范蓓, 王艳霞, 姚泰, 朱依纯
血管内皮细胞中血管内皮生长因子（vascular endothelial growth factor, VEGF）的合成增加在促进血管新生的过程中起着非常重要的作用。然而低氧诱导VEGF分泌的细胞内信号传导机制还不是很清楚。人脐静脉内皮细胞系（ECV304）在低氧或常氧的状态下培养12--24小时后分别用实时定量PCR和Western blot的方法来检测VEGFmRNA的表达及ERK1/2和P38激酶的磷酸化水平。分泌到培养液中的VEGF蛋白用酶联免疫吸附（ELISA）的方法来检测。已报道，ERK的抑制剂PD98059能够抑制低氧诱导的VEGF基因的表达，根据这个报道，我们发现低氧情况下，ECV304细胞的ERK1/2磷酸化水平增高以及VEGF的合成增加等这些变化也能被PD98059所抑制。本次实验的新发现是P38激酶的激活在低氧诱导VEGF合成增加中的作用。P38激酶的抑制剂SB202190能抑制低氧诱导的VEGF合成增加。这些数据首次直接证实了P38激酶在低氧诱导人内皮细胞分泌VEGF增加过程中的作用。
p38 Mitogen-activated protein kinase mediates hypoxia-induced vascular endothelial growth factor release in human endothelial cells
Fan Bei, Wang Yanxia, Yao Tai, Zhu Yichun
Department of Physiology and Pathophysiology, Key Laboratory of Molecular Medicine of The Ministry of Education, Fudan University Shanghai Medical College.Shanghai 200032
Increased vascular endothelial growth factor (VEGF) biosynthesis in vascular endothelial cells has been reported to play an obligatory role in promoting angiogenesis. Nevertheless, the intracellular signaling mechanisms of hypoxia-induced VEGF release remain largely unknown. Human umbilical vein endothelial cell lines (ECV304) were cultured in normoxic or hypoxic conditions for 12-24 hours and harvested for determination of VEGF mRNA expression and phosphorylation of ERK1/2 and p38 MAPK by real-time reverse transcription polymerase chain reaction (RT-PCR) and Western blot analysis, respectively. Secreted VEGF protein was measured by enzyme-linked immunosorbent assay (ELISA). It has reported that PD98059, an ERK inhibitor, was able to blunt the hypoxia-induced activation of the expression of VEGF gene. In accordance with this report, an increase in ERK1/2 phosphorylation and VEGF biosynthesis was observed in ECV304 cells cultured in hypoxia, and this increase was blocked by PD98059. The novel finding of the present study is that an activation of p38 MAP kinase is involved in hypoxia-induced increase in VEGF biosynthesis. SB202190, an inhibitor of p38 MAPK was able to blunt the hypoxia-induced increase in VEGF biosynthesis. These dada provide the first direct evidence for a role of p38 MAPK in mediating hypoxia-induced increase in VEGF biosynthesis in human endothelial cells.
范蓓, 王艳霞, 姚泰, 朱依纯. P38丝裂原素激活的蛋白激酶在调节低氧诱导人内皮细胞分泌血管内皮生长因子过程中的作用[J]. 生理学报 2005; 57 (1): .
Fan Bei, Wang Yanxia, Yao Tai, Zhu Yichun. p38 Mitogen-activated protein kinase mediates hypoxia-induced vascular endothelial growth factor release in human endothelial cells. Acta Physiol Sin 2005; 57 (1): (in Chinese with English abstract).