Integrated analysis of three-dimensional molecular dynamics of actin and myosin, and macro-dynamics of cardiac muscle and left ventricle

本项目旨在用时间分辨的方法分析离体心肌和全心肌动蛋白与肌球蛋白在一个心动周期内的分子相互作用。我们在Spring-8（超级光子环8GeV）的BL45XU光束线上组装了x射线能量12.4keV、波长1a的短角衍射仪。利用x射线像增强器和冷却CCD测量x射线衍射图样。对于左心室衍射图的测量，我们使用了比BL45XU能量更高的BL40XU。在这种情况下，相机长度为3m， x射线能量为16keV。通过改变前端狭缝的宽度，将x射线能量控制在合适的强度。通过激光衍射图测量肌节长度，并与x射线横向衍射图进行比较，分析分子水平和细胞水平的三维动力学。我们对大鼠右心室离体和灌注的乳头肌进行了赤道衍射。通过(1,0)/(1,1)衍射的比值，测定了肌凝蛋白分子向肌动蛋白分子的传质，即形成过桥的指标。我们分析了肌动蛋白和肌球蛋白分子间相互作用随肌节长度和等张收缩后负荷变化的变化。此外，我们还分析了离体langendorff灌注心脏左室壁x射线衍射图。在等长收缩过程中，肌球蛋白分子向肌动蛋白的质量传递随着心肌长度的增加而增加，因此我们得出结论，过桥形成的数量是心脏Starling机制的分子基础。在等张收缩中，发展力也随着桥的形成量的增加而增加。然而，有趣的是，在小的后负荷收缩中，从肌凝蛋白分子到肌动蛋白的传质量比等长收缩中相同发展力下的传质量要小。等容收缩左心室的压力发展与传质一致，说明了全心起搏机制的分子基础。左室壁衍射图随x线照射心内膜下、中壁和心内膜下部位的不同而有规律变化。我们认为，等容收缩离体灌注心脏左室游离壁不同部位的三维分子结构变化可以用时间分辨的方法来估计。少

The purpose of this project is to analyze the molecular interaction between actin and myosin in isolated cardiac muscle and whole heart with time- resolved way in one cardiac cycle.We assembled the short angle diffraction apparatus with x-ray energy 12.4keV and wave length 1 A at BL45XU beam line in Spring-8 (Super Photon Ring 8GeV). We measured x-ray diffraction pattern by using x-ray image intensifier and cooled CCD. For the measurement of diffraction pattern from the left ventricle we used BL40XU, which has higher energy than BL45XU. In this case the camera length was 3m and x-ray energy was 16keV. We controlled the x-ray energy to the suitable strength with changing the width of front-end slit. We measured the sarcomere length by laser diffraction pattern and compared it with lateral diffraction pattern by x-ray, and thus analyze the three-dimensional dynamics between molecular and cellular level.We got the equatorial diffraction from isolated and perfused papillary muscle from rig … More ht ventricle of rat. From the ratio between (1,0)/(1,1) diffraction the mass transfer from the myosin molecule to actin molecule, which is the index of crossbridge formation, was measured. We analyzed the change of molecular interaction between actin and myosin with the change of sarcomere length and afterload in isometric and isotonic contraction. In addition we analyzed the x-ray diffraction pattern from left ventricular wall in isolated Langendorff-perfusion heart.As the mass transfer from myosin molecular to actin increased with cardiac muscle length in isometric contraction, we concluded the amount of crossbridge formation is the molecular basis of the Starling mechanism of the heart. In isotonic contraction the developed force also increased with the amount of crossbridge formation. However, the amount of mass-transfer from the myosin molecular to actin in small afterloaded contraction was interestingly smaller than that at the same developed force in isometric contraction. The developed pressure in isovolumically contracting left ventricle was in accordance with the mass-transfer, indicating the molecular basis of the Staring mechanism of the heart in whole heart. The diffraction pattern from the left ventricular wall changed with regular manner depending on x-ray irradiation portion at subepicardium, mid wall and subendocardium. We concluded that the change of the three-dimensional molecular structure at the different portion of the left ventricular free wall could be estimated in time resolved manner in isovolumically contracting isolated and perfused heart. Less