Matrone Gianfranco, Meng Shu, Gu Qilin, Lv Jie, Fang Longhou, Chen Kaifu, Cooke John P
From the Department of Cardiovascular Sciences, Houston Methodist Research Institute, TX.
Arterioscler Thromb Vasc Biol. 2017 Oct;37(10):1860-1868. doi: 10.1161/ATVBAHA.117.309609. Epub 2017 Aug 3.
(LIM-domain-only)2 transcription factor is involved in hematopoiesis and vascular remodeling. (sphingosine kinase)1 phosphorylates sphingosine to S1P (sphingosine-1-phosphate). We hypothesized that regulates to promote endothelial cell (EC) migration and vascular development. APPROACH AND RESULTS: and knockdown (KD) were performed in ) zebrafish and in human umbilical vein EC. Rescue of phenotypes or overexpression of these factors were achieved using mRNA encoding or . EC proliferation in vivo was assessed by BrdU (bromodeoxyuridine) immunostaining and fluorescence-activated cell sorter analysis of dissociated ) embryos. Cell migration was assessed by scratch assay in human umbilical vein EC and mouse aortic rings. interactions with promoter were assessed by ChIP-PCR (chromatin immunoprecipitation-polymerase chain reaction). or KD reduced number and length of intersegmental vessels. There was no reduction in the numbers of GFP (green fluorescent protein) ECs after KD. However, reduced numbers of BrdUGFP nuclei were observed along the dysmorphic intersegmental vessels, accumulating instead at the sprouting origin of the intersegmental vessels. This anomaly was likely because of impaired EC migration, which was confirmed in migration assays using KD human umbilical vein ECs and mouse aortic rings. Both in vivo and in vitro, KD reduced gene expression, associated with less binding to the promoter as assessed by ChIP-PCR. mRNA rescued the KD phenotype.
Our data showed that is necessary for gene expression in ECs. KD reduced - gene interaction, impaired intersegmental vessels formation, and reduced cell migration. We identified for the first time as downstream effector of .
（仅含LIM结构域）2转录因子参与造血和血管重塑。鞘氨醇激酶1将鞘氨醇磷酸化为S1P（1-磷酸鞘氨醇）。我们推测[此处原文有缺失信息未明确指出推测的主体]调节[此处原文有缺失信息未明确指出被调节的对象]以促进内皮细胞（EC）迁移和血管发育。
在斑马鱼和人脐静脉内皮细胞中进行[此处原文有缺失信息未明确指出具体进行敲低的对象]敲低（KD）。使用编码[此处原文有缺失信息未明确指出具体因子]的mRNA实现这些因子的表型挽救或过表达。通过BrdU（溴脱氧尿苷）免疫染色和对解离的[此处原文有缺失信息未明确指出具体胚胎]胚胎进行荧光激活细胞分选分析来评估体内EC增殖。通过人脐静脉内皮细胞和小鼠主动脉环的划痕试验评估细胞迁移。通过染色质免疫沉淀 - 聚合酶链反应（ChIP - PCR）评估[此处原文有缺失信息未明确指出具体因子]与[此处原文有缺失信息未明确指出具体启动子]启动子的相互作用。[此处原文有缺失信息未明确指出具体因子]或[此处原文有缺失信息未明确指出具体因子]KD减少了节间血管的数量和长度。[此处原文有缺失信息未明确指出具体因子]KD后GFP（绿色荧光蛋白）内皮细胞数量没有减少。然而，在畸形的节间血管中观察到BrdU - GFP细胞核数量减少，反而在节间血管的发芽起源处积累。这种异常可能是由于内皮细胞迁移受损，这在使用[此处原文有缺失信息未明确指出具体因子]KD人脐静脉内皮细胞和小鼠主动脉环的迁移试验中得到证实。在体内和体外，[此处原文有缺失信息未明确指出具体因子]KD均降低了[此处原文有缺失信息未明确指出具体基因]的基因表达，如通过ChIP - PCR评估的那样，这与[此处原文有缺失信息未明确指出具体因子]与[此处原文有缺失信息未明确指出具体启动子]启动子的结合减少有关。[此处原文有缺失信息未明确指出具体因子]mRNA挽救了[此处原文有缺失信息未明确指出具体因子]KD表型。
我们的数据表明[此处原文有缺失信息未明确指出具体因子]对于内皮细胞中[此处原文有缺失信息未明确指出具体基因]的基因表达是必需的。[此处原文有缺失信息未明确指出具体因子]KD减少了[此处原文有缺失信息未明确指出具体因子] - [此处原文有缺失信息未明确指出具体基因]的相互作用，损害了节间血管形成，并减少了细胞迁移。我们首次确定[此处原文有缺失信息未明确指出具体因子]为[此处原文有缺失信息未明确指出具体因子]的下游效应物。
