Open AccessCaveolin‐1 is important for nitric oxide‐mediated angiogenesis in fibrin gels with human umbilical vein endothelial cells 1
Author(s) -
PAN Yiming,
YAO Yongzhong,
ZHU Zhanghua,
SUN Xitai,
QIU Yudong,
DING Yitao
Publication year - 2006
Publication title -
acta pharmacologica sinica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.514
H-Index - 90
eISSN - 1745-7254
pISSN - 1671-4083
DOI - 10.1111/j.1745-7254.2006.00462.x
Subject(s) - umbilical vein , angiogenesis , enos , nitric oxide , fibrin , human umbilical vein endothelial cell , microbiology and biotechnology , vascular endothelial growth factor , caveolin , chemistry , endothelium , nitric oxide synthase , biology , endocrinology , immunology , biochemistry , caveolae , in vitro , cancer research , signal transduction , vegf receptors
Aim: The role of caveolin‐1 (Cav‐1) in angiogenesis remains poorly understood. The endothelial nitric oxide (NO) synthase (eNOS), a caveolin‐interacting protein, was demonstrated to play a predominant role in vascular endothelial growth factor (VEGF) ‐induced angiogenesis. The purpose of our study was to examine the role of Cav‐1 and the eNOS complex in NO‐mediated angiogenesis. Methods: Human umbilical vein endothelial cells (HUVEC) were isolated and cultured in 3 ‐D fibrin gels to form capillary‐like tubules by VEGF stimulation. The expression of Cav‐1 and eNOS was detected by semiquantitative RT‐PCR. The HUVEC were treated with antisense oligonucleotides to downregulate Cav‐1 expression. Both transduced and non‐infected HUVEC were cultured in fibrin gels in the presence or absence of VEGF (20 ng/mL) and N G ‐nitro‐ L ‐arginine methyl ester ( L ‐NAME; 5 mmol/L). NO was measured using a NO assay kit and capillary‐like tubules were quantified by tubule formation index using the Image J program. Results: RT‐PCR analysis revealed that Cav‐1 levels steadily increased in a time‐dependent manner and reached their maximum after 5 d of incubation, but there were no obvious changes in eNOS mRNA expression in response to VEGF in the fibrin gel model. VEGF (20 ng/mL) can promote NO production and the formation of capillary‐like tubules, and this promoting effect of VEGF was blocked by the addition of L ‐NAME (5 mmol/L). When transduced HUVEC with the antisense Cav‐1 oligonucleotides were plated in the fibrin gels, the capillary‐like tubules were significantly fewer than those of the non‐infected cells. The capillary‐like tubules formation and NO production of transduced HUVEC with the antisense Cav‐1 oligonucleotides cultured in fibrin gels showed no responses to the addition of VEGF (20 ng/mL) and L ‐NAME (5.0 mmol/L). Conclusion: NO was a critical angiogenic mediator in this model. Cav‐1 was essential for NO‐mediated angiogenesis and may be an important target of anti‐angiogenesis therapy.