G protein activation in endothelial cells by stretch

Endothelial cells sense and respond to their mechanical environment both in vivo and in vitro. Fluid shear and mechanical stretch have been demonstrated to modulate G protein activation, second messenger activity, as well as gene expression. Here we demonstrate activation of a 42 kDa G protein in human umbilical vein endothelial cells by mechanical stretch, utilizing a radiolabeled (³²P) GTP analogue. Confluent monolayers cultured on collagen coated silastic membranes were incubated with the GTP label and subjected to various regimens of 2, 10, and 25% stretch over 60 seconds. Activation, as assessed by the amount of bound nucleotide, was sensitive to both the strain magnitude as well as the rate at which the strain was applied. A strain of 10% applied at 20% per second (60 cycles) increased activation 1.8-fold relative to identically cultured sham controls, while 10% stretch applied at 0.3% per second did not increase activation. Similarly, 60 cycles of 2% strain did not significantly increase activation. A small G protein of approximately 28 kDa was also modulated by stretch, showing an inverse relationship, with a decrease in bound GTP as either the strain or strain magnitude increased