Elevated endothelial Sox2 causes lumen disruption and cerebral arteriovenous malformations
Jiayi Yao, … , Kristina I. Boström, Yucheng Yao
Jiayi Yao, … , Kristina I. Boström, Yucheng Yao
Published June 24, 2019
Citation Information: J Clin Invest. 2019;129(8):3121-3133. https://doi.org/10.1172/JCI125965.
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Research Article Vascular biology

Elevated endothelial Sox2 causes lumen disruption and cerebral arteriovenous malformations

Abstract

Lumen integrity in vascularization requires fully differentiated endothelial cells (ECs). Here, we report that endothelial-mesenchymal transitions (EndMTs) emerged in ECs of cerebral arteriovenous malformation (AVMs) and caused disruption of the lumen or lumen disorder. We show that excessive Sry-box 2 (Sox2) signaling was responsible for the EndMTs in cerebral AVMs. EC-specific suppression of Sox2 normalized endothelial differentiation and lumen formation and improved the cerebral AVMs. Epigenetic studies showed that induction of Sox2 altered the cerebral-endothelial transcriptional landscape and identified jumonji domain–containing protein 5 (JMJD5) as a direct target of Sox2. Sox2 interacted with JMJD5 to induce EndMTs in cerebral ECs. Furthermore, we utilized a high-throughput system to identify the β-adrenergic antagonist pronethalol as an inhibitor of Sox2 expression. Treatment with pronethalol stabilized endothelial differentiation and lumen formation, which limited the cerebral AVMs.

Authors

Jiayi Yao, Xiuju Wu, Daoqin Zhang, Lumin Wang, Li Zhang, Eric X. Reynolds, Carlos Hernandez, Kristina I. Boström, Yucheng Yao

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Figure 5

Sox2/JMJD5 prompts ECs to undergo mesenchymal transitions, resulting in lumen disorder.

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Sox2/JMJD5 prompts ECs to undergo mesenchymal transitions, resulting in ...
(A) MGP expression and pSMAD1/5/8 levels in HBMECs after depletion of MGP using CRISPR/Cas9 (MGP CRISPR) (n = 5). (B) Coimmunofluorescent staining of CD31 (green) and N-cadherin (red) in MGP CRISPR cells. HBMECs were used as controls (n = 10). Scale bars: 100 μm. (C) Alizarin red staining of MGP CRISPR cells after treatment with osteogenic media (top). Coimmunofluorescent staining of CD31 (green) and neurofilament (red) in MGP CRISPR cells after treatment with neurogenic media (bottom). HBMECs were used as controls (n = 3). Scale bars: 100 μm. (D) Expression of the adipogenic markers PPARγ and C/EBP in MGP CRISPR cells after treatment with adipogenic media (n = 3). (E) Flow cytometric analysis of MGP CRISPR cells. HBMECs were used as controls (n = 3). (F) Increased expression of the lumen-associated genes Par3 and Rasip1 in Sox2 and N-cadherin double-positive cells isolated from MGP CRISPR cells (n = 6). (G) Suppression of Sox2 expression by siRNA (si) decreases expression of JMJD5, N-cadherin, Par3, and Rasip1 in MGP CRISPR cells, as shown by immunoblotting (n = 6). (H) Limiting JMJD5 expression by siRNA decreases the expression of N-cadherin, Par3, and Rasip1 in MGP CRISPR cells, as shown by immunoblotting (n = 3). (I) Coimmunoprecipitation of Sox2 and JMJD5 from lysed MGP CRISPR cells as shown by immunoblotting. HBMECs were used as controls (n = 3). (J) Sox2 DNA–binding site in the promoter of the N-cadherin gene. (K) ChIP assay of Sox2 DNA–binding in the promoter of N-cadherin using enriched genomics DNA from MGP CRISPR cells with or without transfection of Sox2 siRNA. Genomics DNA was enriched by using anti-Sox2 (a-Sox2) or anti-JMJD5 (a-JMJD5) antibodies (n = 3). (L) Schematic diagram: Sox2 induces and interacts with JMJD5 to induce EndMTs. Data shown in A, D, and F were analyzed by Student’s t test. Data shown in K was analyzed by 1-way ANOVA with Tukey’s multiple comparisons test. Data are shown by box and whisker plots. The bounds of the boxes represent upper and lower quartiles. The lines in the boxes represent the median, and the whiskers represent the maximum and minimal values. ***P < 0.001.