ACE2/ACE-Imbalance in Circulating CD34⁺ Cells of Individuals with Diabetic Retinopathy

Abstract

Purpose : Endothelial dysfunction and reduced microvascular repair are the hallmarks of diabetic retinopathy (DR). Bone marrow derived CD34⁺ cells have the propensity of vascular regeneration. Diabetes is associated with decreased number and vasoreparative dysfunction of CD34⁺ cells. Angiotensin-converting enzyme (ACE) and ACE2 are primary enzymes of the vascular-detrimental and protective axes of the renin-angiotensin system (RAS), respectively, and are expressed in the vascular endothelium and CD34⁺ cells. Activation of ACE2 was shown to stimulate vasoreparative functions in CD34⁺ cells. We tested the hypothesis that balance between ACE and ACE2 shifted towards detrimental axis in CD34⁺ cells of individuals with DR.

Methods : Peripheral blood was obtained from healthy (control) and diabetic individuals with DR. Mononuclear cells (MNCs) and plasma were separated and lineage- (Lin^-) and CD34⁺ cells isolated. Activities of ACE and ACE2 were determined in plasma and lysates of Lin^- and CD34⁺ cells by using enzyme-selective fluorogenic substrates and inhibitors. Proliferation and migration of cells were determined by using BrdU-ELISA or by chemotaxis assay, respectively.

Results : RESULTS: DR was associated with decreased number of Lin^- ((2±0.2)x10⁵, P<0.04) and CD34⁺ (0.6±0.1)x10³, P<0.02) cells (n=6) compared to control group (Lin^- (25±4)x10⁵; CD34⁺ (2±0.4)x10³), expressed as per million MNCs. Abundance of both populations is negatively correlated with HbA1C levels (r²=0.9, P<0.01). ACE and ACE2 activities were not different in MNCs. Decreased ACE2 and increased ACE activities were observed in plasma, and CD34⁺ cell-lysates resulting in reduced ACE2/ACE (DR vs control: Plasma - 0.34±0.07 vs 0.8±0.08, P<0.05; CD34⁺ - 0.4±0.07 vs 1.8±0.2, P<0.01, n=6). Proliferation of Lin^- or CD34⁺ cells from DR-group was lower in basal conditions or in response to stromal-derived factor-1a (SDF) or vascular endothelial growth factor (VEGF) compared to control (P<0.05 to 0.001, n=4-8). SDF- or VEGF-induced migration of both populations was impaired in DR-group compared to control (n=4-8).

Conclusions : DR decreased the number of circulating Lin^- and CD34⁺ cells and impaired vascular repair due to imbalance in the protective and detrimental axes of RAS. Future trials would offer a unique, minimally invasive treatment for vascular regeneration to those with blinding DR and other diabetic diseases.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.