Directional Exosome Proteomes Reflect Polarity-Specific Functions in Retinal Pigmented Epithelium Monolayers.
Directional Exosome Proteomes Reflect Polarity-Specific Functions in Retinal Pigmented Epithelium Monolayers.
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DOI：
10.1038/s41598-017-05102-9
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发表时间：
2017-07-07
影响因子：
4.6
通讯作者：
Bowes Rickman C
中科院分区：
文献类型：
--
作者：
Klingeborn M;Dismuke WM;Skiba NP;Kelly U;Stamer WD;Bowes Rickman C
视网膜色素上皮（RPE）形成眼睛中的外部血液-视网膜屏障，并且其极性负责蛋白质、脂蛋白颗粒和细胞外囊泡（EV）的定向分泌和摄取。这种分泌分裂决定了体循环（基底外侧）和视网膜（顶端）之间的直接相互作用。我们的目标是定义从RPE释放的EV的极化蛋白质组和物理特征。猪RPE细胞的原代培养物在可渗透的支持物上分化成极化的RPE单层。从浸泡顶端或基底外侧RPE表面的培养基中分离EV，并纯化和处理小EV的两个亚群，包括外泌体和致密EV，用于蛋白质组学分析。同时，测定EV尺寸分布和浓度。使用蛋白质相关性分析质谱法，共鉴定了631种外泌体制备物中的蛋白质，其中299种独特地在顶部释放，94种独特地在基底外侧释放。通过Western印迹验证所选择的蛋白。这些外泌体和致密EV制剂的蛋白质组表明上皮极性影响定向释放。这些数据作为比较研究的基础，旨在阐明外泌体在视网膜疾病的分子病理生理学中的作用，并帮助确定潜在的治疗靶点和生物标志物。
The retinal pigmented epithelium (RPE) forms the outer blood-retinal barrier in the eye and its polarity is responsible for directional secretion and uptake of proteins, lipoprotein particles and extracellular vesicles (EVs). Such a secretional division dictates directed interactions between the systemic circulation (basolateral) and the retina (apical). Our goal is to define the polarized proteomes and physical characteristics of EVs released from the RPE. Primary cultures of porcine RPE cells were differentiated into polarized RPE monolayers on permeable supports. EVs were isolated from media bathing either apical or basolateral RPE surfaces, and two subpopulations of small EVs including exosomes, and dense EVs, were purified and processed for proteomic profiling. In parallel, EV size distribution and concentration were determined. Using protein correlation profiling mass spectrometry, a total of 631 proteins were identified in exosome preparations, 299 of which were uniquely released apically, and 94 uniquely released basolaterally. Selected proteins were validated by Western blot. The proteomes of these exosome and dense EVs preparations suggest that epithelial polarity impacts directional release. These data serve as a foundation for comparative studies aimed at elucidating the role of exosomes in the molecular pathophysiology of retinal diseases and help identify potential therapeutic targets and biomarkers.
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DOI：
10.1073/pnas.1222142110
发表时间：
2013-06-11
影响因子：
11.1
作者：
Chowanadisai, Winyoo;Graham, David M.;Messerli, Mark A.
通讯作者：
Messerli, Mark A.
影响因子：
4.4
作者：
Ablonczy, Zsolt;Dahrouj, Mohammad;Crosson, Craig E.
通讯作者：
Crosson, Craig E.
影响因子：
4.8
作者：
Gangalum, Rajendra K.;Bhat, Ankur M.;Bhat, Suraj P.
通讯作者：
Bhat, Suraj P.
影响因子：
64.5
作者：
Foster, LJ;de Hoog, CL;Mann, M
通讯作者：
Mann, M
DOI：
10.1136/bjophthalmol-2011-300344
发表时间：
2011-12
期刊：
The British journal of ophthalmology
影响因子：
--
作者：
Curcio CA;Johnson M;Rudolf M;Huang JD
通讯作者：
Huang JD