Harvest Time-Dependent Changes in the Composition and Function of Microglia-Derived Small Extracellular Vesicles

作者：Muhammad Waqas Salim, Wei Zhang, Su Su Thae Hnit, Karthik Shantharam Kamath, Lyndsey E. Collins‐Praino, Andrew Care, Yuling Wang · 发表于：ACS Biomaterials Science & Engineering · 年份：2025 · DOI：10.1021/acsbiomaterials.5c00791 · 被引用次数：4 · 研究领域：Extracellular vesicles in disease、Neuroinflammation and Neurodegeneration Mechanisms、RNA Interference and Gene Delivery

Microglia are dynamic macrophage-like cells that survey the central nervous system (CNS) and mediate neuroinflammatory responses. Even under homeostatic conditions, they exhibit phenotypic plasticity in response to environmental cues. Microglia-derived small extracellular vesicles (sEVs), which are lipid bilayer vesicles encapsulating proteins, lipids, and nucleic acids, reflect the physiological state of their parent cells and influence recipient cell function. Although increasing attention has been given to their roles in neuroinflammation, neurodegeneration, and intercellular communication, the impact of experimental variables, particularly harvest timing, on microglial sEV composition remains poorly understood. This study investigated how different harvest durations (24, 48, and 72 h) influence the molecular and functional properties of sEVs released by BV2 microglial cells. sEVs were isolated via ultracentrifugation and characterized for particle size, yield, RNA and protein content, surface marker expression, and proteomic profile. While sEV size and morphology remained consistent across time points, particle yield and RNA content increased significantly with prolonged harvest, peaking at 72 h. Classical sEV markers (CD9, CD81, TSG101) were consistently expressed, suggesting stable vesicle identity. However, proteomic profiling revealed a time-dependent shift in cargo, with an increase in unique proteins and enrichment of stress- and neurodegeneration-related pathways a...