Generation of transgenic mice with arrythmia expressing a dominant-negative Isk

产生表达显性失活 Isk 的心律失常转基因小鼠

• 批准号: 10557004
• 负责人: TAKUMI Toru
• 金额: $ 5.76万
• 依托单位: Kobe University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10557004/
• 关键词: Arrythmia; Myocyte; Transgenic mice; K channel

To understand physiological function of the Isk channel in heart, we investigated behavior of the Isk channels in three different ways ; in mammalian cultured cells, in isolated ventricular myocytes and in the heart of transgenic mice.(1)In COS7 cells transfected, Isk was associated with KvLQT1 and HERG. The rapidly activating potassium current was abolished by cotransfection of D77N dominant negative Isk and HERG cDNA.(2)In isolated ventricular myocytes, two types of outward KィイD2+ィエD2 currents, the rapidly activating potassium channel (which corresponds to IKr) and the slowly activating potassium channels (which corresponds to IKs), are identified. The outward KィイD2+ィエD2 currents were inhibited by the antisense Isk oligonucleotide or D77N dominant negative Isk mutant. The action potential duration in ventricular myocyte was significantly prolonged by introduction of D77N dominant negative mutant.(3)Transgenic mice expressing a dominant-negative D77N Isk in heart were generated under the control of α-myosin heavy chain promoter. The introduced Isk was strongly expressed in the heart. The electrocardiogram of the mice and electrophysiology using the isolated ventricular myocytes from the mice have been analyzed.

为了了解Isk通道在心脏中的生理功能，我们以三种不同的方式研究了Isk通道的行为;在哺乳动物培养细胞中，在分离的心室肌细胞中和在转基因小鼠的心脏中。(1)In转染COS 7细胞后，Isk与KvLQT 1和HERG相关。D 77 N显性负性Isk和HERG cDNA共转染可阻断快速激活性钾电流。(2)In在分离的心室肌细胞中，鉴定了两种类型的外向钾通道D2+ D2电流，即快速激活钾通道（对应于IKr）和缓慢激活钾通道（对应于IKs）。Isk反义寡核苷酸或D 77 N显性失活Isk突变体均能抑制外向K + D2+ D2电流。引入D 77 N显性负突变体后，心室肌细胞动作电位时程显著延长。（3）在α-肌球蛋白重链启动子的调控下，构建了心脏显性失活的D 77 N Isk转基因小鼠。被介绍的伊斯克在心里强烈地表达出来。对小鼠的心电图和离体心室肌细胞的电生理进行了分析。

项目成果

V.Prevot, S.Bouret, D.Croix, T.Takumi, L.Jennes, V.Mitchell and J.-C.Beauvillain: "Evidence that members of the TGF-βsuperfamily play a key role in the regulation of the GnRH neuroendocrine axis : expression of a type I serine-threonine kinase receptor fo

V.Prevot、S.Bouret、D.Croix、T.Takumi、L.Jennes、V.Mitchell 和 J.-C.Beauvillain：“证据表明 TGF-β 超家族成员在 GnRH 调节中发挥关键作用神经内分泌轴：I 型丝氨酸-苏氨酸激酶受体的表达

Takumi, T., et al.: "A light-independent oscillatory gene mPer3 in mouse SCN and OVLT"The EMBO Journal. 17. 4753-4759 (1998)

Takumi, T. 等人：“小鼠 SCN 和 OVLT 中的光独立振荡基因 mPer3”EMBO 杂志。

Y.Maebayashi, Y.Shigeyoshi, T.Takumi and H.Okamura: "A putative transcription factor with seven zinc-finger motif identified in the developing suprachiasmatic nucleus by the differential display PCR method"J. Neurosci.. 19. 10176-10183 (1999)

Y.Maebayashi、Y.Shigeyoshi、T.Takumi 和 H.Okamura：“通过差异显示 PCR 方法在发育中的视交叉上核中鉴定出具有七个锌指基序的假定转录因子”J.

Takumi,T.et al.: "A light independent oscillatory gene in Per3 in mouse SCN and OVLT" The EMBO Journal. 17. 4753-4759 (1998)

Takumi,T.et al.：“小鼠 SCN 和 OVLT 中 Per3 中的光独立振荡基因”EMBO 杂志。

Maebayashi, Y., et al.: "A putative transcription factor with seven zinc-finger motifs identified in the developing suprachiasmetic nucleus by the differential display PCR method"The Journal of Neuroscience. 19. 10176-10183 (1999)

Maebayashi, Y. 等人：“通过差异显示 PCR 方法在发育中的视交叉上核中鉴定出具有七个锌指基序的推定转录因子”《神经科学杂志》。