Badosa, C. | Search bioRxiv

bioRxiv2025

Generation and Characterization of Col6a1 knock-in mice: A Promising Pre-Clinical Model for Collagen VI-Related Dystrophies

Collagen VI Related Dystrophies (COL6-RD) are congenital muscle diseases, typically inherited as an autosomal dominant trait. A frequent type of mutation involves glycine substitutions in the triple helical domain of collagen VI alpha chains, exerting a dominant-negative effect on the unaltered protein. Despite this, no prior animal model captured this mutation type. Using CRISPR/Cas9, we generated transgenic mice with the equivalent of the human COL6A1 c.877 G>A; p. Gly293Arg mutation. We characterized their skeletal muscle phenotype over time, utilizing computer-aided tools applied to standardized parameters of muscle pathology and function. Knock-in mice exhibited early-onset reduced muscle weight, myopathic histology, increased fibrosis, reduced collagen VI expression, muscle weakness, and impaired respiratory function. These features provide adequate outcome measures to assess therapeutic interventions. The different automated image analysis methods deployed here analyze thousands of features simultaneously, enhancing accuracy in describing muscle disease models. Overall, the Col6a1 Ki Gly292Arg mouse model offers a robust platform to deepen our understanding of COL6-RD and advance its therapeutic landscape. Summary StatementWe generated and characterized over time the first mouse model representing dominant negative glycine substitutions in the alpha chains of collagen VI that are a frequent cause of Collagen VI-Related Dystrophies.

neuroscience↗

bioRxiv2024

Cell-derived matrices mechanics as a functional read-out in Collagen VI-related Congenital Muscular Dystrophies

Atomic force microscopy-force spectroscopy (AFM-FS) applied to biological samples can provide information on the micro- and nanoscale mechanical properties of tissues, which are a determinant of cellular behavior. Mechanical properties are largely determined by the structure and composition of the extracellular matrix (ECM), for which several in vitro models have been developed, including cell-derived matrices (CDMs). CDMs are decellularized natural ECMs assembled by cells, that closely mimic the in vivo stromal fiber organization and molecular content. Here we applied AFM-FS to evaluate the nanomechanical properties of CDMs obtained from primary skin fibroblast cultures of patients affected by collagen VI-related congenital muscular dystrophies (COL6-RDs). COL6-RDs are a set of neuromuscular conditions caused by pathogenic variants in any of the three major COL6 genes, which result in deficiency or dysfunction of the COL6 incorporated into the ECM of connective tissues. Current diagnosis includes the genetic confirmation of the disease and categorization of the phenotype based on maximum motor ability, as no direct correlation exists between genotype and phenotype of COL6-RDs. The development of new tools able to identify phenotype traits can significantly contribute to the diagnosis and prognosis of COL6-RDs. We describe differences in the elastic modulus (E) among CDMs deriving from patients with different clinical phenotypes, as well as the restoration of E in CDMs obtained from genetically edited cells. Results anticipate the potential of the nanomechanical analysis of CDMs as a complementary clinical tool, providing phenotypic information about COL6-RDs and their response to gene therapies.

biophysics↗