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Studies on gap-junctional intercellular communication as a mechanism regulating cell death in the cell society

间隙连接细胞间通讯作为细胞社会中细胞死亡调节机制的研究

基本信息

批准号：
15390129
负责人：
OYAMADA Masahito
金额：
$ 7.04万
依托单位：
Kyoto Prefectural University of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15390129/
关键词：
gap functions connexin cell death apoptosis necrosis ultraviolet light mitochondria GFP caspase

项目摘要

Gap junctions are considered to play an important role in moderating cell death including apoptosis.However, the basic phenomena underlying when and where the alterations of gap junctions occur during apoptosis have not been well documented. In this study, we examined the fate of Cx43 during apoptosis in order to answer the following questions : (1)Does Cx43 localization change during apoptosis? (2)If so, when? (3)Does alteration of Cx43 depend on caspase activation? To answer these questions, we analyzed the spatiotemporal changes of connexin(Cx) during UV light-induced apoptosis using Cx43-enhanced green fluorescent protein(EGFP)-expressing HeLa cells, and compared them with those of mitochondrial membrane potential(MMP) using tetramethylrhodamine ethyl ester(TMRE) and nuclear morphological observation using Hoechst 33342. At 2 hr post-UV-irradiation, a third of the cells became TMRE-negative, i.e., they showed the loss of MMP, but with slight nuclear fragmentation, and high percentages of linear Cx43-EGFP plaques were found among both TMRE-positive and TMRE-negative cells. At 4 hr post-UV-irradiation, the percentage of these linear plaques was decreased, and both punctate and diffuse localization of Cx43-EGFP were noted in the cytoplasm of TMRE-negative cells without nuclear fragmentation. At 8 hr post-irradiation, punctate cytoplasmic localization of Cx43-EGFP was noted in TMRE-negative cells with nuclear fragmentation.Treatment with the caspase inhibitor Z-VAD-FMK blocked nuclear fragmentation and partially preserved both gap junctional plaques and MMP. These results indicate that, during apoptosis, Cx mobilization into the cytoplasm occurs after MMP depolarization but before nuclear fragmentation and that this alteration partly depends on caspase.

间隙连接被认为在调节包括细胞凋亡在内的细胞死亡中发挥重要作用。然而，细胞凋亡过程中间隙连接改变何时何地发生的基本现象尚未得到充分记录。在本研究中，我们检测了细胞凋亡过程中Cx43的命运，以回答以下问题：（1）细胞凋亡过程中Cx43定位是否发生变化？ (2)如果是的话，什么时候？ (3)Cx43的改变是否依赖于caspase的激活？为了回答这些问题，我们利用表达 Cx43 增强型绿色荧光蛋白（EGFP）的 HeLa 细胞，分析了紫外线诱导细胞凋亡过程中连接蛋白（Cx）的时空变化，并与四甲基罗丹明乙酯（TMRE）的线粒体膜电位（MMP）和 Hoechst 的核形态观察进行比较。 33342. 在 UV 照射后 2 小时，三分之一的细胞变为 TMRE 阴性，即它们显示出 MMP 丢失，但有轻微的核碎片，并且在 TMRE 阳性和 TMRE 阴性细胞中发现了高百分比的线性 Cx43-EGFP 斑块。 UV 照射后 4 小时，这些线性斑块的百分比减少，并且在 TMRE 阴性细胞的细胞质中观察到 Cx43-EGFP 的点状和弥漫性定位，而没有核碎片。照射后 8 小时，在 TMRE 阴性细胞中观察到 Cx43-EGFP 的点状细胞质定位，并伴有核碎片。用 caspase 抑制剂 Z-VAD-FMK 处理可阻断核碎片，并部分保留间隙连接斑块和 MMP。这些结果表明，在细胞凋亡过程中，Cx 动员到细胞质中发生在 MMP 去极化之后但在核碎裂之前，并且这种改变部分取决于 caspase。

项目成果

期刊论文数量（43）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Hirakawa, H.et al.: "Regional differences in blood-nerve barrier function and tight-junction protein expression within the rat dorsal root ganglion"NeuroReport. 15. 405-408 (2004)

Hirakawa, H.等人：“大鼠背根神经节内血神经屏障功能和紧密连接蛋白表达的区域差异”NeuroReport。

DOI：
发表时间：
期刊：
影响因子：
0
作者：
通讯作者：

Stimulation of living cells by femtosecond near-infrared laser pulses

DOI：
10.1117/12.478591
发表时间：
2003-06
期刊：
影响因子：
0
作者：
S. Iwanaga;N. Smith;K. Fujita;T. Kaneko;M. Oyamada;T. Takamatsu;S. Kawata;O. Nakamura
通讯作者：
S. Iwanaga;N. Smith;K. Fujita;T. Kaneko;M. Oyamada;T. Takamatsu;S. Kawata;O. Nakamura

Loss and recovery of the blood-nerve barrier in the rat sciatic nerve after crush injury are associated with expression of intercellular junctional proteins