Dataset Information

Hepatitis C Virus NS5A Protein Triggers Oxidative Stress by Inducing NADPH Oxidases 1 and 4 and Cytochrome P450 2E1.

ABSTRACT: Replication of hepatitis C virus (HCV) is associated with the induction of oxidative stress, which is thought to play a major role in various liver pathologies associated with chronic hepatitis C. NS5A protein of the virus is one of the two key viral proteins that are known to trigger production of reactive oxygen species (ROS). To date it has been considered that NS5A induces oxidative stress by altering calcium homeostasis. Herein we show that NS5A-induced oxidative stress was only moderately inhibited by the intracellular calcium chelator BAPTA-AM and not at all inhibited by the drug that blocks the Ca(2+) flux from ER to mitochondria. Furthermore, ROS production was not accompanied by induction of ER oxidoreductins (Ero1), H2O2-producing enzymes that are implicated in the regulation of calcium fluxes. Instead, we found that NS5A contributes to ROS production by activating expression of NADPH oxidases 1 and 4 as well as cytochrome P450 2E1. These effects were mediated by domain I of NS5A protein. NOX1 and NOX4 induction was mediated by enhanced production of transforming growth factor ?1 (TGF?1). Thus, our data show that NS5A protein induces oxidative stress by several multistep mechanisms.

SUBMITTER: Smirnova OA

PROVIDER: S-EPMC4855014 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Hepatitis C Virus NS5A Protein Triggers Oxidative Stress by Inducing NADPH Oxidases 1 and 4 and Cytochrome P450 2E1.

Smirnova Olga A OA Ivanova Olga N ON Bartosch Birke B Valuev-Elliston Vladimir T VT Mukhtarov Furkat F Kochetkov Sergey N SN Ivanov Alexander V AV

Oxidative medicine and cellular longevity 20160420

Replication of hepatitis C virus (HCV) is associated with the induction of oxidative stress, which is thought to play a major role in various liver pathologies associated with chronic hepatitis C. NS5A protein of the virus is one of the two key viral proteins that are known to trigger production of reactive oxygen species (ROS). To date it has been considered that NS5A induces oxidative stress by altering calcium homeostasis. Herein we show that NS5A-induced oxidative stress was only moderately ...[more]

PMID: 27200149

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Project description:HIV-1 glycoprotein 120 (gp120) is known to cause neurotoxicity via several mechanisms including production of proinflammatory cytokines/chemokines and oxidative stress. Likewise, drug abuse is thought to have a direct impact on the pathology of HIV-associated neuroinflammation through the induction of proinflammatory cytokines/chemokines and oxidative stress. In the present study, we demonstrate that gp120 and methamphetamine (MA) causes apoptotic cell death by inducing oxidative stress through the cytochrome P450 (CYP) and NADPH oxidase (NOX) pathways. The results showed that both MA and gp120 induced reactive oxygen species (ROS) production in concentration- and time-dependent manners. The combination of gp120 and MA also induced CYP2E1 expression at both mRNA (1.7±0.2- and 2.8±0.3-fold in SVGA and primary astrocytes, respectively) and protein (1.3±0.1-fold in SVGA and 1.4±0.03-fold in primary astrocytes) levels, suggesting the involvement of CYP2E1 in ROS production. This was further confirmed by using a selective inhibitor of CYP2E1, diallylsulfide (DAS), and CYP2E1 knockdown using siRNA, which significantly reduced ROS production (30-60%). As the CYP pathway is known to be coupled with the NOX pathway, including Fenton-Weiss-Haber (FWH) reaction, we examined whether the NOX pathway is also involved in ROS production induced by either gp120 or MA. Our results showed that selective inhibitors of NOX, diphenyleneiodonium (DPI), and FWH reaction, deferoxamine (DFO), also significantly reduced ROS production. These findings were further confirmed using specific siRNAs against NOX2 and NOX4 (NADPH oxidase family). We then showed that gp120 and MA both induced apoptosis (caspase-3 activity and DNA lesion using TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP nick-end labeling) assay) and cell death. Furthermore, we showed that DAS, DPI, and DFO completely abolished apoptosis and cell death, suggesting the involvement of CYP and NOX pathways in ROS-mediated apoptotic cell death. In conclusion, this is the first report on the involvement of CYP and NOX pathways in gp120/MA-induced oxidative stress and apoptotic cell death in astrocytes, which has clinical implications in neurodegenerative diseases, including neuroAIDS.

Project description:This study investigated the role of ethanol-inducible cytochrome P450-2E1 (CYP2E1) in enhancing CYP2E1 and other P450 proteins in extracellular vesicles (EVs) from alcohol-exposed rodents and human patients with alcoholism and their effects on oxidative hepatocyte injury. Female Fischer rats and wild-type or Cyp2e1-null mice were exposed to three oral doses of binge ethanol or dextrose control at 12-hour intervals. Plasma EV and hepatic proteins from alcohol-exposed rodents, patients with alcoholism, and their respective controls were isolated and characterized. The number of EVs and the amounts of EV CYP2E1, CYP2A, CYP1A1/2, and CYP4B proteins were markedly elevated in both patients with alcoholism and alcohol-exposed rats and mice. The number of EVs and EV P450 proteins were significantly reduced in ethanol-exposed rats fed a diet containing polyunsaturated fatty acids. The increased number of EVs and EV CYP2E1 and other P450 isoforms in alcohol-exposed wild types were significantly reduced in the corresponding Cyp2e1-null mice. EV CYP2E1 amounts depended on increased oxidative and endoplasmic reticulum (ER) stress because their levels were decreased by cotreatment with the antioxidant N-acetylcysteine or the CYP2E1 inhibitor chlormethiazole but increased by ER stress-inducer thapsigargin, which was blocked by 4-phenylbutyric acid. Furthermore, cell death rates were elevated when primary hepatocytes or human hepatoma cells were exposed to EVs from alcohol-exposed rodents and patients with alcoholism, demonstrating that EVs from alcohol-exposed rats and patients with alcoholism are functional and can promote cell death by activating the apoptosis signaling pathway, including phospho-c-Jun N-terminal kinase, proapoptotic Bax, and activated caspase-3. Conclusion: CYP2E1 has an important role in elevating EV CYP2E1 and other P450 isoforms through increased oxidative and ER stress. Elevated EV-CYP2E1 detected after withdrawal from alcohol or exposure to the CYP2E1 inducer pyrazole can be a potential biomarker for liver injury. (Hepatology Communications 2017;1:675-690).

Dataset Information

Hepatitis C Virus NS5A Protein Triggers Oxidative Stress by Inducing NADPH Oxidases 1 and 4 and Cytochrome P450 2E1.

Publications

Hepatitis C Virus NS5A Protein Triggers Oxidative Stress by Inducing NADPH Oxidases 1 and 4 and Cytochrome P450 2E1.

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