Abstract
Purpose::
One of the established roles for the immunoproteasome is the generation of antigenic peptides. However, its substantial expression in the immune-privileged retina implies other non-immune functions are possible. This study investigates a putative role for immunoproteasome in protecting the retina from oxidative stress.
Methods::
The two models of oxidative stress that were investigated include: cells from CuZn superoxide dismutase (SOD) (+/-) knock-out mice and cells chronically exposed to a sublethal dose of H2O2. Interferon-gamma (IFN) was used to specifically upregulate immunoproteasome. Cultured retinal pigment epithelial (RPE) cells from wild-type and CuZn SOD mice were treated with IFN or sublethal doses of H2O2. Proteasome subunit composition was analyzed by Western immunoblotting at intervals following a treatment regimen. RPE cell viability, after preconditioning with either H2O2 or IFN, was evaluated following an acute dose of oxidant. To directly test immunoproteasome’s ability to degrade oxidized proteins compared to constitutive proteasome, 20S proteasome was purified from IFN-treated and untreated RPE cells.
Results::
Under normal culture conditions, CuZn SOD RPE cells contain more immunoproteasome compared with wild-type RPE cells. Following a single exposure to IFN or chronic exposure to sublethal doses of H2O2 total proteasome content did not change. However, treatment caused immunoproteasome upregulation with a corresponding down-regulation in constitutive proteasome subunits. Cells with upregulated immunoproteasome produced by preconditioning with either H2O2 or IFN exhibited greater cell viability when exposed to an acute dose of oxidant compared with untreated cells.
Conclusions::
In both models of chronic oxidative stress (RPE cells from CuZn SOD knock-out mice and RPE preconditioned with H2O2) immunoproteasome expression is upregulated. RPE cells containing more immunoproteasome exhibit greater resistance to an acute dose of oxidant. These results suggest that immunoproteasome plays a role in protecting cells from oxidative stress.
Keywords: retinal pigment epithelium • oxidation/oxidative or free radical damage • cell survival