CRISPR/Cas9 Edited Induced Pluripotent Stem Cell-Based Vascular Tissues to Model Aging and Disease-Dependent Impairment

The discovery of induced pluripotent stem cells (iPSCs) and advancements in genome editing technology introduced a new perspective to disease modeling as genetic factors can now be incorporated to mimic the pathology of interest. Ischemia and age-driven impairment of endothelium is one of the very important factors in the prognosis of many diseases as it leads to decreased angiogenic response and is shown to be related to age-dependent decrease in Hypoxia inducible factor 1 (HIF-1) expression levels in endothelial cells. However, there are no models that show the characteristic age and ischemia-driven deterioration of the endothelium with both the functional and genetic mimicry. In this study, we developed a three dimensional (3D) in vitro tissue model composed of human-origin iPSC-derived endothelial cells (iECs), which were clustered regulatory interspaced short palindromic repeat (CRISPR)/Cas9 edited for HIF-1A knockout. Confirmed with a significant decrease in HIF-1 mRNA and protein content, our CRISPR/Cas9 edited tissue models showed disrupted oxygen-controlled stabilization of HIF-1 evidenced by decreased viability, two dimensional tube formation, and 3D lumen formation along with increased mitochondrial reactive oxygen species accumulation under ischemia, mimicking the age-driven impairment in endothelial function. hiPSC-based tissue and disease models such as the one presented in this study are promising to study human disease in a physiologically and pathologically-relevant manner and to develop new therapies. Impact Statement Modeling human disease as precisely as possible is of upmost importance in understanding the underlying pathology and discovering effective therapies. Therefore, disease models that are highly controlled and composed of human-origin cells that present the disease phenotype are crucial. The human induced pluripotent stem cell (hiPSC)-based tissue model we present in this study is an important example of human-origin tissue model with controlled gene expression. Through CRISPR/Cas9 editing of hypoxia inducible factor 1 in hiPSCs, we developed tissue models that show the age and disease-dependent endothelial deterioration. This model holds promise for various biomedical applications as more realistic disease phenotypes can be created using fully human-origin platforms.