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Biomedical subjects

Pierre Schaaf

Publications and source records attributed to Pierre Schaaf.

At least 37 records · Page 2Linked to original sources
PubMed2005

Effect of thiocyanate counterion condensation on poly(allylamine hydrochloride) chains on the buildup and permeability of polystyrenesulfonate/polyallylamine polyelectrolyte multilayers.

In this study, we investigate the buildup of PEI-(PSS-PAH)(n) polyelectrolyte multilayers at pH 7.4 in the presence of either NaCl or NaSCN as a supporting electrolyte. It appears that in the presence of increasing thiocyanate concentrations (from 0.1 to 0.5 M), the thickness increment, obtained from optical waveguide lightmode spectroscopy experiments, increases whereas it stays practically constant for increasing sodium chloride concentrations (between 0.1 and 0.5 M). The hydration of the films differs also markedly between both electrolyte solutions. The differences in the construction of the polyelectrolyte multilayers in the presence of both supporting electrolytes are rationalized in terms of strong SCN(-) condensation on the PAH chains. The occurrence of this ion condensation is indirectly demonstrated by means of zeta potential measurements and directly demonstrated by means of attenuated total internal reflection infrared spectroscopy on the multilayer films. Moreover when the films are built up in the presence of SCN(-), these ions are only slowly exchanged by the Cl(-) ions introduced in the bulk. Conversely the thick films obtained from 0.5 M NaSCN solutions do not deswell when the buffer solution is replaced by a 0.5 M NaCl containing buffer. The permeability of the films constructed in the presence of both sodium salts is also studied by means of cyclic voltametry and is found to be markedly different in the case of films made from five bilayers at 0.5 M salt concentration. This difference is due to the different morphology and porosity of the films constructed in the presence of 0.5 M NaCl and 0.5 M NaSCN.

Biocompatible Materials↗
PubMed2005

Glassy state of polystyrene sulfonate/polyallylamine polyelectrolyte multilayers revealed by the surface force apparatus.

The interactions between two poly(allylamine)/poly(styrene sulfonate) multilayers made of 4.5 and 5 bilayers are investigated by the surface force apparatus (SFA). As the two surfaces approach, one reaches a threshold point where a repulsion sets in, until they become barely compressible. Repetitive load/unload cycles show that, once compressed, the films remain almost in their compressed state. This indicates that the poly(allylamine)/poly(styrene sulfonate) films are in a glassy state, in marked difference with the SFA findings on poly-(L-lysine)/poly-L-glutamic acid) multilayers. These results are discussed in the light of linearly and exponentially growing films.

Letter↗
PubMed2005

Endothelial cell--interactions with polyelectrolyte multilayer films.

The seeding of endothelial cells (ECs) on biomaterial surfaces became a major challenge, allowing to improve the non-thrombogenic properties of these surfaces. Recently, the use of polyelectrolyte films has been suggested as a new versatile technique of surface modification aimed at tissue engineering. In this study, we evaluate the adhesion properties of ECs on two types of polyelectrolyte films ending either by poly(D-lysine) (PDL), or poly(allylamine hydrochloride) (PAH), and compared them to data obtained on PDL or PAH monolayers, glass and fibronectin (Fn)-coated glass. ECs seeded on polyelectrolyte films showed a good morphology, allowing ECs to resist physiological shear stress better compared to ECs seeded on glass or Fn. The expression of beta1 integrins was slightly lower on polyelectrolyte films than on control surfaces. However, the phosphorylation of focal adhesion kinase, involved in the transduction of adhesion signal, was not modified on PAH ending films compared to control surfaces; whereas it became lower on PDL ending films. Finally, PAH ending films improve strongly ECs adhesion without disturbing the adhesion mechanism, necessary for the development of a new endothelium. These types of films or similar build-ups could thus be used in the future as a way to modify surfaces for vascular tissue engineering.

Actins↗
PubMed2005

Application of fluorescence recovery after photobleaching to diffusion of a polyelectrolyte in a multilayer film.

The diffusion coefficient, D, and the proportion of mobile molecules, p, is measured for a fluorescently labeled polyelectrolyte in a multilayer film using fluorescence recovery after photobleaching (FRAP). The film was composed of poly(L-lysine) (PLL) and hyaluronan (HA). The labeled polyelectrolyte (PLL(FITC)) was either deposited on top of the film or embedded within it. A circular area of diameter approximately 60 microm was bleached using a confocal laser scanning microscope. Because molecules do already diffuse during the bleaching step, the initial light intensity profile is not characteristic of a uniformly bleached circular area. A formalism is developed in which a simple mathematical representation of a measured profile serves as starting profile. This radial distribution is introduced in the equation describing the time evolution of the labeled molecule concentration under the hypothesis that the recovery results from pure, two-dimensional Brownian diffusion of the mobile molecules according to Fick's law. The analysis of a series of images taken at successive times after bleaching (up to approximately one hour) leads to estimates of D of the order of 0.2 microm(2) s(-1) for labeled molecules deposited on top of the film and a 5-fold smaller value for the molecules embedded in the film. However, p is remarkably insensitive to the position in the multilayer architecture (p approximately 0.40).

Diffusion↗
PubMed2005

Effect of functionalization of multilayered polyelectrolyte films on motoneuron growth.

We studied in vitro cell-substrate interaction of motoneurons with functionalized polylectrolyte films. Thin polylectrolyte films were built on glass by alternating polycations, poly(ethylene-imine) PEI, poly(L-lysine) PLL, or poly(allylamine hydrochloride) PAH, and polyanions, poly(sodium-4-styrenesulfonate) PSS or poly(L-glutamic acid) (PGA). These architectures were functionalized with Brain Derived Neurotrophic Factor (BDNF) or Semaphorin 3A (Sema3A). We used Optical Waveguide Lightmode Spectroscopy (OWLS) and Atomic Force Microscopy (AFM) to characterize the architectures. The viability of motoneurons was estimated by the acid phosphatase method, and morphometrical measures were performed to analyse the influence of different architectures on cell morphology. Motoneurons appeared to adhere and spread on all the architectures tested and preferentially on PSS ending films. The viability of motoneurons on polyelectrolyte multilayers was higher compared to polyelectrolyte monolayers. BDNF and Sema3A embedded in the films remained active and thereby create functionalized nanofilms.

Animals↗
PubMed2005

Polyelectrolyte multilayers functionalized by a synthetic analogue of an anti-inflammatory peptide, alpha-MSH, for coating a tracheal prosthesis.

Polyelectrolyte multilayer films made of poly (L-lysine) (PLL) and poly (L-glutamic acid) (PGA) have been functionalized by covalent binding of a synthetic analogue of the anti-inflammatory peptide, alpha-melanocyte-stimulating hormone (alpha-MSH) to PGA to create biologically active coatings for tracheal prostheses. The morphology and in vivo stability of the films were investigated by atomic force microscopy and confocal laser scanning microscopy, respectively. For the in vivo evaluation, 87 rats were implanted and examined for a period superior to 3 months. Histological analysis, performed 1 month after implantation, showed a fibroblast colonization of the periprosthetic side and a respiratory epithelium type on the endoluminal side of the implant for all the polyelectrolyte coatings tested. However, for prostheses modified by PGA ending multilayer films, a more regular and less obstructive cell layer was observed on the endoluminal side compared to those modified by PLL ending films. Systemic anti-inflammatory IL-10 production was only detected in rats implanted with prostheses functionalized by alpha-MSH, demonstrating, in vivo, the anti-inflammatory activity of the embedded peptide into multilayer architectures.

Animals↗
PubMed2005

Natural polyelectrolyte films based on layer-by layer deposition of collagen and hyaluronic acid.

The aim of the present work was to assemble extracellular matrix components into polyelectrolyte multilayers using the layer-by-layer deposition method. The films are constructed with type-I collagen and hyaluronic acid. The construction exhibits the general features observed during polyelectrolyte multilayer buildup: alternate positive and negative values of the zeta potential of the film during its construction and regular increase of the film thickness with the number, n, of deposition step. This increase is shown to be linear with n. As expected for a linearly growing film, the confocal microscopy shows that when the film is brought in contact with a collagen solution, collagen does not diffuse into the film but interacts only with its outer layer. However, the films are not constituted of homogeneously distributed polyanion/polycation complexes as it is usually observed, but they are formed of fibers as imaged by AFM. The typical width of these fibers increases with the number of deposition steps. Finally, it is found that chondrosarcoma cells spread well and synthesize extracellular matrix components only on the collagen ending films, whereas no cellular matrix was found for HA ending ones. Such architectures may be further functionalized by inclusion of active drugs, peptides, proteins..., and could be used as tunable biomaterial interfaces.

Biocompatible Materials↗
PubMed2005

Antifungal coating by biofunctionalized polyelectrolyte multilayered films.

The surface of medical devices is a common site of bacterial and fungal adhesion, first step to the constitution of a resistant biofilm leading frequently to chronic infections. In order to prevent such complications, several physical and chemical modifications of the device surface have been proposed. Here, we experiment a new type of topical antifungal coating using the layer-by-layer technique. The nanometric multilayer film obtained by this technique is functionalized by the insertion of a chromogranin A-derived antifungal peptide (CGA 47-66, chromofungin). We show that the embedded peptide keeps its antifungal activity by interacting with the fungal membrane and penetrating into the cell. In vitro studies demonstrate that such an antifungal coating is able to inhibit the growth of yeast Candida albicans by 65% and completely stop the proliferation of filamentous fungus Neurospora crassa. The cytotoxicity of such a coating was also assessed by growing human gingival fibroblasts at its surface. Finally, the antifungal coating of poly(methylmethacrylate), a widely used material for biomedical devices, is successfully tested in an in vivo oral candidiasis rat model. Taken together, these results assessed the functionalized multilayer films containing a new potent antifungal non-toxic peptide, as a novel and promising technique for local antifungal protection.

Amino Acid Sequence↗
PubMed2004

Measurement of film thickness up to several hundreds of nanometers using optical waveguide lightmode spectroscopy.

Up to now, most studies based on optical waveguide lightmode spectroscopy (OWLS) were dedicated to thin adlayers, assumed to be isotropic and homogeneous, for which data analysis was based on an approximation of the mode equations valid when the thickness is small with respect to the wavelength of the laser light. The aim of the present paper is to extend the use of OWLS to thicker deposited layers (up to approximately 400 nm). Both the simplified and extended models are compared in terms of optical parameters, i.e. the refractive index nA, the thickness dA, and the optical mass QA, for experimental data obtained with polyelectrolyte multilayer films. The deviation of these parameters can be quite large when derived using the simplified model instead of the extended model. This observation evidences that OWLS is well suited for the study of "thick" films if the appropriate model is applied to the data analysis.

Algorithms↗
PubMed2004

Multicompartment films made of alternate polyelectrolyte multilayers of exponential and linear growth.

The layer by layer deposition process of polyelectrolytes is used to construct films equipped with several compartments containing "free polyelectrolytes". Each compartment corresponds to a stratum of an exponentially growing polyelectrolyte multilayer film, and two consecutive compartments are separated by a stratum composed of a linearly growing multilayer that acts as a barrier preventing polyelectrolyte diffusion from one compartment to another. We use hyaluronic acid/poly(L-lysine) as the system to build the compartments and the poly(styrene sulfonate)/poly(allylamine) system for the barrier. Using confocal microscopy, it is shown that poly(L-lysine) diffuses only within the compartment in which it was initially introduced during the film construction and is thus unable to cross the barriers. Using fluorescein isothiocyanate as a pH indicator, it is also shown that although poly(styrene sulfonate)/poly(allylamine) multilayers act as a barrier for polyelectrolytes, they do not prevent proton diffusion through the film. Such films open the route for multiple functionalization of biomaterial coatings.

Journal Article↗
PubMed2004

Giant liposome microreactors for controlled production of calcium phosphate crystals.

Calcium phosphates are among the most important biominerals in living organisms, where they play both a mechanical and a calcium storage role. Their growth in vivo is under strong biological control, and this process occurs in closed spaces. Our aim in this paper is to describe a microreactor system able to control the mineralization process within closed spaces. To this aim we produce giant liposomes containing calcium ions as active ions in the mineralization process, spermine as an activator of crystal growth, and alkaline phosphatase as a catalyst to convert phosphate esters into phosphates. These phosphate esters are provided in the form of p-nitrophenyl phosphate outside of the liposomes. It is demonstrated that these amphiphilic molecules are able to diffuse through the lipidic container and to be subsequently hydrolyzed under enzymatic catalysis into active phosphate species which interact with the already available calcium and spermine to produce calcium phosphates only in the interior of the liposomes. This opens the route to control the calcium phosphate particle size in biomimetic systems.

Alkaline Phosphatase↗
PubMed2004

Human serum albumin self-assembly on weak polyelectrolyte multilayer films structurally modified by pH changes.

Adsorption of proteins onto film surfaces built up layer by layer from oppositely charged polyelectrolytes is a complex phenomenon, governed by electrostatic forces, hydrogen bonds, and hydrophobic interactions. The amounts of the interacting charges, however, both in polyelectrolytes and in proteins adsorbed on such films are a function of the pH of the solution. In addition, the number and the accessibility of free charges in proteins depend on the secondary structure of the protein. The subtle interplay of all these factors determines the adsorption of the proteins onto the polyelectrolyte film surfaces. We investigated the effect of these parameters for polyelectrolyte films built up from weak "protein-like" polyelectrolytes (i.e., polypeptides), poly(L-lysine) (PLL), and poly(glutamic acid) (PGA) and for the adsorption of human serum albumin (HSA) onto these films in the pH range 3.0-10.5. It was found that the buildup of the polyelectrolyte films is not a simple function of the pure charges of the individual polyelectrolytes, as estimated from their respective pKa values. The adsorption of HSA onto (PLL/PGA)n films depended strongly on the polyelectrolyte terminating the film. For PLL-terminated polyelectrolyte films, at low pH, repulsion, as expected, is limiting the adsorption of HSA (having net positive charge below pH 4.6) since PLL is also positively charged here. At high pH values, an unexpected HSA uptake was found on the PGA-ending films, even when both PGA and HSA were negatively charged. It is suggested that the higher surface rugosity and the decrease of the alpha-helix content at basic pH values (making accessible certain charged groups of the protein for interactions with the polyelectrolyte film) could explain this behavior.

Absorption↗
PubMed2004

Layer by layer self-assembled polyelectrolyte multilayers with embedded phospholipid vesicles.

We describe a method to embed phospholipid vesicles into polyelectrolyte multilayers built up by the alternate deposition of polyanions and polycations. Before deposition, the vesicles are rigidified by polycation adsorption onto their surface avoiding their fusion once deposited on the multilayer surface. The vesicles adsorb to form a compact and "hard" monolayer as imaged by atomic force microscopy. The thickness of the adsorbed vesicle layer, of the order of 250 nm, is very close to the diameter of the vesicles in solution. This work should open the route to the buildup of multilayer films containing phospholipid vesicles that could act as "reservoirs" for drugs or enzymatic nanoreactors.

Adsorption↗
PubMed2004

Layer by layer buildup of polysaccharide films: physical chemistry and cellular adhesion aspects.

The formation ofpolysaccharide films based on the alternate deposition of chitosan (CHI) and hyaluronan (HA) was investigated by several techniques. The multilayer buildup takes place in two stages: during the first stage, the surface is covered by isolated islets that grow and coalesce as the construction goes on. After several deposition steps, a continuous film is formed and the second stage of the buildup process takes place. The whole process is characterized by an exponential increase of the mass and thickness of the film with the number of deposition steps. This exponential growth mechanism is related to the ability of the polycation to diffuse "in" and "out" of the whole film at each deposition step. Using confocal laser microscopy and fluorescently labeled CHI, we show that such a diffusion behavior, already observed with poly(L-lysine) as a polycation, is also found with CHI, a polycation presenting a large persistence length. We also analyze the effect of the molecular weight (MW) of the diffusing polyelectrolyte (CHI) on the buildup process and observe a faster growth for low MW chitosan. The influence of the salt concentration during buildup is also investigated. Whereas the CHI/HA films grow rapidly at high salt concentration (0.15 M NaCl) with the formation of a uniform film after only a few deposition steps, it is very difficult to build the film at 10(-4) M NaCl. In this latter case, the deposited mass increases linearly with the number of deposition steps and the first deposition stage, where the surface is covered by islets, lasts at least up to 50 bilayer deposition steps. However, even at these low salt concentrations and in the islet configuration, CHI chains seem to diffuse in and out of the CHI/HA complexes. The linear mass increase of the film with the number of deposition steps despite the CHI diffusion is explained by a partial redissolution of the CHI/HA complexes forming the film during different steps of the buildup process. Finally, the uniform films built at high salt concentrations were also found to be chondrocyte resistant and, more interestingly, bacterial resistant. Therefore, the (CHI/HA) films may be used as an antimicrobial coating.

Animals↗
PubMed2002

Viability, adhesion, and bone phenotype of osteoblast-like cells on polyelectrolyte multilayer films.

The aim of this study was to develop new biocompatible coatings for bone implants by the alternating deposition of oppositely charged polyelectrolytes. Polyelectrolyte films were built up with different terminating layers on which SaOS-2 osteoblast-like cells and human periodontal ligament (PDL) cells were grown. The terminating layer was made of one of the following polyelectrolytes: poly(ethylene imine) (PEI), poly(sodium 4-styrenesulfonate) (PSS), poly(allylamine hydrochloride) (PAH), poly(L-glutamic acid) (PGA), or poly(L-lysine) (PLL). Cell adherence, viability, stability of osteoblast phenotype, and inflammatory response were studied. Adherence and viability were good on all terminating layers except the PEI-terminating layer, which was cytotoxic. Maintenance of osteoblast phenotype marker expression was observed on PSS- and PGA-terminating films for both cell types, whereas downregulation, associated with the induction of Interleukin-8 (IL-8) secretion, was detected on PEI and PAH for both cell types and on PLL for PDL cells. These results suggested a good biocompatibility of PSS- and PGA-ending films for PDL cells and of PSS-, PGA-, and PLL-terminating films for SaOS-2 cells. As a result, polyelectrolyte multilayer films could emerge as new alternatives for implant coatings.

Alkaline Phosphatase↗
PubMed2002

Multi-bead-and-spring model to interpret protein detachment studied by AFM force spectroscopy.

This article deals with the detachment of molecules (fibrinogen) from a surface studied experimentally with an atomic force microscope. The detachment (or rupture) forces are measured as a function of the retraction velocity and exhibit a clear dependence on this parameter, even though the interaction between the molecules and the surface are nonspecific. To interpret these data, a mechanical multi-bead-and-spring model is developed. It consists of one to several parallel, "molecular" springs connected to an extra spring representing the cantilever that is moved at constant velocity. The free end of each molecular spring terminates with a particle that interacts with the surface through a Lennard-Jones potential. This Brownian dynamics model is used to analyze the experimental findings. In the framework of this model, it appears that the fibrinogen molecule must be ascribed a stiffness much smaller than that of the cantilever. In addition, several bonds between the molecule and the surface must be taken into account for the range of the molecule-surface interaction not to be unrealistically small. In future work, this model will be extended to more complex mechanisms such as the detachment of cells from a surface.

Biophysical Phenomena↗
PubMed2002

Protein adsorption onto auto-assembled polyelectrolyte films.

Surface modification by deposition of ordered protein systems constitutes one of the major objectives of bio-related chemistry and biotechnology. In this respect a concept has recently been reported aimed at fabricating multilayers by the consecutive adsorption of positively and negatively charged polyelectrolytes. We investigate the adsorption processes between polyelectrolyte multilayers and a series of positively and negatively charged proteins. The film buildup and adsorption experiments were followed by Scanning Angle Reflectometry (SAR). We find that proteins strongly interact with the polyelectrolyte film whatever the sign of the charge of both the multilayer and the protein. When charges of the multilayer and the protein are similar, one usually observes the formation of protein monolayers, which can become dense. We also show that when the protein and the multilayer become oppositely charged, the adsorbed amounts are usually larger and the formation of thick protein layers extending up to several times the largest dimension of the protein can be observed. Our results confirm that electrostatic interactions dominate protein/polyelectrolyte multilayer interactions.

Adsorption↗