Richard S. Ruiz Department of Ophthalmology and Visual Science, University of Texas Medical School at Houston, Houston, Texas 77030, USA.
Invest Ophthalmol Vis Sci. 2010 Feb;51(2):952-9. doi: 10.1167/iovs.09-4331. Epub 2009 Sep 24.
To establish the identity of a prominent protein, approximately 70 kDa, that is markedly increased in the retina of monkeys with experimental glaucoma compared with the fellow control retina, the relationship to glaucoma severity, and its localization in the retina.
Retinal extracts were subjected to 2-D gel electrophoresis to identify differentially expressed proteins. Purified peptides from the abundant 70 kDa protein were analyzed and identified by liquid chromatography/mass spectrometry/mass spectrometry (LC/MS/MS) separation, and collision-induced dissociation sequencing. Protein identity was performed on MASCOT (Matrix Science, Boston, MA) and confirmed by Western blot. The relationship between the increase in this protein and glaucoma severity was investigated by regression analyses. Protein localization in retina was evaluated by immunohistochemistry with confocal imaging.
The abundant protein was identified as Macaca mulatta serum albumin precursor (67 kDa) from eight non-overlapping proteolytic fragments, and the identity was confirmed by Western blot. The average increase in retinal albumin content was 2.3 fold (P = 0.015). In glaucoma eyes, albumin was localized to some neurons of the inner nuclear layer, in the inner plexiform layer, and along the vitreal surface, but it was only found in blood vessels in control retinas.
Albumin is the abundant protein found in the glaucomatous monkey retinas. The increased albumin is primarily localized to the inner retina where oxidative damage associated with experimental glaucoma is known to be prominent. Since albumin is a major antioxidant, the increase of albumin in the retinas of eyes with experimental glaucoma may serve to protect the retina against oxidative damage.
鉴定一种在实验性青光眼猴子的视网膜中明显增加的、约 70kDa 的显著蛋白的身份,比较其与同窝对照视网膜的关系,以及其在视网膜中的定位。
视网膜提取物进行二维凝胶电泳,以鉴定差异表达的蛋白质。从丰富的 70kDa 蛋白中纯化的肽段通过液相色谱/质谱/质谱(LC/MS/MS)分离和碰撞诱导解离测序进行分析和鉴定。蛋白质鉴定在 MASCOT(马萨诸塞州波士顿的 Matrix Science)上进行,并通过 Western blot 进行确认。通过回归分析研究这种蛋白质的增加与青光眼严重程度之间的关系。通过免疫组织化学和共聚焦成像评估蛋白质在视网膜中的定位。
该丰富的蛋白被鉴定为猕猴血清白蛋白前体(67kDa),来自 8 个非重叠的蛋白水解片段,其身份通过 Western blot 得到确认。视网膜白蛋白含量平均增加 2.3 倍(P = 0.015)。在青光眼眼中,白蛋白定位于内核层的一些神经元、内丛状层和玻璃体表面,但仅在对照视网膜的血管中发现。
白蛋白是在青光眼猴子视网膜中发现的丰富蛋白。增加的白蛋白主要定位于内视网膜,实验性青光眼相关的氧化损伤在此处较为明显。由于白蛋白是一种主要的抗氧化剂,因此实验性青光眼眼中的视网膜中白蛋白的增加可能有助于保护视网膜免受氧化损伤。