences that are possibly related to the different biosynthesis pathways were observed.