TY - JOUR
T1 - CCN1 acutely increases nitric oxide production via integrin αvβ3-Akt-S6K-phosphorylation of endothelial nitric oxide synthase at the serine 1177 signaling axis
AU - Hwang, Soojin
AU - Lee, Hyeon Ju
AU - Kim, Gyungah
AU - Won, Kyung Jong
AU - Park, Yoon Shin
AU - Jo, Inho
N1 - Funding Information:
This work was supported by a Mid-career Research Program grant ( 2012R1A2A2A01004914 and 2015R1A2A2A01003315 ) through the National Research Foundation of Korea funded by the Ministry of Education, Science, and Technology .
Publisher Copyright:
© 2015 Elsevier Inc.
PY - 2015/12/1
Y1 - 2015/12/1
N2 - Although CCN1 (also known as cysteine-rich, angiogenic inducer 61, CYR61) has been reported to promote angiogenesis and neovascularization in endothelial cells (ECs), its effects on endothelial nitric oxide (NO) production have never been studied. Using human umbilical vein ECs, we investigated whether and how CCN1 regulates NO production. CCN1 acutely increased NO production in a time- and dose-dependent manner, which was accompanied by increased phosphorylation of endothelial NO synthase (eNOS) at serine 1177 (eNOS-Ser1177), but not that of eNOS-Thr495 or eNOS-Ser114. The level of total eNOS expression was unaltered. Treatment with either LY294002, a selective inhibitor of phosphoinositide 3-kinase known as an upstream kinase of Akt, or H-89, an inhibitor of protein kinase A, mitogen- and stress-activated protein kinase 1, Rho-associated protein kinase 2, and ribosomal protein S6 kinase (S6K), inhibited CCN1-stimulated eNOS-Ser1177 phosphorylation and subsequent NO production. Ectopic expression of small interfering RNA against Akt and S6K significantly inhibited the effects of CCN1. Consistently, CCN1 increased the phosphorylation of Akt-Ser473 and S6K-Thr389. However, CCN1 did not alter the expression or secretion of VEGF, a known downstream factor of CCN1 and a potential upstream factor of Akt-mediated eNOS-Ser1177 phosphorylation. Furthermore, neutralization of integrin αvβ3 with corresponding antibody completely reversed all of the observed effects of CCN1. Moreover, CCN1 increased acetylcholine-induced relaxation in the rat aortas. Finally, we also found that CCN1-stimulated eNOS-Ser1177 phosphorylation and NO production are true for other types of EC tested. In conclusion, CCN1 acutely increases NO production via activation of a signaling axis in integrin αvβ3-Akt-S6K-eNOS-Ser1177 phosphorylation, suggesting an important role for CCN1 in vasodilation.
AB - Although CCN1 (also known as cysteine-rich, angiogenic inducer 61, CYR61) has been reported to promote angiogenesis and neovascularization in endothelial cells (ECs), its effects on endothelial nitric oxide (NO) production have never been studied. Using human umbilical vein ECs, we investigated whether and how CCN1 regulates NO production. CCN1 acutely increased NO production in a time- and dose-dependent manner, which was accompanied by increased phosphorylation of endothelial NO synthase (eNOS) at serine 1177 (eNOS-Ser1177), but not that of eNOS-Thr495 or eNOS-Ser114. The level of total eNOS expression was unaltered. Treatment with either LY294002, a selective inhibitor of phosphoinositide 3-kinase known as an upstream kinase of Akt, or H-89, an inhibitor of protein kinase A, mitogen- and stress-activated protein kinase 1, Rho-associated protein kinase 2, and ribosomal protein S6 kinase (S6K), inhibited CCN1-stimulated eNOS-Ser1177 phosphorylation and subsequent NO production. Ectopic expression of small interfering RNA against Akt and S6K significantly inhibited the effects of CCN1. Consistently, CCN1 increased the phosphorylation of Akt-Ser473 and S6K-Thr389. However, CCN1 did not alter the expression or secretion of VEGF, a known downstream factor of CCN1 and a potential upstream factor of Akt-mediated eNOS-Ser1177 phosphorylation. Furthermore, neutralization of integrin αvβ3 with corresponding antibody completely reversed all of the observed effects of CCN1. Moreover, CCN1 increased acetylcholine-induced relaxation in the rat aortas. Finally, we also found that CCN1-stimulated eNOS-Ser1177 phosphorylation and NO production are true for other types of EC tested. In conclusion, CCN1 acutely increases NO production via activation of a signaling axis in integrin αvβ3-Akt-S6K-eNOS-Ser1177 phosphorylation, suggesting an important role for CCN1 in vasodilation.
KW - Akt
KW - CCN1
KW - Endothelial nitric oxide synthase
KW - Free radicals
KW - Integrin
KW - Nitric oxide
KW - Phosphorylation
KW - S6K
KW - Vasodilation
UR - http://www.scopus.com/inward/record.url?scp=84943181992&partnerID=8YFLogxK
U2 - 10.1016/j.freeradbiomed.2015.08.005
DO - 10.1016/j.freeradbiomed.2015.08.005
M3 - Article
C2 - 26393424
AN - SCOPUS:84943181992
SN - 0891-5849
VL - 89
SP - 229
EP - 240
JO - Free Radical Biology and Medicine
JF - Free Radical Biology and Medicine
M1 - 12538
ER -