Yan Guijun, Chen Shaoping, You Bei, Sun Jianxin
Department of Cell Biology and Molecular Medicine, University of Medicine and Dentistry of New Jersey , New Jersey Medical School, 185 South Orange Avenue, MGB G-653, Newark, NJ 07103, USA.
Cardiovasc Res. 2008 May 1;78(2):308-14. doi: 10.1093/cvr/cvn006. Epub 2008 Jan 10.
Recent evidence suggests that the epoxyeicosatrienoic acids (EETs), which are products of cytochrome P450 (CYP) epoxygenases, possess mitogenic and angiogenic effects in vascular endothelial cells. However, the mechanisms underlying these effects are not fully elucidated. Because sphingosine kinase (SK) and its product S1P play essential roles in cell growth, survival and migration, we hypothesized that SK activation by EETs may mediate some of its angiogenic effects.
We studied the effects of EETs on SK activity in human umbilical vein endothelial cells (HUVECs). Treatment with EETs, particularly 11,12-EET, markedly augmented SK activity in HUVECs. At the concentration of 1 micromol/L, 11,12-EET increased SK activity by 110% and the maximal effect on SK activation was observed at 20 min after 11,12-EET addition. Furthermore, inhibition of SK by a specific inhibitor, SKI-II, markedly attenuated 11,12-EET-induced EC proliferation. Importantly, 11,12-EET-induced activation of Akt kinase and transactivation of the epidermal growth factor (EGF) receptor was also inhibited by SKI-II. To investigate the isoform-specific role of SK in EET-induced angiogenesis, inhibition of SK1 by expression of dominant-negative SK1(G82D) substantially attenuated 11,12-EET-induced EC proliferation, migration, and tube formation in vitro and Matrigel plug angiogenesis in vivo. Furthermore, knockdown of SK1 expression by specific siRNA also inhibited 11,12-EET-induced EC proliferation and migration, whereas SK2 siRNA knockdown was without effect.
These results suggest that SK1 is an important mediator of the 11,12-EET-induced angiogenic effects in human ECs. Thus, SK1 may represent a novel therapeutic modality for the treatment of angiogenesis-related diseases such as cancer and ischaemia.
最近的证据表明，细胞色素P450（CYP）环氧化酶的产物环氧二十碳三烯酸（EETs）在血管内皮细胞中具有促有丝分裂和血管生成作用。然而，这些作用的潜在机制尚未完全阐明。由于鞘氨醇激酶（SK）及其产物S1P在细胞生长、存活和迁移中起重要作用，我们推测EETs激活SK可能介导其一些血管生成作用。
我们研究了EETs对人脐静脉内皮细胞（HUVECs）中SK活性的影响。用EETs处理，特别是11,12-EET，显著增强了HUVECs中的SK活性。在1微摩尔/升的浓度下，11,12-EET使SK活性增加了110%，并且在添加11,12-EET后20分钟观察到对SK激活的最大作用。此外，一种特异性抑制剂SKI-II对SK的抑制显著减弱了11,12-EET诱导的内皮细胞增殖。重要的是，SKI-II也抑制了11,12-EET诱导的Akt激酶激活和表皮生长因子（EGF）受体的反式激活。为了研究SK在EET诱导的血管生成中的亚型特异性作用，通过表达显性负性SK1（G82D）抑制SK1，在体外显著减弱了11,12-EET诱导的内皮细胞增殖、迁移和管形成，以及在体内基质胶栓血管生成。此外，用特异性siRNA敲低SK1表达也抑制了11,12-EET诱导的内皮细胞增殖和迁移，而敲低SK2 siRNA则没有效果。
这些结果表明，SK1是11,12-EET诱导人内皮细胞血管生成作用的重要介质。因此，SK1可能代表一种治疗癌症和缺血等血管生成相关疾病的新型治疗方式。
