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To date, many studies have been conducted to find out the underlying mechanisms of hyperglycemia-induced complications in diabetes mellitus, attributed to the cellular pathologies of different cells-especially endothelial cells. However, there are still many ambiguities and unresolved issues to be clarified. Here, we investigated the alteration in biophysical and biochemical properties in human umbilical vein endothelial cells exposed to a high-glucose concentration (30mM), comparable to glucose content in type 2 diabetes mellitus, over a course of 120 hours. In addition to a reduction in the rate of cell viability and induction of oxidative stress orchestrated by the high-glucose condition, the dynamic of the fatty acid profile-including polyunsaturated, monounsaturated, and saturated fatty acids-was also altered in favor of saturated fatty acids. Genetic imbalances were also detected at chromosomal level in the cells exposed to the abnormal concentration of glucose after 120 hours. Moreover, the number of tip cells (CD31 /CD34 ) and in vitro tubulogenesis capability negatively diminished in comparison to parallel control groups. We found that diabetic hyperglycemia was associated with a decrease in the cell-cell tight junction and upregulation in vascular endothelial cadherin and zonula occludens (ZO)-1 molecules after 72 and 120 hours of exposure to the abnormal glucose concentration, which resulted in a profound reduction in transendothelial electrical resistance. The surface plasmon resonance analysis of the human umbilical vein endothelial cells immobilized on gold-coated sensor chips confirmed the loosening of the cell to cell intercellular junction as well as stable attachment of each cell to the basal surface. Our findings highlighted the disturbing effects of a diabetic hyperglycemia on either biochemical or biophysical properties of endothelial cells.

Introduction: Under the diabetic condition, sustained production of oxidative/nitrosative stress results in irreversible vascular injuries. A great number of diabetic pathologies, such as inefficient or aberrant neo-angiogenesis emerge following chronic hyperglycemic condition. Lack of enough data exists regarding hydroxychloroquine (HCQ) contribution on angiogenesis during diabetes mellitus. Methods: To better address whether HCQ could blunt or exacerbate oxidative status and angiogenesis under high glucose condition (HCG), human umbilical vein endothelial cells (HUVECs) were exposed to 30 µM HCQ in combination with 30 mM glucose over a course of 72 hours. Viability was measured was evaluated by MTT assay. We used Griess method and TBARS assay to monitor changes in the levels of NO and MDA followed by flow cytometric analysis of ROS using DCFDA. To show the impact of HCQ on cell motility and in vitro angiogenic properties, we exploited routine scratch test and in vitro tubulogenesis, respectively. Results: Our data showed that HCQ diminished cell viability under 5 and 30 mM glucose contents. HCQ significantly decreased the total levels of nitric oxide (NO), malondialdehyde (MDA), and reactive oxygen species (ROS) in both sets of environments. Additionally, inhibitory effects were observed on cell migration after exposure to HCQ (P < 0.001). Anti-angiogenic activity of HCQ was confirmed by the reduction of tube areas under a normal or surplus amount of glucose (P < 0.001). Conclusion: In overall, results suggest that HCQ changes the oxidative/nitrosative status of HUVECs both in 5 and 30 mM conditions. HCQ is able to reduce migration and angiogenic activity of HUVECs irrespective of the glucose content.

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