Shear stress regulates occludin content and phosphorylation

Previous studies determined that shear stress imposed on bovine aortic endo thelial cell (BAEC) monolayers increased the hydraulic conductivity (L-P); however, the mechanism by which shear stress increases L-P remains unknown. This study tested the hypothesis that shear stress regulates paracellular transport by altering the expression and phosphorylation state of the tight junction protein occludin. The effect of shear stress on occludin content was examined by Western blot analysis. Ten dyn/cm(2) significantly reduced occludin content in a time-dependent manner such that after a 3 h exposure to shear, occludin content decreased to 44% of control. Twenty dyn/cm(2) de creased occludin content to 50% of control and increased L-P by 4.7-fold af ter 3 h. Occludin expression and L-P depend on tyrosine kinase activity bec ause erbstatin A (10 muM) attenuated both the shear-induced decrease in occ ludin content and increase in L-P. Shear stress increased occludin phosphor ylation after 5 min, 15 min, and 3 h exposures. The shear-induced increase in occludin phosphorylation was attenuated with dibutyryl (DB) cAMP (1 mM), a reagent previously shown to reverse the shear-induced increase in L-P. W e conclude that shear stress rapidly (less than or equal to5 min) increases occludin phosphorylation and significantly decreases the expression of occ ludin over 1-4 h. Alterations in the occludin phosphorylation state and occ ludin total content are potential mechanisms by which shear stress increase s L-P.