Abstract 21206: Aquaporin-Associated Vascular Development of Human Induced Pluripotent Stem Cells Stimulated with High Levels of Glucose
Background and Objective: In overt diabetes, hyperglycemia increases the plasma osmolarity, leading to induction of hypertonicity-responsive genes, including those coding for the osmosensing water channel aquaporin-1 (AQP1). Induced pluripotent stem (iPS) cells offer a disease-relevant cell model that could may recapitulate the interplay between genetic and environmental risk factors in the vascular complication of diabetes. This study was to test whether hyperglycemia activates AQP1 and induces iPS cells to differentiate into vascular cells.
Methods and results: Human iPS cells generated from skin fibroblasts by transduction with four reprogramming factors (Oct4, SOX2, Klf4, and c-Myc) were characterized for their adaptive responses to high glucose-induced hypertonicity stress. Following 3 weeks of culture in human embryonic stem cell-supporting conditions, compact refractive ES-like colonies emerged. All iPS cell colonies showed expression of embryonic proteins, including OCT4, SSEA3, SSEA4 and SOX2. Cultured in an endothelial differentiating medium, embryoid bodies (EBs) derived of iPS cells developed the endothelial markers, KDR and CD31, detected by flow cytometry and immunoblotting. EBs were incubated with 5.5 mmol/L glucose (normoglycemia), high glucose (HG) at 12.5, 25 and 45 mmol/L, or with the hyperosmolar control mannitol (HM) at 12.5, 25 and 45 mmol/L for 48 hours. Both HG and HM increased expression of AQP1 (immunoblotting). EBs formed tube networks (Fig. a) in matrigel, especially when they were exposed to HG (b) and HM (c) (HG 2.8±0.2 fold; HM 3.3±0.5 fold; n=3, p<0.01). Small interfering RNA to AQP1 could diminish the HG (d) and HM (e) proangiogenic activity.
Conclusions: Human iPS cells are a useful model to test vascularization of iPS-derived vascular progenitor cells through an AQP1-associated hyperosmolar mechanism. Targeting the osmosignaling pathway may represent a novel strategy to reduce vascular complication of diabetes.
- © 2010 by American Heart Association, Inc.