Characterization of recombinant human cementum protein 1 (hrCEMP1): primary role in biomineralization.

Cementum protein 1 (CEMP1) has been recently cloned, and in vitro experiments have shown functions as regulator of cementoblast behavior and inducer of differentiation of non-osteogenic cells toward a cementoblastic/osteoblastic phenotype. In this study, we have produced a full-length human recombinant CEMP1 protein in a human gingival fibroblast cell line. The purified protein (hrCEMP1) has a M(r) 50,000. Characterization of hrCEMP1 indicates that its secondary structure is mainly composed of beta-sheet (55%), where random coil and alpha helix conformations correspond to 35% and 10%, respectively. It was found that hrCEMP1 is N-glycosylated, phosphorylated and possesses strong affinity for hydroxyapatite. Even more important, our results show that hrCEMP1 plays a role during the biomineralization process by promoting octacalcium phosphate (OCP) crystal nucleation. These features make CEMP1 a very good candidate for biotechnological applications in order to achieve cementum and/or bone regeneration.

Osteopontin upregulation in atherogenesis is associated with cellular oxidative stress triggered by the activation of scavenger receptors.

BACKGROUND: Osteopontin (OPN) is a highly phosphorylated sialoprotein and a prominent component of mineralized extracellular matrices of bones and teeth. Although the structure of OPN has begun to be elucidated, the role of OPN overexpression in tissues distant from the bones and teeth remains poorly understood. In the present study, a rabbit model of hypercholesterolemia was employed to analyze the relationship between the vascular calcification process and OPN overexpression in the neointima of atherosclerotic plaques. METHODS: OPN identification in the aorta of experimental animals fed with a high cholesterol diet was carried out by immunohistochemical procedures and Western blot analysis of tissue homogenates. Transmission electron microscopy was employed to localize target-like extracellular structures of atherosclerotic aortas. The human cell line T/G HA-VSMC was employed in the establishment of a ROS generation model employing the internalization of OxLDL particles. RESULTS: Using immunohistochemical and Western blot analysis, OPN overexpression was detected in the aortas of rabbits fed a high-cholesterol diet. Results from the ultrastructural analysis of the rabbit neointima through transmission electron microscopy and from the detection of calcium phosphate precipitates by specific histochemical techniques, suggested that OPN may be functionally important as a regulator of vascular calcification. OPN was dramatically overexpressed by vascular smooth muscle cells in the presence of oxidized and acetylated LDL particles bound to scavenger receptors, thereby promoting cytosolic oxidative stress. CONCLUSIONS: This study establishes the in vivo role of OPN in the intima of the aorta regulating calcium phosphate precipitate deposition in response to oxidative stress.