Abstract:Objective To investigate the effect of plasma membrane-associated sialidase 3 (NEU3) activity on the proliferation and apoptosis of osteosarcoma MG-63 cells in vitro. Methods MG-63 cells were cultured in vitro. Anti-NEU3 antibody (Ab) immunofluorescent staining was used to indicate the cellular localization of NEU3 in MG-63 cells. The cells treated with 0 nmol/L 2-deoxy-2,3didehydro-N-acetyl neuraminic acid (DANA) or 0 μg/ml anti-NEU3 Ab were used as blank control groups. The cells were treated with 10, 20, 50 nmol/L DANA, or 0.5, 1.0, 2.0 μg/ml antiNEU3 Ab for 24 h or 48 h, respectively. The inhibition rates of the cell proliferation and cell apoptosis rates were measured with CCK-8 and flow cytometry. The expression levels of oncogene-related proteins, Ras protein and Bcl-2 protein, were detected by Western blotting. Results The immunofluorescence result showed that NEU3 was located in the cytoplasm of MG-63 cell. After treating with 0, 10, 20, 50 nmol/L DANA for 48 h, the inhibition rates of cell proliferation were 0, 15.10%±3.23%, 41.46%±2.31%, 64.68%±4.12%, with significant statistical difference (F=99.90, P<0.001), and the following contrast between each two groups met the statistical significance (all P<0.05). After treating with 0, 0.5, 1.0, 2.0 μg/ml anti-NEU3 Ab for 48 h, the inhibition rates of cell proliferation were 0, 9.34%±1.53%, 19.66%±4.18%, 42.50%±5.68%, and the difference was statistically significant (F=25.67, P<0.001), and the following contrast between each two groups met the statistical significance (P<0.05), except the difference between 0.5 and 1.0 μg/ml groups (P>0.05). When the MG-63 cells were treated with 0, 10, 20, 50 nmol/L DANA for 24 h, the cell apoptosis rates were 4.05%±0.07%, 4.15%±0.23%, 12.85%±1.48%, 8.29%±0.86%, respectively, and the difference was statistically significant (F=23.21, P<0.001). And the following contrast between each two groups met the statistical significance (P<0.05), except the differences between 0 nmol/L and 10 nmol/L, 20 nmol/L and 50 nmol/L groups (P>0.05). When the MG-63 cells were treated with 0, 0.5, 1.0, 2.0 μg/ml anti-NEU3 Ab for 24 h, the cell apoptosis rates were 4.05%±0.07%, 20.13%±2.97%, 20.29%±2.82%, 20.58%±0.70%, with statistical significant difference (F=15.36, P=0.001). And the following contrast between each two groups showed that the differences between 0 μg/ml and each treated group were statistically significant (P<0.05), while the differences between two treated groups were not statistically significant (P>0.05). Western blotting results showed that the expression levels of Ras and Bcl-2 decreased with the increasing concentrations of DANA and anti-NEU3. Conclusion Inhibition of NEU3 enzyme activity can suppress the survival rate of MG63 cells and increase the cell apoptosis. The possible mechanism may be related to the declined expression of oncogene-related proteins Ras and Bcl-2, which suggests that NEU3 may be a possible target for treating osteosarcoma.

Key words:Neuraminidase,Osteosarcoma,Cell proliferation,Apoptosis,MG-63 cells