RWOP2 Heterogeneous metabolic response of endothelial cells from different vascular beds to hyperglycaemia and metformin

Objectives Endothelial cell (EC) dysfunction is a primary feature of diabetes. Those affected are at an increased risk of vascular bed-specific complications such as retinopathy and small vessel disease. ECs are highly glycolytic with mitochondria primarily serving a signalling function, however the degree to which different ECs rely on mitochondria is currently unknown. This study aimed to characterise mitochondrial function in human ECs in response to hyperglycaemia (HG) to help determine the susceptibility of particular vascular beds to the diabetic milieu and determine the effectiveness of metformin upon EC mitochondrial health. Methods Human primary retinal (HREC), brain microvascular (HBEC), dermal (HDEC), aortic (HAEC), and human umbilical vein endothelial cells (HUVEC) were cultured in HG conditions (25 mM D-Glucose) and treated with metformin (500μM) for 48 hours. Mitochondrial function was interrogated using a 96-well Seahorse XF-Pro. Mitochondrial morphology was assessed using mitotracker deep red and confocal microscopy. Results ECs from differing vascular beds displayed significantly different metabolic profiles (p<0.05). Genes associated with mitochondrial dynamics displayed heterogeneous expression, typically being significantly lower in HREC and HBECs compared to HDEC and HAEC (p<0.05). 48-hour exposure to HG had modest impacts upon oxygen consumption in all ECs, while metformin reduced metabolic function and ATP production, significantly in HREC and HBECs (p<0.05). These changes coincided with phenotypic changes in mitochondrial morphology, where network area and branching were lowered in HG but preserved to varying degrees with metformin. Conclusions ECs from different vascular beds differ significantly in their respiratory profile and mitochondria-related gene expression at baseline. HG significantly impacted mitochondrial morphology. The higher metabolic activity yet reduced mitochondrial dynamics of HRECs may contribute to the increased risk of vascular insult and retinopathy in diabetes. Preserved morphology with metformin treatment may make it effective in maintaining vascular health where mitochondrial turnover is low, such as HREC and HBECs.