| Budget Amount *help |
¥4,020,000 (Direct Cost: ¥3,600,000、Indirect Cost: ¥420,000)
Fiscal Year 2007: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
Fiscal Year 2006: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
Diacylglycerol kinase (DGK) is an enzyme that phosphorylates a second messenger diacylglycerol (DG) and is involved in a variety of pathophysiological cellular responses. We have previously reported that DGKζ, characterized by a nuclear localization signal (NLS), is translocated from the nucleus to the cytoplasm in hippocampal pyramidal neurons after 20 min of transient forebrain ischemia. However, the detailed mechanism of this phenomenon and it's functional significance remains unknown.
First, we used hippocampal slices exposed to oxygen and glucose deprivation (OGD) to simulate an ischemic model of the brain and evaluated the time course of the translocation of DGKζ. As seen in the animal ischemic model, DGKζ-immunoreactivity was gradually shifted from the nucleus to the cytoplasm after 20 min of OGD and was never detected in the nucleus after reperfusion. In addition, we found that while DGKζ was detected in the nucleus by 10 min of OGD, the following 60 min of reperfusion induced c
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omplete translocation of DGKζ from the nucleus to the cytoplasm. These results suggest that 10 min of OGD switches on the signal cascade to cause translocation of DGKζ to the cytoplasm.
Second, we aimed to see whether the subcellular localization of DGKζ would change after the ischemic insult in cardiomyocytes, which are also postmitotic, highly differentiated cells after birth as neurons. We first generated the 10 min-transient ischemic heart model of rat by clipping of left anterior descending branch. In the non-ischemic septal region, DGKζ-immunoreactivity was observed in the nucleus of cardiomyocytes similarly to normal conditions. However, we found that the immunoreactivity almost disappeared from the nucleus of cardiomyocytes in the ischemic region. In the border area of the ischemic region, DGKζ-immunoreactivity was observed mostly in the nucleus and faintly in the cytoplasm. To examine the extent of the severity of ischemia to cause this change in subcellular localization of DGKζ, we generated models of 3 min-, 5 min-, and 20 min-ischemia. Surprisingly, 3 min of ischemia was shown to be enough to cause DGKζ to disappear from the nucleus in cardiomyocytes. Similar results were obtained from the 5 min- and 20 min-ischemic models. Collectively, under ischemic conditions of heart DGKζ quickly disappeared from the nucleus of ischemic cardiomyocytes at least up to 20 min. Less
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