Vitamin D Activation and Function in Human Corneal Epithelial Cells During TLR-Induced Inflammation

PURPOSE. Vitamin D is recognized to be an important modulator of the immune system. In the eye, studies have shown that deficiencies and genetic differences in vitamin D-related genes have a significant impact on the development of various ocular diseases. Our current study examines the ability of human corneal epithelial cells (HCEC) to activate vitamin D and the effect of vitamin D treatment on antimicrobial peptide production and cytokine modulation during inflammation, with the ultimate goal of using vitamin D therapeutically for corneal inflammation. METHODS. Human corneal epithelial cells were treated with 10(-7)M vitamin D-3 (D-3) or 25-hydroxyvitamin D-3 (25D(3)) for 24 hours and 1,25-dihydroxyvitamin D-3 (1,25D(3)) detected by immunoassay. Human cathelicidin (LL-37) expression was examined by RT-PCR, immunoblot, and immunostaining following 1,25D(3) treatment and antimicrobial activity of 1,25D(3)-treated cells was determined. Cells were stimulated with TLR3 agonist polyinosinic-polycytidylic acid (Poly[I:C]) for 24 hours and cytokine levels measured by RT-PCR, ELISA, and Luminex. Immunostaining determined expression of vitamin D receptor (VDR) and retinoic acid inducible gene-1 receptor (RIG-1) as well as NF-kappa B nuclear translocation. RESULTS. When treated with inactive vitamin D metabolites, HCEC produced active 1,25D(3), leading to enhanced expression of the antimicrobial peptide, LL-37, dependent on VDR. 1,25-D-3 decreased the expression of proinflammatory cytokines (IL-1 beta, IL-6, TNF alpha, and CCL20) and MMP-9 induced by Poly(I:C) as well as pattern recognition receptor expression (TLR3, RIG-1, MDA5). However, early activation of NF-kappa B was not affected. CONCLUSIONS. These studies demonstrate the protective ability of vitamin D to attenuate proinflammatory mediators while increasing antimicrobial peptides and antipseudomonas activity in corneal cells, and further our knowledge on the immunomodulatory functions of the hormone.