Journal
INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE
Volume 59, Issue 10, Pages 4218-4227Publisher
ASSOC RESEARCH VISION OPHTHALMOLOGY INC
DOI: 10.1167/iovs.18-24394
Keywords
corneal endothelium; p38 MAPK; tissue engineering
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Funding
- Program for the Strategic Research Foundation at Private Universities from MEXT
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PURPOSE. We have performed clinical research on cell-based therapy for corneal endothelial decompensation since 2013. The purpose of this study was to investigate the usefulness of a p38 MAPK inhibitor for promoting proliferation of human corneal endothelial cells (HCECs). METHODS. HCECs were cultured in media supplemented with various low-molecular-weight compounds to screen for the effect of those compounds on cell proliferation. Activation of substrates of p38 MAPK and cell cycle regulatory proteins were evaluated by western blotting. Corneal endothelial wounds were created in a rabbit model, and p38 MAPK was applied in eye drop form, followed by evaluation of cell proliferation in the corneal endothelium by Ki67-immunostaining. RESULTS. HCECs cultured with SB203580 exhibited hexagonal morphology and similar size and morphology, whereas control HCECs cultured without inhibitor exhibited monolayer morphology and varied in size and morphology. Flow cytometry demonstrated that cell proliferation was significantly increased by SB203580. Western blotting showed activation of ATF2 and HSP27 (substrates of p38 MAPK), and upregulation of cyclin D and downregulation of p27 were induced by inhibiting p38 MAPK. In the rabbit model, promotion of wound healing of the corneal endothelium was associated with significant upregulation of Ki67-positive proliferating cells following topical administration of SB203580 when compared with untreated endothelium (50.9% and 36.1%, respectively). CONCLUSIONS. Activation of p38 MAPK signaling due to culture stress might suppress the proliferation of HCECs, whereas a p38 MAPK inhibitor can counteract this activation and enable efficient in vitro HCEC expansion.
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