Wagner Felix A, Loescher Christine M, Unger Andreas, Kühn Michel, Klotz Annika J, Liashkovich Ivan, Ciechanska Dominika, Schillers Hermann, Koser Franziska, Freundt Johanna K, Hessel Anthony L, Linke Wolfgang A
Institute of Physiology II, University of Münster, Robert-Koch-Str. 27B, 48149, Münster, Germany.
Basic Res Cardiol. 2025 Jun 13. doi: 10.1007/s00395-025-01119-8.
Progressive myocardial dysfunction in patients with heart failure often involves alterations in myocardial passive stiffness, yet the underlying mechanisms remain incompletely understood. While passive stiffness in the longitudinal direction has been extensively characterized via uniaxial tensile stretching of cardiac specimens, transverse stiffness has received far less attention despite its equal mechanical importance. In this study, we combined atomic force microscopy nanoindentation with stretching assays on myocardial preparations to quantify the relative contributions of the three myofilament networks - actin, myosin, and titin - to passive stiffness in both transverse and longitudinal orientations. We employed a transgenic mouse model in which titin's elastic springs contain a tobacco etch virus protease (TEVp) recognition site, enabling selective and acute titin cleavage upon TEVp treatment. Actin filaments were severed using a calcium-independent gelsolin fragment, and myosin filaments were dissociated by high-salt extraction. Along the longitudinal axis, titin accounted for over 50% of total passive stiffness in both cardiac fiber bundles and isolated cardiomyocytes across most physiological strain ranges, whereas actin contributed under 35% overall - and only 15-20% within the collagen-containing fiber bundles. In contrast, in the transverse axis, titin and actin each contributed approximately 20-26% of passive stiffness in cardiac slices under varying compression forces. The myosin-titin composite thick-filament network contributed ~ 55% longitudinally but only ~ 35% transversely. These results reveal pronounced, direction-dependent differences in myofilament contributions to myocardial passive stiffness, with titin exhibiting the greatest disparity. Our findings deepen our understanding of the myocardium's multidimensional mechanics and may inform therapeutic strategies to ameliorate pathological cardiac stiffening.
心力衰竭患者的进行性心肌功能障碍通常涉及心肌被动僵硬度的改变，但其潜在机制仍未完全明确。虽然通过心脏标本的单轴拉伸对纵向被动僵硬度进行了广泛的表征，但横向僵硬度尽管具有同等的力学重要性，却受到的关注要少得多。在本研究中，我们将原子力显微镜纳米压痕与心肌制剂的拉伸试验相结合，以量化三种肌丝网络（肌动蛋白、肌球蛋白和肌联蛋白）对横向和纵向方向被动僵硬度的相对贡献。我们采用了一种转基因小鼠模型，其中肌联蛋白的弹性弹簧包含烟草蚀纹病毒蛋白酶（TEVp）识别位点，使得在TEVp处理后能够进行选择性和急性的肌联蛋白切割。使用不依赖钙的凝溶胶蛋白片段切断肌动蛋白丝，并用高盐提取法解离肌球蛋白丝。沿纵轴，在大多数生理应变范围内，肌联蛋白在心脏纤维束和分离的心肌细胞中占总被动僵硬度的50%以上，而肌动蛋白总体贡献低于35%，在含胶原纤维束中仅占15%-20%。相比之下，在横轴上，在不同压缩力下，肌联蛋白和肌动蛋白在心脏切片中各自对被动僵硬度的贡献约为20%-26%。肌球蛋白-肌联蛋白复合粗丝网络纵向贡献约55%，但横向仅贡献约35%。这些结果揭示了肌丝对心肌被动僵硬度的贡献存在明显的方向依赖性差异，其中肌联蛋白表现出最大的差异。我们的发现加深了我们对心肌多维力学的理解，并可能为改善病理性心脏僵硬的治疗策略提供依据。
