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A Rapid and Specific Microplate Assay for the Determination of Intra- and Extracellular Ascorbate in Cultured Cells
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How does ascorbate improve endothelial dysfunction? - A computational analysis.

Sheetal Panday1, Saptarshi Kar2, Mahendra Kavdia1

  • 1Department of Biomedical Engineering, Wayne State University, Detroit, 48202, MI, USA.

Free Radical Biology & Medicine
|January 26, 2021
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Ascorbate (Asc) supplementation can improve endothelial dysfunction by increasing nitric oxide (NO) production and restoring eNOS coupling. This study models how Asc mitigates oxidative stress and enhances NO bioavailability in cardiovascular and neurovascular diseases.

Keywords:
Endothelial cell computational modelGlutathioneKinetic modelOxidative stressPeroxynitriteSuperoxideTetrahydrobiopterinVitamin CeNOS uncoupling

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Area of Science:

  • Biochemistry
  • Computational Biology
  • Vascular Biology

Background:

  • Low ascorbate (Asc) levels are linked to cardiovascular and neurovascular diseases.
  • Endothelial dysfunction, marked by reduced nitric oxide (NO) and increased oxidative stress, is a key feature.
  • Mechanisms of Asc's therapeutic potential in endothelial dysfunction require further elucidation.

Purpose of the Study:

  • To develop a computational model of endothelial cells to investigate Asc's mechanisms in mitigating endothelial dysfunction.
  • To quantitatively analyze Asc-mediated pathways, including BH4 synthesis, oxidative stress, and eNOS activity.
  • To determine optimal Asc levels for improving endothelial cell function.

Main Methods:

  • Developed an endothelial cell-based computational model.
  • Integrated eNOS biochemical pathway, oxidative stress, BH4 synthesis, and Asc/GSH interactions.
  • Quantitatively analyzed Asc's effects on NO production, biopterin ratio, and oxidative stress markers.

Main Results:

  • Asc (0.1-100 μM) dose-dependently improved NO production, biopterin ratio, and reduced oxidative stress.
  • Asc increased BH4 availability and restored eNOS coupling under oxidative stress.
  • Asc (1-10 mM) acted as an antioxidant, reducing superoxide and peroxynitrite levels.

Conclusions:

  • Asc supplementation is a potential therapeutic strategy for endothelial dysfunction, especially when BH4 is depleted.
  • Asc enhances NO bioavailability by increasing BH4 and restoring eNOS coupling.
  • This study provides mechanistic insights into optimal cellular Asc levels for vascular health.