Biomedical subjects

K Anselme

Publications and source records attributed to K Anselme.

At least 19 recordsLinked to original sources

PubMed2006

Physico-chemical characteristics and protein adsorption potential of hydroxyapatite particles: influence on in vitro biocompatibility of ceramics after sintering.

Through the example of two HA ceramics prepared from two HA powders (HAD and HAL), we explored the relation between the physico-chemical qualities of the initial HA powder and the final HA ceramic and their influence on the protein adsorption and cell response to the final HA ceramics. The powders were characterized by XRD, FT-IR, zeta potential, and specific surface area (SSA). Their protein adsorption potential was tested after immersion in culture medium +15% of fetal calf serum. These results were correlated with the protein adsorption potential of the two ceramics (cHAD and cHAL) prepared from the HAD and HAL powders respectively and to the cell attachment after 4, 24 and 72 h on the ceramics. From our results, it appears that a relation can be established between the physico-chemical characteristics of the initial HA powders and the final biological response to the sintered ceramics prepared from these powders. An inverse relation exists between the SSA and the protein adsorption capacity of HA powders and the protein adsorption and cell attachment on HA ceramics. This inverse relation is related to phenomenon occurring during the sintering phase and the formation of inter-granular micro-porosity.

Adsorption↗

PubMed2006

Statistical demonstration of the relative effect of surface chemistry and roughness on human osteoblast short-term adhesion.

The effects of material composition, surface chemistry or surface topography on cell attachment (short-term adhesion) have been largely studied on bone-derived cells. However, no statistical demonstration of these effects has been performed until now. With this objective, we quantified the attachment after 24 hours of human osteoblasts on pure titanium, titanium alloy and stainless steel substrates presenting 6 different surface morphologies and 2 different roughness amplitude obtained by sand-blasting, electro-erosion, acid etching, polishing and machine-tooling. The coating by a gold-palladium layer of these surfaces allowed determining the relative effect of the surface roughness and of the surface chemistry. By multiple analysis of variance, we demonstrated that neither material composition nor surface roughness amplitude influenced cell attachment except on sandblasted pure titanium substrates. On the contrary, a high significant influence of the process used to produce the surface was observed meaning that the main influent factor on cell attachment could be either the surface morphology or the surface chemistry induced by the process. As the coating of surfaces by a gold-palladium layer decreased significantly the attachment of cells on the majority of substrates, we concluded that attachment is rather influenced by surface chemistry than by surface topography.

Adhesiveness↗

PubMed2005

A kinetic approach to osteoblast adhesion on biomaterial surface.

An incompletely understood question in the field of biomaterials is how eucaryotic cells adhere on material surfaces. The adhesion of cells on materials is generally studied after some hours. Because this evaluation after some hours cannot let us presume about the future of the cells on the material, we have developed a culture model that does allow study in the long term of an elaborate cell/material interface closer to the in vivo situation. For that, we used a progressive trypsin-based detachment method. Here we report on the mathematical modeling of long-term human primary osteoblastic cell adhesion on metallic substrates, which allows us to quantify the real adhesion simultaneously by taking into account the effect of cell proliferation. A time-dependent adhesion index t(d) is proposed, which varies with culture time t according to the power law: t(d)(t) = at(b), a being independent of b. The exponent b is equal to 0.5 +/- 0.03 and is independent of the substrate's characteristics, meaning that the long-term adhesion increases proportionally to the square root of culture time. On the contrary, the parameter a significantly depends on the material's nature, the surface's topography, and the surface chemistry of the substrate and is sufficient to characterize cell adhesion. From this relationship, we suggest that a diffusion-based process related to the kinetic of formation of extracellular matrix should be involved in long-term adhesion on materials.

Biocompatible Materials↗

PubMed2005

Bootstrap analysis of the relation between initial adhesive events and long-term cellular functions of human osteoblasts cultured on biocompatible metallic substrates.

Classically, the evaluation of cellular adhesion of cells on substrates is limited to the evaluation of cell attachment after some hours. We have claimed for several years that this evaluation is incomplete concerning the evaluation of cell adhesion and more precisely of the quality of the in vitro cell/biomaterial interface. With a view to demonstrating this assertion, we develop in this paper statistical correlations between short-term adhesion (STA) evaluating the attachment after 24 h (IA: initial attachment) and long-term adhesion (LTA) evaluating the strength of the cell/matrix substrate interface over 21 days of culture (AP: adhesion power). Additionally, as the adhesion phase is known to influence further growth of cells we proceed to the correlation of STA with the number of cells after 21 days. We demonstrate statistically the expected positive relation existing between STA and cell growth and we show that this relation is totally independent of the substrate's surface topography or chemistry. More surprisingly, we demonstrate the absence of correlation between IA and AP. This illustrates the fact that different mechanisms underlie STA and LTA. Moreover, this study demonstrates that the evaluation of the number of attached cells after some hours cannot let us presume either that cells will survive or that they will adhere at later times by forming a complex cell/substrate interface by synthesis of extracellular matrix proteins. Finally, the originality of this work lies in the extensive statistical correlation analysis performed between biological parameters describing the cell behaviour on a substrate.

Alloys↗

PubMed2005

Statistical correlation between cell adhesion and proliferation on biocompatible metallic materials.

Our ambition for several years is to appreciate and quantify the long-term adhesion of cells on materials at times where the interface between cells and substrate becomes more complex, more closed to the cell/matrix/substrate interface existing in vivo. With this objective, we quantified the long-term adhesion and proliferation of human osteoblasts cultured from 24 h to 21 days on pure titanium, titanium alloy, and stainless-steel substrates presenting six different surface morphologies and two different roughness amplitude. Hence, we did proceed to the statistical correlation of cell adhesion and cell proliferation on 30 different substrates. Additionally, we described surface topography not only by the roughness amplitude but also by the roughness morphology using new specific parameters. By multiple analysis of variance, we demonstrated that nor material composition nor surface roughness amplitude did influence cell proliferation, whereas a very significant influence of the process used to produce the surface was observed meaning that the main influent factor on cell proliferation was the surface morphology. The long-term adhesion and proliferation capacity of cells were positively correlated on 23 types of substrates on 30, this positive correlation being statistically asserted on 13 types of substrates on 23. This study is the first demonstration of the existence of a statistical correlation between long-term adhesion and proliferation capacity of human bone cells on substrates with various chemical composition, surface chemistry, and surface topography.

Biocompatible Materials↗

PubMed2004

Topography effects of pure titanium substrates on human osteoblast long-term adhesion.

Classically various treatments are applied to increase the roughness of titanium implants and improve their integration in the tissues. Many in vitro studies have been performed to better understand the mechanisms underlying the adhesion of cells on materials. Frequently, the adhesion is related to the attachment of cells during the first hours of contact with the substrate. For several years, our objective has been to develop experimental methods to evaluate the long-term adhesion of human osteoblasts from some hours to several weeks in order to model in vitro a tissue-like interface. This culture model allows for the formation over 21 days of a complex osteoblast/extracellular matrix/material interface. We recently developed a new parameter called adhesion power (AP) to evaluate this long-term adhesion. In this study, our objective is to check its efficiency in discriminating the long-term adhesion of human osteoblasts on pure titanium substrates with seven different surface morphologies obtained by electro-erosion, sandblasting, polishing, acid-etching and machine-tooling. By scanning electron microscopy, we observed that the human osteoblasts did spread more intimately on surface with low roughness amplitude than on rough ones. However, the AP was higher on rough isotropic surfaces obtained by electro-erosion, sandblasting or acid-etching and lower on smoother surfaces obtained by polishing and machine-tooling. We demonstrated that the AP was pertinent for evaluating human osteoblast's long-term adhesion on pure titanium surfaces with various roughness parameters. Its correlation with the order parameter, which describes the organization of the roughness, confirmed once more that human osteoblasts are more sensitive to the organization and morphology of the roughness than to its amplitude.

Cell Adhesion↗

PubMed2004

Time-dependent morphology and adhesion of osteoblastic cells on titanium model surfaces featuring scale-resolved topography.

The role of micrometer and submicrometer surface roughness on the interaction of cells with titanium model surfaces of well-defined topography was investigated using human bone-derived cells (MG63 cells). The early phase of interactions was studied using a kinetic morphological analysis of adhesion, spreading and proliferation of the cells. By SEM and double immunofluorescent labeling of vinculin and actin, it was found that the cells responded to nanoscale roughness by a higher cell thickness and a delayed apparition of the focal contacts. A singular behavior was observed on nanoporous oxide surfaces, where the cells were more spread and displayed longer and more numerous filopods. On electrochemically microstructured surfaces with hemispherical cavities, arranged in a hexagonal pattern, the MG63 cells were able to go inside, adhere and proliferate in cavities of 30 or 100 microm in diameter, whereas they did not recognize the 10 microm diameter cavities. Cells adopted a 3D shape when attaching inside the 30 microm diameter cavities. Condensation of actin cytoskeleton correlated with vinculin-positive focal contacts on cavity edges were observed on all microstructured surfaces. Nanotopography on surfaces with 30 microm diameter cavities had little effect on cell morphology compared to flat surfaces with same nanostructure, but cell proliferation exhibited a marked synergistic effect of microscale and nanoscale topography.

Adaptation, Physiological↗

PubMed2004

Effect of dexamethasone on moesin gene expression in rabbit bone marrow stromal cells.

The influence of dexamethasone on rabbit bone marrow stromal cells differentiation was studied by screening the action of dexamethasone on gene expression. Using differential display, we observed some differential amplifications. The use of five of thirteen different primers combination allowed to identify one or more differential bands. One of them was identified as moesin gene. Real-time PCR confirmed a significant reduction of moesin gene expression following dexamethasone treatment. The decrease of expression for this protein, involved in cytoskeletal organization, could explain the effects of dexamethasone treatment on bone marrow stromal cells differentiation.

Animals↗

PubMed2004

Kinetic study of the expression of beta-catenin, actin and vinculin during osteoblastic adhesion on grooved titanium substrates.

Intercellular adhesions are known to play an important role in differentiation of osteoblasts and in the development of bone tissue architecture. However, to our knowledge, they have never been studied during the formation of bone tissue in contact with a biomaterial surface. In an in vitro kinetic study, we followed the expression of proteins involved in cell-cell interactions (beta-catenin), in cell-material interactions (vinculin) and in cytoskeleton (actin) of human osteoblastic cells cultured on grooved titanium-based substrates during 1, 2, 4, 6, 24, 48, and 72 hours. The human osteoblasts aligned themselves in the 150 microm wide grooves only after 24 hours. The distribution of vinculin-positive focal contacts, actin cytoskeleton and beta-catenin positive-adherens junctions was not significantly influenced by the cell alignment. beta-catenin-positive adherens junctions were expressed by human osteoblasts as soon as 1 hour after inoculation. At this time, they showed a patch-like aspect along cytoplasmic processes in contact with an underlying or an adjacent cell. After 2 hours, the patches were more and more numerous underlining the connections between cells. After 4 hours and more, the patches were organised in a parallel arrangement perpendicular to the two connected cells forming a "zip-like" aspect. Additionally, using double immuno-staining, we demonstrated that sometimes beta-catenin and vinculin appeared co-localised and sometimes not. The linkage of catenin/cadherin complex and vinculin-positive focal contacts with actin filaments may explain this apparent co-localisation.

Biocompatible Materials↗

PubMed2002

Effect of grooved titanium substratum on human osteoblastic cell growth.

Various surface treatments have been developed to increase the clinical performance of titanium-based implants. Many in vitro tests have been carried out on substrates with varied surface topography for a complete understanding of osteoblasts. In previous research, we made the observation that surface roughness must be taken into account, not only in terms of amplitude but also in terms of organization. In this study, we tested the adhesion and proliferation of human primary osteoblasts on grooved titanium surfaces with various amplitudes and organizations of topography. The roughness was described at a scale above (macro-roughness) or below (micro-roughness) the cell size. We observed better orientation and proliferation of human osteoblasts on surfaces with a micro-roughness characterized by a lower Order (parameter describing the organization of topography) and by a higher Ra and Rz (parameters describing the amplitude of topography). It appears that cultured human osteoblasts prefer surfaces with relatively high micro-roughness amplitude and with a low level of repeatability.

Biocompatible Materials↗

PubMed2002

Improvement in the morphology of Ti-based surfaces: a new process to increase in vitro human osteoblast response.

Surface roughness has been shown to be an influencing parameter for cell response. In this experience we attempted to compare the effect of roughness organization of Ti6A14V or pure titanium substrates on human osteoblast (hOB) response (proliferation, adhesion). Surface roughness was extensively analyzed at scales above the cell size (macro-roughness) or below the cell size (micro-roughness) by calculation of relevant classic amplitude parameters (Ra, Rt) and original frequency parameters (Order, Delta). We developed a new process to prepare isotropic surfaces (electro-erosion), which were compared to isotropic surfaces obtained by polishing and anisotropic surfaces obtained by machine-tooling. The hOB response on electro-eroded (EE) Ti6A14V surfaces or pure titanium (Ti) surfaces was largely increased when compared to polished or machine-tooled surfaces after 21 days of culture. Moreover, the polygonal morphology of hOB on these EE surfaces was very close to the aspects of hOB in vivo on human bone trabeculae. By a complete description of the surface topography of EE surfaces, we concluded that when the topography was considered below the cell scale, hOB appreciated their isotropic smooth aspect, although when the topography was considered above the cell scale they appreciated their rough isotropic 'landscape' formed by many 'bowl-like nests' favouring cell adhesion and growth. Electro-erosion is a promising method for preparation of bone implant surfaces, as it could easily be applied to preparation of most biomaterials with complex geometries.

Algorithms↗

PubMed2002

An unscaled parameter to measure the order of surfaces: a new surface elaboration to increase cells adhesion.

We present a new parameter to quantify the order of a surface. This parameter is scale-independent and can be used to compare the organization of a surface at different scales of range and amplitude. To test the accuracy of this roughness parameter versus a hundred existing ones, we created an original statistical bootstrap method. In order to assess the physical relevance of this new parameter, we elaborated a great number of surfaces with various roughness amplitudes on titanium and titanium-based alloys using different physical processes. Then we studied the influence of the roughness amplitude on in vitro adhesion and proliferation of human osteoblasts. It was then shown that our new parameter best discriminates among the cell adhesion phenomena than others' parameters (Average roughness (Ra em leader )): cells adhere better on isotropic surfaces with a low order, provided this order is quantified on a scale that is more important than that of the cells. Additionally, on these low ordered metallic surfaces, the shape of the cells presents the same morphological aspect as that we can see on the human bone trabeculae. The method used to prepare these isotropic surfaces (electroerosion) could be undoubtedly and easily applied to prepare most biomaterials with complex geometries and to improve bone implant integration. Moreover, the new order parameter we developed may be particularly useful for the fundamental understanding of the mechanism of bone cell installation on a relief and of the formation of bone cell-material interface.

Alloys↗

PubMed2000

Qualitative and quantitative study of human osteoblast adhesion on materials with various surface roughnesses.

We quantitatively evaluated the adhesion of human osteoblasts on orthopedic metallic substrates (Ti6Al4V alloy) with various surface roughnesses at several times after inoculation and studied its correlation with qualitative changes in the expression of adhesion proteins and with parameters extensively describing the surface topographies. Cells were orientated in a parallel order on polished surfaces. This orientation was not affected by residual grooves after polishing. On sandblasted surfaces the cells never attained confluence and had a stellate shape, and the cell layer had no particular organization. Extracellular matrix (fibronectin, type I collagen, osteopontin) and cytoskeletal protein (actin, vinculin) orientation reflected the cell layer organization. In our experiment human osteoblasts expressed alpha3beta1 integrin but not alpha2beta1 integrin. In addition to currently analyzed roughness magnitude parameters, we calculated roughness organization parameters (fractal dimension parameters) of the substrates. We observed lower adhesion and proliferation on less organized surfaces (i.e., sandblasted ones). The significant statistical correlation observed between fractal dimension parameters (describing surface roughness organization) and cell parameters adds a new concept to the studies of substratum roughness influence on cell behavior. An attempt at modelization of the cell-surface interaction was made that includes the influence of fractal dimensions parameters.

Alloys↗

PubMed2000

The relative influence of the topography and chemistry of TiAl6V4 surfaces on osteoblastic cell behaviour.

Proliferation and adhesion of mouse (MC3T3-E1) osteoblastic cells and primary human osteoblastic cells were carried out on Ti6Al4V titanium alloy samples with varied surface roughnesses. Mechanically or manually polished surfaces were prepared to produce respectively non-oriented or oriented residual polishing grooves. Sand-blasted surfaces were prepared using 500 microm or 3 mm alumina particles. Surface roughness parameters showed a negative correlation in comparison to proliferation and adhesion parameters. X-ray microprobe chemical surface microanalysis showed complete disturbance of the surface element composition of the Ti6Al4V alloy following sand-blasting treatment. An AlOx-enriched layer was observed on sample surfaces. This may lead to the suspicion that the concomittant effect of surface roughness amplitude and AlOx surface concentration has an effect on osteoblastic cell proliferation and adhesion. These findings show the significance of chemical surface analysis after any surface treatment of titanium-based implants before any biological use.

3T3 Cells↗

PubMed2000

Osteoblast adhesion on biomaterials.

The development of tissue engineering in the field of orthopaedic surgery is now booming. Two fields of research in particular are emerging: the association of osteo-inductive factors with implantable materials; and the association of osteogenic stem cells with these materials (hybrid materials). In both cases, an understanding of the phenomena of cell adhesion and, in particular, understanding of the proteins involved in osteoblast adhesion on contact with the materials is of crucial importance. The proteins involved in osteoblast adhesion are described in this review (extracellular matrix proteins, cytoskeletal proteins, integrins, cadherins, etc.). During osteoblast/material interactions, their expression is modified according to the surface characteristics of materials. Their involvement in osteoblastic response to mechanical stimulation highlights the significance of taking them into consideration during development of future biomaterials. Finally, an understanding of the proteins involved in osteoblast adhesion opens up new possibilities for the grafting of these proteins (or synthesized peptide) onto vector materials, to increase their in vivo bioactivity or to promote cell integration within the vector material during the development of hybrid materials.

Animals↗

PubMed2000

Comparative study of the in vitro characteristics of osteoblasts from paralytic and non-paralytic children.

STUDY DESIGN: Cerebral palsy is known to provoke a high loss of bone in children and adults. The potential interest of human osteoblastic cell culture for assessing the osteoblastic function in metabolic bone disorders has been demonstrated by many previous studies. Few studies have attempted to evaluate the capacities of osteoblasts isolated from immobilized or normal bones by in vitro culture methods. Moreover, a few teams did make the distinction between young spastic and flaccid patients. OBJECTIVES: We attempted to characterize mature osteoblasts (OB) and bone marrow-stromal cells (BM) originating from 56 immobile and normal children. Spastic and flaccid patients formed the paralytic group. SETTING: France. METHODS: Osteoblasts and bone marrow cells were isolated from iliac crests obtained during pelvic osteotomies of young control and paralytic patients. The in vitro viability, proliferation and differentiation parameters of the cells from paralytic patients were compared with those of cells coming from normal controls. RESULTS: No significant differences in the cell proliferation parameters were observed between the two groups. Only initial cell viability before inoculation was lower for the paralytic group, compared to the control group. On the other hand, contrary to expectations, we found that fresh and thawed OB cells from flaccid patients synthesized more osteocalcin and more collagen respectively than those of the spastic and control groups. Opposite results were obtained from BM cultures. CONCLUSION: A negative feedback mechanism by systemic or local factors, which is not conserved in vitro but controls the in vivo osteocalcin and collagen synthesis of flaccid paralytic OB cells, is hypothesized. Because these flaccid patients are known to have a high fat/lean mass, we suggest that leptin may be the potential regulating factor implicated in the hypothesized negative feedback mechanism.

Bone Marrow Cells↗

PubMed2000

Tissue engineering and skeletal diseases.

Tissue engineering, a cross between the science of the living organism and that of engineering, aims to replace, maintain or improve human tissue functions, by means of tissue substitutes containing living elements. Thus, it is about production of artificial tissue, using (alone or in combination) cells, matrix or bioactive factors. Their association gives rise to a hybrid biomaterial combining biological components (cells, growth factors or adhesion proteins) and materials (polymers, ceramics). The applications are wide-ranging, from the skin, to the liver, or to the cornea as well as to the locomotor system. Bone tissue engineering has advanced the most in this field, partly because of the progress made by research into bone substitutes, although cartilage and tendons are also concerned. This technology requires cell culture (committed cells or more often bone marrow stem cells), biomaterials (porous materials with controlled architecture and cements), growth factors (such as 'Bone Morphogenetic Proteins'), the proteins implicated in cell adhesion (such as fibronectin or the aminoacid sequences specifically recognised by integrin subunits) or gene therapy (notably using transfected stem cells). Tissue engineering and regenerative stimulation of tissue are now booming on experimental and industrial levels and clinical applications are increasingly numerous. Considering the potential of these technologies, they should continue to develop widely.

Biomedical Engineering↗

PubMed1999

Association of porous hydroxyapatite and bone marrow cells for bone regeneration.

The preparation of hybrid material with osteoinductive capacity may be achieved by association of cultured autologous bone cells with a porous ceramic vehicle. We optimized culture conditions for rabbit marrow stromal stem cells (MSCs), notably by selection from batches of fetal calf serum. Rabbit MSCs formed colony-forming unit-ribroblastic (CFU-Fs) in vitro. Their alkaline phosphatase (ALP) activity was doubled in the presence of dexamethasone. Autologous rabbit serum allowed the formation of ALP-positive CFU-Fs, but results were highly variable depending on the rabbit. We tested the osteogenic potential of autologous cultured (with or without dexamethasone addition in the culture medium) and noncultured rabbit MSCs associated with a porous hydroxyapatite ceramic after a dorsal intramuscular implantation. Nucleated cells (10(7) or 10(8)/mL) were used for the preparation of autologous hybrid material. A significantly higher number of implants containing bone was obtained with a suspension of 10(7) cells/mL cultured in the presence of 10(-8) M dexamethasone. Some positive implants were also obtained with a suspension of 10(8) noncultured cells/mL. We demonstrated the feasibility of preparing rabbit autologous hybrid materials following a process for controlling culture conditions, cell characterization and cell/material association.

Alkaline Phosphatase↗