Overexpression of p120 in HUVECs inhibits PMN transmigration. (A) The level of p120 GFP fluorescence and PMN transmigration was quantified as described in “Image acquisition and analysis” and “Quantitation of p120GFP and VE-cad fluorescence in endothelium,” using live cell fluorescence digital imaging. The p120GFP expression was grouped into 3 categories (low, intermediate, and high). (B) The amount of TEM was determined in each category. Values represent the mean plus or minus SD of 5 different experiments. P values are indicated, comparing each bar with GFP construct. ***P < .001; **P < .01. (C) PMNs remained bound at the cell-cell junctions on monolayers overexpressing p120GFP, but disappeared from the cell-cell junctions and efficiently transmigrated when perfused on HUVECs transduced with GFP. Values represent (number of cells bound to the junctions)/(number of total cells bound) × 100, and are the mean plus or minus SD of 5 different experiments. *P < .01. (D) The inhibitory effect of p120 overexpression was not due to a delay in PMN transmigration, as the block of TEM was sustained for more than 20 minutes. The percentage of TEM was normalized by dividing percentage of TEM at each time point by the number of cells that were bound initially at time 0. P values for p120GFP versus GFP at each time point are indicated. *P < .05; **P < .01. Data represent mean plus or minus SD of 3 different experiments. (E) Total accumulation of PMNs was similar on VE-cad–GFP– or GFP-transduced monolayers. (F) Overexpression of VE-cadherin GFP did not affect PMN TEM. Values in panels E and F represent the mean plus or minus SD from 3 different experiments using HUVECs and PMNs from different donors.