Role of FAK phosphorylation in hypoxia-induced hMSCS migration: involvement of VEGF as well as MAPKS and eNOS pathways.
Here we show that the effect of hypoxia on human umbilical cord blood mesenchymal stem cell (hMSC) migration is via the modulation of focal adhesion kinase (FAK) and its related signaling pathways. Hypoxia increased hMSC migration and cell viability, whereas lactate dehydrogenase (LDH) release was not affected for up to 48 h (data not shown). In addition, hypoxia increased the level of reactive oxygen species (ROS) generation in a time-dependent manner. Hypoxia-induced phosphorylation of p38 mitogen-activated protein kinase (MAPK) and stress-activated protein kinase/c-Jun NH(2)-terminal kinase (SAPK/JNK) were inhibited by the antioxidant (N-acetylcysteine, NAC, 10(-6) M) and (taurine, 4x10(-6) M). Hypoxia-induced endothelial nitric oxide synthase (eNOS) phosphorylation was regulated by p38 MAPK and SAPK/JNK activation. In addition, hypoxia increased the level of hypoxia inducible factor (HIF)-1alpha expression, which was blocked by inhibition of eNOS. Also, hypoxia-induced expression of Flk-1, vascular endothelial growth factor (VEGF), and its secreted form were inhibited by HIF-1alpha small interfering RNA (siRNA). In this hypoxic condition, FAK and Src phosphorylation were increased in a time-dependent manner. Inhibition of Src with specific inhibitor (PP2, 10(-8) M) blocked hypoxia-induced FAK activation. Subsequently, hypoxia-induced FAK phosphorylation was blocked by VEGF siRNA. Finally, hypoxia-induced increase of hMSC migration was inhibited by FAK siRNA. The results indicate that hypoxia increases migration of hMSCs via VEGF-mediated FAK phospholylation and involves the cooperative activity of the ROS, MAPK, eNOS and HIF-1alpha pathways.
Lee
Sang Hun
SH
Dept. of Veterinary Physiology, Biotherapy Human Resources Center, College of Veterinary Medicine, Chonnam National Univ., Gwangju 500-757, Korea.
Lee
Yu Jin
YJ
Song
Chang Hun
CH
Ahn
Young Keun
YK
Han
Ho Jae
HJ
D002465:Cell Movement / Q000502:physiology*; D002478:Cells, Cultured; D005312:Fetal Blood / Q000166:cytology; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000378:metabolism*; D006801:Humans; D000860:Hypoxia / Q000378:metabolism*; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000502:physiology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D010766:Phosphorylation; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*; D019061:src-Family Kinases / Q000378:metabolism
D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D017382:Reactive Oxygen Species; D042461:Vascular Endothelial Growth Factor A; D052250:Nitric Oxide Synthase Type III; D051416:Focal Adhesion Protein-Tyrosine Kinases; D019061:src-Family Kinases; D020928:Mitogen-Activated Protein Kinases