High-Mobility Group Box 1 in Dry Eye Inflammation

PURPOSE. To determine high-mobility group box 1 (1) expression during experimental dry eye (EDE) and dry eye-like culture conditions and elucidate its role in corneal dry eye related inflammation METHODS. EDE was induced in 8- to 12-week-old. C57BL/6 mice. Corneal tissue sections and lysates from EDE and untreated mice were evaluated for HMGB1 expression by immunostaining and quantitative real-time PCR (qPCR). For in vitro studies, human corneal epithelial cells (HCEC) were treated with hyperosmolar media, toll-like receptor (TLR) agonists, or proinflanunatory cytokines to determine HMGB1 expression. HCEC were also treated with human recombinant HMGBI (hrHMGB1) alone or in combination with inflammatory stimuli, and TINF alpha, IL-6, and IL-8 expression evaluated by qPCR and HASA. Nuclear factor-kappa B (NF-kappa B) p65 nuclear translocation was determined by immunostaining. RESULTS. EDE mice had higher corneal HMGB1 RNA and protein expression compared to untreated animals. In HCEC, hyperosmolar stress and. TNF alpha treatment stimulated HMGB1 production and secretion into culture supernatants. However, in vitro stimulation with hrHMGB1 did not induce secretion of TNF alpha,/ IL-6, or IL-8 or NF-kappa B p65 nuclear translocation. In addition, the inflammatory response elicited by TLR agonists fibroblast-stimulating lipopeptide-1 and lipopolysaccharide was not enhanced by hrHMGB1 treatment. CONCLUSIONS. HMGB1 expression was enhanced by dry eye conditions in vivo as well as in vitro, during hyperosmolar stress and cytokine exposure, suggesting an important role for HMGB1 in dry eye disease. however, no direct inflammatory effect was observed with HMGBl treatment. Therefore, under these conditions, HMGBI does not contribute directly to dry eye-induced inflammation and its function at the ocular surface needs to he explored further.