Figure 3.

Morphine increased cell surface expression of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and activated leukocyte cell adhesion molecule (ALCAM). Exposure to morphine (0.1 µM) for 2, 24, 48, and 72 h increased the number of hCMEC/D3 cells expressing ICAM-1 (A); VCAM-1 (B); and ALCAM (C) on the cell surface as determined by flow cytometry on nonpermeabilized hCMEC/D3 cells (panels on left). The levels of expression in untreated cells are shown in blue; individual isotype controls are shown in red for each antibody treatment. A range gate, as shown in these histograms, was used to quantitate the number of cells within a given fluorescence intensity (FI) range. Treatment with IL-1β (20 ng/mL) for 24 h induced an increased level of ICAM-1, VCAM-1, and ALCAM expression (compare the blue and brown lines in the left panels), while it slightly decreased the number of cells expressing the proteins on the cell surface (right panels). Quantitation of hCMEC/D3 cells expressing ICAM-1 (A); VCAM-1 (B); and ALCAM (C) is shown as the percent change in the number of expressing cells as compared with untreated controls (panels on right). The untreated control is shown as 0% change. All bars represent average percent change in the number of cells compared with the untreated control. All bars represent three independent experiments performed in triplicate. p < 0.001 for all treatment conditions compared with untreated controls by Student’s t-test.