血管性血友病因子
脐静脉
内皮干细胞
免疫学
细胞培养
生物
细胞
人脐静脉内皮细胞
细胞生物学
分子生物学
作者
Christopher James Ng,Alice Liu,Sujatha Venkataraman,Katrina J. Ashworth,Christopher D. Baker,Rebecca O’Rouke,Rajeev Vibhakar,Kenneth Jones,Jorge Di Paola
出处
期刊:Blood
[American Society of Hematology]
日期:2022-02-10
标识
DOI:10.1182/blood.2021010683
摘要
Von Willebrand Factor (VWF) plays a key role in normal hemostasis and deficiencies of VWF lead to clinically significant bleeding. We sought to identify novel modifiers of VWF levels in endothelial colony forming cells (ECFCs) using single cell RNA sequencing (scRNA-seq). ECFCs were isolated from patients with Low VWF levels (plasma VWF antigen levels between 30-50 IU/dL) and from healthy controls. Human umbilical vein endothelial cells were used as an additional control cell line. Cells were characterized for their Weibel Palade body (WPB) content and VWF release. scRNA-seq of all cell lines was performed to evaluate for gene expression heterogeneity and for candidate modifiers of VWF regulation. Candidate modifiers identified by scRNA-seq were further characterized with siRNA experiments to evaluate for effects on VWF. We observed that ECFCs derived from patients with Low VWF demonstrated alterations in baseline WPB metrics and exhibit impaired VWF release. scRNA-seq analyses of these endothelial cells revealed overall decreased VWF transcription, mosaicism of VWF expression, and genes that are differentially expressed in Low VWF ECFCs and control endothelial cells (control ECs). A siRNA screen of potential VWF modifiers provided further evidence of regulatory candidates, and one such candidate, FLI1, alters the transcriptional activity of VWF. In conclusion, ECFCs from Low VWF individuals demonstrate alterations in their baseline VWF packaging and release as compared to control ECs. scRNA-seq revealed alterations in VWF transcription and siRNA screening identified multiple candidate regulators of VWF.
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