Skeletal Muscle最新文献

Background: Duchenne muscular dystrophy (DMD) is a lethal pediatric degenerative muscle disease for which there is no cure. Robust preclinical models that recapitulate major clinical features of DMD are required to investigate efficacy of potential DMD therapeutics. Rat models of DMD have emerged as promising small animal models to accomplish this; however, there have been no comprehensive studies investigating the functional skeletal muscle decrements associated with the modeling of DMD in rats.

Methods: CRISPR/Cas9 gene editing was used to generate a dystrophin-deficient Sprague-Dawley muscular dystrophy rat (MDR). Biochemical and immunofluorescent analyses were performed to confirm loss of dystrophin in striated muscles of this rat model. In situ and ex vivo muscle function was assessed in wild-type (WT) and MDR muscles at 3, 6, and 12 months of age, followed by histopathological analyses.

Results: MDR muscle tissues exhibited loss of full-length dystrophin and reduced content of other dystrophin glycoprotein complex members. MDR extensor digitorum longus (EDL) muscles and diaphragms displayed pronounced and progressive muscle weakness beginning at 3 months of age, compared to WT littermates. EDLs also exhibit susceptibility to eccentric contraction-induced damage. Functional deficits in soleus muscles were less severe and were associated with a right shift in force-frequency relationship. MDR muscles display progressive histopathology including degenerative lesions, fibrosis, regenerative foci, and modest adipose deposition.

Conclusions: MDR is a preclinical model of DMD that exhibits many translational features of the human disease, including a large dynamic range of muscle decrements, that has high utility for the evaluation of potential therapeutics for DMD.

Background: Filaminopathies, caused by pathogenic FLNC variants, are rare neuromuscular disorders characterized by protein aggregation, z-disk pathology and lead to progressive muscle weakness and/or cardiomyopathies.

Methods: To address the lack of existing filaminopathy models in skeletal muscle, we developed a patient-specific cellular platform using induced pluripotent stem cells (iPSCs) harboring two truncating filamin C (FLNc) variants (p.Q1662X, p.Y2704X). Employing a developmental human skeletal muscle organoid hSMO model, we enrich for myogenic progenitor cells that are further differentiated into functional myotubes through 2D and 3D approaches (myotubes and musculoids).

Results: The 2D myotubes exhibited poor sarcomeric organization and hallmarks of filaminopathies, including protein aggregation and proteostatic dysfunction, marked by elevated aggresome formation and an increased basal autophagic flux. The 3D musculoids revealed ultrastructural abnormalities and enabled the identification of novel disease-associated proteins involved in ER stress and protein folding (e.g. DNAJC10) through proteomic analysis. Proteomic findings were additionally validated in 2D cultures and in corresponding patient-derived muscle biopsies enhancing the model's translational value.

Conclusions: Our model is suitable to monitor aspects of filaminopathies' pathogenesis and to investigate possible therapeutic interventions with quantitative readouts.

Background: Human primary muscle cell (HPMC) lines derived from skeletal muscle biopsies are potentially powerful tools to interrogate the molecular pathways underlying fundamental muscle mechanisms. HPMCs retain their genome in culture, but many endogenous circulating factors are not present in the in vitro environment, or at concentrations that do not mirror physiological levels. To address the assumption that HPMCs are valid models of age and sex-specificity in human muscle research, we examined to what extent differentiated HPMC lines retain their source phenotype in culture.

Methods: Biopsies from the vastus lateralis muscle were collected from ten males aged 18-30, ten females aged 18-30 and ten males aged 60-75 recruited from a healthy population. A portion of the muscle was used for the establishment of 30 individual HMPC lines. The remaining sample was immediately snap frozen and stored for further analysis. RNA was extracted from muscle tissue samples and their corresponding, fully differentiated HMPCs and analysed using RNA Sequencing. To compare their transcriptomic signature, principal component analysis (PCA), differential expression analysis, single-cell deconvolution and pathway enrichment analysis were conducted in R.

Results: A comparison of the transcriptomic signature of 30 human muscle biopsies and their corresponding differentiated HPMCs indicated a near-complete lack of retention of the genes and pathways differentially regulated in vivo when compared to their in vitro equivalent, with the exception of several genes encoded on the Y-chromosome.

Conclusions: The diversity of resident cell populations in muscle tissue and the lack of sex- and age-dependent circulating factors in the cellular milieu likely contribute to these observations, which call for caution when using differentiated HPMCs as an experimental model of human muscle sex or age.

Accurate fibrosis quantification is essential for understanding muscle and cardiac disease, yet current manual and semi‑automated methods remain slow, subjective, and poorly reproducible. We introduce FibroTrack, a standalone deep learning platform with a graphical user interface (GUI) that streamlines fibrosis analysis across Sirius Red (SR), Masson's Trichrome (MT), and immunohistochemistry (IHC) stainings. FibroTrack uniquely integrates LAB (lightness, green-red, blue-yellow) color space normalization with a You Only Look Once version 11 (YOLOv11) segmentation model trained on 2,034 histological images. This approach achieved 99.5% mask precision for muscle segmentation and demonstrated excellent concordance with blinded pathologists (Spearman correlation, r = 0.87-0.96). Automated outputs include segmented images and structured spreadsheets, ensuring high reproducibility and scalability. By combining advanced color analysis with state‑of‑the‑art segmentation in an accessible tool, FibroTrack provides a novel, accurate, and clinically relevant solution for high‑throughput fibrosis quantification in both preclinical research and pathology practice.

Background: Skeletal muscle plays a crucial role in human life, contributing to posture, movement, nutrient storage, and body temperature regulation. Development and regeneration of skeletal muscles rely on embryonic myogenic progenitors and postnatal satellite cells (MuSCs), respectively. Identification of new molecular markers and elucidating their functions in MuSCs will provide better understanding of muscle development and regeneration.

Methods: We surveyed single cell RNA-seq (scRNA-seq) data (Tabula Muris and GSE150366) to identify ASB5 (Ankyrin repeat and Suppressor of cytokine signaling Box containing 5) as a marker of MuSCs. We also used CRISPR-CAS9 genome editing and oviduct electroporation to generate a germline knockout (KO) mouse line of Asb5. We then analyzed the muscle growth and regeneration of the KO mice. We further analyzed proliferation and differentiation of MuSCs attached on myofibers. We finally performed Realtime PCR (qPCR) to examine how Asb5 KO affects gene expression in the skeletal muscle.

Results: Analysis of data publicly available at Tabula Muris identified Asb5 as a specific marker of MuSCs. Further analysis of scRNA-seq data on FACS-purified MuSCs at various regeneration time points revealed that Asb5 is highly expressed in MuSCs and their progenies across various stages of muscle regeneration. We then generated a novel Asb5 KO mouse line through CRISPR-Cas9 deletion of Exon 4. The Asb5-KO mice were born normally and exhibited normal postnatal growth. In addition, Asb5-KO MuSCs proliferated, differentiated and self-renewed normally on myofiber explants. Furthermore, the skeletal muscles of Asb5-KO mice regenerated normally after acute injury. qPCR analysis showed that Asb5 KO reduces the expression levels of Tnfa (Tumor Necrosis Factor Alpha) in the skeletal muscles.

Conclusion: These data together identify ASB5 as an abundantly expressed and specific marker of MuSCs and myogenic progenitors. However, Asb5 loss-of-function has no effects on embryonic development and postnatal growth of skeletal muscles, or behavior and regenerative functions of MuSCs under normal physiological conditions.