Anti-Proliferative And Differentiation Inducing Activities Of Guanosine 5ˈ-Triphosphate On Osteosarcoma Cell Line

Among differentiating agents, purine bases and their corresponding nucleosides and nucleotideshave been the center of focuses due to their important and diverse effects on many biologicalprocesses and also their physiological roles as extracellular signal molecules. Recent studies haveproved that among purines, guanosine-base purines inhibit the excessive proliferation of a widevariety of cancer cells and guide them toward the maturation pathways. Based on these facts, antiproliferativeand osteogenic differentiation potential of guanosine 5ˈ-triphosphate (GTP) on humanosteosarcoma cell line Saos-2 was evaluated. Cell viability was measured by tetrazolium-basedcolorimetric assay. Cell cycle was analyzed by propidium iodide staining using flow cytometry.Osteogenic differentiation responses were registered by assessment of alkaline phosphataseactivity and the level of extracellular matrix mineralization which is detected and quantified byalizarin red S staining. Single dose treatment with variable concentrations of GTP revealed that GTPinhibits cell growth after 24 h of exposure and continued to increase in a time dependent manner,reaching a maximum value at 72-96 h of treatment. Cell cycle analyses by flow cytometry disclosedthat growth inhibition was accompanied with an accumulation of cells in the S-phase of the cellcycle. Moreover, precise assessment showed that GTP has ability to induce osteogenicdifferentiation which was quantified by monitoring ALP activity and calcium deposition. Asignificant increase was observed in ALP activity in GTP (400 μM) treated cells as compared withcontrol. Osteogenic differentiation was further confirmed by calcium mineralization under theinfluence of GTP. These finding may pave the way for further pharmaceutical evaluation of GTP as asuitable differentiating agent for poorly-differentiated osteosarcoma cells.