Mayberry, Jordan1,2,3; Xian, Shuyu1,2; Kuehn, Markus, H.1,2,3
1Department of Ophthalmology and Visual Sciences, University of Iowa; 2VA Center for the Prevention and Treatment of Visual Loss, Iowa City; 3Interdisciplinary Graduate Program in Genetics, University of Iowa
Purpose: Glaucoma is a neurodegenerative disease that causes loss of retinal ganglion cells (RGC), and subsequently, loss of vision. Previous studies in our lab have demonstrated that adoptive transfer of CD3+ cells from animals with glaucoma into healthy recipients results in progressive loss of RGC, suggesting a functional role of adaptive immunity in the pathology of the disease. A crucial first step in this process is the extravasation of T-cells into the immune-privileged retina. This study was conducted to determine if elevated IOP facilitates extravasation using donor mice with a fixed T-cell receptor against GFP (Just EGFP Death Inducing, JEDI) and Thy1-GFP+ recipient mice that exhibit GFP expression in a subset of RGC.
Methods: Mild IOP elevation was induced in the eyes of Thy1-GFP+ mice through intracameral injection of Ad5.MyocY437H (n=12). Thy1-GFP+ and Thy1-GFP- control animals received intracameral injections of Ad5.empty (n=11-12/per group). Following injection, IOP was monitored by rebound tonometry and GFP positive cells were imaged by fundoscopy for 7 weeks. 4 weeks following injection, spleens from donor JEDI mice were harvested, homogenized, and transferred into recipient Thy1-GFP mice. At week 4.5 following intracameral injection, one group received an intraperitoneal injection of lipopolysaccharide (LPS). At the conclusion of the experiment, retinas were collected, whole-mounted, stained using antibodies directed against CD3+ T cells, and visualized using confocal and widefield fluorescence microscopy.
Results: Following intracameral injection and adoptive transfer of JEDI splenocytes, all mice experienced some degree of RGC loss on fundus imaging. Losses are similar in mice that did not receive splenocytes (-4.14%), those that did receive splenocytes but without elevated IOP (-2.83%), and those receiving splenocytes and LPS (-4.5%). In contrast, mice receiving splenocytes and experienced elevated IOP lost significantly more RGC (-18.43%, p=0.0062). In these animals RGC loss was particularly noticeable in the central retina rather than in the periphery. Detectable levels of retinal CD3+ cells were low and similar in all groups. However, the size of these cells in the retinas of mice with elevated IOP appeared 5-15 μm larger than those observed in all other groups.
Conclusions: Slight elevation in IOP facilitates T cell mediated degradation of RGC. LPS-mediated weakening of the blood-retina barrier by itself is insufficient to achieve RGC loss, suggesting that additional IOP-mediated mechanisms are required to overcome retinal immune privilege.
Last Updated: