Dataset Information

Steatosis-induced proteins adducts with lipid peroxidation products and nuclear electrophilic stress in hepatocytes.

ABSTRACT: Accumulating evidence suggests that fatty livers are particularly more susceptible to several pathological conditions, including hepatic inflammation, cirrhosis and liver cancer. However the exact mechanism of such susceptibility is still largely obscure. The current study aimed to elucidate the effect of hepatocytes lipid accumulation on the nuclear electrophilic stress. Accumulation of intracellular lipids was significantly increased in HepG2 cells incubated with fatty acid (FA) complex (1mM, 2:1 oleic and palmitic acids). In FA-treated cells, lipid droplets were localized around the nucleus and seemed to induce mechanical force, leading to the disruption of the nucleus morphology. Level of reactive oxygen species (ROS) was significantly increased in FA-loaded cells and was further augmented by treatment with moderate stressor (CoCl2). Increased ROS resulted in formation of reactive carbonyls (aldehydes and ketones, derived from lipid peroxidation) with a strong perinuclear accumulation. Mass-spectroscopy analysis indicated that lipid accumulation per-se can results in modification of nuclear protein by reactive lipid peroxidation products (oxoLPP). 235 Modified proteins involved in transcription regulation, splicing, protein synthesis and degradation, DNA repair and lipid metabolism were identified uniquely in FA-treated cells. These findings suggest that steatosis can affect nuclear redox state, and induce modifications of nuclear proteins by reactive oxoLPP accumulated in the perinuclear space upon FA-treatment.

SUBMITTER: Anavi S

PROVIDER: S-EPMC4309854 | biostudies-other | 2015

REPOSITORIES: biostudies-other

ACCESS DATA

Publications

Steatosis-induced proteins adducts with lipid peroxidation products and nuclear electrophilic stress in hepatocytes.

Anavi Sarit S Ni Zhixu Z Tirosh Oren O Fedorova Maria M

Redox biology 20141224

Accumulating evidence suggests that fatty livers are particularly more susceptible to several pathological conditions, including hepatic inflammation, cirrhosis and liver cancer. However the exact mechanism of such susceptibility is still largely obscure. The current study aimed to elucidate the effect of hepatocytes lipid accumulation on the nuclear electrophilic stress. Accumulation of intracellular lipids was significantly increased in HepG2 cells incubated with fatty acid (FA) complex (1mM, ...[more]

PMID: 25560244

Similar Datasets

Project description:Ferroptosis is an iron dependent form of cell death, that is triggered by the discoordination of iron, lipids, and thiols. Its unique signature that distinguishes it from other forms of cell death is the formation and accumulation of lipid hydroperoxides, particularly oxidized forms of polyunsaturated phosphatidylethanolamines (PEs), which drives cell death. These readily undergo iron-catalyzed secondary free radical reactions leading to truncated products which retain the signature PE headgroup and which can readily react with nucleophilic moieties in proteins via their truncated electrophilic acyl chains. Using a redox lipidomics approach, we have identified oxidatively-truncated PE species (trPEox) in enzymatic and non-enzymatic model systems. Further, using a model peptide we demonstrate adduct formation with Cys as the preferred nucleophilic residue and PE(26:2) +2 oxygens, as one of the most reactive truncated PE-electrophiles produced. In cells stimulated to undergo ferroptosis we identified PE-truncated species with sn-2 truncations ranging from 5 to 9 carbons. Taking advantage of the free PE headgroup, we have developed a new technology using the lantibiotic duramycin, to enrich and identify the PE-lipoxidated proteins. Our results indicate that several dozens of proteins for each cell type, are PE-lipoxidated in HT-22, MLE, and H9c2 cells and M2 macrophages after they were induced to undergo ferroptosis. Pretreatment of cells with the strong nucleophile, 2-mercaptoethanol, prevented the formation of PE-lipoxidated proteins and blocked ferroptotic death. Finally, our docking simulations showed that the truncated PE species bound at least as good to several of the lantibiotic-identified proteins, as compared to the non-truncated parent molecule, stearoyl-arachidonoyl PE (SAPE), indicating that these oxidatively-truncated species favor/promote the formation of PEox-protein adducts. The identification of PEox-protein adducts during ferroptosis suggests that they are participants in the ferroptotic process preventable by 2-mercaptoethanol and may contribute to a point of no return in the ferroptotic death process.

Project description:The incidence of obesity and its related morbidities has rapidly increased over the past twenty years. One consequence of this is the increased probability of having to treat obese individuals for breast cancer. An important co-morbidity associated with obesity are the spectrum of fatty liver diseases. As such, an important question is how treatment of breast cancer with standard therapeutic agents may alter in individuals with fatty liver disease. Such impacts could result in an enhanced adverse effect profile over both acute (e.g. increased cytotoxicity against non-malignant tissues) or chronic (e.g. increased progression through the fatty liver disease spectrum) time periods. In this work we examine if lipid loading in human hepatocytes, leading to a steatotic phenotype, results in an altered toxic liability for the cancer therapeutic doxorubicin. We demonstrate that there is a synergistic impact on cytotoxicity, due to increased oxidative stress. Array analysis reveals the cellular changes associated with hepatocyte adaptation to lipid loading or doxorubicin treatment alone, and how these profiles are significantly altered during doxorubicin treatment of lipid-loaded hepatocytes. Such alterations, we believe, not only underlie the enhanced sensitivity of obese individuals to the acute adverse effects of doxorubicin, but may also potentiate transition along the fatty liver disease spectrum.Experimental design: Huh7 cells were exposed to lipid and/or doxorubicin as required. Huh7 cells were treated with vehicle control or 300 μM FFA mixture for 24 h in T25 culture flasks. The following day, the cells were treated with either 300 μM FFA mixture, 0.1 μM or 3.6 μM DOX and incubated for 4 hrs or 12 hrs. By the end of incubation time, Total RNA was extracted from cells using the RNeasy Plus Mini Kit (QIAGEN-UK) according to the instructions of manufacturer. RNA samples were sent to the Central Biotechnology Services (Cardiff University, UK), for quality control, synthesis of biotin-labeled cRNA, and hybridisation against Illumina Human-HT12 (Illumina, Inc., Hayward, CA) chips. Washed chips were scanned with Bead Station 500x (Illumina) and the signal intensities quantified with BeadStudio (Illumina). To ensure high quality results from the microarray data, RNA samples were extracted from three independent treatments, giving a biological n = 3 per treatment which gives statistical power to increase the confidence of the results/conclusions generated from the microarray experiment.

Dataset Information

Steatosis-induced proteins adducts with lipid peroxidation products and nuclear electrophilic stress in hepatocytes.

Publications

Steatosis-induced proteins adducts with lipid peroxidation products and nuclear electrophilic stress in hepatocytes.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets