Ultrasound-Mediated BBB Opening Promotes Microglial Reprogramming and Enhanced Endothelial Interactions in the Human Peritumoral Brain [Visium]

The blood–brain barrier (BBB) restricts entry of therapeutics and shapes immune surveillance in the central nervous system, while microglia maintain barrier integrity and coordinate repair. Low-intensity pulsed ultrasound with intravenously administered microbubbles (LIPU/MB) transiently opens the BBB in patients, with restoration mostly occurring within about an hour; however, how microglia respond to LIPU/MB during this acute repair window in the human brain remains unknown. We report on intraoperative LIPU/MB studies where the peritumoral brain of glioblastoma patients was biopsied 45 minutes after sonication and analyzed, as part of phase I/II clinical trials (NCT04528680). We integrated single-cell RNA-seq, single-cell ATAC-seq, spatial transcriptomics, and multiplex immunofluorescence. We observe that LIPU/MB modulates perivascular microglia by promoting extension of processes toward the endothelium, inducing pro-inflammatory and chemoattractant transcriptional programs, and remodeling of endothelial–myeloid ligand–receptor communication. We find that microglial reprogramming is accompanied by coordinated changes in chromatin accessibility and heightened activity of the AP-1 transcription factor complex. These results reveal that LIPU/MB-based BBB opening in humans does not merely permit drug passage but reprograms resident myeloid cells to engage the vasculature and amplify innate and adaptive immune pathways, providing a mechanistic basis to leverage immune activation by LIPU/MB with immunotherapies for brain tumors. These findings also warrant consideration in the context of the pathophysiology and therapeutics of CNS diseases where there is acute BBB disruption, or where LIPU/MB is under investigation.