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- Insight to severe muscular disorders by multiscale imaging: from atomic force microscopy to second harmonic generation microscopy study hal link

Auteur(s): Gergely C.

Conference: 2nd BE-OPTICAL Workshop (La Grande Motte, Montpellier, FR, 2018-03-18)


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Résumé:

The search for predictive indicators of disease has largely focused on molecular markers. However, biophysical markers, which can integrate multiple pathways, may provide a more global picture of pathophysiology. The adequacy of the chosen, mostly combined, advanced imaging techniques is often determined by their ability for multi-scale follow-up of molecular processes. I will present through two studies the different level of information we have obtained on the pathophysiological processes of two muscular illnesses that are the amyotrophic lateral sclerosis (ALS) and the Duchenne muscular dystrophy (DMD). Early symptoms of ALS usually include muscle weakness or stiffness. Therefore, mechanical response of differentiated myotubes from primary cultures of mice expressing the ALS-causing SOD1 mutation was examined by atomic force microscopy. Simultaneous acquisition of topography and cell elasticity of ALS myotubes was performed by force mapping method, compared with healthy myotubes and supplemented with immunofluorescence and qRT-PCR studies. Wild type myotubes reveal a significant difference in elasticity between a narrow and a wide population, consistent with maturation occurring with higher actin expression. However, this is not true for SOD1 expressing myotubes, where a significant shift of thin population towards higher elastic modulus values was observed indicating that SOD1 mutation induces structural changes that occurs very early in muscle development.Duchenne muscular dystrophy (DMD) is a severe and lethal disease linked to mutations in the dystrophin gene. However, the link between the dystrophin deficiency and the contractile dysfunction of the cardiomyocytes is unclear. Second harmonic generation (SHG) microscopy was used to acquire images on a mouse model of DMD (mdx mice) and control alive cardiomyocytes at different ages and in different experimental conditions to study the cellular cytoskeleton and contractile apparatus organization. SHG makes functional imaging possible as without need of fluorescent staining, by measuring the SHG anisotropy important results are extracted on the myosin filament conformation and the organization of key sarcomeric cardiac proteins.