Limbal stem cell deficiency approaches and limbal niche restoration.

Approaches to limbal stem cell deficiency remain challenging, especially in bilateral cases, where healthy limbal stem cells are not accessible. While living-related allogeneic and allogeneic limbal stem cell sources have been utilized, their dependence on immunosuppression and its associated side effects pose significant limitations. Mucosal and mesenchymal stem cells have shown potential for differentiation into limbal stem cells and promoting corneal healing, primarily when cultured on the amniotic membrane or fibrin. However, none can fully replicate the original limbus. Innovations in surgical techniques, such as simple oral mucosal transplantation and subconjunctival or intrastromal mesenchymal stem cell injections, are emerging approaches. For successful limbal regeneration, both appropriate cells and suitable scaffolds are essential. Recent studies on decellularized and acellularized limbus models have demonstrated the potential to provide a three-dimensional native structure for cell seeding, retention, and differentiation. Creating a thin, evenly decellularized scaffold is a critical step in ensuring proper corneo-limbal slope formation, facilitating cell migration to the ocular surface. Harvesting the limbus, decellularization, and cell seeding are the three main steps in limbal reconstruction. Recent studies focus on microkeratome-assisted limbal harvesting to create a thin, even, and 360-degree limbal graft. This technique helps form an attached corneo-limbal interface, facilitating limbal stem cell migration. In the second step, acellularization is performed to preserve the extracellular matrix as much as possible, maintaining hemostasis and supporting paracrine interactions. The final steps involve recellularization and transplantation onto the eye. We summarize various limbal decellularization methods, their outcomes, and their potential in limbal reconstruction. More clinical studies are needed to validate this phase of limbal deficiency treatment.