CT-1 induces angiogenesis by regulating the ADMA/DDAH Pathway.
BACKGROUND
Cardiotrophin-1 (CT-1), a member of the IL-6 superfamily, is elevated in the serum of patients with ischemic and valvular heart disease. In this study, we hypothesized that CT-1 induces endothelial cell angiogenesis and that the ADMA/DDAH pathway plays an important role in the process.


METHODS
pEGFP-N1-CTF1-GFP and pEGFP-N1 were constructed and used to transiently transfect to HUVECs, mediated by LipofectamineTM 2000. After transfection, the expression of CT-1 was examined by qRT-PCR and western blotting. Endothelial cell proliferation assay was evaluated using the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide (MTT) method. Migration assay was performed using transwell, tube formation test was examined on Matrigel, eNOSmRNA expression was assayed by qRT-PCR, DDAH I, DDAHII and VEGF expression were detected by western blotting, the level of ADMA and the activity of DDAH were measured by High Performance Liquid Chromatography, NOS activity and the concentration of NO were assayed by L-[3H] citrulline production from L-[3H]arginine.


RESULTS
Overexpression of CT-1, increased endothelial cell proliferation, migration and formation of blood vessels, upregulated the expression of eNOSmRNA, DDAHI, DDAHII and VEGF, elevated the activity of DDAH and NOS, decreased the level of ADMA and promoted NO synthesis. In contrast, ADMA partially inhibited the effects of CT-1 induction.


CONCLUSIONS
Overexpression of CT-1 increases cell proliferation, migration and formation of blood vessels. This result also suggests that CT-1 may regulate angiogenesis through the ADMA/DDAH pathway.
Department of Cardiology, The First Afficiated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China.
Zhen Zhong
ZZ
Zheng
Department of Cardiology, The First Afficiated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China.|Department of Electrocardiography, Hubei Provincial Maternal and Child Health Institute, Hubei, Wuhan, 430000, China.
Xiao
X
Tian Fu
Department of Cardiology, The First Afficiated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China.
Jin
J
Liang
Department of Cardiology, The First Afficiated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China.
Zhi
Z
Bing Guo
D000581:Amidohydrolases / Q000378:metabolism*; D001120:Arginine / Q000031:analogs & derivatives* / Q000378:metabolism; D054443:Cell Migration Assays; D002465:Cell Movement / Q000502:physiology; D049109:Cell Proliferation; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism / Q000502:physiology*; D004730:Endothelium, Vascular / Q000166:cytology; D061307:Human Umbilical Vein Endothelial Cells / Q000166:cytology; D006801:Humans; D018919:Neovascularization, Physiologic / Q000502:physiology*; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D060185:Tetraspanin 24 / Q000378:metabolism; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
D016207:Cytokines; D012333:RNA, Messenger; D060185:Tetraspanin 24; D042461:Vascular Endothelial Growth Factor A; D009569:Nitric Oxide; C018524:N,N-dimethylarginine; D001120:Arginine; C092073:cardiotrophin 1; D052250:Nitric Oxide Synthase Type III; D000581:Amidohydrolases; C059999:dimethylargininase
ADMA; DDAH; angiogenesis; cardiotrophin-1 (CT-1); eNOS