The Krebs Cycle and Mitochondrial Mass Are Early Victims of Endothelial Dysfunction

Abstract

Endothelial cell dysfunction is associated with bioavailable nitric oxide deficiency and an excessive generation of reactive oxygen species. We modeled this condition by chronically inhibiting nitric oxide generation with subpressor doses of N G -monomethyl-Larginine (L-NMMA) in C57B6 and Tie-2/green fluorescent protein mouse strains. L-NMMA-treated mice exhibited a slight reduction in vasorelaxation ability , as well as detectable abnormalities in soluble adhesion molecules (soluble intercellular adhesion molecule-1 and vascular cellular adhesion molecule-1, and matrix metalloproteinase 9), which represent surrogate indicators of endothelial dysfunction. Proteomic analysis of the isolated microvasculature using 2-dimensional gel electrophoresis and matrixassisted laser desorption/ionization time-of-flight mass spectroscopy revealed abnormal expression of a cluster of mitochondrial enzymes, which was confirmed using immunodetection. Aconitase-2 and enoyl-CoA-hydratase-1 expression levels were decreased in L-NMMA-treated animals; this phenotype was absent in nitric oxide synthase-1 and -3 knockout mice. Depletion of aconitase-2 and enoyl-CoA-hydratase-1 resulted in the inhibition of the Krebs cycle and enhanced pyruvate shunting toward the glycolytic pathway. To assess mitochondrial mass in vivo, co-localization of green fluorescent protein and MitoTracker fluorescence was detected by intravital microscopy. Quantitative analysis of fluorescence intensity showed that L-NMMA-treated animals exhibited lower fluorescence of MitoTracker in microvascular endothelia as a result of reduced mitochondrial mass. These findings provide conclusive and unbiased evidence that mitochondriopathy represents an early manifestation of endothelial dysfunction, shifting cell metabolism toward "metabolic hypoxia" through the selective depletion of both aconitase-2 and enoyl-CoAhydratase-1. These findings may contribute to an early preclinical diagnosis of endothelial dysfunction. (Am J

Resveratrol induces mitochondrial biogenesis in endothelial cells

“…Impairment of mitochondrial biogenesis is frequently observed in diabetes and the metabolic syndrome (28) and is thus likely to contribute to cellular energetic imbalance, oxidative stress, and endothelial dysfunction in these pathological conditions (16). Previous studies show that dysregulation of mitochondrial biogenesis represents an early manifestation of endothelial dysfunction, shifting cell metabolism toward metabolic hypoxia in animals with impaired nitric oxide (NO) bioavailability (1). Since increased mitochondrial production of reactive oxygen species (ROS) due to impaired mitochondrial biogenesis also appears to be a key event in the development of aging-induced vascular pathologies (6,41,42), identification of mechanisms that promote mitochondrial biogenesis in the endothelial cells may contribute to the development of improved pharmacological approaches to promote vascular health in both patients with diabetes (20) and the elderly.…”

“…vasoprotection; histone deacetylase; endothelial dysfunction; diabetes; obesity ENDOTHELIAL MITOCHONDRIA HAVE a crucial role in vascular pathophysiology (1,12,25,34). Mitochondria are highly dynamic organelles, and their biogenesis is likely to be involved in the regulation of endothelial cell metabolism, redox regulation, and signal transduction.…”

Photoluminescence intensity ratio of Eu‐conjugated lactates—A simple optical imaging technique for biomarker analysis for critical diseases

“…In the present study, a unique dependence of visible photoluminescence of Eu 3+ ions resulting from 5 D 0 to 7 F J(J = 0,1,2,3,4) transitions, which can be exploited for rapid detection of biomarkers, both in vitro and ex vivo, has been reported. It is observed that the integrated intensity ratio of photoluminescence signals dominating at 591 and 616 nm originating from 5 D 0 to 7 F 2 and 5 D 0 to 7 F 1 transitions in Eu 3+ ions can be used as a biosensing and bioimaging tool for detection of biomarkers released at disease states.…”

“…So as to fathom the severity of disease, hence to provide earlier treatments, it is essential to determine the elevated levels of biomarkers [4]. Lactate plays a significant role in various metabolic cycles in the body including Cori cycle and Krebs cycle [5][6][7]. The normal lactate concentration in human blood plasma is 0.3 to 1 mM and increased lactate levels indicate possible error in cellular metabolism [3].…”

Characterization of a cellular denitrase activity that reverses nitration of cyclooxygenase

“…The mobile phases were 0.1% formic acid in water (solvent A) and 0.1% formic acid in 90% acetonitrile (solvent B). The protein digest (8 l; preparation described in the Supplemental Material) 1 was injected onto a 4-mm 40-nl Zorbax 300SB-C18 enrichment column at a flow rate of 5 l/min, and peptides were resolved on a 0.075 ϫ 43-mm Zorbax 300SB-C18 analytic column (3.5-m particle size) at a flow rate of 0.3 l/min with a gradient of 3-50% solvent B for 20 min and 50 -90% solvent B for 2 min. Mass spectra were acquired in the automated MS/MS mode, in which MS/MS scans were performed on the four most intense ions from each MS scan.…”