Human coronary arterial SMCs were cultured in聽vitro. Boyden microchemotaxis assays of migration were performed in response to S-1-P in the presence and absence the src inhibitor (PP2, 10 渭M) and a dominant negative src construct (DNsrc). siRNA to S-1-P receptors was used to down-regulate the S-1-P receptors. Western blotting was performed for src and MAPK phosphorylation.
Inhibition of src with PP2 but not PP3 partially blocked S-1-P-mediated cell migration. S-1-P induced time-dependent activation of src, which was inhibited by PP2 and adenoviral DNsrc. PP3 or an empty vector had no effect. Activation of src by S-1-P was inhibited by siRNA to S-1-PR1 and S-1-PR3 but not by S-1-PR2. When the VSMC were transfected with adenovirus containing 尾ARKCT, an inhibitor to G尾纬, src activation was significantly attenuated. Src inhibition with PP2 reduced p38MAPK and JNK activation but did not alter ERK1/2 activation.
S-1-P mediated VSMC migration is modulated by a G-protein-coupled src pathway partially through src-mediated p38MAPK and JNK signaling and requires S-1-PR1 and S-1-PR3 receptors.