Hoegsbjerg, C. | bioRxiv Science

bioRxiv2024

Fibre type differences in the organisation of mononuclear cells and myonuclei at the tips of human myofibres

The myotendinous junction (MTJ) is a weak link in the musculoskeletal system. Here, we isolated the tips of single myofibres from healthy human hamstring muscles for confocal microscopy (n=6) and RNAscope in situ hybridization (n=6) to gain insight into the profiles of cells and myonuclei in this region. A marked presence of mononuclear cells was observed coating the fibre tips, with a median of 29 (range 16-63) and 16 (9-23) cells per fibre for type I and II myofibres, respectively (p<0.05). The number and density of myonuclei gradually increased from the myofibre proper towards the tip (p<0.05), similarly for both fibre types, and a greater number of COL22A1-expressing nuclei was seen in type II vs type I myofibres (p<0.05). These divergent fibre type-specific characteristics of the MTJ reflect the respective demands for remodelling of the tendon and myofibre sides of the junction according to loading patterns. This insight refines our fundamental understanding of the human MTJ at the cell and structural levels. Summary statementAt the site of attachment to tendon, type I and II human myofibre tips display divergent numbers of mononuclear cells and COL22A1+ nuclei, changing our understanding of myotendinous junction biology.

physiology↗

bioRxiv2022

Distinct myofibre domains of the human myotendinous junction revealed by single nucleus RNA-seq

The myotendinous junction (MTJ) is a specialized domain of the multinucleated myofibre, faced with the challenge of maintaining robust cell-matrix contact with the tendon under high mechanical stress and strain. Here, we profiled 24,161 nuclei in semitendinosus muscle-tendon samples from 3 healthy males by single nucleus RNA-sequencing (snRNA-seq), alongside spatial transcriptomics, to gain insight into the genes characterizing this specialization in humans. We identified a cluster of MTJ myonuclei, represented by 47 enriched transcripts, of which the presence of ABI3BP, ABLIM1, ADAMTSL1, BICD1, CPM, FHOD3, FRAS1 and FREM2 was confirmed at the MTJ at the protein level by immunofluorescence. Four distinct subclusters of MTJ myonuclei were apparent and segregated into two COL22A1-expressing subclusters and two lacking COL22A1 but with a clear fibre type profile expressing MYH7 or MYH1/2. Our findings reveal distinct myonuclei profiles of the human MTJ, a weak link in the musculoskeletal system, which is selectively affected in pathological conditions, from muscle strains to muscular dystrophies.

molecular biology↗