Comparison of Hydrostatic Filtration Dialysis with Ultracentrifugation Methods for the Identification and Proteomic Profiling of Urinary Extracellular Vesicles
Comparison of Hydrostatic Filtration Dialysis with Ultracentrifugation Methods for the Identification and Proteomic Profiling of Urinary Extracellular Vesicles
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DOI：
10.7754/clin.lab.2018.180922
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发表时间：
2019-01-01
影响因子：
0.7
通讯作者：
Zou, Hequn
中科院分区：
文献类型：
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作者：
Chen, Yunying;Hong, Guobao;Zou, Hequn
背景资料：尿细胞外囊泡（UEVs）携带丰富的标记物，他们的父母细胞，所以他们可以作为可能的生物标志物的肾脏疾病。方法：在这项研究中，我们分离尿细胞外囊泡从5个人使用一个简单的，临床上适用的方法称为静水过滤透析（HFD），并比较它的金标准超滤（UC）与透射电子显微镜（TEM）。我们还采用了一种蛋白质组学方法，使用来自相同五个个体的合并人尿液样本来分析UEV的蛋白质组成，以评估这两种方法的有效性。值得注意的是，使用TEM，我们发现所有分离物都含有具有传统报道形态的0 - 400 nm囊泡，尽管TEM结果显示从HFD分离的UEV比从UC分离的UEV更大和更广泛。两种方法共获得2,564个UEV蛋白。我们显示了通过两种分离方法鉴定的蛋白质之间的大重叠（2，185> 85%）。结果还表明，所获得的蛋白质的细胞外囊泡，这是用这些方法分离，与目前可用的数据库中的结果一致。然而，在相关的基因本体，富集的蛋白质发现这两种方法显示出一定的differentiation.Conclusions：HFD方法在临床上是可行的，并允许大规模的蛋白质谱UEV生物标志物。本研究的结果也提供了有价值的UEV蛋白质数据的方法学比较，这可能是有价值的其他研究人员。
Background: Urinary extracellular vesicles (UEVs) carry rich markers of their parent cells, so they can serve as possible biomarkers of kidney diseases.Methods: In this study, we isolated urinary extracellular vesicles from five individuals using a simple, clinically applicable method called hydrostatic filtration dialysis (HFD) and compared it to the gold-standard ultracentrifugation (UC) with transmission electron microscopy (TEM). We also employed a proteomic approach using pooled human urine samples from the same five individuals to profile the protein composition of UEVs to evaluate the effectiveness of these two methods.Results: Notably, using TEM, we found that all isolations contained 0 - 400 nm vesicles with the traditionally reported morphology, although the TEM results showed that the UEVs isolated from HFD compared to those from UC are larger and more extensive. We obtained a total of 2,564 UEV proteins in the two methods. We showed a large overlap (2,185 > 85%) between the proteins identified by both isolation methods. The result also showed that the obtained proteins in extracellular vesicles, which are isolated with these methods, are consistent with the results in currently available databases. However, in the associated gene ontologies, the enriched proteins found by the two methods showed some differences.Conclusions: The HFD method is clinically feasible and allows large-scale protein profiling of UEV biomarkers. The results of this study also provide valuable UEV protein data from the methodological comparison, which might be valuable to other researchers.