Honeycomb form ß-tricalcium phosphate induces osteogenesis by geometrical property with BMSC.

To establish an effective method for bone augmentation, we introduced a new honeycomb-like ß-tricalcium phosphate (H-ß-TCP) with BMP-2 as a scaffold, whose unique geometrical properties induce osteoblastic differentiation of autologous bone marrow mesenchymal stem cells (BMSCs). A total of six beagle dogs from 6 to 7 years old were used for this study. BMSCs were cultured with autologous serum and BMP-2 on H-ß-TCP. Differentiation to osteoblasts was demonstrated in vitro and exo vivo. Scanning electron microscopy revealed formation and calcification of a matrix-like structure within the H-ß-TCP tunnels in BMSC culture. Moreover, treatment of BMP-2 promoted osteoblastic differentiation of BMSCs in H-ß-TCP in a diffusion chamber. These results indicated that H-ß-TCP may be a useful tool for construction of functional artificial bone.

Transcriptional regulation of the salivary histatin gene: finding of a strong positive regulatory element and its binding protein.

Histatins are salivary proteins found and expressed in human salivary glands. They play a role in the non-immune system of antimicrobial defense, for instance, against Candida albicans. The transcriptional regulatory sequences of the histatin gene, HIS1, have remained obscure for a long time. Here, we cloned the putative promoter from human genomic DNA and tested it in a luciferase reporter system. This promoter is much more active in salivary gland cells than in other cell types. Analysis of deletion mutants revealed that the region encompassing -2254 to -1748 is a strong positive transcriptional element, and its functional core sequence (termed HTN27 box) works in correct and reverse orientations in synergy with downstream sequences, the region spanning -680 to +28 and a proximal promoter. The plus single-stranded HTN27 box is specifically bound by a 100 kDa protein that is present in HSG cells, but not in HeLa cells. These findings indicate that the regulation of the histatin gene expression may be intricate, and it seems to have a cell-type preference in the salivary gland cells.

Regulation of osteoclast polarization.

Osteoclast function consists of several processes: recognition of mineralized tissues, development of ruffled borders and sealing zones, secretion of acids and proteolytic enzymes into the space beneath the ruffled border, and incorporation and secretion of bone degradation products using the transcytosis system. One of the most important questions concerning osteoclast function is how osteoclasts recognize bone and polarize. During the past decade, new approaches have been taken to investigate the regulation of osteoclast polarization. Attachment of osteoclasts to some proteins containing the Arg-Gly-Asp sequence motif through vitronectin receptors is the first step in inducing the polarization of osteoclasts. Physical properties of bone such as hardness or roughness are also required to induce osteoclast polarity. Osteoclasts cultured even on plastic dishes secrete protons toward the dish surface, suggesting that osteoclasts recognize plastic as a mineralized matrix and secrete protons. This notion was supported by the recent findings that bisphosphonates and reveromycin A were specifically incorporated into polarized osteoclasts cultured even on plastic dishes. On the other hand, a sealing zone, defined as a thick band of actin, is induced in osteoclasts adherent only on an apatite-containing mineralized matrix. These results suggest that osteoclasts recognize physical properties of the mineralized tissue to secrete protons, and also sense apatite itself or components of apatite to form the sealing zone. Here, we review recent findings on the regulation of osteoclast polarization. We also discuss how osteoclasts recognize mineralized tissues to form the sealing zone.