Lee Gyeong Won, Koo KyoIck, Sung Soo-Eun, Kim Young-In, Seo Min-Soo, Park Wook-Tae, Yang Seung Yun, Lee Gun-Woo
Department of Biomaterials Science (BK21 Four Program), Pusan National University, Miryang, 50463, South Korea.
Preclinical Research Center, Daegu-Gyeongbuk Medical Innovation Foundation (K-MEDI hub), Dong-gu, Daegu, 41061, South Korea.
Biomed Microdevices. 2025 Jul 10;27(3):35. doi: 10.1007/s10544-025-00761-2.
Extracellular vesicles (EVs) are nanosized particles secreted by most cells for information transmission, which affects the microenvironment. EVs are known to follow the characteristics and conditions of their mother cells and have attracted considerable attention for disease diagnosis and therapeutic effects. In particular, mesenchymal stem cell (MSC)-derived EVs have shown potential for facilitating regenerative wound healing, modulating immune responses, and inhibiting inflammatory diseases. However, previous isolation methods demonstrated limited EV yield, purity, and filter capacity. Here, we report a two-step tangential flow filtration (TFF) system using track-etched membranes with uniform cylindrical nanopores for effectively isolating EVs with high purity and yield. Using two different uniform nanoporous track-etched membranes (50 and 200 nm), only the particles in the small EV (sEV) size range were separated through a size-exclusion mechanism. Comparative analysis with the existing ultrafiltration membrane-based TFF system revealed that the nanoporous membrane-based TFF (Nano-TFF) system exhibited a separation efficiency (yield) exceeding twofold, achieving sEVs purity surpassing 90%. The efficacy of the highly purified sEVs was validated by incorporating them into wound dressing material and applying them to a wound animal model. Notably, the sEVs-loaded wound dressing group demonstrated enhanced wound recovery compared to control groups. The Nano-TFF system, which provides precise separation and high efficiency, can be applied to separate various bioactive agents, including sEVs, that require high-purity isolation.
细胞外囊泡（EVs）是大多数细胞分泌的纳米级颗粒，用于信息传递，影响微环境。已知EVs具有其母细胞的特征和条件，并在疾病诊断和治疗效果方面引起了相当大的关注。特别是，间充质干细胞（MSC）衍生的EVs已显示出促进再生性伤口愈合、调节免疫反应和抑制炎症性疾病的潜力。然而，以前的分离方法显示出EVs产量、纯度和过滤能力有限。在这里，我们报告了一种两步切向流过滤（TFF）系统，该系统使用具有均匀圆柱形纳米孔的径迹蚀刻膜，以有效地分离出高纯度和高产量的EVs。使用两种不同的均匀纳米多孔径迹蚀刻膜（50和200纳米），只有小细胞外囊泡（sEV）尺寸范围内的颗粒通过尺寸排阻机制被分离出来。与现有的基于超滤膜的TFF系统进行比较分析表明，基于纳米多孔膜的TFF（Nano-TFF）系统的分离效率（产量）超过两倍，实现了sEVs纯度超过90%。通过将高度纯化的sEVs掺入伤口敷料材料并将其应用于伤口动物模型，验证了其功效。值得注意的是，与对照组相比，装载sEVs的伤口敷料组显示出伤口恢复增强。提供精确分离和高效率的Nano-TFF系统可应用于分离各种需要高纯度分离的生物活性剂，包括sEVs。
