Abstract 491: Protein Kinase D2 Mediates LPA-induced Tissue Factor Expression and Activity through the MAPK Pathway in Mouse Aortic Smooth Muscle Cells

Abstract

Lysophosphatidic acid (LPA), a potent bioactive lipid in oxidized-LDL, modulates vascular cell functions in vitro and in vivo via regulating the expression of specific functional genes. We previously reported that LPA highly induces the expression of tissue factor (TF) in vascular smooth muscle cells. TF is the principal initiator of blood coagulation and is important for thrombosis. However, the intracellular signaling pathway leading to LPA induction of TF is not well understood. In the current study, we found that LPA markedly induced protein kinase D (PKD) activation in mouse primary aortic smooth muscle cells (MASMCs). PKD consists of three isoforms (PKD1, PKD2 and PKD3), which belong to a family of serine/threonine protein kinases. We observed that all three PKD isoforms were expressed in MASMCs. LPA-induced PKD phosphorylation is via a PKC-dependent pathway. Employing a small-interfering RNA approach to knock down the expression of the specific isoforms of PKD in MASMCs, we found that knockdown of PKD2 expression blocked LPA-induced TF expression and activity, indicating that PKD2 is the key intracellular mediator for the expression and surface activity of TF. The role of PKD2 was further confirmed by overexpression of the dominant-negative PKD2. We observed that overexpression of the dominant-negative isoform of PKD2 blocked LPA-induced TF expression. Our data also demonstrate that PKD2 mediated LPA-induced TF expression via the mitogen-activated protein kinase pathway. Using primary MASMCs isolated from LPA receptor knockout mice, our data revealed that LPA receptor 1 was responsible for LPA-induced TF expression. Therefore, these data provide the first evidence that PKD is a new component in LPA signaling in vascular SMCs, leading to gene expression, as evidenced by TF expression. Our data provide new insights into the mechanisms of coagulation and possible therapeutic targets for atherothrombosis.