Investigation of cardiac cell-to-cell interaction using on-tip single-cell-based cell cultivation and analysis method

使用基于单细胞的尖端细胞培养和分析方法研究心脏细胞间相互作用

• 批准号: 17590749
• 负责人: YANO Katsusuke
• 金额: $ 1.86万
• 依托单位: Nagasaki University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2005
• 资助国家: 日本
• 起止时间: 2005 至 2006
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-17590749/
• 关键词: Cardiomyocyte; Synchronization; Connexin 43; Single-cell-based culture assaysystem; Beating; Isoprotelenol; オンチップ; 心筋ネットワーク; 心筋拍動; 細胞間相互作用

We developed a new method that can analyze single-cell cardiomyocytes, such as contraction, synchronization. We constructed 30 um-diameter microchambers and narrow microtunnels by melting of thin agar layer using 1,480 nm the infrared focused laser beam. Next, we put a cardiomyocytes into each microchamber by glass micropipette. Cardiomyocytes in microchamber attached to the dish and began to contract spontaneously. Then, cardiomyocytes connected to neighboring cells through the microtunnels, and began to contract synchronously. Connexin43 expression was confirmed in the cell-connection sites by immunostaining just after myocyte-to-myocyte connection. Beating rate of cardiomyocytes was increased by the addition of isoproterenol dose-dependently (10^9M 〜 10^6 M) and returned to control level rapidly after wash out of isoproterenol. This system enables us to observe the interaction of cardiomyocytes.

我们开发了一种新的方法，可以分析单细胞心肌细胞，如收缩，同步化。我们使用1,480 nm的红外聚焦激光束通过熔化薄琼脂层构建了直径为30 μ m的微腔和狭窄的微通道。然后，我们用玻璃微吸管将心肌细胞放入每个微室中。微室中的心肌细胞贴壁并开始自发收缩。然后，心肌细胞通过微通道连接到相邻的细胞，并开始同步收缩。连接蛋白43的表达在肌细胞连接后通过免疫染色证实。加入异丙肾上腺素后，心肌细胞的搏动率呈剂量依赖性增加（10^9 M × 10^6 M），并在洗脱异丙肾上腺素后迅速恢复到对照水平。该系统使我们能够观察心肌细胞的相互作用。