Human umbilical tissue-derived cells protect retinal pigment epithelial cells from oxidative damage

Abstract

Purpose : Retinal pigment epithelium (RPE) cells perform many functions crucial for retinal homeostasis and vision. RPE cell dysfunction results in various retinal degenerative diseases including age-related macular degeneration (AMD) for which there are currently no effective treatment. Cell therapies targeting RPE cells are being developed in the clinic for the treatment of retinal degeneration. Human umbilical tissue-derived cell (hUTC) is an allogeneic cell-based medicinal product for the treatment of geographic atrophy, an advanced form of dry AMD. We previously demonstrated that subretinal injection of hUTC preserves visual function and retinal architecture in the RCS rat with reduced apoptosis in the retina, suggesting that hUTC may have a protective effect on retina. The retina is particularly susceptible to oxidative stress and this plays an important role in the pathogenesis of AMD and contributes to RPE damage. Here, we aimed to determine whether hUTC could improve the health of RPE cells exposed to oxidative stress.

Methods : The ARPE-19 cell line was obtained from American Type Culture Collection. Primary human RPE (hRPE) cultures were prepared from postmortem human eyes obtained from donors and the purity was confirmed by cytokeratin staining. The MTT and crystal violet assays were performed to evaluate the cell viability of ARPE-19 and hRPE exposed to a lethal dose (1500 µM) of hydrogen peroxide (H₂O₂) after co-culturing with hUTC for 72 hours in transwell inserts. Western blots were preformed to examine the level of protein carbonylation, a key marker of oxidative stress induced damage, in ARPE-19 cells co-cultured with hUTC and then subject to sublethal doses of H₂O₂ (200 and 400 µM) for 3 and 6 hours, respectively.

Results : ARPE-19 co-cultured with hUTC for 72 hours showed significantly improved viability following treatment with 1500 µM H₂O₂ compared to untreated control cells, as assessed by both MTT and crystal violet assays. Similar results were obtained in hRPE. Moreover, a significantly lower level of protein carbonylation was observed in hUTC pretreated ARPE-19 cells exposed to 400 µM H₂O₂ for 6 hours as determined by Western blot analysis.

Conclusions : These findings suggest that hUTC may secrete factors that can potentially protect RPE from oxidative damage. Investigation is currently underway to examine the mechanisms of action.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.