No abstract

Blood vessels of the vertebrate circulatory system typically exhibit tissue-specific patterning. However, the cues that guide the development of these patterns remain unclear. We investigated the effect of cyclic uniaxial strain on vascular endothelial cell dynamics and sprout formation in vitro in two-dimensional (2D) and three-dimensional (3D) culture systems under the influence of growth factors. Cells preferentially aligned and moved in the direction perpendicular to the major strain axis in monolayer culture, and mechanical strain also regulated the spatial location of cell proliferation in 2D cell culture. Cells in 3D cell culture could be induced to form sprouts by exposure to appropriate growth factor combinations (vascular endothelial growth factor and hepatocyte growth factor), and the strain direction regulated the directionality of this process. Moreover, cyclic uniaxial strain inhibited branching of the structures formed by endothelial cells and increased their thickness. Taken together, these data support the importance of external mechanical stimulation in the regulation of endothelial cell migration, proliferation, and differentiation into primitive vessels.

To promote the osteogenic properties of osteoblasts, we synthesized a hydroxyapatite (HAp) with beta-tricalcium phosphate (beta-TCP) biphasic calcium phosphate containing Nb ions (NbTCP/HAp). NbTCP/HAp was prepared by annealing precipitates obtained by coprecipitation of an aqueous solution of Ca(NO(3))(2) and a mixture of (NH(4))(2)HPO(4) and aqueous Nb solution. The precipitates can be regarded as a calcium-deficient HAp, the PO(4) sites of which are partly occupied by Nb ions. NbTCP/HAp was successfully synthesized by thermal decomposition of the precipitates. NbTCP/HAp enhanced the calcification of normal human osteoblasts (NHOst), and the amount of calcified tissue increased in proportion to the Nb ion concentration in the NbTCP/HAp. The alkaline phosphatase (ALP) activity of NHOst was also enhanced by NbTCP/HAp. Because Nb ions significantly enhance the ALP activity of NHOst, calcification by NbTCP/HAp is considered to be due to enhancement of ALP activity induced by Nb ions dissolved from NbTCP/HAp. These results indicate that NbTCP/HAp can be an effective bone repair material.

A surface of biomaterials is known to affect the behavior of cells after their adhesion on the surface, indicating that surface characteristics of biomaterials play an important role in cell adhesion, proliferation, and differentiation. To assess the effects of functional groups on biomaterial surface, normal human osteoblasts (NHOsts) were cultured on surfaces coated with self-assembled monolayers (SAMs) containing various functional groups, and the adhesion, proliferation, differentiation, and gap junctional intercellular communication (GJIC) of the NHOsts were investigated. In the case of SAM with terminal methyl groups (hydrophobic surface), NHOst adhesion and proliferation was less prevalent. In contrast, NHOsts were adhered well on SAMs with hydroxyl, carboxyl, amino, phosphate, and sulfate group, which are relatively hydrophilic, their proliferation and differentiation level were dependent on the type of functional groups. Especially, when they were cultured on either SAMs with phosphate or sulfate group, both their alkaline phosphate activity and the calcium deposition by them were enhanced more than those cultured on a collagen-coated dish. More interestingly, GJIC of NHOsts, which has been reported to play a role in cell differentiation as well as homeostasis of cells, were not significantly different among the SAM surfaces tested. These suggest that a specific functional group on a material surface can regulate NHOst adhesion, proliferation, and differentiation via cell-functional group interaction without influencing their homeostasis.

Alginate is a polysaccharide that can be crosslinked by divalent cations, such as calcium ions, to form a gel. Chemical modification is typically used to improve its cell adhesive properties for tissue engineering applications. In this study, alginates were modified with peptides containing RGD (arginine-glycine-aspartic acid) or PHSRN (proline-histidine-serine-arginine-asparagine) sequences from fibronectin to study possible additive and synergistic effects on adherent cells. Alginates modified with each peptide were mixed at different ratios to form gels containing various concentrations and spacing between the RGD and PHSRN sequences. When normal human osteoblasts (NHOsts) were cultured on or in the gels, the ratio of RGD to PHSRN was found to influence cell behaviors, especially differentiation. NHOsts cultured on gels composed of RGD- and PHSRN-modified alginates showed enhanced differentiation when the gels contained >33 % RGD-alginate, suggesting the relative distribution of the peptides and the presentation to cells are important parameters in this regulation. NHOsts cultured in gels containing both RGD- and PHSRN-alginates also demonstrated a similar enhancement tendency of calcium deposition that was dependent on the peptide ratio in the gel. However, calcium deposition was greater when cells were cultured in the gels, as compared to on the gels. These results suggest that modifying this biomaterial to more closely mimic the chemistry of natural cell adhesive proteins, (e.g., fibronectin) may be useful in developing scaffolds for bone tissue engineering and provide three-dimensional cell culture systems which more closely mimic the environment of the human body.