["The Journal of Physiology, Volume 604, Issue 11, Page 4208-4224, 1 June 2026. ", "\nAbstract figure legend Diabetes mellitus is associated with vascular pathology that leads to vascular complications in several tissues, such as retinopathy of the eye. Endothelial dysfunction plays a pivotal role in the progression of each complication. This study examines how human endothelial cells (ECs) from different vascular beds differ, specifically in their mitochondrial function both at baseline and in response to experimental hyperglycaemia (HG), aiming to determine whether basal metabolism influences the response to HG. We find that ECs exhibit metabolic heterogeneity at baseline where respiratory function is associated with nitric oxide (NO) signalling potential. In HG conditions, mitochondrial network morphology was perturbed with a concomitant impact in glycolytic function that may be dependent upon basal metabolic profile.\n\n\n\n\n\n\n\n\n\nAbstract\nEndothelial cells (ECs) are highly glycolytic, with mitochondria primarily serving a signalling function. Metabolic disruptions are early contributors to endothelial dysfunction, a primary feature of diabetic vascular complications, such as retinopathy, impaired wound healing and cerebral small vessel disease. The degree to which metabolism varies amongst such different vascular beds is unknown. Mitochondrial function was therefore characterised in human aortic, dermal, retinal and cerebral ECs in vitro, aiming to determine whether basal metabolism influences the response and susceptibility of vascular beds experimental hyperglycaemia (HG). Furthermore, the potential of metformin to maintain endothelial function independent of glycaemic control was assessed. Using a Seahorse analyser, metabolic function of human primary ECs from different vascular beds was compared under basal conditions, as well as HG and metformin treatment. ECs differed significantly in respiratory profile and glycolytic function. For example aortic ECs were preferentially aerobic, whereas dermal ECs were glycolytic. HG significantly lowered mitochondrial network area but elicited modest effects upon respiratory function at the same time as influencing glycolytic function in a manner that was possibly conditional upon basal utilisation. Metformin inhibited basal respiratory function at the same time as significantly enhancing glycolysis in retinal and brain ECs. These data suggest that EC responses to HG and metformin are influenced by the basal metabolic profile, highlighting the potential of targeting EC metabolism to preserve function in a diabetic condition. A nuanced approach is needed to address diabetic vascular complications and endothelial metabolic health in diabetes, both in the investigation of pathophysiology and in prospective therapeutics.\n\n\n\n\n\n\n\n\n\nKey points\n\nEndothelial dysfunction is an early feature of diabetes‐associated cardiovascular complications\nEndothelial cells (ECs) are highly glycolytic, with mitochondria serving a signalling function\nECs are known to be heterogeneous in function, but how this is reflected in metabolism is not fully understood, in addition to how this influences their response to hyperglycaemia\nUsing experimental hyperglycaemia (HG) in vitro, we demonstrate that ECs differed significantly in respiratory profile and glycolytic function. Their response to HG is possibly contingent upon this basal utilisation.\nThese results suggest a nuanced approach is needed when investigating diabetic vascular complications, both in the investigation of pathophysiology and in prospective therapeutics.\n\n\n"]