Biochmechanics of MFM: a multi-scale approach
Final Report Abstract
The Z-Project ‘Biomechanics of Myofibrillar Myopathies’ entered FOR1228 in the second funding phase with the aim to provide a nationwide unique platform to assess passive and active biomechanical properties in muscle of MFM disease models within the consortium, covering various organ levels from whole muscle to multicellular fibre bundle preparations, single muscle fibres/myocytes and subcellular multi-myofibrillar bundles. Technologies available were automated force transducer MyoRobot technology (MBT, OF), optical tweezers, traction microscopy and cell stretcher technologies (ZMPT, BF) and laser-cantilever force transducers (Physiology, GP). The MyoRobot technology at the Erlangen labs was further developed and driven into automated biomechanics recordings and applied to small fibre bundles of DesR349P expressing fast- and slowtwitch muscle from young (17-23 wks) animals with hetero-/homogenous background. Main finding was a markedly increased axial stiffness of bundles that limit their resistance to mechanical stretch and renders them more susceptible to stretch-induced damage and rupture. The studies were complemented by high-end label-free multiphoton Second Harmonics Generation microscopy in conjunction with quantitative morphometry that documented a vast ultrastructural myofibrillar disorganization already at pre-clinical stages of desminopathy before the onset of muscle weakness. We also tested the mechanical stress vulnerability of desmin mutant cells under uniaxial static and cyclic stretch, and found significantly increased cell death in homozygous desmin mutant myoblasts upon stretch amplitudes of 15% and larger. Traction force microscopy measurements showed that both heterozygous and homozygous desmin mutant myoblasts were less contractile compared to wildtype control cells. These data at the single cell level recapitulate previously reported data obtained at the tissue level. Finally, we succeeded to culture desmin mutant myoblasts in 3-dimensional microtissues in betwen two force sensors on a stretchable substrate. These cells differentiate into myotubes, and when electrically excited, the microtissues contract promptly. GSD and STED superresolution microscopy was performed in Cologne to visualize the 3D-architecture of desmin and plectin linkages in thin myofibrillar bundles isolated from fast psoas and slow soleus muscles. Both proteins are detected in the space between the Z-discs of single myofibrils corroborating the hypothesized role of plectin and desmin as intermyofibrillar structural linkers, even in very thin (< 2 µm diameter) bundles. The effect of loss of these linkages studied by force measurements of myofibrillar bundles and skinned single skeletal muscle fibers isolated from plectin-KO mouse models, however, revealed a relatively mild biomechanical phenotype. Subtle effects on relaxation kinetics could be provoked by increased mechanical stress.
Publications
- Chemical chaperone ameliorates pathological protein aggregation in plectin-deficient muscle. J Clin Invest 2014; 124: 1144-1157
Winter L, Staszewska I, Mihailovska E, Fischer I, Goldmann WH, Schröder R, Wiche G
(See online at https://doi.org/10.1172/JCI71919) - Determining the mechanical properties of plectin in mouse myoblasts and keratinocytes. Exp Cell Res 2014; 331: 331-337
Bonakdar N, Schilling A, Spörrer M, Lennert P, Mainka A, Winter L, Walko G, Wiche G, Fabry B, Goldmann WH
(See online at https://doi.org/10.1016/j.yexcr.2014.10.001) - Biomechanical characterization of myofibrillar myopathies. Cell Biol Int 2015; 39361-39363
Winter L, Goldmann WH
(See online at https://doi.org/10.1002/cbin.10384) - Plectin reinforces vascular integrity by mediating vimentinactin network crosstalk. J Cell Sci 2015; 1284138-4150
Osmanagic-Myers S, Rus S, Wolfram M, Brunner D, Goldmann WH, Bonakdar N, Fischer I, Reipert S, Zuzuarregui A, Walko G, Wiche G
(See online at https://doi.org/10.1242/jcs.172056) - The toxic effect of R350P mutant desmin in striated muscle of man and mouse. Acta Neuropathol 2015; 129: 297-315
Clemen CS, Stöckigt F, Strucksberg KH, Chevessier F, Winter L, Schütz J, Bauer R, Thorweihe JM, Wenzel D, Schlötzer-Schrehardt U, Rasche V, Ksmanovic P, Katus HA, Rottbauer W, Just S, Müller OJ, Friedrich O, Meyer R, Herrmann H, Schrickel JW, Schröder R
(See online at https://doi.org/10.1007/s00401-014-1363-2) - Early signs of architectural and biomechanical failure in isolated myofibers and immortalized myoblasts from desmin-mutant knock-in mice. Sci Rep 2017; 7: 1391
Diermeier S, Iberl J, Vetter K, Haug M, Pollmann C, Reischl B, Buttgereit A, Schürmann S, Spörrer M, Goldmann WH, Fabry B, Elhamine F, Stehle R, Pfitzer G, Winter L, Clemen CS, Herrmann H, Schröder R, Friedrich O
(See online at https://doi.org/10.1038/s41598-017-01485-x) - Pre-aged remodelling of myofibrillar cytoarchitecture in skeletal muscle expressing R349P mutant R349P desmin. Neurobiol Aging 2017; 58: 77-87
Diermeier S, Buttgereit A, Schürmann S, Xu H, Murphy RM, Clemen CS, Schröder R, Friedrich O
(See online at https://doi.org/10.1016/j.neurobiolaging.2017.06.001)
