Aims: Atropine eye drops are widely used to treat myopia (short-sight) progression in children. However, the optimal concentration and the mechanism by which atropine regulates scleral shell to slow axial eye growth and myopia, remains unclear. Here, we investigate the effects of atropine on the proteomic profiles of exosomes derived from scleral fibroblasts in vitro.
Methods: Primary scleral fibroblasts cultured from the sclera of normally developing juvenile tree shrew (Tupaia belangeri) eyes were characterized using phenotypic markers and treated with four concentrations of atropine (AT, 0.01, 0.05, 0.1%) for 24 hours. Scleral fibroblasts cultured without atropine treatment (NT) were used as controls. After treatment, exosomes were isolated and purified from the conditioned media using double ultracentrifugation. Exosomal proteins were then quantified by mass spectrometry and compared across experimental conditions.
Results: Of the 242 exosomal proteins identified, 57 were differentially expressed between AT and NT cells (fold change≥1.5, p<0.05), including proteins previously implicated in experimental myopia models, such as COL1A1, COL6A2, COL6A1, MMP2, THBS2, ITGB1, THBS1, GRP78, PRDX1, AQP1, HBA, HSP90AA1, and HSP90AB1. Atropine produced concentration-dependent upregulation of AQP1, PKM, HSP90AA1, and ANXA1 and concentration-dependent downregulation of COL1A1, COL6A2, COL6A1, MMP2, THBS2, EHD2, DHX9, PCBP1, and HNRH2 (p<0.05). Functional enrichment analysis revealed integrin cell surface interactions, extracellular matrix organization, and collagen degradation and biosynthesis as top canonical pathways.
Conclusion: Atropine produced differential expression of scleral fibroblast-derived exosomal proteins with concentration-dependent changes, suggesting that atropine might modulate scleral remodeling through alterations of exosomal proteomic signatures to slow axial eye growth and myopia.