Development of In Vitro Dry Eye Models to Study Proliferative and Anti-Inflammatory Effects of Allogeneic Serum Eye Drops

This study aimed to develop valid in vitro models for preclinical evaluation of proliferative and anti-inflammatory effects of human allogeneic serum eye drops for dry eye disease (DED) treatment. A DED wound healing model was developed by analyzing the influence of coating and serum concentrations on human corneal epithelial (HCE-T) wound closure. Further, intralaboratory variance, freeze–thaw cycle effects, donor variability and stability assays were conducted. Interleukin-1β (IL-1β) and tumor necrosis factor α (TNFα) were used to induce the gene expression of matrix metalloproteinase 9 (MMP9), cyclooxygenase 2 (COX2), transforming growth factor-β (TGFβ) and IL-1β. MMP9 induction was optimized using a design-of-experiments (DoE) approach and applied to examine serum under static and dynamic conditions. MMP9 protein expression was analyzed by ELISA. The DED wound healing model detected proliferative effects of serum down to 1% with a small intralaboratory variance. Serum stability was shown over six months, donor variance could be detected, and freeze–thaw cycle effects did not affect wound closure. Serum decreased MMP9 expression on the gene and protein levels. The induction method was successfully optimized using DoE modeling and transferred to a dynamic setting mimicking tear film fluidics. The DED wound healing and inflammatory DED model present useful in vitro models for the preclinical evaluation of allogeneic serum eye drops without the use of animal experiments.