Loss of endothelial ALK1 signaling induces the emergence of a KIT+ angiogenic endothelial cluster driving brain arteriovenous malformations
Abstract
Background
Hereditary Hemorrhagic Telangiectasia type 2 (HHT2) is a genetic disorder caused by mutations in theALK1(ACVRL1) gene, encoding a receptor for Bone Morphogenetic Proteins 9 and 10 (BMP9/BMP10). HHT2 patients frequently develop brain arteriovenous malformations (bAVMs), which are abnormal connections between arteries and veins. Currently, surgical resection is the only treatment, associated with significant risks and complications. Despite evidence suggesting endothelial cell (EC) heterogeneity in bAVMs, it remains poorly characterized, limiting our ability to identify new therapeutic avenues.
Methods
We employed endothelial cell-specific and inducibleAlk1knockout mice (Alk1iECKO) with tamoxifen-induced deletion at postnatal day 6 (P6). We separately analyzed the P8 perineural (PNVP) and intraneural (INVP) vascular plexuses, which differ in vessel composition and flow dynamics. Single-cell RNA sequencing (scRNAseq) was performed to characterize EC heterogeneity and identify transcriptomic changes in both vascular plexuses of mutant versus wild type mice.
Results
Loss of endothelial ALK1 signaling triggered bAVM formation predominantly in the PNVP vascular network. scRNAseq revealed thatAlk1deletion promoted brain capillaries differentiation into angiogenic-1 ECs, whereas it drove PNVP venules EC proliferation and the emergence of the unique angiogenic-2 cluster. The latter shares transcriptomic features with human AVM ECs, including angiogenic tip cell markers and a strong glycolytic signature. Among its defining markers,Kitemerged as a direct downstream target of BMP9-ALK1 signaling. Pharmacological KIT inhibition using Masitinib, Imatinib, or KIT-blocking antibodies prevented bAVM formation inAlk1iECKOmice.
Conclusion
Our study uncovers a previously unrecognized EC population, the angiogenic-2 cluster, as a key contributor to bAVM development. We identifyKitas a central regulator of this cluster, establishing it as a promising therapeutic target for preventing bAVMs in HHT2.
Clinical Perspective
What is new?
Using endothelial-specificAlk1knockout mouse models and single-cell transcriptomics, we identified a novel angiogenic endothelial cell population as a key driver of brain AVM formation.
This angiogenic EC cluster shares molecular features with human bAVM cells, including high expression ofKIT, which we identified as a new direct transcriptional target of BMP9-ALK1 signaling.
Pharmacological inhibition of KIT using small molecules or blocking antibodies effectively prevents AVM formationin vivo, establishing KIT as a promising therapeutic target.
What are the clinical implications?
Our findings support the therapeutic potential of targeting KIT with FDA-approved drugs such as Imatinib to treat HHT2-associated brain AVMs, offering a non-invasive alternative to surgical intervention.
Characterizing AVM-specific endothelial subtypes may enable the development of targeted and personalized therapies, improving patient outcomes and minimizing treatment-associated risks in HHT.
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