Role of Lumican on TgfβR Signaling in The Healing of Deep Corneal Epithelial Debridement

Abstract

Purpose : Previous studies demonstrated that Lumican, the last C-terminal 13 amino acid peptide (YECLRVANEVTLN) and Lumikine (YEALRVANEVTLN) bind Alk5 (type 1 Tgfβ receptor) to promote healing of corneal epithelium debridement. The present study attempts to examine if Lumikine can promote healing of deep corneal epithelium debridement (DCED).

Methods : Boyden chamber method is used to examine if Lumikine can promote migration of cultured human cornea stromal fibroblasts. Wild type and Smad4^CK△/△ (Kera-rtTA/tetO-Cre/Smad4^f/f fed Doxycycline chow) mice (8~12 weeks old) were subjected to DCED (removal of epithelium plus basement membrane). Eye drops of Lumikine (0.3 µM in PBS) are administered to injured corneas every 15 min for 2 hours, then 3 times daily for two weeks. Alternatively, the injured corneas are immediately treated with 20 µl fibrin gel containing 0.3 µM Lumikine daily for 3 days after keratectomy. Thereafter, the injured corneas are treated with eye drop up to two weeks. The experimental mouse corneas are examined by stereo microscope and/or OCT periodically after 1, 2, 3 days, 1, 2, 4 weeks of keratectomy. Eye drops containing LumC18_△C5 (LPPDMYECLRVAN) peptide and PBS are used as control.

Results : Boyden assays demonstrates that Lumikine induces cell migration of cultured human corneal stroma fibroblasts. Two distinct healing processes are involved in DCED: 1). Epithelium migration to resurface denuded corneas, 2). Remodeling of extracellular matrix of damaged corneal stroma. In wild type mice, Lumikine promote re-epithelization of denude corneal surface in 2 days, in contrast treatment of LumC18_△C5 and PBS reepithelization is delayed for more than 3 days after DCED. Interestingly, Lumikine prevents scar tissue formation whereas the prominent and progressive scar tissue formation is observed in cornea treated with LumC18_△C5 and PBS. Similar to observation seen in wild type mice, Lumikine promote epithelium debridement healing and suppress scar tissues formation in both Smad4^CK△/△ and Smad4^CKf/f corneas and suppresses scar tissues formation. However, LumC18_△C5 failed to do the same.

Conclusions : Lumikine can promote epithelium healing and suppress stroma scar tissue formation in DCED mouse corneas. The Smad4-dependent TgfβR signaling is dispensable for Lumikine efficacy in the healing of keratectomy corneas.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.