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PIERRE COLOMBE, Deepti Singh, Myron Spector, Michael J Young; 3D hydrogels protect human retinal progenitor cells from stress exerted during transplantation. Invest. Ophthalmol. Vis. Sci. 2019;60(9):3328.
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© ARVO (1962-2015); The Authors (2016-present)
The current challenges faced in cell therapy is a lack of approved polymeric cell carriers that could aid in delivery of cells and would enhance cell survival and degrade within an acceptable time frame without triggering an immune response from the host. This study is aimed at exploring the use of a hydrogel as a “protective envelope” for cells that experience high stress exerted during transplantation by doing, first, an invitro study to replicate the conditions of transplantation and, secondly, an invivo study to confirm those results.
Human retinal progenitor cells (hRPCs) were cultured in different condition on a 2D layer until reaching confluence (PBS with endothelial growth factor (EGF), cells encapsulated in gel with PBS and EGF). We have developed a gelatin-hydroxyphenyl propionic acid (GA-HPA) hydrogel with a gelation time of 20 seconds. To check viability, proliferation, apoptosis, stemness, retinal progenitor and cones markers immunohistostaining and flow cytometry were performed. Live and dead assay were used to study viability, Caspase 9 and Ki67 were used for apoptosis and proliferation. Finally flow cytometry was performed using MACSQuant to study stemness, retinal progenitor and cones markers. The Invivo study was performed on Long Evans rats by injecting in the subretinal space different conditions (hRPCs alone, cells encapsulated in gel with or without EGF).
Live-dead assay shows cells encapsulated in gel have higher viability in comparison with non-gel cultures. Cells cultured with gel show higher viability and proliferation in comparison to the monolayer culture. Apoptosis quantification using CAS9 clearly indicated the protective properties of the gel. Flow cytometry assay shows cells encapsulated in gel express higher percentage of stemness and retinal progenitor markers. More viable cells were found in the 3D culture compared to saline-delivered controls. Furthermore, the in-vivo study shows higher integration and viability of cell injected in the 3D culture after 3 days.
The invitro and invivo results with gel have shown great promise and we intend to use it in the degenerative animal model to ehance the chances of cell survival and integration for restoring vision. This study demonstrates GA-HP injected in-situ crosslinking biodegradable hydrogel has the potential to be used as cell carrier for in-vivo delivery of stem and progenitor cells.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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