Modeling GLUT1 deficiency syndrome in a Petri Dish using induced pluripotent stem cells: A preliminary report

Background: Glucose is an important source of energy for the central nervous system. The entry of glucose is tightly regulated by the blood-brain barrier (BBB), mediated mostly through the presence of glucose transporter 1 (GLUT1), a facilitated transporter belonging to the solute carrier superfamily (SLCs). GLUT1 Deficiency Syndrome (GLUT1DS) is a haploinsufficiency characterized by mutations in the SLC2A1 gene and resulting in impaired glucose uptake at the BBB. This preliminary study provides the characterization of a novel GLUT1DS in vitro model based on an established human induced pluripotent stem cell line (iPSC) genetically edited by CRISPR/Cas9. Methods: : The human iPS(IMR90)-c4 iPSC cell line was genetically edited via CRISPR/Cas9, resulting in a SLC2A1 +/- genotype (GLUT1DS-iPSC). Cells were differentiated in brain microvascular endothelial cells (iBMECs) as previously reported. Cells were characterized in terms of SLC2A1 expression, changes in the barrier function, and glucose uptake. Results: : Two GLUT1DS-iPSC clones were generated and characterized in this study: C7 and E8. In undifferentiated iPSCs, both showed comparable phenotype than their parental control, and both differentiated into iBMECs. Both C7- and E8-iBMECs showed a decrease in GLUT1 expression, with C7 showing accentuated decrease. No major disruption in the barrier function was reported. However, we noted a significant decrease in glucose uptake in both clones. Conclusions: : Our study provides the first documented characterization of GLUT1DS-iBMECs generated by CRISPR-Cas9, showing little effect on SLC2A1 deletion on the ability of iPSCs to differentiate into iBMECs and their ability to form tight monolayers. However, an impaired glucose uptake associated with increased mitochondrial respiration and impaired angiogenesis suggests that such deletion may affect other functions in endothelial cells. Hence, a further investigation of how SLC2A1 depletion impacts brain endothelial cells and other cellular components of the neurovascular unit is needed.