Elucidating the Role of Limbal Niche Cell Secretome Fractions in Corneal Epithelial Homeostasis

• Applicant: Naresh Polisetti, Ph.D.
• Subject Area: Ophthalmology
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 556984092

Limbal epithelial stem/progenitor cells (LEPC) are essential for corneal epithelial homeostasis and regeneration. They reside in the corneoscleral limbus, where they interact with melanocytes (LM) and mesenchymal stromal cells (LMSC) in the limbal stem cell niche. Damage to LEPC or their surrounding niche results in limbal stem cell deficiency (LSCD), leading to corneal vascularization, opacity, and vision loss. The success rate of LEPC transplantation is limited to 50 to 70% if the tissue structure of the limbal stem cell niche is damaged. Therefore, both the restoration of a healthy niche structure and a source of syngenic cells to colonize the limbal niche are necessary to improve the treatment. Based on our previous research showing that the secretome of LMSC and LM contains factors that promote epithelial regeneration, immune- and angio-modulation, we hypothesize that specific fractions of the limbal niche cell secretome may harbor distinct regenerative, immunomodulatory, and angiomodulatory factors involved in maintaining limbal stem cell niche homeostasis. We propose that certain secretome fraction(s) may represent a novel source of therapeutic agents for treating LSCD and other corneal injuries challenging the limbal stem cell niche. Therefore, this proposal aims to fractionate LMSC- and LM-conditioned media into large EVs, small EVs, and non-vesicle fractions using tangential flow filtration, size exclusion chromatography and characterize each fraction at protein and RNA levels. It also seeks to evaluate their effects on LEPC phenotype, wound healing, immune modulation, and angiogenesis through functional in vitro assays and in a rabbit cornea injury model. This systematic evaluation of bioactive limbal niche secretome components/fractions will aid the development of cell-free therapies for treating LSCD as well as other corneal injuries. Identifying the key niche factors regulating LEPC function could enable novel approaches to pharmacologically manipulate the limbal niche microenvironment and stimulate endogenous stem cell activity to improve corneal epithelial regeneration.

• DFG Programme: Research Grants