Activation of the Protective Arm of Renin Angiotensin System (RAS) Corrects the Reparative Dysfunction of Diabetic CD34+ Cells.

Abstract 2637 Purpose: RAS plays a vital role in regulating many physiological processes of the vascular system. Angiotensin II (Ang II), a product of angiotensin converting enzyme (ACE), mediates its effects through activation of either the AT 1 receptor – to induce vasoconstriction, proliferation, fibrosis, and inflammation – or the AT 2 receptor to promote NO generation. The protective arm of RAS involves ACE2, which produces angiotensin-(1-7) [Ang-(1-7)]. Ang-(1-7) activates the MAS receptor to promote vascular health. Because diabetic endothelial progenitor cells are dysfunctional and this limits their utility in autologous cell therapy, we asked whether angiotensin (Ang)-(1-7) could restore the vasoreparative function of diabetic CD34 + cells. Methods: Healthy nondiabetic (ND) and diabetic (D) Lin − CD45 mid CD34 + cells were obtained from peripheral blood mononuclear cells (PB-MNCs) by FACS. The effect of Ang-(1-7) on migration, proliferation, NO bioavailability, reactive oxygen species (ROS) levels and NADPH oxidase activity were evaluated in ND- and D-CD34 + cells. The effect of Ang-(1-7) on the formation of ECFCs from ND- and D-MNCs was evaluated. Ang-(1-7) production by cells was analyzed and the expression of ACE2 and Mas-receptor were assessed by real-time PCR and flow cytometry. Effects of ACE2 activators XNT and DIZE in CD34 + cells were also evaluated. D-CD34 + cells were genetically modified to overexpress Ang-(1-7) by lentiviral Ang-(1-7)-fusion transgene and their function was evaluated in vitro . The effect of transduction on the surface expression of CD133, CD34 and CD309 was assessed. In vivo homing function was assessed in a mouse model of ischemia-reperfusion (I/R). After one week of I/R insult, when retinal capillary damage was appreciable, CD34 cells were intravitreally injected. Neural retinas were harvested after 48 hours and human cells within the mouse vasculature were localized by immunohistochemistry. Results: Migration to SDF1- and VEGF-were impaired in D-CD34 + cells. In contrast, Ang-(1-7)-induced migration in both D-CD34 + and ND-CD34 + cells was dependent on Mas receptor expression and eNOS. ROS levels and NADPH oxidase activity were reduced and proliferation and NO bioavailability were restored in D-CD34 + cells by Ang-(1-7). ECFCs from D-MNCs were appeared only in the presence of Ang-(1-7). Migration, NO release and Ang-(1-7) release by ACE2 activators, XNT and DIZE, were significantly decreased in D-CD34 + cells. Ang-(1-7) release and ACE2 expression were decreased in D-cells while Mas-receptor expression was similar that in ND-cells. Ang-(1-7) gene-modified cells showed reduced ROS levels, increased NO bioavailability, enhanced migration to SDF-1 and proliferation. Lentiviral transduction did not alter surface expression of CD133, CD34 and CD309. Ang-(1-7)-overexpression restored the homing efficiency of D-CD34 + cells similar to that of ND-CD34 + cells in vivo . Conclusions: Ang-(1-7) stimulates the vasoreparative functions in CD34 + cells. Ang-(1-7) bypasses the reduced ACE2 seen in diabetic CD34 + cells and restores the vasoreparative potential of these cells by decreasing oxidative stress and normalizing NO bioavailability. Pharmacological strategies that either increase ACE2 or Ang-(1-7) in diabetic CD34+ cells will improve their therapeutic utility for autologous cell therapy in treatment of diabetic complications. Disclosures: No relevant conflicts of interest to declare