Comparative investigation of exosome extraction from rat bone marrow mesenchymal stem cells using three different methodologies

IF 3 3区 生物学 Q2 BIOCHEMICAL RESEARCH METHODS
Na Wang, Mingyue Yin, Jiaqi Yu, Jing Zhang, Xueli Pan
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Abstract

Exosomes have been identified as crucial mediators in numerous physiological and pathological processes, emerging as a focal point of scientific inquiry. This study aims to compare three methods for isolating exosomes from rat bone marrow mesenchymal stem cells: ultracentrifugation (UC), ultrafast separation system (EXODUS), and commercial precipitation kit (EXO-kit). First, the investigation compared exosomal morphology, particle size distribution, and expression of marker proteins. Subsequently, the RNA content, protein concentration, and purity of exosomes were evaluated. Finally, the impact of these exosomes on cellular metabolic viability and migration capacity was assessed. Results indicated that exosomes exhibited spherical or elliptical membrane structures, and most of the exosomes extracted by the three methods were in the range of 30 to 200 nm. UC-extracted exosomes demonstrated the least impurities and clearest background, followed by EXODUS-extracted exosomes, and lastly EXO-kit-extracted exosomes. The EXO-kit-extracted exosomes yielded the highest RNA and protein content, whereas those isolated through UC exhibited superior purity. Furthermore, exosomes extracted from EXODUS and EXO-kit methods effectively enhanced the metabolic viability and migratory ability of osteoblast precursor cells compared to UC-extracted exosomes. In conclusion, each of the three methodologies presents advantages and limitations. Therefore, the selection of an appropriate exosome extraction technique should be based on specific experimental objectives and requirements.

使用三种不同方法从大鼠骨髓间充质干细胞中提取外泌体的比较研究。
外泌体已被确定为众多生理和病理过程中的关键介质,成为科学研究的焦点。本研究旨在比较从大鼠骨髓间充质干细胞中分离外泌体的三种方法:超速离心法(UC)、超快速分离系统(EXODUS)和商业沉淀试剂盒(EXO-kit)。首先,研究人员比较了外泌体的形态、粒度分布和标记蛋白的表达。随后,对外泌体的 RNA 含量、蛋白质浓度和纯度进行了评估。最后,评估了这些外泌体对细胞代谢活力和迁移能力的影响。结果表明,外泌体呈现球形或椭圆形膜结构,三种方法提取的外泌体大多在30至200纳米之间。UC提取的外泌体杂质最少,背景最清晰,其次是EXODUS提取的外泌体,最后是EXO-kit提取的外泌体。EXO-kit提取的外泌体的RNA和蛋白质含量最高,而通过UC分离的外泌体的纯度更高。此外,与 UC 提取的外泌体相比,EXODUS 和 EXO-kit 方法提取的外泌体能有效提高成骨细胞前体细胞的代谢活力和迁移能力。总之,这三种方法各有优势和局限性。因此,应根据具体的实验目的和要求选择合适的外泌体提取技术。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
ELECTROPHORESIS
ELECTROPHORESIS 生物-分析化学
CiteScore
6.30
自引率
13.80%
发文量
244
审稿时长
1.9 months
期刊介绍: ELECTROPHORESIS is an international journal that publishes original manuscripts on all aspects of electrophoresis, and liquid phase separations (e.g., HPLC, micro- and nano-LC, UHPLC, micro- and nano-fluidics, liquid-phase micro-extractions, etc.). Topics include new or improved analytical and preparative methods, sample preparation, development of theory, and innovative applications of electrophoretic and liquid phase separations methods in the study of nucleic acids, proteins, carbohydrates natural products, pharmaceuticals, food analysis, environmental species and other compounds of importance to the life sciences. Papers in the areas of microfluidics and proteomics, which are not limited to electrophoresis-based methods, will also be accepted for publication. Contributions focused on hyphenated and omics techniques are also of interest. Proteomics is within the scope, if related to its fundamentals and new technical approaches. Proteomics applications are only considered in particular cases. Papers describing the application of standard electrophoretic methods will not be considered. Papers on nanoanalysis intended for publication in ELECTROPHORESIS should focus on one or more of the following topics: • Nanoscale electrokinetics and phenomena related to electric double layer and/or confinement in nano-sized geometry • Single cell and subcellular analysis • Nanosensors and ultrasensitive detection aspects (e.g., involving quantum dots, "nanoelectrodes" or nanospray MS) • Nanoscale/nanopore DNA sequencing (next generation sequencing) • Micro- and nanoscale sample preparation • Nanoparticles and cells analyses by dielectrophoresis • Separation-based analysis using nanoparticles, nanotubes and nanowires.
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