After centrifuging twice at 10 000 g for 10 min, the supernatants of the reaction mixtures were designated PBS-platelet reactant and osteosarcoma-platelet reactant, respectively.
We found that the addition of the supernatant of an osteosarcoma-platelet reactant, but not that of the PBS-platelet reactant, significantly enhanced the growth of MG63/ZsGreen and HOS/ZsGreen cells (Fig. 1c).
These results indicate that the proliferation of osteosarcoma cells is increased in the presence of platelets as well as by supernatants of osteosarcoma-platelet reactant.
To assess the contribution of PDGFs to the activation of PDGFR-Akt axis, we treated MG63/ZsGreen and HOS/ZsGreen cells with the supernatant of the osteosarcoma-platelet reactant in the absence or presence of PDGFRs inhibitor, sunitinib.
In some experiments, the supernatant harvested from osteosarcoma-platelet reactants was added to the cultured osteosarcoma cells instead of platelets.
The addition of supernatants of osteosarcoma-platelet reactants also increased the growth of MG63 and HOS cells as well as the level of phosphorylated-PDGFR and -Akt.
To determine whether PDGF-BB was released during the osteosarcoma cell-mediated platelet aggregation, we measured the amount of PDGF-BB in the supernatants of the osteosarcoma-platelet reactants using an enzyme linked immunosorbent assay (ELISA).
(8) To identify the mechanism that mediated the effects of platelets and supernatants of osteosarcoma-platelet reactants on osteosarcoma cell proliferation, we used arrays comprising a panel of antibodies specific for cytokines, growth factor receptors, or downstream signaling components.
Objective measures included platelet count (acute phase reactant), white cell count, serum glucose, HDL-cholesterol, triglycerides, BMI, and waist circumference.
Among these are acute-phase reactants such as platelet factor 4, histidine-rich glycoprotein, vitronectin, fibronectin and lipopolysaccharide-binding protein, which increase in sepsis [ 19, 20].
Among these are acute-phase reactants such as platelet factor 4, histidine-rich glycoprotein, vitronectin, fibronectin and lipopolysaccharide-binding protein, all of which increase in sepsis and other forms of inflammation [ 2, 7- 9].
