Abstract
Purpose :
Chemical trauma or burns to the cornea can result in limbal stem cell deficiency (LSCD), which is a debilitating condition that can lead to conjunctivalisation and neovascularisation. Regeneration and long-term repair of the cornea is a significant clinical challenge, and whilst corneal transplantation is possible, it does have some significant limitations. Dental pulp stem cells (DPSC) have shown some promise in ocular regenerative medicine and this experimental study investigated the potential for DPSC to transdifferentiate into cornea epithelial like cells (CEC), and if effective proliferation and maturation can be achieved using an acid burnt cornea model.
Methods :
A homogenous population of DPSC were isolated and selectively sorted using CD29 and CD90 markers. Keratinocyte growth factor (KGF) and Collagen Type 4 (from human placenta) were then used to facilitate the transdifferentiation of DPSC into CEC. Light microscopy was then used to assess morphological change whilst PCR and immunocytochemistry were used to identify the expression of KRT3 and KRT12 along with CD44 and CD90 to confirm that the CEC had originated from the DPSC. The transdifferentiated cells were then transferred onto the acid burnt corneas (porcine corneas burnt with 5M Hydrochloric acid to stimulate LSCD) using a soft contact lens delivery system, and attachment and proliferation was assessed under confocal microscopy. Transdifferentiated DPSCs were labelled with a Q-tracker® labelling kit and following removal of the contact lenses, immunohistochemistry was carried out to determine KRT3/12 and KRT19 expression.
Results :
There was evidence of KRT 3/12 expression and organised morphology in the DPSCs cultured with KGF and Collagen type 4. There was also evidence of reduced CD44 and CD90 expression in some DPSCs in these groups. Following successful transfer via the contact lens delivery system, DPSC had shown enhanced expression of KRT 3 and 12.
Conclusions :
This study has shown that there is potential in using KGF and Collagen Type 4 in transdifferentiating DPSCs into CEC. Transdifferentiated DPSCs can also be successfully delivered onto acid-burnt corneas via using soft contact lens. However, further work is required to determine if DPSC can form a strong and long-lasting adherence to the damaged cornea and proliferate further into mature corneal-epithelial cells.
This is a 2020 ARVO Annual Meeting abstract.