Endothelial dysfunction due to the inhibition of the synthesis of nitric oxide: Proposal and characterization of an in vitro cellular model

Abstract

The vascular endothelium plays a pivotal role in the maintenance of vascular homeostasis, mediated by vasoactive molecules produced by endothelial cells. The balance between vasoconstrictor and vasodilator biomolecules is what guarantees this equilibrium. Therefore, an increase in the bioavailability of vasoconstrictors along with a reduction in vasodilators may indicate a condition known as endothelial dysfunction. Endothelial dysfunction is marked by an inflammatory process and reduced activity of vasoprotective enzymes, being characterized by some factors like the reduction of the bioavailability of nitric oxide (NO) and increase in the production of reactive oxygen species (ROS), pro-inflammatory and vasoconstrictor molecules. This condition is a predictive marker of several cardiovascular diseases (e.g., atherosclerosis, hypertension, and diabetes). Research is affected by the scarcity of suitable in vitro models that simulate endothelial dysfunction. The goal of this study was to induce an in vitro condition to mimic endothelial dysfunction by inhibiting NO synthesis in cells. Thymus-derived endothelial cells (tEnd.1) were treated with different concentrations of L-NAME (from 1 to 1,000 μM) for different times (12, 24, 48, 72, 96, and 120 h without and with retreatment every 24 h). Cell viability, nitrite concentration, p22phox, NOX2, NOX4, IL-6, and ACE genes expression and lipid peroxidation were evaluated. The results indicate that the treatment with 100 μM L-NAME for 72 h without retreatment reduced NO concentration and NOX4 gene expression while increasing ACE expression, thus mimicking reduced vascular protection and possibly increased vasoconstriction. On the other hand, treatment with 100 μM L-NAME for 96 h with retreatment reduced the concentration of NO and the expression of the p22phox gene while increasing the expression of the IL-6 and ACE genes, mimicking the increase in inflammation and vasoconstriction parameters. Based on these results, we thus propose that both 100 μM L-NAME for 72 h without retreatment and 100 μM L-NAME for 96 h with retreatment may be used as models for in vitro endothelial dysfunction according to the purpose of the study to be conducted.

Nitrosation of CD36 regulates endothelial function and serum lipids

“…To test the hypothesis the Cav1 loss is causing a compensatory increase in NO and disrupting lipid uptake, we used HAMECs genetically devoid of Cav1, a condition that previously inhibited lipid uptake (Figure 3). We treated these cells with L-NAME, a well-known inhibitor of nitric oxide synthase 49, 50 , to examine the role of NO in endothelial lipid uptake. After lipid treatment, cells devoid of Cav1 were indistinguishable from control treated cells ( Figure 4A ).…”

Tracer-based metabolomics for profiling nitric oxide metabolites in a 3D microvessel-on-a-chip model

“…We utilized the detected isotopologues to verify the uptake of 13 C 6 , 15 N 4 -L-arginine and reconstruct the metabolic pathways controlled by eNOS and arginase in HCAECs. This analysis encompassed four conditions: 1) control, 2) treatment with the stimulatory compound VEGF, known for inducing NO release from endothelial cells (Feliers et al, 2005); 3) treatment with the eNOS inhibitory compound L-NAME (Rees et al, 1990; Silva et al, 2022); and 4) treatment with the arginase enzyme inhibitor BEC (Caldwell et al, 2015). The pathway and resulting metabolite expression for each treatment are depicted in ( Fig.…”

Dynamics of the Level of Endothelial Monocyte Activating Polypeptide-ІI in Patients with Acute Myocardial Infarction with Concomitant Diabetes Mellitus Type 2

“…Зрештою це призводить до зниження біодоступності оксиду азоту, погіршення судинного тонусу та інших фенотипових змін ендотелію, які спільно називають ендотеліальною дисфункцією. Розуміння механізмів, що лежать в основі розвитку ендотеліальної дисфункції, є важливою базою знань для запобігання пошкодженню судин при метаболічних і серцево-судинних захворюваннях [1,2].…”