Multiphoton microscopy method could improve diagnosis of muscle diseases

The method allows muscle function to be assessed using multiphoton microscopy without the need for biomechanical recordings.

A team of researchers at Friedrich–Alexander University Erlangen–Nürnberg (FAU; Erlangen, Germany) has developed a system that, when added to multiphoton microscopy, can accurately measure muscle weakness caused by structural changes in muscle tissue. The method allows muscle function to be assessed using imaging without the need for sophisticated biomechanical recordings, and could someday make taking tissue samples for diagnosing myopathy unnecessary.

The muscle is a highly ordered and hierarchically structured organ. This is reflected not only in the parallel bundling of muscle fibers, but also in the structure of individual cells. The myofibrils responsible for contraction consist of hundreds of identically structured units connected one after another. This orderly structure determines the force that is exerted and the strength of the muscle. Inflammatory or degenerative diseases or cancer can lead to a chronic restructuring of this architecture, causing scarring, stiffening, or branching of muscle fibers and resulting in a dramatic reduction in muscular function. Although such changes in muscular morphology can already be tracked using noninvasive multiphoton microscopy, it has not yet been possible to assess muscle strength accurately on the basis of imaging alone.

So, the research team has now developed a system that allows muscular weakness caused by structural changes to be measured at the same time as optically assessing muscular architecture. The researchers engineered a miniaturized biomechatronics system and integrated it into a multiphoton microscope, allowing them to directly assess the strength and elasticity of individual muscle fibers at the same time as recording structural anomalies, explains Oliver Friedrich, head of the Chair of Medical Biotechnology at Friedrich–Alexander University Erlangen–Nürnberg.

To prove the muscle's ability to contract, the researchers dipped the muscle cells into solutions with increasing concentrations of free calcium ions. Calcium is also responsible for triggering muscle contractions in humans and animals. The viscoelasticity of the fibers was also measured, by stretching them little by little. A highly sensitive detector recorded mechanical resistance exercised by the muscle fibers clamped on the device.

The technology is, however, merely the first step towards being able to diagnose muscle disorders much more easily in future. "Being able to measure isometric strength and passive viscoelasticity at the same time as visually showing the morphometry of muscle cells has enabled us, for the first time, to obtain direct structure-function data pairs," Friedrich says. "This allows us to establish significant linear correlations between the structure and function of muscles at the single-fiber level."

The data pool will be used in the future to reliably predict forces and biomechanical performances in skeletal muscle exclusively using optical assessments based on second-harmonic generation (SHG) images, without the need for complex strength measurements. At present, muscle cells still have to be removed from the body before they can be examined using a multiphoton microscope. However, it is plausible that this may become unnecessary if the technology can continue to be miniaturized, making it possible for muscle function to be examined, for example, using a microendoscope.

Full details of the work appear in the journal Light: Science & Applications.

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