Journalartikel

Jahr der Veröffentlichung: 2023

Zeitschrift: Cells

Bandnummer: 12

Heftnummer: 12

eISSN: 2073-4409

Open Access Status: Gold

DOI Link: https://doi.org/10.3390/cells12121600

Verlag: MDPI

Abstract: 
Hereditary hemorrhagic telangiectasia (HHT) type 2 is an autosomal dominant disease in which one allele of the ACVRL1 gene is mutated. Patients exhibit disturbances in TGF-beta/BMP-dependent angiogenesis and, clinically, often present with severe nosebleeds as well as a reduced quality of life. The aim of our study was to use CRISPR/Cas9 to knockout ACVRL1 in normal induced pluripotent stem cells (iPSCs) and evaluate the effects on TGF-beta- and BMP-related gene expression as well as angiogenesis. The CRISPR/Cas9 knockout of the ACVRL1 gene was carried out in previously characterized wild-type (ACVRL1(wt/wt)) iPSCs. An HHT type 2 iPS cell line was generated via a single-allele knockout (ACVRL1(wt/mut)) in wild-type (ACVRL1(wt/wt)) iPSCs, resulting in a heterozygous 17 bp frameshift deletion in the ACVRL1 gene [NG_009549.1:g.13707_13723del; NM_000020.3:c.1137_1153del]. After the generation of embryoid bodies (EBs), endothelial differentiation was induced via adding 4 ng/mL BMP4, 2% B27, and 10 ng/mL VEGF. Endothelial differentiation was monitored via immunocytochemistry. An analysis of 151 TGF-beta/BMP-related genes was performed via RT-qPCR through the use of mRNA derived from single iPS cell cultures as well as endothelial cells derived from EBs after endothelial differentiation. Differential TGF-beta/BMP gene expression was observed between ACVRL1(wt/wt) and ACVRL1(wt/mut) iPSCs as well as endothelial cells. EBs derived from CRISPR/Cas9-designed ACVRL1 mutant HHT type 2 iPSCs, together with their isogenic wild-type iPSC counterparts, can serve as valuable resources for HHT type 2 in vitro studies.

Harvard-Zitierstil: Xiang-Tischhauser, L., Bette, M., Rusche, J., Roth, K., Kasahara, N., Stuck, B., et al. (2023) Generation of a Syngeneic Heterozygous ACVRL1^(wt/mut) Knockout iPS Cell Line for the In Vitro Study of HHT2-Associated Angiogenesis, Cells, 12(12), Article 1600. https://doi.org/10.3390/cells12121600

APA-Zitierstil: Xiang-Tischhauser, L., Bette, M., Rusche, J., Roth, K., Kasahara, N., Stuck, B., Bakowsky, U., Wartenberg, M., Sauer, H., Geisthoff, U., & Mandic, R. (2023). Generation of a Syngeneic Heterozygous ACVRL1^(wt/mut) Knockout iPS Cell Line for the In Vitro Study of HHT2-Associated Angiogenesis. Cells. 12(12), Article 1600. https://doi.org/10.3390/cells12121600