Bone Marrow-Derived Alk1 Mutant Endothelial Cells and Clonally Expanded Somatic Alk1 Mutant Endothelial Cells Contribute to the Development of Brain Arteriovenous Malformations in Mice

Homozygous mutation in arteriovenous malformation (AVM) causative genes in a fraction of endothelial cells (ECs) causes brain AVMs (bAVMs). Heterozygous mutation of an AVM causative gene, endoglin, in bone marrow (BM) caused capillary dysplasia in mouse brain. We hypothesize that homozygous mutation of activin receptor-like kinase 1 (Alk1, another AVM causative gene) in BM derived ECs (BMDECs) is sufficient to cause bAVM in brain angiogenic region, Alk1(−) ECs can clonally expand in bAVM, and the burden of Alk1(−) ECs correlates with bAVM severity. The BMs of PdgfbiCreER;Alk1(2f/2f);Ai14 mice with EC-specific tamoxifeninducible Cre and Alk1 floxed alleles were transplanted into wild-type mice to analyze the role of BMDECs in bAVM development. PdgfbiCreER;Alk1(2f/2f);confetti(+/−) mice with Cre-regulated confetti transgene were used to study EC clonal expansion. BAVMs were induced by intra-brain injection of an adeno-associated viral vector expressing vascular endothelial growth factor followed with intra-peritoneal injection of tamoxifen. Wild-type mice transplanted with PdgfbiCreER;Alk1(2f/2f);Ai14 BM developed bAVMs after bAVM induction. Recombined BMDECs were detected in bAVMs. The presence of clusters of ECs expressing same confetti color in bAVMs suggests that Alk1(−) ECs have been clonally expanded. Increasing tamoxifen dose increased the number of Alk1(−) ECs and abnormal vessels in bAVMs of PdgfbiCreER;Alk1(2f/2f) mice. Our data indicated that homozygous mutation of Alk1 in BMDECs is sufficient to cause bAVM development in brain angiogenic region; clonal expansion of Alk1(−) ECs provides a likely mechanism for how a fraction of mutant ECs causes bAVM; and the burden of Alk1(−) ECs correlates with AVM severity.