CD8+CD103+ Tregs Attenuate Dry Eye Disease Through Suppression of CD4+ T Cell-Mediated Inflammation and Protection of Ocular Surface Integrity in Mice

CD8+CD103+ regulatory T cells represent a specialized immunosuppressive population with demonstrated therapeutic potential in autoimmune disorders; however, their role in ocular surface inflammation remains unexplored. This protocol describes a systematic approach for isolating CD8+CD103+ T cells from mice exposed to desiccating stress and evaluating their therapeutic efficacy through adoptive transfer in experimental dry eye disease. The methodology encompasses magnetic-activated cell sorting for CD8+CD103+ T-cell purification, standardized desiccating stress induction, and comprehensive functional assessments, including tear production measurement, corneal barrier permeability testing, and molecular analysis of inflammatory mediators. Representative results demonstrate that CD8+CD103+ T cell therapy significantly restored tear production, preserved conjunctival goblet cell populations, and maintained corneal epithelial barrier integrity. Treatment achieved substantial suppression of matrix metalloproteinase expression, reduced apoptotic cell death, and modulated cytokine profiles from pro-inflammatory to tissue-protective patterns. The protocol effectively reduced CD4+ T cell infiltration in ocular tissues, while simultaneously suppressing pathogenic IL-17A and IFN-γ production and enhancing protective IL-13 levels. This approach provides a robust experimental framework for investigating CD8+CD103+ T cell-mediated immunomodulation in ocular surface disorders and establishes a foundational methodology for developing novel cell-based therapeutics for dry eye disease management.