Short-Term Ultraviolet A Irradiation Leads to Dysfunction of the Limbal Niche Cells and an Antilymphangiogenic and Anti-inflammatory Micromilieu

PURPOSE. We analyzed the effects of short-term ultraviolet A (UVA) irradiation on the putative limbal stem cell phenotype, limbal fibroblasts, corneal inflammation, and corneal (lymph) angiogenic privilege. METHODS. Primary human limbal epithelial cells and fibroblasts were irradiated with 5.2 J/cm(2) of UVA. The limbal epithelial cell phenotype was assessed using P63a, cytokeratin 15, integrin b1 (marking stem and transient amplifying cells), and cytokeratin 3 (a differentiation marker) as well as by a colony-forming efficiency (CFE) assay. An epithelial-fibroblast coculture model was used to compare the ability of irradiated and nonirradiated fibroblasts to support the putative limbal stem cell phenotype. The effects of the conditioned media of irradiated and nonirradiated cells on proliferation and tube formation of human lymphatic and blood endothelial cells also were tested. The levels of factors related to angiogenesis and inflammation were assessed in a protein array and using ELISA. RESULTS. Ultraviolet A induced phenotypical changes of limbal epithelial cells, as their CFE and putative stem cell/transient amplifying marker expression decreased. Limbal epithelial cells cocultured with UVA-irradiated limbal fibroblasts also exhibited differentiation and CFE decrease. Conditioned media from irradiated limbal epithelial cells and fibroblasts inhibited lymphatic endothelial cell proliferation and tube network complexity. Levels of monocyte chemoattractant protein 1 (MCP1) were reduced following UVA irradiation of both cell populations, while levels of IFN-gamma increased in irradiated limbal epithelial cells. CONCLUSIONS. These data imply a key role of cellular components of the limbal niche following short-term UVA irradiation. Overall, UVA irradiation leads to dysfunction of these cells and a anti(lymph) angiogenic and anti-inflammatory micromilieu.