基于原子力显微镜技术研究不同激活态下肌节超微结构与压弹性

Ultrastructure and compression elasticity of sarcomere in different activated states using atomic force microscopy

目的搭建研究不同激活态下昆虫飞行肌纤维的超微结构与力学特性的系统,并以此开展近生理环境下的心肌生物力学研究,进而推动对心肌结构、力学特征和生理功能间关系的认识,为心肌病的基础研究和临床治疗提供更多的线索。方法采用轻敲模式的原子力显微镜技术研究僵直态、松弛态和活化态下昆虫飞行肌肌原纤维的超微精细结构;采用纳米压痕技术研究不同生理状态下肌纤维的定点弹性特性。结果肌原纤维在僵直态、松弛态和3种活化态的肌节长度分别为:(2.10±0.05)(、3.10±0.10)(、2.50±0.15)μm(2 mmol/L Ca2+)(,2.60±0.25)μm(5 mmol/L Ca2+)和(2.55±0.15)μm(10 mmol/L Ca2+),A带长度始终保持在1.50μm左右,而I带的长度则有较大的变化(0.7～1.6μm);力学实验发现,在同一生理状态,肌原纤维的弹性参数的大小满足Z线>M线>重叠区>I带;在不同活化状态下,钙离子浓度变化对Z线、M线和I带的压弹性影响的程度很接近。结论 5 mmol/L的Ca2+浓度是合适的肌纤维活化浓度,肌节长度的分布符合肌动蛋白与肌球蛋白在重叠区相对滑移的力学和空间结构模型;AFM是肌纤维超微结构与力学特性高分辨研究的潜力工具。

Objective To construct a system for studying the ultrastructure and mechanical properties of insect flight muscle fiber in different activated states so as to carry out cardiac biomechanics study in physiological environment,and further promote understanding of the relationship between cardiac structure,mechanical properties and physiological function,and provide more clues for the basic and clinical research on cardiac diseases.Methods The ultrastructure of insect flight muscle fibrils in rigor,relaxed and activated state was investigated using the tapping mode of atomic force microscopy（AFM）,and the elasticity of muscle fibers in different physiological states was studied using the nanoindentation.Results Sarcomere lengths of insect flight muscle fiber in rigor,relaxed and activated state were（2.10±0.05）,（3.10±0.10）,（2.50±0.15） μm（2 mmol/L Ca2＋）,（2.60±0.25） μm（5 mmol/L Ca2＋） and（2.55±0.15） μm（10 mmol/L Ca2＋）,respectively,while the A-band length maintained at 1.50 μm and I-band changed from 0.7-1.6 μm.Mechanical test found that the elasticity of different bands or lines in the same physiological state varied in the order of Z-lineM-lineoverlapI-band.Conclusions Critical Ca2＋ concentration for muscle fiber activation was 5 mmol/L,and sarcomere length distributions were in line with the relative slip theory and structure model,and AFM was the potential tool for the high-resolution study on ultrastructure and mechanical properties of the muscle fibers.