血管内皮生长因子（VEGF）表达对氨基酸剥夺及内质网应激诱导剂的反应。

Response of VEGF expression to amino acid deprivation and inducers of endoplasmic reticulum stress.

作者信息

Abcouwer Steve F, Marjon Philip L, Loper Robyn K, Vander Jagt David L

机构信息

Department of Biochemistry and Molecular Biology, University of New Mexico, School of Medicine, Albuquerque, New Mexico 87131, USA.

出版信息

Invest Ophthalmol Vis Sci. 2002 Aug;43(8):2791-8.

PMID:12147617

Abstract

PURPOSE

Vascular endothelial growth factor (VEGF) plays an important role in initiation of the angiogenesis that leads to proliferative retinopathy. Several environmental conditions and chemical agents that influence the expression of VEGF can also cause endoplasmic reticulum (ER) stress. The hypothesis for the current study was that expression of VEGF is responsive to conditions that cause ER stress, including amino acid deprivation.

METHODS

Confluent cultures of a human retinal pigmented epithelial cell line (ARPE-19) were deprived of amino acids or treated with chemical inducers of ER stress. Treatment with cobalt was used to mimic hypoxia-induced expression of VEGF. Northern blot analysis was used to measure intracellular VEGF mRNA, and ELISA was used to measure secreted VEGF protein. Glucose-regulated protein 78 (GRP78) mRNA levels were compared with those of VEGF. Glyceraldehyde-phosphate dehydrogenase (GAPDH) mRNA was used as a control.

RESULTS

Conditions and chemical agents known to activate ER stress response (ERSR) pathways also induced the expression of VEGF. Deprivation of amino acids in the culture medium increased VEGF mRNA expression by 1.3- to 6-fold. Glucose deprivation or treatment of ARPE-19 cells with tunicamycin, brefeldin A, the calcium ionophore A23187, or thapsigargin increased the expression of VEGF mRNA in these cells by 8- to 10-fold. Expression of GRP78 mRNA was well correlated with that of VEGF mRNA under all conditions. These treatments also increased the secretion of VEGF protein by up to twofold. The increase in VEGF mRNA level in response to glutamine deprivation was rapid (greater than 10-fold) and was observed in a physiologically relevant range of glutamine concentrations. The half-life of VEGF mRNA was increased 2.5-fold by glutamine starvation.

CONCLUSIONS

These results indicate that VEGF is an ER stress-responsive gene and suggest that cells can respond to nutrient deprivation by increasing VEGF expression through both transcriptional and posttranscriptional mechanisms.

摘要

目的

血管内皮生长因子（VEGF）在导致增殖性视网膜病变的血管生成起始过程中起重要作用。几种影响VEGF表达的环境条件和化学试剂也可引起内质网（ER）应激。本研究的假设是VEGF的表达对引起ER应激的条件有反应，包括氨基酸剥夺。

方法

人视网膜色素上皮细胞系（ARPE-19）的汇合培养物被剥夺氨基酸或用ER应激的化学诱导剂处理。用钴处理以模拟缺氧诱导的VEGF表达。Northern印迹分析用于测量细胞内VEGF mRNA，ELISA用于测量分泌的VEGF蛋白。将葡萄糖调节蛋白78（GRP78）mRNA水平与VEGF的水平进行比较。甘油醛-3-磷酸脱氢酶（GAPDH）mRNA用作对照。

结果

已知激活内质网应激反应（ERSR）途径的条件和化学试剂也诱导了VEGF的表达。培养基中氨基酸的剥夺使VEGF mRNA表达增加1.3至6倍。葡萄糖剥夺或用衣霉素、布雷菲德菌素A、钙离子载体A23187或毒胡萝卜素处理ARPE-19细胞，使这些细胞中VEGF mRNA的表达增加8至10倍。在所有条件下，GRP78 mRNA的表达与VEGF mRNA的表达密切相关。这些处理还使VEGF蛋白的分泌增加了两倍。谷氨酰胺剥夺后VEGF mRNA水平的增加迅速（大于10倍），并且在生理相关的谷氨酰胺浓度范围内观察到。谷氨酰胺饥饿使VEGF mRNA的半衰期增加了2.5倍。