Regulation of endothelial matrix metalloproteinase-2 by hypoxia/reoxygenation.
Among the consequences resulting from the exposure of endothelial cells (ECs) to ischemia/reperfusion is angiogenesis, involving degradation of vascular basement membrane and extracellular matrix. Matrix metalloproteinase (MMP)-2, a member of the MMP family, partakes in this process. MMP-2, secreted as a proenzyme, undergoes activation through interaction with membrane type (MT)1-MMP and the endogenous tissue inhibitor of MMPs (TIMP)-2. Although hypoxia and reoxygenation (H/R) are major constituents of ischemia/reperfusion processes, their direct effects on endothelial MMP-2 have been scarcely investigated. This study examined the in vitro effects of H/R on human macrovascular ECs (EAhy 926). The level of MMP-2 mRNA (Northern blot) and protein (zymography, ELISA) and the mRNA of its activator (MT1-MMP) and inhibitor (TIMP-2) were analyzed. Short (6-hour) hypoxia inhibited the mRNA expression of MMP-2, MT1-MMP, and TIMP-2, culminating in reduced latent and active MMP-2 protein. Prolonged (24-hour) hypoxia further suppressed MT1-MMP and TIMP-2 mRNA, whereas it enhanced MMP-2 mRNA and enzyme secretion (after 48-hour hypoxia). Reoxygenation did not influence the inhibited TIMP-2 but upregulated MMP-2 and MT1-MMP mRNA expression, leading to enhanced secretion of active MMP-2 protein. These results demonstrate H/R-mediated modulation of EC MMP-2 at both transcriptional and posttranscriptional levels. Prolonged hypoxia of ECs appears to enhance MMP-2 production and secretion, whereas reoxygenation further increases its level. These H/R-mediated effects on MMPs have the potential of enabling EC migration and possible angiogenesis.
Ben-Yosef
Yaara
Y
Neuroimmunology Unit, Carmel Medical Center, Haifa, Israel.
Lahat
Nitza
N
Shapiro
Sarah
S
Bitterman
Haim
H
Miller
Ariel
A
D015152:Blotting, Northern; D015687:Cell Hypoxia / Q000502:physiology*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D004789:Enzyme Activation / Q000502:physiology; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D020778:Matrix Metalloproteinase 2 / Q000235:genetics / Q000378:metabolism*; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases / Q000235:genetics / Q000378:metabolism; D008833:Microcirculation / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism / Q000494:pharmacology*; D020871:RNA Stability / Q000187:drug effects / Q000502:physiology; D012333:RNA, Messenger / Q000378:metabolism; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects
D012333:RNA, Messenger; D019716:Tissue Inhibitor of Metalloproteinase-2; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases; D020778:Matrix Metalloproteinase 2; D010100:Oxygen