Functional significance of nuclear diacylglycerol kinase in neurons and cardioocytes under ischemic stress

二酰基甘油激酶（DGK）是一种磷酸化第二信使二酰基甘油（DG）的酶，参与多种病理生理细胞反应。我们之前报道过DGKζ，其特征是核定位信号（NLS），在短暂的前脑缺血20分钟后从核转移到海马金字塔神经元的细胞质。然而，这一现象的具体机制及其功能意义尚不清楚。首先，我们使用暴露于氧和葡萄糖剥夺（OGD）的海马切片来模拟脑缺血模型，并评估DGKζ易位的时间过程。在动物缺血模型中，dgk - ζ免疫反应性在OGD 20 min后逐渐从细胞核转移到细胞质中，再灌注后细胞核中未检测到dgk - ζ免疫反应性。此外，我们发现，虽然DGKζ在OGD 10 min时在细胞核中检测到，但随后的60 min再灌注诱导了DGKζ从细胞核到细胞质的更完整的易位。这些结果表明，10分钟的OGD打开信号级联，导致DGKζ向细胞质易位。其次，我们的目的是观察DGKζ的亚细胞定位是否会在心肌细胞缺血损伤后发生变化，心肌细胞也是有丝分裂后的高度分化细胞，出生后成为神经元。首先采用左前降支夹持法建立大鼠10 min短暂性缺血心脏模型。在非缺血间隔区，心肌细胞核中观察到dgk - ζ免疫反应性与正常情况相似。然而，我们发现缺血区心肌细胞核的免疫反应性几乎消失。在缺血区边缘区，dgk - ζ免疫反应主要出现在细胞核中，细胞质中较弱。为了研究引起DGKζ亚细胞定位变化的缺血严重程度，我们建立了3分钟、5分钟和20分钟缺血模型。令人惊讶的是，3分钟的缺血足以导致DGKζ从心肌细胞的细胞核中消失。在5分钟和20分钟缺血模型中也得到了类似的结果。总的来说，在心脏缺血条件下，DGKζ迅速从缺血心肌细胞的细胞核中消失至少20分钟。少

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 … More 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