Abstract: We present a new bio-photonic method based on Raman spectroscopy able to
characterize living cells in in-vitro cultures. The main advantages of this technology are: no labels or other contrast enhancers are required; provides real-time analysis; cells can be maintained in physiological conditions during the measurements; no cell-damage is induced during the measurements; it is rich in information about the biochemical composition of the cell. The results show that this spectroscopic method can be used to study the most important cellular functions involved in cell-biomaterial interactions, such as cell death, differentiation, de-differentiation and
mineralization. The method offers the potential for studying cell-bioceramics interaction and reduce the need of animal testing until the final steps of proving efficacy prior to clinical trials.
Abstract: Osteoblasts constitutively release glutamate and this release appears to be regulated by calcium entry. In this work we investigated if the bioactive glass with 60% of silicon (BG60S) could alter glutamate release by osteoblasts. We demonstrated that osteoblasts incubated with medium containing ionic products from the dissolution of BG60S showed lower release of glutamate when compared to control. Since intracellular calcium (Cai 2+) increase is required for glutamate release we investigated the subcellular distribution of the calcium channel inositol
triphosphate receptors (InsP3Rs) in the presence of BG60S compared to control. We found that the type-III InsP3R was not expressed in osteoblast, while the type-II InsP3R was expressed mainly in the cytosol. We also found that the expression of type-II InsP3R decreased in BG60S treated osteoblasts compared to control. On the other hand, we found that the type-I InsP3R was expressed
mainly in the nucleus and its expression increased in the presence of the biomaterial.
Abstract: Poor cell adhesion to orthopaedic and dental implants results in implant failure.
Establishing and maintaining mature bone at the bone/device interface is critical to the long-term success of the prostheses. Considerable effort has been devoted to alter the surface characteristics of these biomaterials in order to improve the initial interlocking of device and skeleton in the noncemented joint prosthesis. Previously we demonstrated that surface chemistry modification of bioceramics induced osteogenesis. In the present work, we investigate the effect of surface chemistry modification of titanium alloy (Ti-6Al-4V) with alkoxide-derived carbonate
hydroxyapatite (CHAp) using sol-gel coating methods on human bone derived cell (HBDC)behaviour. Western blotting demonstrated that sol gel coating of Ti-6Al-4V with CHAp upregulated the expression of key signalling protein Shc isoforms (p46, p52, p66) and phosphorylated Erk1/2. CHAp-modification of Ti-6Al-4V is associated with signal transduction pathways involving the key signalling protein Shc and ERK1/2 which may lead to enhanced gene expression of extracellular
matrix proteins at the skeletal tissue/device interface.
Abstract: The Classical Least Square (CLS) fitting method was used to analyze the Raman spectra of living cells with the aim of identification of new phenotype-specific spectral markers for osteoblasts. The following chemicals were used for the CLS model: DNA, RNA, serum albumin, chymotrypsin and phosphatidyl choline. In this study we analyzed primary mature osteoblasts as well as two other cell types used as potential sources of osteoblasts: embryonic stem cells and fetal bone cells. The results obtained suggest that the Raman spectra of the cell types can be well
approximated with a linear combination of the Raman spectra of the biopolymers used in the CLS model. The relative concentrations of the CLS components varied significantly between cell types, indicating that this analytical method could be used for phenotypic identification of osteoblasts.
Abstract: Different silica and carbonate containing calcium phosphate (CaP) layers were prepared on bioactive glass S53P4 in conventional C-SBF and revised R-SBF. In R-SBF the CaP layer formed faster compared to C-SBF, and the CaP layer formed in R-SBF was amorphous compared to the poorly crystalline bonelike HCA formed in C-SBF. In addition, the influence of chemical composition, dissolution and structure of biomimetically processed CaP layers on osteoclast and osteoblast activity was studied. In general, biomimetic CaP layers on bioactive glass S53P4 did not
affect so much on bone cell activity as it was expected compared to the untreated glass. Additionally, it was observed that the mechanism for good osteoclast activity is multifactorial. The optimal surface for osteoclast adhesion and growth was an amorphous CaP having mesoporous nanotopography and proper dissolution rate of calcium and silica. Also osteoblasts grew well on such surface.
Abstract: The biocompatibility of Zeolite was evaluated, in vitro, compared to a control
and to three different biomaterials: hydroxyapatite from bovine bone, calcium
phosphate and a commercial eugenol paste. The Zeolite did not affect cellular
proliferation neither the alkaline phosphatase and collagen production. The apoptosis
index of the zeolite groups were similar to control and optical microscopy observations did not show any morphological cell change, except the some cytoplasmatic vacuole formation.
Abstract: Based upon the CaO-P2O5 glass system, two glass ceramics were prepared in the meta-, pyro- and orthophosphate regions. The present work describes results concerning the biological activity of MT13B (45CaO-37P2O5-5MgO-13TiO2, in mol %) and MK5B (45CaO-45P2O5-5MgO-5K2O, in mol %) using MG63 osteoblast-like cells cultured for 6 days. The influence of a 24h pretreatment
with culture medium before cell culture was also evaluated (MT13Bi; MK5Bi). Both “as received” and pre-treated material samples supported cell growth, which increased with the culture time. Pre-treatment resulted in an increase of adhered cells on both materials (day 1). Throughout the culture time (days 3 and 6), MT13Bi presented higher cell numbers than that of MT13B, but no differences were found between MK5Bi and MK5B. In addition, cell growth was significantly lower in the K-containing glass ceramic in both experimental situations comparing with Ticontaining glass ceramic. SEM and confocal microscopy observation showed that cell adhesion and spreading were hampered on MK5B samples. Evident macroscopic and microscopic surface instability was observed on this material with simultaneous dissolution/precipitation processes. By contrast, MT13B presented a relatively stable surface throughout the culture time. Results suggest that the pre-incubation with culture medium improves the adhesion of the osteoblast cells to both
glass ceramics and that the high degradation rate of MK5B hinders the in vitro biological performance of this material.
Abstract: Good quality crystalline sol-derived Hydroxyapatite (HA) thin films of thickness ~350 nm on titanium substrates with underlying micro-channels were prepared by microwave annealing at a temperature of 400 °C for 30 min. A comprehensive in-vitro study was conducted to assess the osteoblast adhesive response to the well-defined micro-scale topographical features on HA coated and uncoated surfaces. The study confirmed the osteoblast’s response to the HA coated micro-channels with an elongated morphology. The effects to morphology suggest that osteoblast adhesion can be improved through optimal micro-channel design and may provide baseline design cues for improving the longterm performance of prosthetic orthopedic and dental implants.