Anti-inflammatory effects of topical supernatant from human amniotic membrane cell culture after human amniotic membrane transplantation in canine deep corneal ulcer

Abstract

The objective of this study was to examine the effect of topically applied human amniotic epithelial cell (HAEC) culture supernatant on corneal inflammatory reaction in dogs. Twenty five dogs were randomly assigned into 5 groups. The control group consisted of 5 dogs with normal cornea. Inductions of corneal ulcer were performed using 0.45 cm trephine and human amniotic membrane were transplanted in 20 dogs. These 20 dogs were assigned into 4 groups and treated with topical antibiotic, topical corticosteroid, topical mock media and topical culture supernatant from HAEC, respectively. Administrations of the testing agents started at 24 h after transplantation and continued every 4 h for 9 consecutive days. Tear was collected before operation (day 0), 24 h after transplantation, but before application of the testing agents (day1), and day 3, 5, 7 and 9 after transplantation. The concentrations of interleukin-1 beta (IL-1[beta]) and nitric oxide (NO) in tear fluid were measured using canine IL-1[beta] ELISA kit and Griess assay, respectively. Elevations of IL-1[beta] and NO concentrations are associated with inflammatory conditions within the eyes. IL-1[beta] is a major cytokine involved in ocular inflammation and it may induce NO production from many cell types, such as fibroblasts, macrophages and epithelium of the iris-ciliary body. Corticosteroid, a reference anti-inflammatory drug, and the culture supernatant from HAEC significantly decreased IL-1[beta] and NO concentrations. In addition, the clinical signs such as conjunctivitis and neovascularization were improved in both topical corticosteroid and supernatant from HAEC treated groups. Mock and antibiotic solutions failed to decrease NO and IL-1[beta] concentrations. In conclusion, topical application of the culture supernatant from HAEC alleviated inflammation in induced-corneal ulcer of dogs, possibly via inhibition of IL-1[beta] and NO production