OBJECTIVETo investigate the effects of 20% fluor-hydroxyapatite (FHA) on proliferation and osteogenic differentiation of human MG63 osteosarcoma cells.

METHODSFHA was prepared by chemical precipitation method, and its structure and surface features were tested by scanning microscope, X-ray diffraction (XRD) and Fourier transform infrared spectroscopy. MG63 cells were cultured and divided into FHA, hydroxyapatite (HA) and control groups (n = 3). The proliferation of the cells was evaluated using methylthiazol tetrazolium (MTT) assay. ALP activity of the cells was assessed. Osteogenic differentiation was evaluated based on the reverse transcription PCR (RT-PCR) of differentiation-related genes, namely, collagen type I (Col I), alkaline phosphatase (ALP), osteocalcin (OCN) and core binding factor α1 (Cbfα1). The data were analyzed statistically by one-way analysis of variance using SPSS 13.0 software.

RESULTSXRD test showed that the main crystalline phase of FHA was similar to that of HA. Absorptance value of cells exposed to FHA(1.87±0.06) measured by MTT was higher than that of the control(1.25±0.02) on the third day(P < 0.05), and there was no statistically significant difference between the cells exposed to FHA and HA(1.84±0.03) (P > 0.05). ALP activity of the cells exposed to FHA(4.62±0.09)was higher than that of the control (1.92 ± 0.05) (P < 0.05). RT-PCR tests showed that compared with the control, FHA up-regulated the expression of Col I, ALP and OCN mRNA, down-regulated the expression of Cbfα1 mRNA.

CONCLUSIONSFHA enhances the proliferation and osteogenic differentiation-related gene expression, and has good biocompatibility.

Alkaline Phosphatase ; genetics ; metabolism ; Analysis of Variance ; Biocompatible Materials ; Bone Neoplasms ; metabolism ; pathology ; Cell Differentiation ; drug effects ; genetics ; Cell Proliferation ; drug effects ; physiology ; Collagen Type I ; genetics ; metabolism ; Core Binding Factor Alpha 1 Subunit ; genetics ; Durapatite ; chemistry ; pharmacology ; Humans ; Hydroxyapatites ; Osteocalcin ; genetics ; metabolism ; Osteogenesis ; Osteosarcoma ; metabolism ; pathology ; Spectroscopy, Fourier Transform Infrared ; X-Ray Diffraction

Expressions of p53 protein, a product of the tumor suppressor gene were studied in osteosarcomas relating to various prognostic factors. Thirty-four osteosarcomas were investigated immunohistochemically with a monoclonal antibody clone PAb240, which recognizes a common conformational epitope of mutant p53 proteins and another clone PAb1801, which reacts with both wild- and mutant-type p53 proteins. The results were compared with expressions of proliferating cell nuclear antigen (PCNA) and Ki-67 providing a simple method for the assessment of growth fractions of tumors. PAb240 stained nuclei and cytoplasm of tumor cells in 8 of 34 osteosarcomas (23.5%), whereas PAb1801 reacted in all 34 osteosarcomas (100%). Fifteen tumors (44.1%) showed positivity for PAb1801 in more than half of the tumor cells. Twelve patients were alive and thirteen were dead. Tumors from 9 patients (75%) who survived revealed only focal positive immunoreactions with PAb1801 and tumors from 6 patients (46.1%) who died revealed diffuse reactions. Twelve cases (35.3%) showed a high PCNA index (> 40%) and fibroblastic osteosarcomas revealed the highest PCNA positivity. Twenty-two cases (64.7%) revealed a very low Ki-67 index (less than 10%) and Ki-67 index showed a good correlation with PCNA positivity (r = 0.6247). Expressions of both wild-and mutant-type p53 protein, PCNA, and Ki-67 were not correlated with other clinical or pathological parameters.
Adolescent ; Adult ; Aged ; Antibodies, Monoclonal ; Bone Neoplasms/*chemistry/genetics/pathology ; Cell Cycle/physiology ; Child ; Child, Preschool ; Genes, p53 ; Human ; Immunohistochemistry ; Ki-67 Antigen ; Male ; Middle Age ; Mutation ; Neoplasm Proteins/*analysis ; Nuclear Proteins/*analysis ; Osteosarcoma/*chemistry/genetics/pathology ; Proliferating Cell Nuclear Antigen/*analysis ; Protein p53/*analysis ; Support, Non-U.S. Gov't