Brain damage diseases such as cerebral ischemia brain trauma

Brain damage diseases, such as cerebral ischemia, brain trauma and infection are characterized by the increase in oxygen free radicals. Therefore, deeper understanding of the character and mechanism of free radical damage in the brain is important to ultimately relieve diseases and promote recovery (Yang et al., 2008). In the course of ischemia, free radical chemical mediators play a key role. Arachidonic Cyclosporin H is an important class of mediators, which originates from membrane lipids and can generate prostaglandins, thromboxane A2 (cyclooxygenase pathway), and leukotrienes (lipoxygenase pathway) depending on different enzymes. 5-lipoxygenase (5-LOX) is a rate-limiting enzyme in the synthesis of leukotrienes (LTs). The lipoxygenase pathway has been widely studied in recent years because of the important biological actions of leukotrienes. However, the relationship between 5-LOX, neuronal mitochondria injury and ROS has not been fully clarified. Welt et al. (Welt et al., 2000) determined that lipoxygenase inhibitor FLM 5011 was an effective protectant of myocardial microvessels against ischaemia-reperfusion injury, but the change in ultrastructural morphology was still not clear. Quiescent 5-LOX is predominately localized in the cytosol and nucleus. After stimulation by OGD injury, 5-LOX translocate to the nuclear membrane, and then binds with 5-LOX activating protein (FLAP) and cytosolic PLA2 (cPLA2). Once the 5-LOX-cPLA2-FLAP complex forms, arachidonic acid catalyses leukotriene A4 (LTA4) (Funk, 2001). Therefore, 5-LOX’s translocation to the nuclear envelope and connection with FLAP is a critical factor to trigger an inflammatory response. Amazingly, our western blotting and immunohistochemistry results in PC12 cells indicated that 5-LOX can translocate to the nuclear membrane and be activated after stimulation. These findings reveal that the arachidonic acid lipoxygenase pathway is involved in the OGD process of neuronal cells. Caffeic acid, an inhibitor of 5-LOX metabolism, was used to further determine the relationship of the 5-LOX pathway and mitochondria. In our results, caffeic acid significantly decreased the production of arachidonic acid by lipoxygenase metabolism, maintained the mitochondrial ultrastructure integrity of neuronal cells, enhanced mitochondrial membrane potential, and finally decreased ROS and prevented neuronal cells from undergoing apoptosis after OGD/ODG-R. All of our data determined that the instability of mitochondria may be the critical factor of 5-LOX activation. Fitzl et al. (1999) found that lipoxygenase inhibitor FLM 5011 had a protective effect on mitochondrial ultrastructure in ischemic and reperfused cardiomyocytes. Based on the above data, our research verified the evidence and further explained the detailed mechanism on a cellular level.
Materials and methods
Acknowledgements This study was supported by grants from Zhejiang Provincial Natural Science Foundation of China (No. LY17H090018) and Chinese Clinical Pharmaceutical Science development (No. L2011079). We would like to thank Prof. Er-Qing Wei and Dr. Wei-Ping Zhang for advice on experimental design.
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