Decorin regulates collagen fibrillogenesis during corneal wound healing in mouse in vivo

Abstract

A characteristic rigid spatial arrangement of collagen fibrils in the stroma is critical for corneal transparency. This unique organization of collagen fibrils in corneal stroma can be impacted by the presence and interactions of proteoglycans and extracellular matrix (ECM) proteins in a corneal microenvironment. Earlier studies revealed that decorin, a leucine-rich proteoglycan in stroma, regulates keratocyte-collagen matrix assembly and wound healing in the cornea. This study investigated the role of decorin in the regulation of stromal fibrillogenesis and corneal transparency in vivo employing a loss-of-function genetic approach using decorin null (dcn−/−) and wild type (dcn+/+) mice and a standard alkali-injury model. A time-dependent ocular examinations with Slit lamp microscope in live animals assessed corneal clarity, haze, and neovascularization levels in normal and injured eyes. Morphometric changes in normal and injured dcn+/+ and dcn−/− corneas, post-euthanasia, were analyzed with Masson's Trichrome and Periodic Acid-Schiff (PAS) histology evaluations. The ultrastructure changes in all corneas were investigated with transmission electron microscopy (TEM). Injury to eye produced clinically relevant corneal haze and neovascularization in dcn−/− and dcn+/+ mice while corneas of uninjured eyes remained clear and avascular. A clinically significant haze and neovascularization appeared in injured dcn−/- corneas compared to the dcn+/+ corneas at day 21 post-injury and not at early tested times. Histological examinations revealed noticeably abnormal morphology and compromised collagen levels in injured dcn−/− corneas compared to the injured/normal dcn+/+ and uninjured dcn−/− corneas. TEM analysis exhibited remarkably uneven collagen fibrils size and distribution in the stroma with asymmetrical organization and loose packing in injured dcn−/− corneas than injured/normal dcn+/+ and uninjured dcn−/− corneas. The minimum and maximum inter-fibril distances were markedly irregular in injured dcn−/− corneas compared to all other corneas. Together, results of clinical, histological, and ultrastructural investigations in a genetic knockout model suggested that decorin influenced stromal fibrillogenesis and transparency in healing cornea.