Natural hydroxyapatite (HA) was derived from pig bones (PBs) for tissue engineering applications through heat treatment. Fourier transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD), X-ray fluorescence (XRF), energy-dispersive Xray spectroscopy (EDX), and field emission scanning electron microscopy (FE-SEM) were employed for the analysis of heat-treated PB powder. In addition, inductively coupled plasma (ICP) mass spectroscopy was applied to examine the phase organizations and chemical composition of PB powder. The Ca/P ratio in bone powder was between 1.5 and 1.6. Moreover, in vitro and in vivo studies were performed to assess the biocompatibility of long-term soaked bone powder. These results can be used to (i) establish chemical characterization by analyzing SEM microstructures and undertaking XRD, XRF, EDX, FTIR, and ICP atomic emission spectrometry; (ii) perform cytotoxicity evaluations using human mesenchymal stem cells; and (iii) conduct assessments using an in vivo animal model and histological observations. No significant changes in chemical composition were observed in long-term soaked samples with respect to the original HA. Cells properly adhered to the surface of soaked samples, and rapid hilling of the defected skull was observed in the presence of the soaked sample, indicating its potential for use as a tissue engineering biomaterial. These results suggested that heat treatment at 1200 °C could produce natural bioceramics based on calcium phosphate, which retained their osteogenic potential after 36 months of soaking.

Odontogenic ameloblast-associated protein (ODAM) contributes to cell adhesion. In human cancer, ODAM is down-regulated, and the overexpression of ODAM results in a favourable prognosis; however, the molecular mechanisms underlying ODAM-mediated inhibition of cancer invasion and metastasis remain unclear. Here, we identify a critical role for ODAM in inducing cancer cell adhesion. ODAM induced RhoA activity and the expression of downstream factors, including Rho-associated kinase (ROCK). ODAM-mediated RhoA signalling resulted in actin filament rearrangement by activating PTEN and inhibiting the phosphorylation of AKT. When ODAM is overexpressed in MCF7 breast cancer cells and AGS gastric cancer cells that activate RhoA at high levels, it decreases motility, increases adhesion and inhibits the metastasis of MCF7 cells. Conversely, depletion of ODAM in cancer cells inhibits Rho GTPase activation, resulting in increased cancer migration and invasion. These results suggest that ODAM expression in cells maintains their adhesion, resulting in the prevention of their metastasis via the regulation of RhoA signalling in breast cancer cells. SIGNIFICANCE Breast cancer represents the first most frequent cancer, and the ratio of mortality is high in women. Of utmost importance for reducing risk by breast cancer are their anti-invasion mechanisms, particularly in the non-invasive cancer cells because metastasis is the principal cause of death among cancer patients. ODAM induced RhoA activity. ODAM-mediated RhoA signalling resulted in actin filament rearrangement, increased cell adhesion and inhibited the migration/invasion of MCF7 cells. These results suggest that ODAM expression maintains their adhesion, resulting in the prevention of their metastasis via the regulation of RhoA signalling in breast cancer cells.