High-throughput data-driven analysis of myofiber composition reveals muscle-specific disease and age-associated patterns

Raz, Vered; Raz, Yotam; Vijver, Davy van de; Bindellini, Davide; Putten, M. van; Akker, Erik B. van den

doi:10.1101/461608

Cited by 4 publications

(16 citation statements)

References 34 publications

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“…1C). Immunofluorescence of the longitudinal and cross-sections confirmed asymmetry in myofiber type distribution: MyHC-2A myofibers were mainly found at the interior side, and major architectural differences between distal and proximal sides (Raz et al , 2018a). Moreover, we observed differences in myofiber orientation over the longitudinal axis: at the proximal side, myofiber length was parallel to the longitudinal axis, but on the distal side their axis switched (Fig.…”

Section: Resultsmentioning

confidence: 85%

“…The immunohistochemical procedure was performed with a staining protocol that is detailed in (Raz et al , 2018a). Primary antibodies that were used in this study are: Rabbit-anti-Laminin (1:1000, Abcam, UK).…”

Section: Methodsmentioning

confidence: 99%

“…Alterations in MyHC isoform expression occur in physiological conditions like training or aging, and in pathological conditions (Pette & Staron, 2000). Staining for MyHC isoforms combined with high-throughput image quantification advanced the understanding of myofiber composition and myofiber transition in disease and age(ing) conditions (Riaz et al , 2015; Riaz et al , 2016; Raz et al , 2018a). Studies on myofiber composition and changes in disease animal models are carried out in cross-sections taken from a median region of the muscle.…”

Section: Introductionmentioning

confidence: 99%

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Asymmetrical myofiber architecture along the murine tibialis anterior suggests distinct functional regions

Raz

2020

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Skeletal muscle function is inferred from the spatial arrangement of myofiber architecture and the molecular and metabolic features of myofibers. Features of myofiber types can be distinguished by the expression of myosin heavy chain (MyHC) isoforms, indicating contraction properties. In most studies, a local sampling, typically obtained from the median part of the muscle, is used to represent the whole muscle. It remains largely unknown to what extent this local sampling represents the entire muscle. Here we studied myofiber architecture over the entire wild type mouse tibialis anterior muscle, using a high-throughput procedure combining automatic imaging and image processing analyses. We reconstructed myofiber architecture from consecutive cross-sections stained for laminin and MyHC isoforms. The data showed a marked variation in myofiber geometric features, as well as MyHC expression and the distribution of neuromuscular junctions, and suggest that muscle regions with distinct properties can be defined along the entire muscle. We show that in these muscle regions myofiber geometric properties align with biological function and propose that future studies on muscle alterations in pathological or physiological conditions should consider the entire muscle.

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Section: Resultsmentioning

confidence: 85%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Asymmetrical myofiber architecture along the murine tibialis anterior suggests distinct functional regions

Raz

2020

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“…Two TA muscles were excised from two 8-weeks-old male C57BL/6J wild type mice and immediately snap froze in liquid nitrogen in the presence of iso-pentane, as described in (Raz et al, 2019;Riaz et al, 2015). One TA muscle was sectioned in a transverse axis and the other one in a longitudinal axis.…”

Section: Mouse and Immunohistochemistry Proceduresmentioning

confidence: 99%

“…Methods to stain individual MyHC isoforms using a single antibody mix (Bloemberg and Quadrilatero, 2012;Kammoun et al, 2014), led to the development of image quanti cation protocols, resulting in large myo ber datasets from which myo ber composition and spatial arrangement could be studied using data-driven analyses (Mayeuf-Louchart et al, 2018; Raz et al, 2019; Y et al, 2020). The high-throughput analysis can further improve understanding of myo ber transitions in disease and physiological conditions (Raz et al, 2019;Riaz et al, 2015;Riaz et al, 2016). Those studies on myo ber composition and changes in disease animal models are most often carried out in cross-sections taken from a median region of the muscle.…”

Section: Introductionmentioning

confidence: 99%

A data-driven analysis of fiber type architecture over the entire muscle

Bindellini

Voortman

Olie

et al. 2020

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Background Skeletal muscle function is inferred from the spatial arrangement of muscle fibers architecture, which corresponds to myofiber molecular and metabolic features. Myofiber types can be distinguished by the expression of myosin heavy chain (MyHC) isoforms, representing contraction properties. In most studies, myofiber typing is determined from a local sampling, typically obtained from the muscle median region. This median region is assumed to represent the entire muscle. However, it remains largely unknown to what extent this local sampling represents the entire muscle. Methods We present here a pipeline to study the architecture of muscle fiber type over the entire muscle, from sectioning, staining, imaging to image quantification and data-driven analysis. Results We reconstructed muscle architecture from consecutive cross-sections stained for laminin and MyHC isoforms. Examining the entire muscle using consecutive cross-sections is extremely laborious, we provide consideration to reduce dataset and yet to cover the entire muscle. Analyses of over 15,000 myofibers, showed spatial variations in myofiber geometric features, myofiber type and the distribution of neuromuscular junctions along the entire muscle. Conclusions We suggest that asymmetric spatial distribution of myofiber types, geometric features of myofibers and the neuromuscular junctions along the muscle could affect muscle function. Therefore, instead of a single sampling from a median region, representative regions covering the entire muscle should be investigated in future studies.

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A data‐driven methodology reveals novel myofiber clusters in older human muscles

et al. 2020

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Skeletal muscles control posture, mobility and strength, and influence whole‐body metabolism. Muscles are built of different types of myofibers, each having specific metabolic, molecular, and contractile properties. Fiber classification is, therefore, regarded the key for understanding muscle biology, (patho‐) physiology. The expression of three myosin heavy chain (MyHC) isoforms, MyHC‐1, MyHC‐2A, and MyHC‐2X, marks myofibers in humans. Typically, myofiber classification is performed by an eye‐based histological analysis. This classical approach is insufficient to capture complex fiber classes, expressing more than one MyHC‐isoform. We, therefore, developed a methodological procedure for high‐throughput characterization of myofibers on the basis of multiple isoforms. The mean fluorescence intensity of the three most abundant MyHC isoforms was measured per myofiber in muscle biopsies of 56 healthy elderly adults, and myofiber classes were identified using computational biology tools. Unsupervised clustering revealed the existence of six distinct myofiber clusters. A comparison with the visual assessment of myofibers using the same images showed that some of these myofiber clusters could not be detected or were frequently misclassified. The presence of these six clusters was reinforced by RNA expressions levels of sarcomeric genes. In addition, one of the clusters, expressing all three MyHC isoforms, correlated with histological measures of muscle health. To conclude, this methodological procedure enables deep characterization of the complex muscle heterogeneity. This study opens opportunities to further investigate myofiber composition in comparative studies.

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High-throughput data-driven analysis of myofiber composition reveals muscle-specific disease and age-associated patterns

Cited by 4 publications

References 34 publications

Asymmetrical myofiber architecture along the murine tibialis anterior suggests distinct functional regions

Asymmetrical myofiber architecture along the murine tibialis anterior suggests distinct functional regions

A data-driven analysis of fiber type architecture over the entire muscle

A data‐driven methodology reveals novel myofiber clusters in older human muscles

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