Ultrahigh-Uniformity Nanopore Size Filter for Extracellular Vesicle Isolation and in Vitro Dermatological Assessment

Abstract

Extracellular vesicles (EVs), including exosomes, are abundant in bovine milk and Lactobacillus culture media but difficult to isolate with high efficiency and purity. In response, a micro-electro-mechanical systems (MEMS)-based membrane filter was developed to address these limitations. Under equivalent conditions, the developed filter outperformed commercial filters, achieving up to 66% recovery of EVs, as confirmed by nanoparticle tracking analysis. The enhanced performance is likely due to the formation of uniform 168 nm diameter pores on a 318 nm thick membrane, which reduces fouling as confirmed by blocking-model assessments. Biological evaluations showed that EVs isolated using the developed filter retained notable purity and bioactivity. Specifically, milk-derived EVs increased the proliferation of human fibroblasts (Hs68) and human follicle dermal papilla cells (HFDPCs) by up to 25% and 50%, respectively, while Lactobacillus-derived EVs increased proliferation by up to 11% and 53% at certain concentrations. Furthermore, co-treatment with an anti-aging peptide (AIMP1-derived peptide) had a synergistic effect on both cell types. Similar trends were seen in canine and feline fibroblasts. Milk-derived EVs boosted proliferation by up to 25% in canine and 31% in feline cells, while Lactobacillus-derived EVs increased it by up to 46% and 34%, respectively. These results show the filter’s potential for large-scale EVs isolation and dermatological applications, requiring high purity and yield.

Note:
Funding Information: This work was supported by the Technology Innovation Program (Materials & Components Technology Development Program) (20023179, Development of a 150nm-Scale Mass-Production Filtering System for Functional Cosmetic Ingredients Based on Raw Milk-Derived Micro-vesicles) funded by the Ministry of Trade Industry & Energy (MOTIE, Korea) and also supported by Brain Pool program funded by the Ministry of Science and ICT through the National Research Foundation of Korea (RS-2024-00405682). In addition, a Lactobacillus strain provided by SHEBAH BIOTECH was applied in this study.

Conflict of Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Keywords: Extracellular vesicles, MEMS-based membrane filter, Blocking filtration model, Skin regeneration