Diversity and Heterogeneity of Extracellular Vesicles and Brain-Penetrating Amembranous Non-Vesicular Nanoparticles in Glioblastoma

Advances in understanding intercellular communication through extracellular vesicles (EVs) and non-vesicular extracellular nanoparticles (NVEPs) have been limited by their cellular heterogeneity and unclear origins. Using sequential ultracentrifugation, we identified high- and low-density EVs and performed multi-omics analyses on four EV/NVEP categories, including supermeres. Our findings reveal that extracellular RNA, RNA-binding proteins, and other cellular proteins are differentially expressed among EVs and NVEPs. Notably, supermeres can penetrate the blood-brain barrier, in a manner dependent on their intact RNA and protein structure, and distribute throughout the CNS. Supermeres preferentially interact with microglia at higher levels than other CNS cell types. These interactions lead to decreased TGFß expression, suggesting an immunomodulatory role. This study highlights supermeres as a promising platform for CNS therapeutic delivery. Overall design: Total RNA were collected from 1) cultured cells, and 2) large extracellular vesicles (LEV), 3) small extracellular vesicles (SEV), 4) large nanoparticles (exomeres), and 5) small nanoparticles (supermeres) secreted by the cells. The collection were done in 4 series (A, C, D, E). RNA-seq were performed in two batches (A series and others). In total there are 20 samples.