Development of a method to measure viscoelasticity of cardiomyocytes using scanning probe microscope

开发使用扫描探针显微镜测量心肌细胞粘弹性的方法

• 批准号: 12557066
• 负责人: SUGIMACHI Masaru
• 金额: $ 4.61万
• 依托单位: National Cardiovascular Center Research Institute
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12557066/
• 关键词: scanning probe microscope; minute vibration; time-varying elastance; viscosity; cardiomyocyte; laminin; collagenase; biopsy specimen

We developed a method to measure viscoelasticity of cardiomyocytes from animals. Rats were anesthetized with pentobarbital, and the hearts were retrogradely perfused with collagenase to isolate cells. The cells were incubated for 4 hour (37℃, 5 %CC_2) in a laminin-coated dish, during which the cells firmly attached to the dish. We made a circuit and modified a scanning probe microscope so that we were able to control the vertical position of the probe with a piezoelectric device according to external command. We also developed a software to impose command and to measure the position of the piezoelectric device and the bending of the cantilever. When we imposed a sinusoidal vibration, the bending of the cantilever was in phase, indicating the viscosity of the superfusate is negligible.We measured the changes in elasticity of cells during cardiac cycle by subjecting the cell to electrical stimulation, but the results indicated no significant changes. As it is known osmotic stress do change elasticity, we conjectured that the probe detects the local elasticity, and the changes in probe size seem necessary to detect the myocardial cellular elasticity changes. The lack of preload seems to contribute the small changes in elasticity.We were not able to collect sufficient healthy cells either by superfusion with collagenase with or without ultrasonic agitation, or super fusion with protease.

我们建立了一种测量动物心肌细胞粘弹性的方法。用戊巴比妥麻醉大鼠，用胶原酶逆行灌注心脏以分离细胞。将细胞在层粘连蛋白包被的培养皿中孵育4小时（37℃，5%CC_2），在此期间细胞牢固地附着在培养皿上。我们制作了一个电路，并修改了一个扫描探针显微镜，使我们能够控制与压电器件根据外部命令的探针的垂直位置。我们还开发了一个软件来施加命令，并测量压电器件的位置和悬臂梁的弯曲。当我们施加正弦振动时，悬臂梁的弯曲是同相的，这表明灌流液的粘度可以忽略不计。我们通过使细胞受到电刺激来测量细胞在心动周期中弹性的变化，但结果表明没有显著变化。由于渗透压确实改变了心肌细胞的弹性，我们证实了探针检测的是局部的弹性，而探针大小的改变似乎是检测心肌细胞弹性变化所必需的。缺乏前负荷似乎有助于弹性的小变化。我们不能收集足够的健康细胞，无论是通过超声波搅拌或不超声波搅拌，或超融合与蛋白酶。

项目成果

Sugimachi M et al: "A new model-based method of reconstructing central aortic pressure from peripheral arterial pressure"Jpn J Physiol. 51. 217-222 (2001)

Sugimachi M 等人：“一种基于模型的从外周动脉压重建中心主动脉压的新方法”Jpn J Physiol。

Shishido T et al: "Single-Beat Estimation of End-systolic Elastance Using Bilinearly Approximated Time-Varying Elastance Curve"Circulation. 102. 1983-1989 (2000)

Shishido T 等人：“使用双线性近似时变弹性曲线对收缩末期弹性进行单搏估计”循环。

Shishido T: "Single-beat estimation of end-systolic elastance using bilinearly approximated time-varying elastance curve."Circulation.. 102. 1983-1989 (2000)

Shishido T：“使用双线性近似时变弹性曲线对收缩末期弹性进行单次估计。”Circulation.. 102. 1983-1989 (2000)

Hayashi K et al: "Single-beat estimation of Ees/Ea as an index of ventricular mechano-energrtic performance"Anesthesiologyh. 92. 1769-1776 (2000)

Hayashi K 等人：“Ees/Ea 的单次估计作为心室机械能量性能的指标”麻醉学。

Sugimachi M: "Lower compliance rather than high reflection of arterial system decreases stroke volume in arteriosclerosis : A simulation."Jpn J Physiol. (In press).

Sugimachi M：“动脉系统的低顺应性而不是高反射会降低动脉硬化中的每搏输出量：模拟。”Jpn J Physiol。