FURIN Stimulates NOTCH2 and NOTCH3 Pathways, Leading to Return of Function in Aged Cells

Aging is accompanied by a progressive decline in skeletal muscle regeneration, largely due to impaired myogenic differentiation. The proprotein convertase FURIN is a key protease responsible for activating several signaling molecules, including precursors of NOTCH receptors, which regulate cell fate and differentiation. In this study, we investigated whether age-associated downregulation of FURIN contributes to impaired NOTCH2/3 signaling and myogenic function.

An initial bioinformatics analysis of public scRNA-seq data from Genotype-Tissue Expression (GTEx) project indicated age-related expression of genes in the NOTCH signaling pathway. In vitro verification used early- and late-passage C2C12 myoblasts as a model of muscle cell aging to compare the expression of these genes. Late-passage C2C12 cells were transiently transfected with FURIN plasmid to assess restoration of differentiation potential, quantified by the fusion index, myogenic marker expression, and morphology.

Expression of FURIN, NOTCH2 and NOTCH3 was negatively correlated with age, whereas GZMB increased with age in GTEx dataset. Late-passage myoblasts exhibited impaired myotube formation, reflecting age-associated loss of myogenic capacity. Restoration of FURIN expression in aged myoblasts was associated with reduced GZMB levels, increased expression of embryonic myosin heavy chain IGF1, and partial recovery of myogenic differentiation and myotube formation.

These findings suggest that age-associated loss of FURIN contributes to impaired NOTCH2/3 pathways and myogenic dysfunction. Overexpression of FURIN partially rescues the myogenic phenotype and increases expression of early myogenic markers in aged cells, identifying FURIN as a potential regulator of muscle regenerative capacity during aging. We suggest FURIN as a promising candidate target for further investigation into the mechanisms driving aging or age-related decline.