RESUMO
AIM: The aim of this study was to test the hypothesis that the proliferation of hydroxyapatite (HA)-induced human osteoblast cell line (HOS cells) may be up-regulated by exogenous nitric oxide (NO). METHODS: HOS cells were cultured on the surface of HA with or without the presence of a NO donor, S-nitroso acetyl penicillamine (SNAP) or nitroso acetyl penicillamine (NAP). 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide) known as carboxy PTIO, a NO scavenger, was added in the cell cultures with or without the presence of SNAP. The cells were also pre-treated with L-N5-(1-iminoethyl)orthinine hydrochloride (L-NIO), an endothelial nitric oxide synthase (eNOS) inhibitor, or anti-integrin alphaV antibody before culturing on HA surfaces with or without the presence of SNAP. Medium, cells alone or cells pretreated with these inhibitors or antibodies was used as the controls. Cell proliferation was assessed by colorimetric assay. RESULTS: The results showed that SNAP, but not NAP, augmented HA-induced HOS cell proliferation. This modulatory effect of SNAP on HA-induced HOS cell proliferation was abolished by carboxy PTIO or anti-integrin alphaV antibody, but only partially reduced by L-NIO. CONCLUSION: Therefore, the results of this study suggest that exogenous NO alone may up-regulate the proliferation of HOS cells attached on HA surfaces via integrin alphaV molecules.
RESUMO
The aim of the present study was to determine the effect of nitric oxide (NO) on the production of cyclic AMP (cAMP) by a human osteoblast cell line (HOS cells) stimulated with hydroxyapatite. Cells were cultured on the HA surfaces with or without the presence of NO donors (SNAP and NAP) for 3 days. The effect of adenylyl cyclase inhibitor (SQ22536), NO scavenger (carboxy PTIO) or endothelial nitric oxide synthase (eNOS) inhibitor (L-NIO), was assessed by adding these to the cultures of HA-stimulated HOS cells with or without the presence of SNAP. Furthermore, HOS cells were pre-treated with anti-human integrin alphaV antibody prior to culturing on HA surfaces with or without the presence of SNAP. The levels of cAMP and cGMP were determined from the 3-day culture supernatants. The results showed that the production of cAMP but not cGMP by HA-stimulated HOS cells was augmented by SNAP. SQ22536 and carboxy PTIO suppressed but L-NIO only partially inhibited the production of cAMP by HA-stimulated HOS cells with or without the presence of exogenous NO. Pre-treatment of the cells with anti-human integrin alphaV antibody suppressed the production of cAMP by HA-stimulated HOS cells with or without the presence of NO. Therefore, the results of the present study suggest that NO may up-regulate the production of cAMP, perhaps, by augmenting adenylyl cyclase activity initiated by the binding between HOS cell-derived integrin alphaV and HA surface.
