We developed a novel method for titanium coating material using modified thermal decomposition technique, specifically focusing on presence of CaTiO3 and carbon C. Two layers of thin film coating composed of CaTiO3-amorphous carbon compound (CaTiO3-aC) and hydroxyapatite (HA) were generated on titanium surface. In this method, ratios of Ca/P as well as Ca/Ti, sintering temperature and sintering velocity were carefully planned. Within crystal structure of CaTiO3 granules, 4 atomic % of carbon in amorphous state was included. By our method, inclusion of carbon in HA suggested formation of carbonate apatite. Regarding with attachment-detachment experiment at titanium surface, adhesion strength was 2.5 times stronger in CaTiO3-aC/HA coating substrate as compared to HA only coating material. Results of the novel developed modified thermal decomposition method suggested that the 2 layers biomaterial composed of CaTiO3-aC (0.6 µm) and carbonate apatite-included HA (2.4 µm) can be used as a coating material on titanium surface.

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Bone and dentin resemble each other in composition and mechanism of formation and mineralization. Preliminary analysis of osteoblastic and odontoblastic cells before using them in tissue engineering is mandatory. Thus, in order to evaluate the process of calcification, we evaluated osteoblastic-like (KUSA/A1cells) and odontoblastic-like (MDPC-23 cells) cells seeded in cell culture and in intraperitoneal diffusion chamber. Our results indicated that KUSA/ A1 cells differentiated into osteoblasts-like cells and induced bone tissue inside the chamber. Whereas, MDPC-23 cells showed odontoblastic phenotype without ability to induce dentin formation, suggesting that MDPC-23 cells are special cells, which lost the capacity to induce mineralized dentin matrix after long period of time.This study showed the significance of basic information of calcification process by osteoblastic-like and odontoblastic-like cells before using them in tissue engineering.

It has not been determined the best scaffold-structure for bone engineering. Because of this, we compared Honeycomb porous structure (HPS) and Interconnected porous structure (IPS) with or without KUSA/ A1 cells implanted in mice. The transplants were subjected to radiological and histological examinations after 1,2,4 and 8 weeks of implantation. KUSA/A1 cells alone showed small islands of new bone. Both scaffolds alone did not reveal any bone induction. KUSA/A1-HPS presented the scaffold partially filled with new bone. In contrast, KUSA/A1-IPS showed the whole scaffold filled with new bone. Our results indicated that cotton structure plays an important role in carrying the cells giving the precise size, shape and comfortable environment.

The basic principle of bone tissue engineering is to use seeded stem cells in porous scaffold. Stem cells can proliferate and differentiate into various types of mature cells. A kind of stem cell called KUSA/A1 is a marrow stromal cell, capable of differentiating into three mesenchymal phenotypes: osteocyte, adipocyte, and myocyte by treating with 5-azacytidine in cell culture. Moreover, it has been reported that the mechanism of bone induction by KUSA/A1 cells is similar to intramembranous ossification.In order to clarify the origin of osteoblasts implicated in new bone formation, KUSA/A1 cells alone and combined with Honeycomb carrier were implanted in Transgenic Green Fluorescent Protein mice (GFP) mice. The presence of GFP positive host cells with osteoblastic morphology as well as GFP negative cells, clearly of KUSA/A1 cells in origin were observed around the bony trabeculae. These results indicated that the new bone was not only produced by KUSA/A1 cells but also by host cells from the surrounding connective tissues.To our knowledge, this is the first study to describe that host cells play an important role in ectopic bone induced by implanted marrow stromal cells, which would need special attention in bone tissue engineering.