| Co-Investigator(Kenkyū-buntansha) |
II Kazuo IWAKI GRASS LIFE SCIIENCE CENTER DIRECTOR, ライフサイエンスセンタ, 所長
KIMURA Ichiro WASEDA UNIV.DEPT OF BASIC HUMAN SCIENCES PROFESSOR, 人間科学部, 教授 (10011595)
MATSUDA Ryoichi UNIV OF TOKYO DEPT OF LIFE SCI GRAD SCH ARTS & SCI ASSOCIATE PROFESSOR, 大学院・総合文化, 助教授 (90165837)
SATO Naruki CHIBA UNIV.DEPT OF BIOLOGY,RESEARCH ASSOCIATE, 理学部, 助手 (40261896)
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| Research Abstract |
1.With an aim to clarify the effects of mechanical stress on cells in an in vitro system, we manufactured a culture system which permits unidirectional stretch of a cell culture substrate repeatedly over long periods. Thin elastic silicon membrane with fine parallel grooves as a substrate for cell culture and an appropriate culture medium were also devised.
2.Since muscle cells are formed and maintained under a mechanically active environment, the effects of mechanical stress on cultured cells were examined with muscle cells as a model and observed two remarkable phenomena caused by stretching : 1) stretch-dependent cell arrangement and 2) hypertrophy of the muscle cells as follows :
1) When rat aortic smooth muscle cells (A10) and mouse C2 myoblasts were cultured under a standard condition and then the substrate was stretched repetitively at a frequency of 1 Hz with an amplitude of 20%, the cells became oriented at an angle of about 70o to the direction of stretching. Actin filaments al
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so oriented in the same direction, but filament orientation preceded cell orientation. We found that this stretch-dependent cell orientation was completely blocked by 100 mM gadolinium, an inhibitor of a stretch-activated ion channel, and cycloheximide, but filament reorientation was not affected by these agents at all. From these results, it is concluded that orientation of filaments and the orientation of the cells are closely related but regulated differently.
2) In the case of muscle cells (myotubes), stretching in a longitudinal cell axis is important from a physiological view point. Since rearrangement of cell orientation did not occur during stretching of myotubes aligned along the grooves, it was possible to stretch myotubes for more than a week in the direction of the longitudinal cell axis. We observed that myotube diameters increased significantly as compared with the control myotubes cultured without stretch. We further found that a protein of about 67 kDa accumulated in the stretched myotubes. We identified this protein with albumin based on antigenicity, size and pI.The albumin was the same as that in the culture medium and in addition, biotinized albumin added to the medium was also detected in the stretched myotubes. Therefor, we conclude that albumin in the medium was incorporated into myotubes in a stretching-dependent manner. Physiological implication of stretch-dependent albumin uptake remains for future investigation. Less
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