Patient-specific and Gene-corrected Induced Pluripotent Stem Cell-derived Endothelial Cells Elucidate Single-cell Phenotype of Pulmonary Veno-occlusive Disease

PVOD is a rare form of pulmonary hypertension characterized by the preferential remodeling of the pulmonary venules. Hereditary PVOD is caused by biallelic variants of the EIF2AK4 gene. Three PVOD patients who carried the compound heterozygous variants of EIF2AK4 and two healthy controls were recruited and iPSCs were generated from human peripheral blood mononuclear cells (PBMCs). The EIF2AK4 c.2965C>T variant (PVOD#1), c.3460A>T variant (PVOD#2) and c.4832_4833insAAAG variant (PVOD#3) were corrected by CRISPR/Cas9 in PVOD-iPSCs to generate isogenic controls and gene-corrected-iPSCs (GC-iPSCs). PVOD-iPSC-ECs exhibited a decrease in GCN2 protein and mRNA expression, when compared to control and GC-ECs. PVOD-ECs exhibited an abnormal endothelial cell phenotype featured by excessive proliferation and angiogenesis. The abnormal phenotype of PVOD-ECs was normalized by AKT inhibitors AZD5363 and MK2206. These findings help elucidate the underlying molecular mechanism of PVOD in humans, and identify promising therapeutic drugs for treating the disease Overall design: Comparative gene expression profiling analysis of RNA-seq data for PVOD-endothelial cells and GC-PVOD-endothelial cells.