A Simple Method to Systematically Study Oxidatively Modified Proteins in Biological Samples and Its Applications

Song, Byoung–Joon; Suh, Soo-Kyung; Moon, Kyung Ho

doi:10.1016/s0076-6879(10)73013-5

Cited by 16 publications

(18 citation statements)

References 40 publications

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“…Further studies are necessary to identify the thiol-oxidized proteins that may contribute to the MPT and cell death. The specific protein targets can be purified by biotin-conjugated maleimide and identified by mass spectrometric analysis (Song et al, 2010). We anticipate that the identification of specific proteins will be important to developing novel therapeutic targets that may restore protein sulfhydryl and prevent APAP-induced liver toxicity.…”

Section: Discussionmentioning

confidence: 99%

Mouse Liver Protein Sulfhydryl Depletion after Acetaminophen Exposure

Yang

Greenhaw

Shi

et al. 2012

J Pharmacol Exp Ther

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Acetaminophen (APAP)-induced liver injury is the leading cause of acute liver failure in many countries. This study determined the extent of liver protein sulfhydryl depletion not only in whole liver homogenate but also in the zonal pattern of sulfhydryl depletion within the liver lobule. A single oral gavage dose of 150 or 300 mg/kg APAP in B6C3F1 mice produced increased serum alanine aminotransferase levels, liver necrosis, and glutathione depletion in a dose-dependent manner. Free protein sulfhydryls were measured in liver protein homogenates by labeling with maleimide linked to a near infrared fluorescent dye followed by SDS-polyacrylamide gel electrophoresis. Global protein sulfhydryl levels were decreased significantly (48.4%) starting at 1 hour after the APAP dose and maintained at this reduced level through 24 hours. To visualize the specific hepatocytes that had reduced protein sulfhydryl levels, frozen liver sections were labeled with maleimide linked to horseradish peroxidase. The centrilobular areas exhibited dramatic decreases in free protein sulfhydryls while the periportal regions were essentially spared. These protein sulfhydryl-depleted regions correlated with areas exhibiting histopathologic injury and APAP binding to protein. The majority of protein sulfhydryl depletion was due to reversible oxidation since the global-and lobule-specific effects were essentially reversed when the samples were reduced with tris(2-carboxyethy)phosphine before maleimide labeling. These temporal and zonal pattern changes in protein sulfhydryl oxidation shed new light on the importance that changes in protein redox status might play in the pathogenesis of APAP hepatotoxicity.

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Section: Discussionmentioning

confidence: 99%

Mouse Liver Protein Sulfhydryl Depletion after Acetaminophen Exposure

Yang

Greenhaw

Shi

et al. 2012

J Pharmacol Exp Ther

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“…Cytosol, mitochondrial and nuclear fractions were also prepared by differential centrifugations, as described previously [21,22], and all extraction buffers were pre-equilibrated with nitrogen gas to remove the dissolved oxygen. Protein concentrations of liver homogenates were determined using the bicinchoninic acid protein assay reagent (Pierce, Rockford, IL, USA).…”

Section: Methodsmentioning

confidence: 99%

Binge alcohol promotes hypoxic liver injury through a CYP2E1–HIF-1α-dependent apoptosis pathway in mice and humans

Yun

Son

Abdelmegeed

et al. 2014

Free Radical Biology and Medicine

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Binge drinking, a common pattern of alcohol ingestion, is known to potentiate liver injury caused by chronic alcohol abuse. This study was aimed to investigate theeffects of acute binge alcohol onhypoxia-inducible factor-1α (HIF-1α)-mediated liver injury and rolesof alcohol metabolizing enzymes in alcohol-induced hypoxia and hepatotoxicity. Mice and human specimens assigned as binge or non-binge group were analyzed for blood alcohol concentration (BAC), alcohol metabolizing enzymes, HIF-1α-related protein nitration and apoptosis. Binge alcohol promoted acute liver injury in mice with elevated levels of ethanol-inducible cytochrome P450-2E1 (CYP2E1) and hypoxia, both of which were co-localized in centrilobular areas. We observed positive correlations among elevated BAC, CYP2E1, and HIF-1α in mice and humans exposed to binge alcohol. TheCYP2E1 protein levels(r=0.629, p=0.001) and activities (r=0.641, p=0.001) showed significantly positive correlation with BACs in human livers. HIF-1α levels were also positively correlated with BACs (r=0.745, p<0.001) or CYP2E1activities(r=0.792, p<0.001) in humans. Binge alcoholpromotedprotein nitration and apoptosis with significant correlations observed between inducible nitric oxide synthase and BACs, CYP2E1, or HIF-1α in human specimens. Binge alcohol-induced HIF-1α activation and subsequent protein nitration orapoptosisseen in wild-type were significantly alleviatedin the corresponding Cyp2e1-null mice while pretreatment with an HIF-1α inhibitor PX-478 prevented HIF-1α elevation with a trend of decreased levels of 3-nitrotyrosine and apoptosis, supporting the roles of CYP2E1and HIF-1α in binge alcohol-mediatedprotein nitration and hepatotoxicity. Thus binge alcohol promotes acute liver injury in mice and humans at least partly through a CYP2E1-HIF-1α-dependent apoptosis pathway.

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“…ER functions encompass protein biosynthesis, folding and trafficking as well as calcium homeostasis among others [78]. Hence, the ER plays a critical and elegant role in maintaining cellular and organismic homeostasis [79].…”

Section: The Unfolded Protein Responsementioning

confidence: 99%

Modulation of mitochondrial dysfunction and endoplasmic reticulum stress are key mechanisms for the wide-ranging actions of epoxy fatty acids and soluble epoxide hydrolase inhibitors

Inceoglu

Bettaieb

Haj

et al. 2017

Prostaglandins & Other Lipid Mediators

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The arachidonic acid cascade is arguably the most widely known biologic regulatory pathway. Decades after the seminal discoveries involving its cyclooxygenase and lipoxygenase branches, studies of this cascade remain an active area of research. The third and less widely known branch, the cytochrome P450 pathway leads to highly active oxygenated lipid mediators, epoxy fatty acids (EpFAs) and hydroxyeicosatetraenoic acids (HETEs), which are of similar potency to prostanoids and leukotrienes. Unlike the COX and LOX branches, no pharmaceuticals currently are marketed targeting the P450 branch. However, data support therapeutic benefits from modulating these regulatory lipid mediators. This is being approached by stabilizing or mimicking the EpFAs or even by altering the diet. These approaches lead to predominantly beneficial effects on a wide range of apparently unrelated states resulting in an enigma of how this small group of natural chemical mediators can have such diverse effects. EpFAs are degraded by soluble epoxide hydrolase (sEH) and stabilized by inhibiting this enzyme. In this review, we focus on interconnected aspects of reported mechanisms of action of EpFAs and inhibitors of soluble epoxide hydrolase (sEHI). The sEHI and EpFAs are commonly reported to maintain homeostasis under pathological conditions while remaining neutral under normal physiological conditions. Here we provide a conceptual framework for the unique and broad range of biological activities ascribed to epoxy fatty acids. We argue that their mechanism of action pivots on their ability to prevent mitochondrial dysfunction, to reduce subsequent ROS formation and to block resulting cellular signaling cascades, primarily the endoplasmic reticulum stress. By stabilizing the mitochondrial – ROS – ER stress axis, the range of activity of EpFAs and sEHI display an overlap with the disease conditions including diabetes, fibrosis, chronic pain, cardiovascular and neurodegenerative diseases, for which the above outlined mechanisms play key roles.

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A Simple Method to Systematically Study Oxidatively Modified Proteins in Biological Samples and Its Applications

Cited by 16 publications

References 40 publications

Mouse Liver Protein Sulfhydryl Depletion after Acetaminophen Exposure

Mouse Liver Protein Sulfhydryl Depletion after Acetaminophen Exposure

Binge alcohol promotes hypoxic liver injury through a CYP2E1–HIF-1α-dependent apoptosis pathway in mice and humans

Modulation of mitochondrial dysfunction and endoplasmic reticulum stress are key mechanisms for the wide-ranging actions of epoxy fatty acids and soluble epoxide hydrolase inhibitors

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