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.2014 May;47(5):292-7.
doi: 10.5483/bmbrep.2014.47.5.193.

Increased 26S proteasome non-ATPase regulatory subunit 1 in the aqueous humor of patients with age-related macular degeneration

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Increased 26S proteasome non-ATPase regulatory subunit 1 in the aqueous humor of patients with age-related macular degeneration

Hyungwoo Lee et al. BMB Rep.2014 May.

Abstract

Age-related macular degeneration (AMD) is the leading cause of blindness in the world. Evidence indicates that the suppression of the ubiquitin-proteasome system (UPS) contributes to the accumulation of toxic proteins and inflammation in retinal pigment epithelium (RPE), the functional abnormalities and/or the degeneration of which are believed to be the initiators and major pathologies of AMD. To identify new protein associations with the altered UPS in AMD, we used LC-ESI-MS/MS to perform a proteomic analysis of the aqueous humor (AH) of AMD patients and matched control subjects. Six UPS-related proteins were present in the AH of the patients and control subjects. Four of the proteins, including 26S proteasome non-ATPase regulatory subunit 1 (Rpn2), were increased in patients, according to semi-quantitative proteomic profiling. An LC-MRM assay revealed a significant increase of Rpn2 in 15 AMD patients compared to the control subjects, suggesting that this protein could be a biomarker for AMD.

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Figures

Fig. 1.
Fig. 1.. (A) Box plots of results from LC-MRM analysis in an independent sample set of 15 AH samples of AMD before and after treatment with ranibizumab and 15 matched AH samples of control. The relative abundance of Rpn2 was elevated in patient samples compared to control samples and was decreased after ranibizumab treatment, as determined by a t-test analysis. The relative level of Rpn2 was adjusted by setting the level of the matched control to a value of 1. (B) ROC curve of Rpn2. The AUC value was 0.610 (95% CI, 0.505-0.708).
Fig. 2.
Fig. 2.. Rpn2 secretion is increased by oxidative stress compared to control cultures in ARPE-19 cells. (A) ARPE-19 cells were cultured with or without 400 μM paraquat for 24 hours. Equal amounts of proteins (15 μg) from conditioned media (CM) were subjected to Western blot with an Rpn2 antibody. Rpn2 levels in the CM treated with paraquat increased compared to the CM from control cultures. Ponceau S staining was used as a loading control for CM. (B) Cell lysates from the ARPE-19 cells that produced the CM (15 μg) were immunoblotted with Rpn2. There was no significant difference between the control and paraquat lane. α-Tubulin was the loading control.
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