Fei, Mengdi

Abstract Aberrant vasculature is a hallmark of glioblastoma (GBM), contributing to both blood–brain barrier (BBB) dysfunction and frequent intracranial hemorrhage. Myeloid cells, including microglia and macrophages, are known to co-opt tumor vasculature during tumor progression, but the nature and consequences of their physical interactions with blood vessels require clearer definition. To address this, we characterize vessel-associated myeloid cells in murine gliomas with high spatial and temporal resolution, leveraging in vivo two-photon imaging, MRI, and transgenic mouse lines. To investigate the role of microglia in glioma vasculature, we pretreated glioma-bearing mice with CSF1R inhibitors (PLX3397 & PLX5622) to ablate microglia prior to tumor inoculation. Strikingly, over half of the microglia-depleted mice developed acute, lethal paralysis between days 11–15, whereas control tumor-bearing mice remained asymptomatic. MRI revealed extensive intracranial hemorrhage and edema, and two-photon imaging showed nodular, abnormal vasculature. This vascular dysfunction led to a dramatic reduction in median survival—from 30 days in control glioma-bearing mice to 15 days in PLX-treated mice with gliomas. To identify the protective microglial population involved, we examined vascular–myeloid interactions. We found that the peritumoral region harbored dense capillary networks supporting tumor expansion, along with a marked increase in myeloid cells with somas directly contacting blood vessels. Using TMEM119CreER/+:R26tdTomato/+ mice, we confirmed these vessel-associated cells as brain-resident microglia, not infiltrating macrophages. Quantitatively, ~60% of peritumoral microglia were vessel-associated, compared to ~30% in naïve or contralateral hemispheres. These microglia displayed elongated somata and processes aligned with vessels or wrapping around vessels, which is a phenotype also observed following single-vessel photothrombosis, suggesting a response to impaired blood flow and BBB leakage. Our findings uncover a critical neurovascular role for vessel-associated microglia in maintaining vascular integrity during glioma progression and highlight the importance of preserving, reprogramming, and further investigating this population in GBM therapies.