Briefly, 2 L of a 5% solution of fluorogold in PBS was injected into the superior colliculus of mice immobilized in a stereotaxic apparatus

Briefly, 2 L of a 5% solution of fluorogold in PBS was injected into the superior colliculus of mice immobilized in a stereotaxic apparatus. wild-type mice. Both pattern-ERGs (42%, p?=?0.03) and RGC numbers (37%, p?=?0.0001) were reduced in h3T-A2 mice when compared with wild-type mice. The level of CD3 expression was increased in h3T-A2 mice (h3T-A2: 17427% vs. HLA-A2: 100%; p?=?0.04). The levels of effector cytokine IFN- were also increased significantly in h3T-A2 mice (h3T-A2: 18911% vs. HLA-A2: 100%; p?=?0.023). Both CD3 and IFN- immunostaining were increased in nerve fiber (NF) and RGC layers of h3T-A2 mice. In addition, we have seen a robust increase in GFAP staining in h3T-A2 mice (mainly localized to NF layer), which was substantially reduced in IFN- (-/-) knockout h3T-A2 mice. We also have seen an up-regulation of caspase-3 and -9 in h3T-A2 mice. Based on our data we conclude that h3T-A2 transgenic mice exhibit visual defects that are mostly associated with the inner retinal layers and RGC function. This novel h3T-A2 transgenic mouse model provides opportunity to understand RGC pathology and test neuroprotective strategies to rescue RGCs. Introduction Retinal ganglion cell (RGC) death is a common event in numerous retinopathies and optic neuropathies including glaucoma. Several theories have been proposed for RGC death; however; the pathological process 2-NBDG for RGC death has remained poorly defined. Numerous factors have been identified that contribute directly or indirectly in the RGC death process. These factors include: biomechanical stress (e.g., elevated Mouse monoclonal to CD16.COC16 reacts with human CD16, a 50-65 kDa Fcg receptor IIIa (FcgRIII), expressed on NK cells, monocytes/macrophages and granulocytes. It is a human NK cell associated antigen. CD16 is a low affinity receptor for IgG which functions in phagocytosis and ADCC, as well as in signal transduction and NK cell activation. The CD16 blocks the binding of soluble immune complexes to granulocytes intraocular pressure), oxidative stress, neuroinflammation, alteration in neurotrophic signaling, excitotoxicity, protein misfolding, glial activation, mitochondrial dysfunction, hypoxia/ischemia, genetic mutation, and auto-immunity [1]C[5]. RGCs are highly vulnerable in numerous retinopathies [6]C[10], but there is no effective therapy to prevent/delay RGC death 2-NBDG in such retinopathies where RGCs are at high risk. Thus, a better understanding of the complex network of RGC death 2-NBDG mediators is needed. Although eyes are arguably the most vulnerable, but also the most immune privileged organ; paradoxically, eyes remain subject to destructive autoimmunity that may result after inflammatory reaction triggered by environmental (microbial, stress) and autologous (tissue damage) danger signals [11]. The healthy eye is sequestered behind an efficient blood-retina barrier to the entry of unwanted molecules, while remaining under a profoundly immunosuppressive microenvironment [12], [13]. However, under certain pathological conditions these barriers and the protective microenvironment can be compromised and certain non-tolerant T cells can infiltrate the retina. The discovery of activated T cells and macrophages in the brain parenchyma of neurodegenerative disease confirms the involvement of a cellular immune response within the central nervous system 2-NBDG [14]C[16]. While some studies have also suggested that immune cells (particularly T cells) may play key roles in RGC death, a precise role for T cells in retinopathies remains poorly defined. Our data herein provides evidence for a pathological role for T cells and a cytokine (interferon-gamma, IFN-) in RGC death in a spontaneously depigmentating T cell receptor (TCR) transgenic mouse model h3T-A2 [17], [18]. Our data suggests that the activation of tyrosinase epitope YMDGTMSQV reactive TCR transgenic T cells triggers the release of inflammatory cytokines and thereby initiates neurodegenerative responses. The availability of this unique mouse model provided us an opportunity to understand the cellular events in which T cells and IFN- play crucial roles in deciding the fate of RGCs. Materials and Methods Animals C57BL/6 mice, HLA-A2, and h3T-A2 mice (11C13 months-of-age; 30C40 grams) were used in this study. Mice were kept under a cycle of 12-hours light and 12-hours dark for all the studies. Animal handling was performed in accordance with the Association for Research in Vision and Ophthalmology Statement for the Use of Animals in Ophthalmic and Vision Research; and the study protocol was approved by the Animal Care and Use Committee at the Medical University of South Carolina. The development of h3T-A2 mice has been described previously [17]. Effector-molecule deficient h3T-A2 mice were obtained by crossing h3T-A2 mice with TNF–/- (Stock No. 003008), IFN–/- (Stock No..