Biomimetic layer-by-layer templates for calcium phosphate biomineralization.

Carboxylated, sulfated and/or phosphorylated surfaces are admitted as potential optimal templates for biomimetic deposition of calcium phosphate (CaP) coatings in view of improving implants' osseointegration. Layer-by-layer films were built up consisting of anionic chondroitin sulfate (ChS), a biological carboxylated and sulfated polysaccharide and cationic poly(l-lysine) (PLL). The films were used as soft matrices to immobilize a model phosphoprotein, phosvitin (PhV). The respective roles of ChS, PLL and PhV terminal layers on the heterogeneous nucleation kinetics and the structure of CaP deposits obtained from supersaturated solutions were inspected. Critical supersaturation ratios and induction times preceding heterogeneous nucleation were precisely determined and interpreted within the framework of classical nucleation theory in order to derive the effective interfacial energies of CaP crystals. It was found that the potency of terminal layers toward CaP nucleation increased in the order: PLL

Practice of a testing bench to study the effects of cyclic stretching on osteoblast-orthopaedic ceramic interactions.

A new experimental method has been used to study the behaviour of human osteoblasts cultured on bioceramics subjected to mechanical strains. The ceramics were alumina, hydroxyapatite (HA) and a duplex system composed of hydroxyapatite-covered alumina. The system applied 400 microdeformations for a 6-h period with a cycle frequency of 0.5 Hz to osteoblasts growing on ceramic-covered disks. The effects of strains on short-term cell viability, cell growth, alkaline phosphatase (ALP) activity, and collagen biosynthesis were assessed. When possible, the parameters (lactate dehydrogenase) were studied along the experiment in samples of the culture medium, in the other cases by comparison of stretched and unstretched cultures on the same ceramics with the same cell line. In relationship with the coating, mechanical strains resulted in a decrease in DNA corresponding to cell number, an LDH release during straining, an unchanged (alumina) or decreased (HA and duplex) ALP activity, a decrease (HA and duplex) of collagen and total protein synthesis or an increase of it (alumina). The stress-producing device and its associated protocol are shown to be suitable for investigating the behaviour of cells, cultured on biomaterials subjected to mechanical strain.

Interaction of a plasma-sprayed hydroxyapatite coating in contact with human osteoblasts and culture medium.

The loss of calcium from plasma-sprayed calcium phosphate ceramics (CPCs) on bioinert metal substrate (Ti-6Al-4V) immersed in cell culture medium with or without human osteoblast culture was measured. The ceramics were a CPC and a duplex system composed of a CPC layer on an alumina coating. The dissolution of calcium compounds was monitored by measuring the calcium leaked from the coatings into the culture medium in 15 days. Calcium was measured by flame photometry. The surfaces of the ceramics exposed to the culture medium and in contact with osteoblasts were analysed by X-ray diffraction (XRD). The dissolution process occurred in the first 6 days of contact, but the calcium released into the culture medium was only a small fraction of the calcium content of the coatings. The presence or absence of osteoblasts on the surface of the ceramics did not make significant difference for the calcium release. The XRD spectra of the ceramics before and after immersion and in contact with cells did not show a significant change in the compounds of the coatings.

A standardized parameter for the distribution of individual red blood cell deformability with the cell transit analyzer.

The purpose of this study was to identify a parameter allowing the standardization of the Cell Transit Analyzer (CTA) in order to study the individual deformability of each explored Red Blood Cell (RBC). Using theoretical arguments based on the principle of the CTA, we calculated the thickness "x" of the crown of fluid surrounding each RBC during its entry phase into the micropore. A mathematical equation (x = 62.5/magnitude of dU) was established between the difference of potential (dU, mV) that occurs during this phase and the corresponding thickness ("x", micron) of the crown. By exploring fresh control RBCs of healthy subjects and assuming that the rigid RBCs proportion in a fresh blood sample of healthy subject is less than 3.5%, we performed a thresholding of "x" to distinguish rigid RBC from deformable ones. That thresholding was necessary to stamp the variability of counts linked to polycarbonate filters (PF) used to carry out measures. According to the PF, the value of the threshold "Tx" provided by the thresholding ranged between 0.222 and 0.246 micron. Using the values of "Tx", we counted approximately 10-25% rigid RBCs in the pathological samples of four patients with sickle cells SS disease and of one diabetic patient with splenomegaly. We observed in addition that the percentages of rigid RBCs counted after thresholding are identical from a filter to another one with an absolute error less than 2% in fresh RBCs (normal or pathological) samples. We concluded that the method of standardization by thresholding presented here could be used in clinic routine to count the rigid RBCs percentage contained in a given sample.

Effects of gamma-alumina and hydroxyapatite coatings on the growth and metabolism of human osteoblasts.

The behavior of human osteoblasts cultivated on hydroxyapatite or alumina-coated disks of Ti6AL4V was studied in vitro. Cell anchorage and spreading were observed by scanning electron microscopy. Cell growth was monitored by counting cells and measuring DNA at 5 h and 2,5, and 10 days after cell seeding. Cells grown for 10 days were labeled with 14C-proline and total protein and collagen synthesis were measured; the type of collagen was also determined. Both ceramics showed excellent biocompatibility. At 10 days of culture the cells showed a higher rate of proliferation on alumina than on hydroxyapatite. Neither ceramic altered the collagen biosynthesis or the osteoblast-like properties of the cells, as indicated by the high percentage of type I collagen.

Association of polyacrylamide beads to polyethylene terephthalate prostheses.

A method for the coupling of polyacrylamide beads to polyethylene terephthalate (PET) vascular prostheses is described. The reactional procedure used is performed along several steps; acrylic acid grafting on PET textile fibres, in order to introduce reactive carboxylic groups, introduction of terminal primary amine groups onto the beads, and then, attachment method which consists in coupling carboxylic groups of prostheses with amine groups of modified beads. The relative weight increase of the samples before and after the coupling reaction and, microscopic observations of beads distribution onto the prostheses surface demonstrate the binding feasibility of polyacrylamide matrices to PET prostheses. In the near future, authors expect to replace these beads by microcapsules with polyacrylamide wall and containing active compounds to improve the vascular prostheses biocompatibility.