BackgroundIt is known that physico/chemical alterations on biomaterial surfaces have the capability to modulate cellular behavior, affecting early tissue repair. Such surface modifications are aimed to improve early healing response and, clinically, offer the possibility to shorten the time from implant placement to functional loading. Since FAK and Src are intracellular proteins able to predict the quality of osteoblast adhesion, this study evaluated the osteoblast behavior in response to nanometer scale titanium surface texturing by monitoring FAK and Src phosphorylations.MethodologyFour engineered titanium surfaces were used for the study: machined (M), dual acid-etched (DAA), resorbable media microblasted and acid-etched (MBAA), and acid-etch microblasted (AAMB). Surfaces were characterized by scanning electron microscopy, interferometry, atomic force microscopy, x-ray photoelectron spectroscopy and energy dispersive X-ray spectroscopy. Thereafter, those 4 samples were used to evaluate their cytotoxicity and interference on FAK and Src phosphorylations. Both Src and FAK were investigated by using specific antibody against specific phosphorylation sites.Principal FindingsThe results showed that both FAK and Src activations were differently modulated as a function of titanium surfaces physico/chemical configuration and protein adsorption.ConclusionsIt can be suggested that signaling pathways involving both FAK and Src could provide biomarkers to predict osteoblast adhesion onto different surfaces.

In degenerative diseases or lesions, bone tissue replacement and regeneration are important clinical goals. The most used bone substitutes today are hydroxyapatite (HA) scaffolds. These scaffolds, developed over the last few decades, present high porosity and good osteointegration, but haven't completely solved issues related to bone defects. Moreover, the exact intracellular mechanisms involved in the response to HA have yet to be addressed. This prompted us to investigate the protein networks responsible for signal transduction during early osteoblast adhesion on synthetic HA scaffolds. By performing a global kinase activity assay, we showed that there is a specific molecular machinery responding to HA contact, immediately triggering pathways leading to cytoskeleton rearrangement due to activation of Adducin 1 (ADD1), protein kinase A (PKA), protein kinase C (PKC), and vascular endothelial growth factor (VEGF). Moreover, we found a significantly increased phosphorylation of the activating site Ser-421 in histone deacetylase 1 (HDAC1), a substrate of Cyclin-Dependent Kinase 5 (CDK5). These phosphorylation events are hallmarks of osteoblast differentiation, pointing to HA surfaces ability to promote differentiation. We also found that AKT was kept active, suggesting the maintenance of survival pathways. Interestingly, though, the substrate sequence of CDK5 also presented higher phosphorylation levels when compared to control conditions. To our knowledge, this kinase has never before been related to osteoblast biology, opening a new avenue of investigation for novel pathways involved in this matter. These results suggest that HA triggers a specific intracellular signal transduction cascade during early osteoblast adhesion, activating proteins involved with cytoskeleton rearrangement, and induction of osteoblast differentiation.

To investigate the modulatory effects of sigma ligands on the N-methyl- D-aspartate (NMDA) receptor-ion channel complex in vivo, we examined the intact cell binding of 3H-N-[1-(2-thienyl)cyclohexyl]piperidine (3H- TCP) to cultured neuronal cells prepared from fetal rat telencephalon. The 3H-TCP binding was saturable, reversible, and inhibited by a selective NMDA receptor antagonist, D-amino-5-phosphonovaleric acid. MII-limolar Mg2+ inhibited 3H-TCP binding both in the absence and presence of L-glutamate. 5-Methyl-10,11-dihydro-5H-dibenzo [a,d]cyclohepten-5,10-imine maleate (MK801) inhibited 3H-TCP intact cell binding in a competitive manner, while haloperidol inhibited it in a noncompetitive manner. The effect of the test drugs to inhibit 3H-TCP intact cell binding was in the order of dextromethorphan, haloperidol > (+/-)MK 801 > (+)pentazocine > (-)pentazocine > DTG > PCP > (+)-N- allylnormetazocine [(+)SKF 10047] > (+)3-(3-hydroxyphenyl)-N- (1- propyl)piperidine [(+)3-PPP] > (-)SKF 10047 > (-)3-PPP. The IC50 values of the six sigma ligands for 3H-TCP binding were closely correlated with the Ki values of the corresponding drugs for DTG site 1 in the guinea pig brain reported by Rothman et al. (1991). These findings suggest that the sigma ligand indirectly modulates the NMDA receptor ion channel complex, presumably through sigma 1 sites in vivo as well as in vitro.

No abstract

The aim of this study was to investigate the adhesion (4 and 24 h) and the morphology of fibroblast Balb/c 3T3 seeded onto polystyrene, partially stabilized (ZrO(2)Y(2)O(3)), stabilized zirconia ceramic (3YTZP), and pure titanium (Ti, grade 2). Initial cell adhesion (4 h) was greater (P < 0.05, analysis of variance and Tukey's Multiple Comparisons Test) onto ZrO(2)Y(2)O(3) and polystyrene than in Ti and 3YTZ. After 24 h, the number of adhered cells was similar between the biomaterials tested, but smaller than onto polystyrene (P < 0.05). Cells were more spread onto glass surface after 4 h, but after 24 h, the morphology and density of the cells were similar in all groups (SEM). Profilometry showed distinct Ra values for each material: glass coverslips and ZrO(2)Y(2)O(3) (0.09 microm), Ti (0.88 microm), and 3YTZP (30.93 microm). It was concluded that ZrO(2)Y(2)O(3) promoted the best initial adhesion, thus indicating that surfaces with Ra values smaller than 0.1 microm could be more favorable to initial adhesion.