Lack of effect of OPG on the expression of adhesion molecules. HUVECs were exposed to OPG or TNF-α (used as positive control) for 16 hours and expression levels of adhesion molecules (ICAM-1, VCAM-1, and E-selectin) were evaluated at both (A) mRNA and (B) protein levels by ELISA and (C) flow cytometry. (A) RNA from HUVECs either left untreated or exposed to OPG or TNF-α were quantitatively analyzed by ICAM-1, VCAM-1 and E-selectin RT-PCR after normalization to the level of GAPDH mRNA. Each sample was determined in duplicate. (B) Adhesion molecules expression was determined by ELISA and data are shown as average OD ± SD of 3 independent experiments. In panel C, the control (unshadowed) histograms represent the background fluorescence obtained from the staining of the same cultures with isotype-matched control antibodies. One of 6 experiments with similar results is shown.