Objectives The transparent surface of the eye, called the cornea, plays a crucial role in transmitting light to the retina and in protecting the eye. On its external surface, the cornea is composed of a constantly renewing multistratified epithelium. This mechanism is fueled by stem cells located in the limbus (the transition zone between the cornea and the sclera). Limbal Stem Cell Deficiency (LSCD) is characterized by a decrease or cessation of epithelial renewal and neovascularization of the cornea. Consequently, the cornea loses its integrity and transparency. This visually impairing condition is currently in a therapeutic impasse as only autologous limbal or allogeneic corneal grafts are viable options, but they pose significant risks to patients.
Studies have shown that the oral cavity contains stem cells that can be isolated, cultured, and transdifferentiated into limbal stem cells (LSCs). However, to date, these studies are limited, and no protocol has been validated. In this study, the advantage of the accessibility of the oral cavity is used to develop a protocol for differentiating cells from the oral mucosa into limbal stem cells (LSCs) for use in a future clinical trial with patients.
Methodology This prospective monocentric study will be conducted on patients from the ophthalmology department of the Montpellier University Hospital who have an indication for conjunctival reconstruction. After obtaining consent from the patients, cells from the oral wall will be sent to the tissue bank of the Montpellier University Hospital, where they will be cultured. Finally, the transdifferentiation steps will be analyzed by the "Eye" research team at the Institute of Neuroscience in Montpellier, which is collaborating on the project. The investigators have established objective factors to assess the success of the developed protocol based on the literature : (i) \> 3% of stem cells in primary cultures, (ii) \<10% aborted colonies, (iii) Expression of LSC markers (Pax6, Krt14, p63).
Presenting an indication for conjunctival reconstruction using oral mucosa (for example because of a recurrent pterygium, conjunctival carcinoma or ocular burn)
Written consent to participate in this research
Chronic infection (HIV, hepatitis, tuberculosis, Lyme disease)
Oral pathology which could affect the health of the oral mucosa tissue according to the opinion of the investigating doctor
Uncontrolled systemic disease
Pregnant or breastfeeding
Single group
Conjunctival reconstruction
Procedure/SurgeryConjunctival reconstruction using buccal mucosa taken from the patient's mouth. Following the reconstruction operation, an additional biopsy of buccal mucosa will be taken to develop a laboratory protocol for the transdifferentiation of these buccal cells into corneal epithelial stem cells.
Successful Transdifferentiation
Successful differentiation, defined by the percentage of stem cells on primary cultures ≥ 3%
Successful Transdifferentiation
Successful differentiation, defined by the percentage of aborted colonies \< 10%.
Successful Transdifferentiation
The quantitative real-time polymerase chain reaction, (qPCR) technique will be used to monitor expression levels of limbal stem cell markers such as Krt14 and Krt5. The qPCR technique will be used to monitor expression levels of transcription factors.
Successful Transdifferentiation
Successful differentiation, defined by the presence of transcription factors p63 and Pax6 The qPCR technique will be used to monitor expression levels of transcription factors.
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University Hospital, Montpellier
Lead sponsor
Institute for Neuroscience Montpellier
Collaborator
