Purpose: Mesenchymal stromal cells (MSCs) have been shown to enhance tissue repair as a cell-based therapy. In preparation for a phase I clinical study, we evaluated the safety, dosing, and efficacy of bone marrow–derived MSCs after subconjunctival injection in preclinical animal models of mice, rats, and rabbits.

Methods: Human bone marrow–derived MSCs were expanded to passage 4 and cryopreserved. Viability of MSCs after thawing and injection through small-gauge needles was evaluated by vital dye staining. The in vivo safety of human and rabbit MSCs was studied by subconjunctivally injecting MSCs in rabbits with follow-up to 90 days. The potency of MSCs on accelerating wound healing was evaluated in vitro using a scratch assay and in vivo using 2-mm corneal epithelial debridement wounds in mice. Human MSCs were tracked after subconjunctival injection in rat and rabbit eyes.

Results: The viability of MSCs after thawing and immediate injection through 27-and 30-gauge needles was 93.1%±2.1% and 94.9%±1.3%, respectively. Rabbit eyes demonstrated mild self-limiting conjunctival inflammation at the site of injection with human but not rabbit MSCs. In scratch assay, the mean wound healing area was 93.5%±12.1% in epithelial cells co-cultured with MSCs compared with 40.8%±23.1% in controls. At 24 hours after wounding, all MSC-injected murine eyes had 100% corneal wound closure compared with 79.9%±5.5% in controls. Human MSCs were detectable in the subconjunctival area and peripheral cornea at 14 days after injection.

Conclusions: Subconjunctival administration of MSCs is safe and effective in promoting corneal epithelial wound healing in animal models.

Translational Relevance: These results provide preclinical data to support a phase I clinical study.

Introduction

An intact corneal epithelium provides a critical defensive barrier and is essential for clear vision. After nonpenetrating trauma, the cornea typically re-epithelializes promptly, minimizing the risk of infection, opacification, and perforation. Epithelial defects can persist in the presence of certain pathologic conditions, such as limbal stem cell deficiency, exposure keratopathy, and neurotrophic keratitis. Management of patients with nonhealing corneal epithelial wounds can be challenging. Several strategies are currently available in treatment of these conditions including limbal stem cell transplantation, amniotic membrane transplantation, and soft contact lenses. Despite many advances, there is still an unmet need for effective clinical strategies to promote corneal repair in these patients in which the outcomes of current therapies are suboptimal. 1 For instance, in the setting of severe chemical injuries in which there are very few to no live cells remaining in the cornea, none of the standard treatments can alter the disease course. The emergent cell-based therapies using mesenchymal stromal cells (MSCs) may potentially support the corneal structure and prove useful in addressing these unmet clinical needs in severe ocular surface disease. 2