A resealed-cell system for analyzing pathogenic intracellular events: perturbation of endocytic pathways under diabetic conditions.
A resealed-cell system for analyzing pathogenic intracellular events: perturbation of endocytic pathways under diabetic conditions.
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DOI：
10.1371/journal.pone.0044127
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Murata M
中科院分区：
文献类型：
--
作者：
Kano F;Nakatsu D;Noguchi Y;Yamamoto A;Murata M
基于细胞的检测系统可以作为致病器官中异常功能的细胞模型，这将是筛选药物和阐明各种疾病的分子机制的新颖和有用的工具。我们构建了模型细胞，复制糖尿病肝细胞的条件下，通过使用细胞再密封技术，使细胞溶质的交换。用链球菌毒素链球菌溶血素O使HeLa细胞的质膜透化，并将从野生型或db/db糖尿病小鼠制备的胞质溶胶引入所得的半完整细胞中。通过暴露于Ca 2+重新密封质膜，我们创建了WT或Db模型细胞，其中细胞质条件复制了健康或糖尿病肝脏的条件。有趣的是，在Db细胞中，p38 MAPK的磷酸化被促进，而内体磷脂酰肌醇-3-磷酸的水平降低。我们研究了WT和Db细胞中的几种内吞途径，发现Db细胞中逆行内体至高尔基体的转运以p38 MAPK依赖的方式延迟。此外，在Db细胞中，EGF受体从内体到溶酶体的降解途径增强，并且这不依赖于p38 MAPK的激活。疾病模型细胞系统应该成为检测致病条件下细胞中异常过程和用于治疗应用的有力工具。
Cell-based assay systems that can serve as cellular models of aberrant function in pathogenic organs would be novel and useful tools for screening drugs and clarifying the molecular mechanisms of various diseases. We constructed model cells that replicated the conditions in diabetic hepatocytes by using the cell resealing technique, which enables the exchange of cytosol. The plasma membrane of HeLa cells was permeabilized with the streptococcal toxin streptolysin O, and cytosol that had been prepared from wild-type or db/db diabetic mice was introduced into the resulting semi-intact cells. By resealing the plasma membrane by exposure to Ca2+, we created WT or Db model cells, in which the cytosolic conditions replicated those of healthy or diabetic liver. Interestingly, phosphorylation of p38 MAPK was promoted, whereas the level of endosomal phosphatidylinositol-3-phosphate was decreased, in Db cells. We investigated several endocytic pathways in WT and Db cells, and found that retrograde endosome-to-Golgi transport was delayed in a p38 MAPK-dependent manner in Db cells. Furthermore, the degradation pathway of the EGF receptor from endosomes to lysosomes was enhanced in Db cells, and this did not depend on the activation of p38 MAPK. The disease model cell system should become a powerful tool for the detection of aberrant processes in cells under pathogenic conditions and for therapeutic applications.
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影响因子：
7.7
作者：
Hemi R;Yochananov Y;Barhod E;Kasher-Meron M;Karasik A;Tirosh A;Kanety H
通讯作者：
Kanety H
DOI：
10.1083/jcb.200708010
发表时间：
2008-03-10
期刊：
The Journal of cell biology
影响因子：
--
作者：
Idone V;Tam C;Goss JW;Toomre D;Pypaert M;Andrews NW
通讯作者：
Andrews NW
影响因子：
64.8
作者：
Itzhaki, Ilanit;Maizels, Leonid;Gepstein, Lior
通讯作者：
Gepstein, Lior
影响因子：
4
作者：
Kano, Fumi;Yamauchi, Shinobu;Murata, Masayuki
通讯作者：
Murata, Masayuki
影响因子：
20.3
作者：
Cho, Hyun-Jai;Lee, Choon-Soo;Kim, Hyo-Soo
通讯作者：
Kim, Hyo-Soo