Nitric oxide stimulates vascular endothelial growth factor production in cardiomyocytes involved in angiogenesis.
BACKGROUND
Hypoxia-inducible factor (HIF)-1alpha regulates the transcription of lines of genes, including vascular endothelial growth factor (VEGF), a major gene responsible for angiogenesis. Several recent studies have demonstrated that a nonhypoxic pathway via nitric oxide (NO) is involved in the activation of HIF-1alpha. However, there is no direct evidence demonstrating the release of angiogenic factors by cardiomyocytes through the nonhypoxic induction pathway of HIF-1alpha in the heart. Therefore we assessed the effects of an NO donor, S-Nitroso-N-acetylpenicillamine (SNAP) on the induction of VEGF via HIF-1alpha under normoxia, using primary cultured rat cardiomyocytes (PRCMs).


RESULTS
PRCMs treated with acetylcholine (ACh) or SNAP exhibited a significant production of NO. SNAP activated the induction of HIF-1alpha protein expression in PRCMs during normoxia. Phosphatidylinositol 3-kinase (PI3K)-dependent Akt phosphorylation was induced by SNAP and was completely blocked by wortmannin, a PI3K inhibitor, and NG-nitro-L-arginine methyl ester (L-NAME), a NO synthase inhibitor. The SNAP treatment also increased VEGF protein expression in PRCMs. Furthermore, conditioned medium derived from SNAP-treated cardiomyocytes phosphorylated the VEGF type-2 receptor (Flk-1) of human umbilical vein endothelial cells (a fourfold increase compared to the control group, p < 0.001, n = 5) and accelerated angiogenesis.


CONCLUSIONS
Our results suggest that cardiomyocytes produce VEGF through a nonhypoxic HIF-1alpha induction pathway activated by NO, resulting in angiogenesis.
Kuwabara
Masanori
M
Department of Cardiovascular Control, Kochi Medical School, Nankoku, Japan.
Kakinuma
Yoshihiko
Y
Ando
Motonori
M
Katare
Rajesh G
RG
Yamasaki
Fumiyasu
F
Doi
Yoshinori
Y
Sato
Takayuki
T
D000109:Acetylcholine / Q000494:pharmacology; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000737:chemistry / Q000166:cytology; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000032:analysis / Q000235:genetics / Q000502:physiology; D032383:Myocytes, Cardiac / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D018919:Neovascularization, Physiologic / Q000187:drug effects* / Q000502:physiology; D009569:Nitric Oxide / Q000494:pharmacology*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D010396:Penicillamine / Q000031:analogs & derivatives / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D015398:Signal Transduction / Q000502:physiology; D014471:Umbilical Veins / Q000737:chemistry / Q000166:cytology; D042461:Vascular Endothelial Growth Factor A / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000032:analysis / Q000378:metabolism; D000077191:Wortmannin
D000730:Androstadienes; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D020030:Nitric Oxide Donors; C110881:S-nitro-N-acetylpenicillamine; D042461:Vascular Endothelial Growth Factor A; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D019869:Phosphatidylinositol 3-Kinases; D040301:Vascular Endothelial Growth Factor Receptor-2; D051057:Proto-Oncogene Proteins c-akt; D010396:Penicillamine; D000109:Acetylcholine; D019331:NG-Nitroarginine Methyl Ester; D000077191:Wortmannin