Combination of Reactive Oxygen Species and Tissue-Type Plasminogen Activator Enhances the Induction of Gelatinase B in Brain Endothelial Cells

ABSTRACT

Tissue-type plasminogen activator (t-PA) increases the risk of intracranial hemorrhage by gelatinase B (matrix metalloprotease-9; MMP-9) production in brain endothelial cells. It was recently reported that the free-radical scavenger edaravone significantly decreases t-PA-mediated MMP-9 production. Therefore, by using cultured brain endothelial cells (b.End3), we investigated whether t-PA-mediated MMP-9 production was enhanced by reactive oxygen species (ROS) and its signaling pathways. Moreover, we also investigated that whether this combined enhancement is reduced by edaravone. The b.End3 cells were exposed to t-PA (10 μg/mL), followed by H₂O₂ (30 μM); further, the MMP-9 protein level was measured. ROS enhanced MMP-9 production, and ROS plus t-PA significantly increased MMP-9 production more than t-PA or ROS alone. The results showed that H₂O₂ or t-PA alone caused a significant increase in NF-κB translocation to the nucleus, whereas the combination of t-PA and H₂O₂ increased the translocation of NF-κB to an even greater extent. Moreover, the combination of t-PA and ROS significantly increased I-κB degradation as well as NF-κB expression. Edaravone completely decreased the ROS plus t-PA-mediated MMP-9 enhancement. In conclusion, ROS enhanced t-PA-mediated MMP-9 production in brain endothelial cells; this MMP-9 production was decreased by the addition of edaravone, which inhibited the NF-κB pathway, specifically by enhancing I-κB degradation.

KEYWORDS::

• edaravone
• endothelial cells
• matrix metalloprotease-9
• reactive oxygen species
• tissue-type plasminogen activator