Lee Chang-Kwon, Lee Hwan Myung, Kim Hyo Jin, Park Hyo-Jun, Won Kyung-Jong, Roh Hui Yul, Choi Wahn Soo, Jeon Byeong Hwa, Park Tae-Kyu, Kim Bokyung
Department of Physiology, College of Medicine, Konkuk University, Danwol-dong 322, Chungju 380-701, Republic of Korea.
Cardiovasc Res. 2007 Apr 1;74(1):159-68. doi: 10.1016/j.cardiores.2007.01.012. Epub 2007 Jan 18.
Here we investigated the role of spleen tyrosine kinase (Syk) in the migration induced by platelet-derived growth factor (PDGF) in rat aortic smooth muscle cells (RASMC).
Cell migration was determined using a Boyden chamber, by wound-healing, and by aortic ring assays. Activity of Syk, mitogen-activated protein kinase (MAPK), and heat shock protein 27 (HSP27) were tested using immunoblotting with kinase inhibitors and small interference RNAs.
PDGF-BB induced binding of Syk to the PDGFbeta receptor and increased the phosphorylation of Syk and migration in RASMC. These effects of PDGF-BB were inhibited by piceatannol, an inhibitor of Syk. PDGF-BB increased the phosphorylation of extracellular signal-regulated kinase (ERK) 1/2, p38 MAPK, and HSP27, which were significantly inhibited by piceatannol and in Syk-knockdown cells. The p38 MAPK inhibitor SB203580 and ERK1/2 inhibitor PD98059 inhibited the migration, which was further inhibited by the combination of these inhibitors. SB203580, but not PD98059, inhibited the phosphorylation of HSP27 induced by PDGF-BB in RASMC. PDGF-BB-induced migration was attenuated in HSP27-knockdown cells. Kinase inhibitors and Syk-knockdown diminished PDGF-BB-induced sprout outgrowth in the aortic ring assay.
These results imply that Syk is an upstream signal of the p38 MAPK/HSP27 and ERK1/2 pathways that contributes to PDGF-BB-mediated migration in RASMC.
本研究旨在探讨脾酪氨酸激酶（Syk）在血小板衍生生长因子（PDGF）诱导大鼠主动脉平滑肌细胞（RASMC）迁移中的作用。
采用博伊登小室法、伤口愈合实验和主动脉环实验检测细胞迁移。使用激酶抑制剂和小干扰RNA通过免疫印迹法检测Syk、丝裂原活化蛋白激酶（MAPK）和热休克蛋白27（HSP27）的活性。
PDGF-BB诱导Syk与PDGFβ受体结合，并增加RASMC中Syk的磷酸化和细胞迁移。Syk抑制剂白皮杉醇可抑制PDGF-BB的这些作用。PDGF-BB增加细胞外信号调节激酶（ERK）1/2、p38 MAPK和HSP27的磷酸化，白皮杉醇和Syk基因敲低细胞可显著抑制这种作用。p38 MAPK抑制剂SB203580和ERK1/2抑制剂PD98059可抑制细胞迁移，二者联合使用时抑制作用更强。SB203580可抑制PDGF-BB诱导的RASMC中HSP27的磷酸化，而PD98059则不能。在HSP27基因敲低细胞中，PDGF-BB诱导的细胞迁移减弱。激酶抑制剂和Syk基因敲低可减少主动脉环实验中PDGF-BB诱导的血管生成。
这些结果表明，Syk是p38 MAPK/HSP27和ERK1/2信号通路的上游信号，参与PDGF-BB介导的RASMC迁移。
