Background Extracellular vesicles (EVs) can freely cross the blood-brain barrier (BBB). With the tumor development, BBB is destroyed and its permeability increases. We aim to clarify the influences of Nmdar2a+ EVs on antitumor immunity.

Materials and methods We established the in-vitro BBB system using the co-culture of mouse brain microvascular endothelial cells and astrocytes, and identified Nmdar2a as a marker of the neuron-derived plasma EVs. We detected neuron-derived EVs (Nmdar2a+ EVs), oligodendrocyte-derived EVs (CNPase+ EVs), microglia-derived EVs (CX4CR1+ EVs), and total EVs (CD9+ EVs and CD64+ EVs) using flow cytometry. We purified and identified Nmdar2a+ EVs using immunomagnetic beads.

Results We found that tumor mouse serum can significantly inhibit BBB transepithelial resistance TEER in vitro (Figure 1A). The Evans blue dye reinfused into the tail vein can enter more into the brain tissue (Figure 1B). The levels of Nmdar2a+ EVs gradually increased with tumor progression, while the levels of CNPase+ EVs and CX4CR1+ EVs did not change significantly(Figure 1C). Both the Ctrl/B-EVs and Tumor/B-Evs  could promote tumor progression. The average particle diameters of B-EVs and Nmdar2a+ EVs were 154.4 ± 40.7 nm and 154.6 ± 47.8 nm, respectively (Figure 2C). Nmdar2a+ EVs contained higher levels of Nmdar2a compared with B-EVs (Fig. 2D). The Nmdar2a+ EVs promoted tumor growth significantly stronger than B-EVs (Fig. 2E).

Conclusion With the increase of BBB permeability, the neuron-derived Nmdar2a+ EVs increased progressively in the peripheral blood of tumor mice and inhibit antitumor immunity through the neurotransmitters (NTs) they carry.