Fate of intrastromal injected adipose-derived mesenchymal stem cell and its effects on corneal nerve regeneration: An in vivo confocal microscopy study.

Abstract

Corneal nerve damage reduces corneal sensitivity and compromises ocular surface health. This study investigated the fate of adipose-derived mesenchymal stem cells (AD-MSCs) following intrastromal injection and evaluated their effects on nerve regeneration in a rabbit model of corneal nerve injury. AD-MSCs were first characterized for MSC surface markers using immunocytochemistry staining (IHC), as well as for multipotency with trilineage and neurogenic differentiation assays. The cells expressed CD73, CD90, and CD105, and successfully differentiated into chondrogenic, osteogenic, adipogenic, and neurogenic lineages. A rabbit corneal nerve injury model was created by epithelial debridement. AD-MSCs were detectable in the central cornea up to 3 days post-injection based on in vivo confocal microscopy (IVCM) and H&E staining. TUNEL staining revealed apoptotic cells as early as 0 h after injection, indicating rapid apoptosis despite the persistence of cellular remnants for several days. MSC markers (CD73, CD90, CD105) and NGF were detected in the stroma from 0 h to day 3, and NGF was positive at the epithelium on day 28. Epithelial wound healing did not differ between AD-MSC-treated eyes and controls. However, AD-MSC injection accelerated corneal nerve regeneration, with earlier recovery observed at day 14 compared with day 21 in controls, as assessed by IVCM. Immunohistochemistry for neuronal markers (SMI-312, βIII-tubulin) further confirmed enhanced sub-basal nerve regeneration. Additionally, the AD-MSC culture medium exhibited elevated levels of neurotrophic factors. These findings suggest that AD-MSCs promote corneal nerve regeneration and may represent a promising therapeutic strategy for ocular surface diseases associated with corneal nerve injury.