The importance of Fyn tyrosine kinase in Ca²⁺-sensitization of vascular smooth muscle contraction induced by a sphingosylphosphorylcholine and Rho-kinase pathway.

Abstract

Whereas the Ca²⁺-dependent contraction of vascular smooth muscle (VSM) which regulates physiological vascular tone, the Rho-kinase (ROK)-mediated Ca²⁺-sensitization of VSM contraction contributes to abnormal VSM contraction such as vasospasm. We previously found that sphingosylphosphorylcholine (SPC) is an upstream messenger for the ROK-mediated Ca²⁺ sensitization and that inhibitors of Src family tyrosine kinase (Src-TK) blocked the SPC-induced contraction and activation of ROK. In the present study, we attempted to determine the enzyme molecule in a family of Src-TK which contributes to the Ca²⁺-sensitization mediated by a SPC/ROK pathway. In order to accomplish this purpose, we performed knockdown of the target molecule by using siRNA which was transfected into the human coronary artery smooth muscle cells (CASMCs) with the efficiency of about 100%. The siRNA-mediated knockdown of Fyn inhibited the SPC-induced contraction of CASMC, whereas non-silencing control siRNA lacked any effect. These results provide the first direct evidence that Fyn mediates the Ca²⁺-sensitization of VSM contraction induced by a SPC/ROK pathway. In addition, Fyn constructs (wild, constitutively active, and dominant negative types) were transfected to CASMCs with high efficiency (> 50%), although CASMCs were well-differentiated contractile cells. In poster presentation, the effects of transient overexpression of Fyn constructs on the contraction of CASMCs will be also discussed. [J Physiol Sci. 2006;56 Suppl:S116]