The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform
Aldo Milzani - One of the best experts on this subject based on the ideXlab platform.
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Protein Carbonylation in human bronchial epithelial cells exposed to cigarette smoke extract
Cell Biology and Toxicology, 2019Co-Authors: Graziano Colombo, Daniela Giustarini, Ranieri Rossi, Aldo Milzani, Maria Lisa Garavaglia, Emanuela Astori, Isabella Dalle-donneAbstract:Cigarette smoke is a well-established exogenous risk factor containing toxic reactive molecules able to induce oxidative stress, which in turn contributes to smoking-related diseases, including cardiovascular, pulmonary, and oral cavity diseases. We investigated the effects of cigarette smoke extract on human bronchial epithelial cells. Cells were exposed to various concentrations (2.5–5–10–20%) of cigarette smoke extract for 1, 3, and 24 h. Carbonylation was assessed by 2,4-dinitrophenylhydrazine using both immunocytochemical and Western immunoblotting assays. Cigarette smoke induced increasing Protein Carbonylation in a concentration-dependent manner. The main carbonylated Proteins were identified by means of two-dimensional electrophoresis coupled to MALDI-TOF mass spectrometry analysis and database search (redox proteomics). We demonstrated that exposure of bronchial cells to cigarette smoke extract induces Carbonylation of a large number of Proteins distributed throughout the cell. Proteins undergoing Carbonylation are involved in primary metabolic processes, such as Protein and lipid metabolism and metabolite and energy production as well as in fundamental cellular processes, such as cell cycle and chromosome segregation, thus confirming that reactive carbonyl species contained in cigarette smoke markedly alter cell homeostasis and functions.
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Protein Carbonylation in human smokers and mammalian models of exposure to cigarette smoke focus on redox proteomic studies
Antioxidants & Redox Signaling, 2017Co-Authors: Isabella Dalledonne, Graziano Colombo, Daniela Giustarini, Ranieri Rossi, Rosalba Gornati, Giovanni Bernardini, Maria Lisa Garavaglia, Nicola Portinaro, Aldo MilzaniAbstract:Abstract Significance: Oxidative stress is one mechanism whereby tobacco smoking affects human health, as reflected by increased levels of several biomarkers of oxidative stress/damage isolated from tissues and biological fluids of active and passive smokers. Many investigations of cigarette smoke (CS)-induced oxidative stress/damage have been carried out in mammalian animal and cellular models of exposure to CS. Animal models allow the investigation of many parameters that are similar to those measured in human smokers. In vitro cell models may provide new information on molecular and functional differences between cells of smokers and nonsmokers. Recent Advances: Over the past decade or so, a growing number of researches highlighted that CS induces Protein Carbonylation in different tissues and body fluids of smokers as well as in in vivo and in vitro models of exposure to CS. Critical Issues: We review recent findings on Protein Carbonylation in smokers and models thereof, focusing on redox proteomic s...
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a step by step protocol for assaying Protein Carbonylation in biological samples
Journal of Chromatography B, 2016Co-Authors: Graziano Colombo, Marco Clerici, Maria Elisa Garavaglia, Daniela Giustarini, Ranieri Rossi, Aldo Milzani, Isabella DalledonneAbstract:Protein Carbonylation represents the most frequent and usually irreversible oxidative modification affecting Proteins. This modification is chemically stable and this feature is particularly important for storage and detection of carbonylated Proteins. Many biochemical and analytical methods have been developed during the last thirty years to assay Protein Carbonylation. The most successful method consists on Protein carbonyl (PCO) derivatization with 2,4-dinitrophenylhydrazine (DNPH) and consequent spectrophotometric assay. This assay allows a global quantification of PCO content due to the ability of DNPH to react with carbonyl giving rise to an adduct able to absorb at 366 nm. Similar approaches were also developed employing chromatographic separation, in particular HPLC, and parallel detection of absorbing adducts. Subsequently, immunological techniques, such as Western immunoblot or ELISA, have been developed leading to an increase of sensitivity in Protein Carbonylation detection. Currently, they are widely employed to evaluate change in total Protein Carbonylation and eventually to highlight the specific Proteins undergoing selective oxidation. In the last decade, many mass spectrometry (MS) approaches have been developed for the identification of the carbonylated Proteins and the relative amino acid residues modified to carbonyl derivatives. Although these MS methods are much more focused and detailed due to their ability to identify the amino acid residues undergoing Carbonylation, they still require too expensive equipments and, therefore, are limited in distribution. In this protocol paper, we summarise and comment on the most diffuse protocols that a standard laboratory can employ to assess Protein Carbonylation; in particular, we describe step-by-step the different protocols, adding suggestions coming from our on-bench experience.
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Protein Carbonylation in human endothelial cells exposed to cigarette smoke extract
Toxicology Letters, 2013Co-Authors: Rosalba Gornati, Graziano Colombo, Marco Clerici, Isabella Dalledonne, Roberto Colombo, Federica Rossi, Nicoletta Gagliano, C Riva, Giovanni Bernardini, Aldo MilzaniAbstract:Cigarette smoke is a significant independent risk factor for vascular diseases and is a leading cause of structural and functional alterations of the vascular endothelium. In this study, we show Protein Carbonylation in the human umbilical vein endothelial cell line (ECV-304) exposed to whole-phase cigarette smoke extract. The main carbonylated Proteins, including cytoskeletal Proteins, glycolytic enzymes, xenobiotic metabolizing and antioxidant enzymes, and endoplasmic reticulum Proteins, were identified by means of two-dimensional electrophoresis and Matrix-Assisted Laser Desorption/Ionization-Time of Flight (MALDI-TOF) mass spectrometry (redox proteomics). Morphological analyses by fluorescence microscopy evidenced alterations in the microtubule cytoskeleton, especially at longer exposure time to cigarette smoke extract. Morphological analyses by transmission electron microscopy showed vacuolisation of the cytoplasm, alteration of mitochondria ultrastructure, and some enlargement of the perinuclear space. The possible role played by Protein Carbonylation caused by reactive species contained in cigarette smoke in the cigarette smoke-induced endothelial injury is discussed.
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intervention strategies to inhibit Protein Carbonylation by lipoxidation derived reactive carbonyls
Medicinal Research Reviews, 2007Co-Authors: Giancarlo Aldini, Aldo Milzani, Isabella Dalledonne, Roberto Maffei Facino, Marina CariniAbstract:Protein Carbonylation induced by reactive carbonyl species (RCS) generated by peroxidation of polyunsaturated fatty acids plays a significant role in the etiology and/or progression of several human diseases, such as cardiovascular (e.g., atherosclerosis, long-term complications of diabetes) and neurodegenerative diseases (e.g., Alzheimer's disease, Parkinson's disease, and cerebral ischemia). Most of the biological effects of intermediate RCS, mainly alpha,beta-unsaturated aldehydes, di-aldehydes, and keto-aldehydes, are due to their capacity to react with the nucleophilic sites of Proteins, forming advanced lipoxidation end-products (ALEs). Because of the emerging deleterious role of RCS/Protein adducts in several human diseases, different potential therapeutic strategies have been developed in the last few years. This review sheds focus on fundamental studies on lipid-derived RCS generation, their biological effects, and their reactivity with Proteins, with particular emphasis to 4-hydroxy-trans-2-nonenal (HNE)-, acrolein (ACR)-, malondialdehyde (MDA)-, and glyoxal (GO)-modified Proteins. It also discusses the recently developed pharmacological approaches for the management of chronic diseases in which oxidative stress and RCS formation are massively involved. Inhibition of ALE formation, based on carbonyl-sequestering agents, seems to be the most promising pharmacological tool and is reviewed in detail.
David A. Bernlohr - One of the best experts on this subject based on the ideXlab platform.
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obesity induced Protein Carbonylation in murine adipose tissue regulates the dna binding domain of nuclear zinc finger Proteins
Journal of Biological Chemistry, 2018Co-Authors: Amy K Hauck, Wendy S. Hahn, Tong Zhou, Raphael Petegrosso, Rui Kuang, Yue Chen, David A. BernlohrAbstract:In obesity-linked insulin resistance, oxidative stress in adipocytes leads to lipid peroxidation and subsequent Carbonylation of Proteins by diffusible lipid electrophiles. Reduction in oxidative stress attenuates Protein Carbonylation and insulin resistance, suggesting that lipid modification of Proteins may play a role in metabolic disease, but the mechanisms remain incompletely understood. Herein, we show that in vivo, diet-induced obesity in mice surprisingly results in preferential Carbonylation of nuclear Proteins by 4-hydroxy-trans-2,3-nonenal (4-HNE) or 4-hydroxy-trans-2,3-hexenal (4-HHE). Proteomic and structural analyses revealed that residues in or around the sites of zinc coordination of zinc finger Proteins, such as those containing the C2H2 or MATRIN, RING, C3H1, or N4-type DNA-binding domains, are particularly susceptible to Carbonylation by lipid aldehydes. These observations strongly suggest that Carbonylation functionally disrupts Protein secondary structure supported by metal coordination. Analysis of one such target, the nuclear Protein estrogen-related receptor γ (ERR-γ), showed that ERR-γ is modified by 4-HHE in the obese state. In vitro Carbonylation decreased the DNA-binding capacity of ERR-γ and correlated with the obesity-linked down-regulation of many key genes promoting mitochondrial bioenergetics. Taken together, these findings reveal a novel mechanistic connection between oxidative stress and metabolic dysfunction arising from Carbonylation of nuclear zinc finger Proteins, such as the transcriptional regulator ERR-γ.
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mitochondrial oxidative stress induced by downregulation of antioxidant enzymes leads to nuclear Protein Carbonylation by retrograde signaling in 3t3 l1 adipocytes
Free Radical Biology and Medicine, 2016Co-Authors: Rocio Fonce, Abby Axelson, Madeleine Hart, Amy K Hauck, David A. BernlohrAbstract:Obesity-linked insulin resistance is mechanistically connected to local inflammation of adipose tissue, which produces a metabolic state characterized by oxidative stress and mitochondrial dysfunction. Antioxidants enzymes such as Glutathione S-transferase A4 (GSTA4), peroxiredoxin 3 (Prdx3) and glutathione peroxidase 4 (GPx4) expressions are selectively downregulated in adipose tissue of obese insulin-resistant mice and in human obesity-linked insulin resistance. Also, TNFα treatment of 3T3-L1 adipocytes resulted in decreased expression of GSTA4, GPx4, and Prdx3 and increased Protein Carbonylation. In addition, Protein Carbonylation is implicated as an initiating factor in mitochondrial dysfunction and ER-stress, providing a mechanistic connection between oxidative stress and metabolic disease. Histones are the primary components of chromatin and are notably susceptible to Carbonylation because of their long lysine-rich tails. These modifications may have effects on histone code, leading to long lasting implications of human health, including insulin resistance. In this study GSTA4-Prdx3-GPx4-silenced 3T3-L1 adipocytes were evaluated for reactive oxygen species production (ROS), mitochondrial function and histones Carbonylation. Downregulation of GSTA4, Prdx3 and GPx4 led to an significant increase in ROS, a significant increase in H3 and H4 histones Carbonylation, and mitochondrial dysfunction. These results indicate that a downregulation of antioxidant enzymes in adipocytes leads to increased ROS production, mitochondrial dysfunction and nuclear Protein Carbonylation, and may contribute to the development of insulin resistance and type 2 diabetes.
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roux en y gastric bypass acutely decreases Protein Carbonylation and increases expression of mitochondrial biogenesis genes in subcutaneous adipose tissue
Obesity Surgery, 2015Co-Authors: Cyrus Jahansouz, Brigitte I. Frohnert, Federico J. Serrot, Rocio Foncea, David A. Bernlohr, Robert B Dorman, Bridget Slusarek, Daniel B Leslie, Sayeed IkramuddinAbstract:Background Mitochondrial dysfunction in adipose tissue has been implicated as a pathogenic step in the development of type 2 diabetes mellitus (T2DM). In adipose tissue, chronic nutrient overload results in mitochondria driven increased reactive oxygen species (ROS) leading to Carbonylation of Proteins that impair mitochondrial function and downregulation of key genes linked to mitochondrial biogenesis. In patients with T2DM, Roux-en-Y gastric bypass (RYGB) surgery leads to improvements in glycemic profile prior to significant weight loss. Consequently, we hypothesized that improved glycemia early after RYGB would be paralleled by decreased Protein Carbonylation and increased expression of genes related to mitochondrial biogenesis in adipose tissue.
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detecting Protein Carbonylation in adipose tissue and in cultured adipocytes
Methods in Enzymology, 2014Co-Authors: Wendy S. Hahn, David A. BernlohrAbstract:Reactive oxygen species-mediated attack of the acyl chains of polyunsaturated fatty acids and triglycerides leads to the formation of lipid hydroperoxides. Lipid hydroperoxides are subject to nonenzymatic Fenton chemistry producing a variety of reactive aldehydes that covalently modify Proteins in a reaction referred to as Protein Carbonylation. Given the significant content of triglycerides in fat tissue, adipose Proteins are among the most heavily carbonylated. The laboratory has utilized two methodologies for the detection of Protein Carbonylation in tissue- and cell-based samples. The first utilizes biotin coupled to a hydrazide moiety and takes advantage of the numerous biotin detection systems. The second method utilizes an anti 4-hydroxy-trans-2,3-nonenal (4-HNE)-directed antibody that can detect both 4-HNE and the corresponding 4-oxo derivative when the samples are reduced. Using such methods, we have evaluated the profile of carbonylated Proteins in epididymal white adipose tissue and 3T3-L1 adipocytes using both methods. In addition, we have investigated the effects of two antidiabetic drugs, pioglitazone and metformin, on Protein Carbonylation in 3T3-L1 adipocytes. Overall, the biotin hydrazide method is rapid, inexpensive, and easy to use, but its usefulness is limited because it detects a wide variety of carbonylated derivatives, which makes assignments of individual Proteins difficult. Compared to the biotin hydrazide method, the anti-HNE antibody method detects specific Proteins more readily but identifies only a subset of carbonylated Proteins. As such, the combination of both methods allows for a comprehensive evaluation of Protein Carbonylation plus provides a means towards identification of specific Carbonylation targets.
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oxidative stress and Protein Carbonylation in adipose tissue implications for insulin resistance and diabetes mellitus
Journal of Proteomics, 2013Co-Authors: Tatjana Ruskovska, David A. BernlohrAbstract:While historically considered simply as a depot for excess energy, white adipose tissue is a dynamically active endocrine organ capable of responding to a variety of efferent stimuli resulting in the synthesis and secretion of peptides, Proteins and metabolites that serve as signal transducers to the peripheral and central circulation. Such regulation controls a variety of physiological processes including energy expenditure, food intake, reproductive capacity and responsiveness to insulin. Indeed, the accumulation of inflammatory cells in white adipose tissue is considered to be causative in the development of insulin resistance and eventually type 2 diabetes mellitus. A large body of evidence suggests that oxidative stress in adipose tissue not only correlates with insulin resistance but is also causative in its development. Moreover, using the available plasma oxidative stress biomarkers, many clinical studies have shown the presence of systemic oxidative stress in obese insulin resistant subjects, and its decrease after the successful treatment of obesity. In this review we emphasize the role of Protein Carbonylation in dysfunctional obese white adipose tissue and its metabolic implications. We focus on glutathione S-transferase A4 as the key enzyme for trans-4-hydroxy-2-nonenal and trans-4-oxo-2-nonenal removal from the cell, thus preventing Protein Carbonylation. This article is part of a Special Issue entitled: Posttranslational Protein modifications in biology and Medicine.
Isabella Dalledonne - One of the best experts on this subject based on the ideXlab platform.
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Protein Carbonylation in human smokers and mammalian models of exposure to cigarette smoke focus on redox proteomic studies
Antioxidants & Redox Signaling, 2017Co-Authors: Isabella Dalledonne, Graziano Colombo, Daniela Giustarini, Ranieri Rossi, Rosalba Gornati, Giovanni Bernardini, Maria Lisa Garavaglia, Nicola Portinaro, Aldo MilzaniAbstract:Abstract Significance: Oxidative stress is one mechanism whereby tobacco smoking affects human health, as reflected by increased levels of several biomarkers of oxidative stress/damage isolated from tissues and biological fluids of active and passive smokers. Many investigations of cigarette smoke (CS)-induced oxidative stress/damage have been carried out in mammalian animal and cellular models of exposure to CS. Animal models allow the investigation of many parameters that are similar to those measured in human smokers. In vitro cell models may provide new information on molecular and functional differences between cells of smokers and nonsmokers. Recent Advances: Over the past decade or so, a growing number of researches highlighted that CS induces Protein Carbonylation in different tissues and body fluids of smokers as well as in in vivo and in vitro models of exposure to CS. Critical Issues: We review recent findings on Protein Carbonylation in smokers and models thereof, focusing on redox proteomic s...
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a step by step protocol for assaying Protein Carbonylation in biological samples
Journal of Chromatography B, 2016Co-Authors: Graziano Colombo, Marco Clerici, Maria Elisa Garavaglia, Daniela Giustarini, Ranieri Rossi, Aldo Milzani, Isabella DalledonneAbstract:Protein Carbonylation represents the most frequent and usually irreversible oxidative modification affecting Proteins. This modification is chemically stable and this feature is particularly important for storage and detection of carbonylated Proteins. Many biochemical and analytical methods have been developed during the last thirty years to assay Protein Carbonylation. The most successful method consists on Protein carbonyl (PCO) derivatization with 2,4-dinitrophenylhydrazine (DNPH) and consequent spectrophotometric assay. This assay allows a global quantification of PCO content due to the ability of DNPH to react with carbonyl giving rise to an adduct able to absorb at 366 nm. Similar approaches were also developed employing chromatographic separation, in particular HPLC, and parallel detection of absorbing adducts. Subsequently, immunological techniques, such as Western immunoblot or ELISA, have been developed leading to an increase of sensitivity in Protein Carbonylation detection. Currently, they are widely employed to evaluate change in total Protein Carbonylation and eventually to highlight the specific Proteins undergoing selective oxidation. In the last decade, many mass spectrometry (MS) approaches have been developed for the identification of the carbonylated Proteins and the relative amino acid residues modified to carbonyl derivatives. Although these MS methods are much more focused and detailed due to their ability to identify the amino acid residues undergoing Carbonylation, they still require too expensive equipments and, therefore, are limited in distribution. In this protocol paper, we summarise and comment on the most diffuse protocols that a standard laboratory can employ to assess Protein Carbonylation; in particular, we describe step-by-step the different protocols, adding suggestions coming from our on-bench experience.
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Protein Carbonylation in human endothelial cells exposed to cigarette smoke extract
Toxicology Letters, 2013Co-Authors: Rosalba Gornati, Graziano Colombo, Marco Clerici, Isabella Dalledonne, Roberto Colombo, Federica Rossi, Nicoletta Gagliano, C Riva, Giovanni Bernardini, Aldo MilzaniAbstract:Cigarette smoke is a significant independent risk factor for vascular diseases and is a leading cause of structural and functional alterations of the vascular endothelium. In this study, we show Protein Carbonylation in the human umbilical vein endothelial cell line (ECV-304) exposed to whole-phase cigarette smoke extract. The main carbonylated Proteins, including cytoskeletal Proteins, glycolytic enzymes, xenobiotic metabolizing and antioxidant enzymes, and endoplasmic reticulum Proteins, were identified by means of two-dimensional electrophoresis and Matrix-Assisted Laser Desorption/Ionization-Time of Flight (MALDI-TOF) mass spectrometry (redox proteomics). Morphological analyses by fluorescence microscopy evidenced alterations in the microtubule cytoskeleton, especially at longer exposure time to cigarette smoke extract. Morphological analyses by transmission electron microscopy showed vacuolisation of the cytoplasm, alteration of mitochondria ultrastructure, and some enlargement of the perinuclear space. The possible role played by Protein Carbonylation caused by reactive species contained in cigarette smoke in the cigarette smoke-induced endothelial injury is discussed.
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intervention strategies to inhibit Protein Carbonylation by lipoxidation derived reactive carbonyls
Medicinal Research Reviews, 2007Co-Authors: Giancarlo Aldini, Aldo Milzani, Isabella Dalledonne, Roberto Maffei Facino, Marina CariniAbstract:Protein Carbonylation induced by reactive carbonyl species (RCS) generated by peroxidation of polyunsaturated fatty acids plays a significant role in the etiology and/or progression of several human diseases, such as cardiovascular (e.g., atherosclerosis, long-term complications of diabetes) and neurodegenerative diseases (e.g., Alzheimer's disease, Parkinson's disease, and cerebral ischemia). Most of the biological effects of intermediate RCS, mainly alpha,beta-unsaturated aldehydes, di-aldehydes, and keto-aldehydes, are due to their capacity to react with the nucleophilic sites of Proteins, forming advanced lipoxidation end-products (ALEs). Because of the emerging deleterious role of RCS/Protein adducts in several human diseases, different potential therapeutic strategies have been developed in the last few years. This review sheds focus on fundamental studies on lipid-derived RCS generation, their biological effects, and their reactivity with Proteins, with particular emphasis to 4-hydroxy-trans-2-nonenal (HNE)-, acrolein (ACR)-, malondialdehyde (MDA)-, and glyoxal (GO)-modified Proteins. It also discusses the recently developed pharmacological approaches for the management of chronic diseases in which oxidative stress and RCS formation are massively involved. Inhibition of ALE formation, based on carbonyl-sequestering agents, seems to be the most promising pharmacological tool and is reviewed in detail.
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intervention strategies to inhibit Protein Carbonylation by lipoxidation derived reactive carbonyls
Medicinal Research Reviews, 2007Co-Authors: Giancarlo Aldini, Aldo Milzani, Isabella Dalledonne, Roberto Maffei Facino, Marina CariniAbstract:Protein Carbonylation induced by reactive carbonyl species (RCS) generated by peroxidation of polyunsaturated fatty acids plays a significant role in the etiology and/or progression of several human diseases, such as cardiovascular (e.g., atherosclerosis, long-term complications of diabetes) and neurodegenerative diseases (e.g., Alzheimer's disease, Parkinson's disease, and cerebral ischemia). Most of the biological effects of intermediate RCS, mainly α,β-unsaturated aldehydes, di-aldehydes, and keto-aldehydes, are due to their capacity to react with the nucleophilic sites of Proteins, forming advanced lipoxidation end-products (ALEs). Because of the emerging deleterious role of RCS/Protein adducts in several human diseases, different potential therapeutic strategies have been developed in the last few years. This review sheds focus on fundamental studies on lipid-derived RCS generation, their biological effects, and their reactivity with Proteins, with particular emphasis to 4-hydroxy-trans-2-nonenal (HNE)-, acrolein (ACR)-, malondialdehyde (MDA)-, and glyoxal (GO)-modified Proteins. It also discusses the recently developed pharmacological approaches for the management of chronic diseases in which oxidative stress and RCS formation are massively involved. Inhibition of ALE formation, based on carbonyl-sequestering agents, seems to be the most promising pharmacological tool and is reviewed in detail. © 2006 Wiley Periodicals, Inc. Med Res Rev, 27, No. 6, 817–868, 2007
Giancarlo Aldini - One of the best experts on this subject based on the ideXlab platform.
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key factors regulating Protein Carbonylation by α β unsaturated carbonyls a structural study based on a retrospective meta analysis
Biophysical Chemistry, 2017Co-Authors: Giulio Vistoli, Chiara Mantovani, Silvia Gervasoni, Alessandro Pedretti, Giancarlo AldiniAbstract:Protein Carbonylation represents one of the most important oxidative-based modifications involving nucleophilic amino acids and affecting Protein folding and function. Protein Carbonylation is induced by electrophilic carbonyl species and is an highly selective process since few nucleophilic residues are carbonylated within each Protein. While considering the great interest for Protein Carbonylation, few studies investigated the factors which render a nucleophilic residue susceptible to Carbonylation. Hence, the present study is aimed to delve into the factors which modulate the reactivity of cysteine, histidine and lysine residues towards α,β unsaturated carbonyls by a retrospective analysis of the available studies which identified the adducted residues for Proteins, the structure of which was resolved. Such an analysis involved different parameters including exposure, nucleophilicity, surrounding residues and capacity to attract carbonyl species (as derived by docking simulations). The obtained results allowed a meaningful clustering of the analyzed Proteins suggesting that on average Carbonylation selectivity increases with Protein size. The comparison between adducted and unreactive residues revealed differences in all monitored parameters which are markedly more pronounced for cysteines compared to lysines and histidines. Overall, these results suggest that cysteine's Carbonylation is a finely (and reasonably purposely) modulated process, while the Carbonylation of lysines and histidines seems to be a fairly random event in which limited differences influence their reactivity.
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REACTIVE CARBONYL SPECIES AS POTENTIAL DRUG TARGETS IN PREVENTING Protein Carbonylation AND RELATED CELLULAR DYSFUNCTION
2008Co-Authors: Giancarlo AldiniAbstract:Protein Carbonylation induced by reactive carbonyl species (RCS) generated by peroxidation of polyunsaturated fatty acids plays a significant role in the etiology and/or progression of several human diseases, such as cardiovascular (e.g., atherosclerosis, long-term complications of diabetes) and neurodegenerative diseases (e.g., Alzheimer's disease, Parkinson's disease, and cerebral ischemia)(1). Most of the biological effects of intermediate RCS, mainly alpha,beta-unsaturated aldehydes, di-aldehydes, and keto-aldehydes, are due to their capacity to react with the nucleophilic sites of Proteins, forming advanced lipoxidation end-products (ALEs). Because of the emerging deleterious role of RCS/Protein adducts in several human diseases, different potential therapeutic strategies have been developed in the last few years. The talk will focus on the fundamental studies on lipid-derived RCS generation, their biological effects, and their reactivity with Proteins, with particular emphasis to 4-hydroxy-trans-2-nonenal (HNE)-, acrolein (ACR)-, malondialdehyde (MDA)-, and glyoxal (GO)-modified Proteins (2). It will also consider the recently developed pharmacological approaches for the management of chronic diseases in which oxidative stress and RCS formation are massively involved. The most promising strategy to neutralize/reduce these pathogenetic factors is based on nucleophilic compounds capable to form covalent and unreactive adducts with RCS (RCS sequestering agents) such as pyridoxamine (PYR), hydralazine (HY), dihydralazine (di-HY), aminoguanidine (AG), and metformin (MF)(2). However these compounds are characterized by a severe aspecificty since they react also with physiological aldehydes such as pyridoxal (3). The talk will also describe a new class of RCS-sequestering agents developed in our laboratory, derived form the endogenous peptide Carnosine (\u3b2-alanyl-L-hisitidine), characterized by a significant quenching activity towards electrophilic aldehydes, and high selectivity. The efficacy of this new class of compounds in preventing dyslipidemia, hypertension and kidney damage in Zucker obese rats will be also presented
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Carnosine derivatives as novel RCS sequestering agents in preventing Protein Carbonylation and related cellular dysfunction
2008Co-Authors: Giancarlo AldiniAbstract:Several evidences strongly support a pathogenic role for Reactive Carbonyl Species (RCS), such as in the case of diabetic-related diseases, age-dependent tissue dysfunction, and metabolic distress syndrome. Hence, RCS can be considered a potential biological target for drug discovery. The most promising pharmacological strategy to neutralize/reduce RCS is based on nucleophilic compounds capable to form covalent and unreactive adducts with RCS (RCS sequestering agents) such as pyridoxamine (PYR), hydralazine (HY), dihydralazine (di-HY), and aminoguanidine (AG). However these compounds are characterized by a lack of selectivity since they also react with physiological aldehydes, such as pyridoxal. We recently found that the endogenous dipeptide carnosine (beta-alanyl-L-histidine, CAR) is a specific quencher of alfa,beta-unsaturated aldehydes due to its peculiar mechanism involving the Schiff base formation between the beta-alanine amino group and the RCS aldehyde followed by the Michael adduction between the C3 of the aldehyde and the Ntau of the histidine group. We also found that CAR exogenously given to Zucker obese rats (30 mg/Kg die for 24 weeks) greatly reduces dyslipidemia, hypertension, albuminuria and Protein Carbonylation. However, the therapeutic use of CAR is limited since it is unstable in human plasma due to serum carnosinase. Hence, our interest was to derive carnosine analogues characterized by (i) carnosinase stability and (ii) a grater reactivity towards RCS, even maintaining the same selectivity. The stability was reached by the isomerization of L- to D-histidine, leading to \u3b2-alanyl-D-histidine (D-CAR), which is not recognized by carnosinase, but maintains the same quenching activity of L-CAR. The increase of reactivity was reached by modulating the conformational profile of the Schiff\u2019s base intermediate, in order to favour a close conformation in which the imidazole ring approaches enough the C3 of the Schiff\u2019s base to form the corresponding Michael adduct. A series of D-CAR derivatives was analyzed by in silico approaches to find out those characterized by a favorable folded conformational profile. The most promising were synthesized and the stability and quenching ability evaluated. By this way a set of phenyl derivatives was identified, characterized by high stability in human plasma, and by a three fold HNE-quenching ability increase compared to D-CAR
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intervention strategies to inhibit Protein Carbonylation by lipoxidation derived reactive carbonyls
Medicinal Research Reviews, 2007Co-Authors: Giancarlo Aldini, Aldo Milzani, Isabella Dalledonne, Roberto Maffei Facino, Marina CariniAbstract:Protein Carbonylation induced by reactive carbonyl species (RCS) generated by peroxidation of polyunsaturated fatty acids plays a significant role in the etiology and/or progression of several human diseases, such as cardiovascular (e.g., atherosclerosis, long-term complications of diabetes) and neurodegenerative diseases (e.g., Alzheimer's disease, Parkinson's disease, and cerebral ischemia). Most of the biological effects of intermediate RCS, mainly alpha,beta-unsaturated aldehydes, di-aldehydes, and keto-aldehydes, are due to their capacity to react with the nucleophilic sites of Proteins, forming advanced lipoxidation end-products (ALEs). Because of the emerging deleterious role of RCS/Protein adducts in several human diseases, different potential therapeutic strategies have been developed in the last few years. This review sheds focus on fundamental studies on lipid-derived RCS generation, their biological effects, and their reactivity with Proteins, with particular emphasis to 4-hydroxy-trans-2-nonenal (HNE)-, acrolein (ACR)-, malondialdehyde (MDA)-, and glyoxal (GO)-modified Proteins. It also discusses the recently developed pharmacological approaches for the management of chronic diseases in which oxidative stress and RCS formation are massively involved. Inhibition of ALE formation, based on carbonyl-sequestering agents, seems to be the most promising pharmacological tool and is reviewed in detail.
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intervention strategies to inhibit Protein Carbonylation by lipoxidation derived reactive carbonyls
Medicinal Research Reviews, 2007Co-Authors: Giancarlo Aldini, Aldo Milzani, Isabella Dalledonne, Roberto Maffei Facino, Marina CariniAbstract:Protein Carbonylation induced by reactive carbonyl species (RCS) generated by peroxidation of polyunsaturated fatty acids plays a significant role in the etiology and/or progression of several human diseases, such as cardiovascular (e.g., atherosclerosis, long-term complications of diabetes) and neurodegenerative diseases (e.g., Alzheimer's disease, Parkinson's disease, and cerebral ischemia). Most of the biological effects of intermediate RCS, mainly α,β-unsaturated aldehydes, di-aldehydes, and keto-aldehydes, are due to their capacity to react with the nucleophilic sites of Proteins, forming advanced lipoxidation end-products (ALEs). Because of the emerging deleterious role of RCS/Protein adducts in several human diseases, different potential therapeutic strategies have been developed in the last few years. This review sheds focus on fundamental studies on lipid-derived RCS generation, their biological effects, and their reactivity with Proteins, with particular emphasis to 4-hydroxy-trans-2-nonenal (HNE)-, acrolein (ACR)-, malondialdehyde (MDA)-, and glyoxal (GO)-modified Proteins. It also discusses the recently developed pharmacological approaches for the management of chronic diseases in which oxidative stress and RCS formation are massively involved. Inhibition of ALE formation, based on carbonyl-sequestering agents, seems to be the most promising pharmacological tool and is reviewed in detail. © 2006 Wiley Periodicals, Inc. Med Res Rev, 27, No. 6, 817–868, 2007
V K Dimitriadis - One of the best experts on this subject based on the ideXlab platform.
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effects of organic contaminants in reactive oxygen species Protein Carbonylation and dna damage on digestive gland and haemolymph of land snails
Chemosphere, 2011Co-Authors: A Itziou, Martha Kaloyianni, V K DimitriadisAbstract:The present study focused on early responses of land snails Eobania vermiculata to organic environmental contaminants, by investigating the use of a newly-established method for the measurement of Protein Carbonylation as a new biomarker of terrestrial pollution, as well as by measuring the ROS production and the DNA damage. Land snails were exposed to different concentrations of chlorpyrifos, parathion-methyl or PAHs in vivo or in vitro in the laboratory. The susceptibility of exposed snails was increased in relation to oxidative stress induced by contaminants tested. A statistically significant increase in ROS production, Protein Carbonylation and DNA damage was revealed in the snails treated with pollutants, compared to the untreated ones. The results indicated the effectiveness of measuring ROS production and DNA damage and reinforce the application of the present ELISA method in organic terrestrial pollution biomonitoring studies.
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in vivo and in vitro effects of metals in reactive oxygen species production Protein Carbonylation and dna damage in land snails eobania vermiculata
Archives of Environmental Contamination and Toxicology, 2011Co-Authors: A Itziou, Martha Kaloyianni, V K DimitriadisAbstract:Heavy metals are known to induce oxidative damage by way of enhancement of intracellular reactive oxygen species (ROS) production, which often preludes the onset of alterations, such as Protein Carbonylation and DNA damage. In this study, our aim was to examine the early responses of land snails Eobania vermiculata to environmental contaminants by investigating the use of a modified enzyme-linked immunosorbent assay (ELISA) assay for the measurement of Protein Carbonylation as a new biomarker of terrestrial pollution as well as by measuring ROS production and DNA damage. Land snails were treated with heavy metals―cadmium, lead, or copper―in vivo (15 or 40 ppm) for 25 days or in vitro (0.5, 5, 50 or 500 μM) for 30 min in the laboratory, and the previously mentioned biomarkers were determined in digestive gland and haemolymph of the treated animals. A statistically significant increase in ROS production, Protein Carbonylation, and DNA damage was shown in the snails treated with pollutants compared with the untreated snails. The results indicate the effectiveness of measuring ROS production and DNA damage, as well as using the present ELISA method, as sensitive tools of terrestrial pollution biomonitoring studies. Statistically significant correlations among the previously mentioned techniques further enhance their role as promising biomarkers in terrestrial pollution studies.
