高壓氧處理之人類臍靜脈內皮細胞所誘導之血管內皮細胞生長因子 VEG
F 表現的分子機制探討
高壓氧治療可促進血管新生，進而加速和缺氧相關的傷口癒合，而血
管內皮細胞生長因子 (VEGF) 及其接受器系統無論在生理或病理性
的血管新生都是最主要的調控者。然而高壓氧調控血管內皮細胞活化
的機制尚未明瞭。本實驗即是在探討高壓氧活化血管內皮細胞生長所
扮演的角色及其可能的調控機制。我們由 Real-time PCR 及北方墨點
分析數據證實，高壓氧可誘發血管內皮細胞生長因子基因的表現，並
於 4 小時達到最高峰，且高壓氧所誘發 VEGF mRNA 的表現可被 ME
K 的抑制劑 PD98059 及 JNK 的抑制劑 SP600125 所抑制。另一方面，
西方墨點分析法顯示， VEGF 蛋白最大表現量在 8 小時， ERK1/2 的
磷酸化則在 15 分鐘最強。此外， VEGF 的 promoter 上有 hypoxia ind
ucible factor-1 (HIF-1) 和 Activator protein-1 (AP-1) 轉錄因子的結合位
，由 electrophoretic mobility shift assay (EMSA) 結果顯示，高壓氧處
理 1 小時可明顯活化此二轉錄因子，且其 DNA-protein 的結合能力皆
會被 PD98059 及 SP600125 所抑制。因此推測高壓氧可能是透過 JN
K-mitogenic-activated protein kinase 及 ERK-mitogenic-activated protein
kinase 的訊息傳遞路徑，活化 HIF-1 和 AP-1 轉錄因子而啟動 VEGF
基因的表現，進而促進血管的新生。
Mechanisms of The Induction of Vascular Endothelial Growth Factor
Expression by Hyperbaric Oxygen in Human Umbilical Vein Endothelial
Cells
One of the biological effects of hyperbaric oxygen (HBO) therapy in enhancing ischemia-relat ed wound healing is the induction of angiogenesis. The vascular endothelial growth factor VE GF/VEGF receptor system plays a central regulatory role in physiological and pathological an giogenesis. During embryogenesis, the VEGF/VEGF receptor system is critically involved in t he formation of the vascular system by regulating both the growth and the survival of blood ve ssels. The mechanism by which hyperbaric oxygen regulates angiogenesis is presently unclear . The aim of the present study was to investigate the mechanism of the activation of VEGF ge ne activation in human umbilical vein endothelial cells (HUVECs) exposed to HBO. Prelimina ry Real-time quantitative PCR studies revealed that the expression of VEGF mRNA was induc ed after 4 hours of exposure. The induction of VEGF mRNA was further confirmed by Northe rn blot analysis. Western blot analysis showed that HBO enhanced the VEGF protein level an d increased the ERK1/2 activity, with a maximal level after 8 hours and 15 min, respectively.
Whereas, JNK inhibitor SP600125 and MEK inhibitor PD98059 blocked the HBO-induced V
EGF mRNA expression. Since VEGF promoter region contains AP-1 and HIF-1 binding sites,
stimulation of HUVECs with HBO enhanced AP-1 and HIF-1 binding to their DNA binding si
tes as demonstrated by electrophoretic mobility shift assay(EMSA). This effect was also inhibi
ted by SP600125 and PD98059. These data suggested that HBO-induced VEGF expression mi
ght be through AP-1 and HIF-1 stimulation, and required the involvement of JNK and ERK-m
itogenic-activated protein kinase signaling pathways, which may play an important role in HB
O-induced angiogenesis.
