GFP‐actin dynamics during histamine‐induced endothelial barrier dysfunction

Abstract

We tested the hypothesis that histamine‐induced endothelial barrier dysfunction is associated with disruption of normal actin dynamics at the endothelial cell periphery. Human umbilical vein endothelial cells (HUVEC) were transfected with GFP‐actin (500 ng vector/5 × 105 cells). Transendothelial resistance (TER) served as an index of barrier function. Time‐lapse image sets were acquired before and after the addition of 10 μM histamine. GFP‐actin expression slightly enhanced histamine‐induced endothelial barrier dysfunction, but did not affect the time course. GFP‐actin was found in cortical fibers moving centrally at a mean rate of 63 nm/min, and in membrane ruffles protruding outward at a mean rate of 1462 nm/min. New cortical fibers formed at a frequency of 0.59/min., and membrane ruffling occurred at a frequency of 0.61 events/min. GFP‐actin also localized in small lamellipodia on the cell periphery. Histamine treatment caused a sudden, coordinated formation of lamellipodia around the cell perimeter, followed by inactivity during the time frame of histamine‐induced barrier dysfunction. Shortly after, lamellepodia and membrane ruffling were restored. The data show that histamine‐indcued barrier dysfunction is associated with a loss of normal cortical actin dynamics in endothelial cells. Supported by NIH RR‐018766 and a grant from the American Heart Association.