Assessing dystrophies and other muscle diseases at the nanometer scale by atomic force microscopy

Aim: Atomic force microscopy nanoindentation of myofibers was used to assess and quantitatively diagnose muscular dystrophies from human patients. Materials & methods: Myofibers were probed from fresh or frozen muscle biopsies from human dystrophic patients and healthy volunteers, as well as mice models, and Young's modulus stiffness values were determined. Results: Fibers displaying abnormally low mechanical stability were detected in biopsies from patients affected by 11 distinct muscle diseases, and Young's modulus values were commensurate to the severity of the disease. Abnormal myofiber resistance was also observed from consulting patients whose muscle condition could not be detected or unambiguously diagnosed otherwise. Discussion & conclusion: This study provides a proof-of-concept that atomic force microscopy yields a quantitative read-out of human muscle function from clinical biopsies, and that it may thereby complement current muscular dystrophy diagnosis.

Diagnosis of muscular dystrophies at the nanometer scale

Andrzej Kulik, Thibault Adrien Kuntzer, Stefania Puttini

The diagnosis of muscular dystrophies or the assessment of the functional benefit of gene or cell therapies can be difficult, especially for poorly accessible muscles, and it often lacks a single-fiber resolution. In the present study, we evaluated whether muscle diseases can be diagnosed from small biopsies using atomic force microscopy (AFM). AFM was shown to provide a sensitive and quantitative description of the resistance of normal and dystrophic myofibers within live muscle tissues explanted from Duchenne mdx mice. The rescue of dystrophin expression by gene therapy approaches led to the functional recovery of treated dystrophic muscle fibers, as probed using AFM and by in situ whole-muscle strength measurements. Comparison of muscles treated with viral or non-viral vectors indicated that the efficacy of the gene transfer approaches could be distinguished with a single myofiber resolution. This indicated full correction of the resistance to deformation in nearly all of the muscle fibers treated with an adeno-associated viral vector that mediates exon-skipping on the dystrophin mRNA. Having shown that AFM can provide a quantitative assessment of the expression of muscle proteins and of the muscular function in animal models, we assessed myofiber resistance in the context of human muscular dystrophies and myopathies. Thus, various forms of human Becker syndrome can also be detected using AFM in blind studies of small frozen biopsies from human patients. Interestingly, it also allowed the detection of anomalies in a fraction of the muscle fibers from patients showing a muscle weakness that could not be attributed to a known molecular or genetic defect. Overall, we conclude that AFM may provide a useful method to complement current diagnosis tools of known and unknown muscular diseases, in research and in a clinical context.

World Muscular Society / Elsevier2009

Diagnosis of muscular dystrophies at the nanometer scale

Andrzej Kulik, Thibault Adrien Kuntzer, Stefania Puttini

World Muscular Society / Elsevier2009