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Analyse protéomique et propriétés de ré-épithélialisation des membranes amniotiques humaines en vue d'une greffe de la surface oculaire

Abstract : The Human Amniotic Membrane (HAM) graft allows the healing of corneal ulcers and rescues a significant number of eyes with chemical burn. HAM is a biological material, its use for the treatment of ocular surface diseases gives good results because of its ability to reduce inflammation and promote rapid epithelialization. For its clinical use, the HAM must of course be sterile, but also easy to transport from the sampling center to the transplant center, and easily storable and for a long time. Currently on routine in the tissue bank of Rouen, the amniotic membrane is separated from the amnion and denuded of its spongy layer. Subsequently this membrane is stored by cryopreservation (freezing at -80 ° C) which potentially complicates the delivery of membranes. Consequently, as part of this study with the Banque Normande de Cornées of Rouen University Hospital, we have developed freeze-drying of HAM to facilitate its use and distribution. The HAM mapping study will also determine whether the level of growth factors is homogeneous in the HAM or whether it depends on its distance from the umbilical cord. The study of the in vivo biocompatibility of a second material composed of collagen also allows us to consider an alternative for implantation at the level of the stroma. Our protein analyzes (ELISA and Label-free) of freeze-dried HAM do not show any significant difference in terms of quantity and protein quality. The proteomic approach is complemented by the analysis of the ability of human corneal epithelial cells (CECs) to multiply on the freeze-dried amniotic membrane in vitro. We did not observe any difference between the epithelial cells growths on freeze-dried or frozen HAM. The analysis of the extracted protein total also shows that freeze-drying does not degrade the HAM at the protein level. At the structural level the electron microscopy results showed that the structure of the MAH stroma is impacted by freeze-drying. The MAH transplant performed on corneal ulcers in rabbits was performed. During the experiment the rabbits did not show any sign of inflammation, the histological analyzes highlighted the epithelialization of the ocular surface.This project is in collaboration with OSF association for the development of the clinical use of MAHL responding in particular to humanitarian needs (Cameroon). Our study of HAM mapping also showed that variability in terms of amount of protein exists between different donors. We have also shown that the spongy layer is an important source of important growth factor in the healing process of corneal ulcers.
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Submitted on : Tuesday, January 28, 2020 - 12:13:47 PM
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Parvin Sadat Nazari Hashemi. Analyse protéomique et propriétés de ré-épithélialisation des membranes amniotiques humaines en vue d'une greffe de la surface oculaire. Organes des sens. Normandie Université, 2019. Français. ⟨NNT : 2019NORMR085⟩. ⟨tel-02457769⟩