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This abstract is assigned to session FRO: Fund for Research in Ophthalmology

Title	FRO: Identification of the gene signature of retinal endothelial cells during classical experimental autoimmune uveitis, Th1- and Th17-dependent uveitis
Abstract Nr.	1034

Purpose	Retinal endothelial cell activation by autoreactive T cells plays a critical role in non infectious uveitis development. The aim of this work is to identify key genes regulated in endothelial cells during experimental autoimmune uveitis (EAU).
Methods	Transgenic FVB/N-Tie2-GFP mice, in which vascular endothelial cells express GFP, are backcrossed with EAU-susceptible C57Bl/6 mice. The expression of GFP is characterized by immunofluorescence and FACS in order to validate the animal model. Retinal endothelial cells are isolated from dissociated retinas by FACSAria and their RNA purified for microarray analysis.
Results	Four generations of transgenic C57BL/6-Tie2-GFP mice have already been successfully generated. Immunofluorescence stainings confirm the expression of GFP on retinal endothelial cells, co-expressed with endoglin and CD31. FACS data indicate that up to 5% of dissociated retinal cells express CD31 but no GFP. Less than 1% of retinal cells are GFP+/CD31+. These cells can be efficiently sorted by FACS-Aria with an output of approximately 5.000 cells per animal. Quality RNA could be obtained from 10.000 cells in quantities compatible with microarray analysis.
Conclusion	We have validated our transgenic model, which seems to be more specific than retinal endothelial cell isolation based on CD31 expression. The next part of the project, which is the analysis of retinal endothelial cell mRNA modulation during EAU, is in progress.

Authors 1

Last name	LIPSKI
Initials	D.
Department	IRIBHM - Erasme Hospital
City	Brussels

Authors 2

Last name	Dewispelaere
Initials	R.
Department	IRIBHM - CHU Saint-Pierre
City	Brussels

Authors 3

Last name	Foucart
Initials	V.
Department	IRIBHM - CHU Brugmann
City	Brussels

Authors 4

Last name	Caspers
Initials	L.
Department	CHU Saint-Pierre
City	Brussels

Authors 5

Last name	Bruyns
Initials	C.
Department	IRIBHM
City	Brussels

Authors 6

Last name	Willermain
Initials	F.
Department	IRIBHM - CHU Saint-Pierre
City	Brussels