CDK1 inhibition reduces osteogenesis in endothelial cells in vascular calcification
Yan Zhao, Yang Yang, Xiuju Wu, Li Zhang, Xinjiang Cai, Jaden Ji, Sydney Chen, Abigail Vera, Kristina I. Boström, Yucheng Yao
Yan Zhao, Yang Yang, Xiuju Wu, Li Zhang, Xinjiang Cai, Jaden Ji, Sydney Chen, Abigail Vera, Kristina I. Boström, Yucheng Yao
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Research Article Cell biology Vascular biology

CDK1 inhibition reduces osteogenesis in endothelial cells in vascular calcification

Abstract

Vascular calcification is a severe complication of cardiovascular diseases. Previous studies demonstrated that endothelial lineage cells transitioned into osteoblast-like cells and contributed to vascular calcification. Here, we found that inhibition of cyclin-dependent kinase (CDK) prevented endothelial lineage cells from transitioning to osteoblast-like cells and reduced vascular calcification. We identified a robust induction of CDK1 in endothelial cells (ECs) in calcified arteries and showed that EC–specific gene deletion of CDK1 decreased the calcification. We found that limiting CDK1 induced E-twenty-six specific sequence variant 2 (ETV2), which was responsible for blocking endothelial lineage cells from undergoing osteoblast differentiation. We also found that inhibition of CDK1 reduced vascular calcification in a diabetic mouse model. Together, the results highlight the importance of CDK1 suppression and suggest CDK1 inhibition as a potential option for treating vascular calcification.

Authors

Yan Zhao, Yang Yang, Xiuju Wu, Li Zhang, Xinjiang Cai, Jaden Ji, Sydney Chen, Abigail Vera, Kristina I. Boström, Yucheng Yao

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Figure 5

Limiting CDK1 upregulates ETV2 to prevent endothelial lineage cells from undergoing osteogenic differentiation.

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Limiting CDK1 upregulates ETV2 to prevent endothelial lineage cells from...
(A and B) Expression of ETV2 in tdTomato+ aortic cells isolated from VE-cadherincre/ERT2 RosatdTomato Mgp–/– mice and VE-cadherincre/ERT2 RosatdTomato mice, as shown by real-time PCR (A) and immunoblotting (B) (n = 6). Positive control (Pos Ctr) for ETV2 blotting: Mouse ECs induced from embryonic stem cells (66). Negative control (Neg Ctr) for ETV2 blotting: HAECs transfected with ETV2 siRNA. Each lane represents an independent experimental group. (C) Expression of ETV2 in CD31+CD45– aortic cells isolated from VE-cadherincre/ERT2 Cdk1fl/fl Mgp–/– mice and VE-cadherincre/ERT2 Cdk1fl/fl mice after tamoxifen administration (n = 6). (D) ETV2 expression in HAECs after transfection of MGP siRNA in combination with AT5719 treatment (1–10 μM), as shown by real-time PCR (n = 6). (E) Immunoblotting of ETV2, osterix, osteopontin (OPN), eNOS, and FLK1 in HAECs after transfection of MGP siRNA in combination with infection of lentiviral vectors containing CMV-driven ETV2 cDNA and treatment with osteogenic induction media. Positive and negative controls are the same as described in B and Figure 3. Each lane represents an independent experimental group. (F and G) Micro-CT imaging and Alizarin red and von Kossa staining (F), and total aortic calcium with ETV2 expression (G) of descending aortas of Mgp–/– mice treated with adeno-associated viral (AAV) vectors containing CMV-promoter-driven ETV2 cDNA (n = 8). Empty vectors were used as controls. Scale bars: 5 mm (micro-CT) and 100 μm (staining). (H) Expression of osteogenic and endothelial markers in CD31+CD45– aortic cells from Mgp–/– mice treated with AAV vectors containing CMV-promoter-driven ETV2 cDNA (n = 8). Empty vectors were used as controls. Data were analyzed for statistical significance by 1-way ANOVA with Tukey’s multiple-comparison test (A, C, D, and H) or unpaired, 2-tailed Student’s t test (G). The bounds of the boxes are upper and lower quartiles with data points, the line in the box is the median, and whiskers are maximal and minimal values. ***P < 0.0001.