Superoxide induced by a high-glucose concentration attenuates production of angiogenic growth factors in hypoxic mouse mesenchymal stem cells.
Previous reports have shown that the paracrine system may be an important mediator in bone-marrow-derived mesenchymal stem cell (MSC) therapy for ischemic diseases. Hyperglycemia and hypoxia have been associated with increased levels of reactive oxygen species; oxidative stress may therefore influence the paracrine effects of MSCs under hypoxic conditions in diabetic patients, although the mechanism underlying this effect remains unknown. Hypoxia-inducible factor 1α (HIF-1α) regulates the transcription of hypoxia-inducible genes. We determined the effect of high-glucose concentrations on the production of angiogenic growth factors via HIF-1α induction in hypoxic MSCs. MSCs were cultured with different glucose concentration (5.6, 11, 20, or 30 mM) for 24 h. The cells were then incubated in a hypoxic chamber (5% O₂) or under normoxia (21% O₂) for 6 or 24 h. Protein levels of HIF-1α, vascular endothelial growth factor A₁₆₅ (VEGF-A₁₆₅), and platelet-derived growth factor B (PDGF-B) were attenuated by glucose in hypoxic MSCs in a dose-dependent manner. Treatment with MG132, a specific inhibitor of proteasome activity, significantly reversed the inhibitory effect of high-glucose concentrations in hypoxic MSCs. 4-Hydroxyl-tetramethylpiperidin-oxyl (a cell-permeable superoxide scavenger) or Apocynin (a NADPH oxidase inhibitor) significantly reversed glucose-induced attenuation of VEGF-A₁₆₅, PDGF-B, and HIF-1α protein levels. Stimulation with a high-glucose concentration (30 mM) significantly increased intracellular superoxide levels in hypoxic MSCs. Our results suggest that in hypoxic MSCs the increase in intracellular superoxide levels induced by high-glucose concentrations may attenuate hypoxia-induced HIF-1α expression, which in turn attenuates hypoxia-induced VEGF-A₁₆₅ and PDGF-B transcription.
Department of Pharmacology, National Defense Medical College, 3-2, Namiki, Tokorozawa, Saitama 359-8513, Japan. tishizu@ndmc.ac.jp
Toshiaki
T
Ishizuka
Takashi
T
Hinata
Yasuhiro
Y
Watanabe
D000818:Animals; D001853:Bone Marrow / Q000473:pathology; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D005947:Glucose / Q000008:administration & dosage*; D000860:Hypoxia / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology*; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis*; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D008297:Male; D059630:Mesenchymal Stem Cells / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D018919:Neovascularization, Physiologic / Q000187:drug effects*; D009994:Osmolar Concentration; D018384:Oxidative Stress; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects; D020574:Proto-Oncogene Proteins c-sis / Q000235:genetics; D012720:Severity of Illness Index; D013481:Superoxides / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism
C497443:Hif1a protein, mouse; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D036341:Intercellular Signaling Peptides and Proteins; D020574:Proto-Oncogene Proteins c-sis; D042461:Vascular Endothelial Growth Factor A; C467480:vascular endothelial growth factor A, mouse; D013481:Superoxides; D046988:Proteasome Endopeptidase Complex; D005947:Glucose