Song Ji Yeon, Lim Joo Weon, Kim Hyeyoung, Kim Kyung Hwan
Department of Pharmacology and Institute of Gastroenterology, Brain Korea 21 Project for Medical Science, Yonsei University College of Medicine, Seoul 120-752, Korea.
Ann N Y Acad Sci. 2003 Dec;1010:259-63. doi: 10.1196/annals.1299.044.
Reactive oxygen species have been known to cause DNA damage and induce apoptosis. During DNA damage, DNA repair proteins Ku70 and Ku80 prevent cell death, but severe DNA damage beyond the repair capacity of the DNA repair proteins triggers necrosis or apoptosis. Recent reports have shown that NF-kappaB plays a critical role in protecting the cells from apoptosis. We investigated whether glucose oxidase acting on beta-D-glucose (G/GO), which continuously produces H(2)O(2), induces apoptosis, and whether NF-kappaB and Ku are involved in G/GO-induced apoptosis in pancreatic acinar AR42J cells. Electron microscopic observation showed that apoptotic cells with characteristic nuclear condensation and shrinkage as well as large vacuoles were detected after G/GO treatment. G/GO treatment induced apoptotic cell death, as determined by viable cell count and DNA fragmentation. G/GO-induced apoptosis was increased in the cells transfected with the Ku-dominant negative mutant (Ku D/N) and mutated IkappaBalpha gene (IkappaB mt) as compared to the wild-type cells (Wild) and the cells transfected with the control pcDNA3 vector (pcN-3). G/GO treatment caused nuclear loss of both Ku70 and Ku80 in Wild cells and pcN-3 cells. Even without G/GO treatment, nuclear loss of Ku proteins was observed in IkappaB mt cells. These results suggest that oxidative stress-induced reduction of nuclear Ku proteins may cause loss of defense against DNA damage and thus induce apoptosis in pancreatic acinar cells. The novel finding is that nuclear translocation of Ku proteins may be mediated by NF-kappaB.
已知活性氧会导致DNA损伤并诱导细胞凋亡。在DNA损伤过程中，DNA修复蛋白Ku70和Ku80可防止细胞死亡，但超出DNA修复蛋白修复能力的严重DNA损伤会引发坏死或凋亡。最近的报告表明，核因子κB在保护细胞免受凋亡方面起着关键作用。我们研究了作用于β-D-葡萄糖（G/GO）的葡萄糖氧化酶持续产生H₂O₂是否会诱导细胞凋亡，以及核因子κB和Ku是否参与G/GO诱导的胰腺腺泡AR42J细胞凋亡。电子显微镜观察显示，G/GO处理后检测到具有特征性核浓缩、核收缩以及大液泡的凋亡细胞。通过活细胞计数和DNA片段化检测确定，G/GO处理诱导了凋亡性细胞死亡。与野生型细胞（Wild）和转染对照pcDNA3载体（pcN-3）的细胞相比，用Ku显性负突变体（Ku D/N）和突变的IkappaBα基因（IkappaB mt）转染的细胞中，G/GO诱导的凋亡增加。G/GO处理导致Wild细胞和pcN-3细胞中Ku70和Ku80的核丢失。即使没有G/GO处理，在IkappaB mt细胞中也观察到Ku蛋白的核丢失。这些结果表明，氧化应激诱导的核Ku蛋白减少可能导致对DNA损伤的防御丧失，从而诱导胰腺腺泡细胞凋亡。新发现是Ku蛋白的核转位可能由核因子κB介导。
