Ubiquitin carboxyl-terminal esterase L1 promotes proliferation of human choroidal and retinal endothelial cells
Ashander, Liam M
Smith, Justine R
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Purpose: We aimed: (1) to establish endothelial expression of ubiquitin carboxyl-terminal esterase L1 (UCHL1) in human choroid and retina and; (2) to investigate a role for UCHL1 in basic processes involved in intraocular neovascularization. Design: Controlled translational experimental study. Methods: Ethanol-fixed human choroid and retina (n = 3 eyes) were indirectly immunostained with rabbit anti-human UCHL1 antibody. Endothelial proliferation and migration assays were performed using cultured human choroidal and retinal endothelial cells (n = 6 isolates/assay). Cells were transfected with UCHL1-targeted or non-targeted small interfering (si)RNA and a commercially available transfection system, and used 48 hours later in experiments. Cell proliferation was evaluated using an assay in which cellular DNA was fluorescently tagged for quantification by microplate reader. Cell migration was examined in an assay that involved counting the number of endothelial cells moving across a perforated membrane. Transcript silencing was verified by Western blot for all assays. Results: Immunohistochemistry confirmed expression of UCHL1 by endothelium in human choroid and retina in vivo. UCHL1-specific knockdown resulted in significantly less proliferation (p G 0.0001) for 3 human choroidal endothelial isolates and 3 human retinal endothelial isolates, and significantly less migration (p e 0.016) for 2 of 3 human choroidal endothelial isolates and 1 of 3 human retinal endothelial isolates. Conclusions: Our results suggest that UCHL1 may be involved in choroidal and retinal endothelial proliferation in most persons, and endothelial migration in some persons. UCHL1 may be a suitable target for a new treatment of intraocular neovascularisation.