Hypoxia confers protection against apoptosis via the PI3K/Akt pathway in endothelial progenitor cells.
OBJECTIVE
The recruitment of endothelial progenitor cells (EPC) to ischemia has recently been suggested as an important mechanism of tissue repair. Although tissue ischemia can facilitate EPC mobilization, recruitment, and retention at the hypoxic site, the effects of hypoxia on EPC survival are not well known. In the present study, we examined whether hypoxia (2% O2) would suppress apoptosis induced by serum withdrawal and whether survival signals, such as the phosphatidylinositol 3-kinase (PI3K)/Akt and extracellular signal-regulated protein kinase (ERK) pathways, were involved in this process.


METHODS
After being serum-starved for 24 h, EPC were cultured under normoxic or hypoxic conditions (2% O2) for 24 h. Cell survival was assessed by 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, annexin V-propidium iodide dual-color flow cytometry, and terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP nick-end labeling assay. The expressions of signaling proteins were evaluated by Western blot analysis.


RESULTS
Under hypoxic conditions, EPC were resistant to apoptosis induced by serum starvation. The inhibition of the PI3K/Akt pathway using the LY294002 inhibitor prevented hypoxia-inhibited apoptosis in EPC and altered the phosphorylation state of glycogen synthase kinase-3beta, an effector protein involved in regulation of EPC apoptosis. However, ERK inhibitor PD98059 had no significant effect on cell survival.


CONCLUSIONS
Our data demonstrated that hypoxia inhibited serum withdrawal-induced apoptosis in EPC, which might be associated with the activation of the PI3K/Akt pathway.
Dai
Tao
T
Department of Cardiology, Biomedical Research (Therapy) Center, Sir Run Run Shaw Hospital, College of Medicine, Zhejiang University, Hangzhou 310016, China.
Zheng
Hao
H
Fu
Guo-sheng
GS
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002478:Cells, Cultured; D002867:Chromones / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology / Q000502:physiology*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005419:Flavonoids / Q000378:metabolism; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D000860:Hypoxia / Q000378:metabolism*; D009025:Morpholines / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D051381:Rats; D015398:Signal Transduction / Q000502:physiology*; D013234:Stem Cells / Q000166:cytology / Q000502:physiology*
D002867:Chromones; D004791:Enzyme Inhibitors; D005419:Flavonoids; D009025:Morpholines; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D019869:Phosphatidylinositol 3-Kinases; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605508:Gsk3b protein, rat; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases; D038362:Glycogen Synthase Kinase 3; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one