Hayakawa T, Noda M, Yasuda K, Yorifuji H, Taniguchi S, Miwa I, Sakura H, Terauchi Y, Hayashi J, Sharp G W, Kanazawa Y, Akanuma Y, Yazaki Y, Kadowaki T
Third Department of Internal Medicine, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo 113, Japan.
J Biol Chem. 1998 Aug 7;273(32):20300-7. doi: 10.1074/jbc.273.32.20300.
Recently, a mitochondrial mutation was found to be associated with maternally inherited diabetes mellitus (Kadowaki, T., Kadowaki, H., Mori, Y., Tobe, K., Sakuta, R., Suzuki, Y., Tanabe, Y, Sakura, H., Awata, T., Goto, Y., Hayakawa, T., Matsuoka, K., Kawamori, R., Kamada, T., Horai, S., Nonaka, I., Hagura, R., Akanuma, Y., and Yazaki, Y. (1994) N. Engl. J. Med. 330, 962-968). In order to elucidate its etiology, we have investigated the involvement of mitochondrial function in insulin secretion. Culture of the pancreatic beta-cell line, betaHC9, with low dose ethidium bromide (EB) (0.4 microg/ml) for 2-6 days resulted in a substantial decrease in the transcription level of mitochondrial DNA (to 10-20% of the control cells) without changing its copy number, whereas the transcription of nuclear genes was grossly unaffected. Electron microscopic analysis revealed that treatment by EB caused morphological changes only in mitochondria and not in other organelles such as nuclei, endoplasmic reticula, Golgi bodies, or secretory granules. When the cells were treated with EB for 6 days, glucose (20 mM) could no longer stimulate insulin secretion, while glibenclamide (1 microM) still did. When EB was removed after 3- or 6-day treatment, mitochondrial gene transcription recovered within 2 days, and the profiles of insulin secretion returned to normal within 7 days. Studies with fura-2 indicated that in EB-treated cells, glucose (20 mM) failed to increase intracellular Ca2+, while the effect of glibenclamide (1 microM) was maintained. Our system provides a unique way to investigate the relationship between mitochondrial function and insulin secretion.
最近，人们发现一种线粒体突变与母系遗传的糖尿病有关（门胁敏郎、门胁博、森恭子、户部贵史、坂田良、铃木阳一、田辺洋、桜井秀人、粟田哲、后藤洋、早川智、松冈启、川森良一、镰田哲、堀井史、野中郁次郎、羽仓良、赤沼洋、矢崎义雄，（1994年）《新英格兰医学杂志》第330卷，第962 - 968页）。为了阐明其病因，我们研究了线粒体功能在胰岛素分泌中的作用。用低剂量溴化乙锭（EB）（0.4微克/毫升）培养胰腺β细胞系βHC9 2 - 6天，导致线粒体DNA转录水平大幅下降（降至对照细胞的10 - 20%），而其拷贝数不变，而核基因的转录基本未受影响。电子显微镜分析显示，EB处理仅导致线粒体形态发生变化，而细胞核、内质网、高尔基体或分泌颗粒等其他细胞器未受影响。当细胞用EB处理6天时，葡萄糖（20毫摩尔）不再能刺激胰岛素分泌，而格列本脲（1微摩尔）仍能刺激。在3天或6天处理后去除EB，线粒体基因转录在2天内恢复，胰岛素分泌模式在7天内恢复正常。用fura - 2进行的研究表明，在EB处理的细胞中，葡萄糖（20毫摩尔）未能增加细胞内钙离子浓度，而格列本脲（1微摩尔）的作用得以维持。我们的系统为研究线粒体功能与胰岛素分泌之间的关系提供了一种独特的方法。
