Docosahexaenoic acid (DHA) is an n-3 polyunsaturated fatty acid that is highly enriched in the brain, and the oxidation products of DHA are present or increased during neurodegenerative disease progression. The characterization of the oxidation products of DHA is critical to understanding the roles that these products play in the development of such diseases. In this study, we developed a sensitive and specific analytical tool for the detection and quantification of twelve major DHA hydroperoxide (HpDoHE) and hydroxide (HDoHE) isomers (isomers at positions 4, 5, 7, 8, 10, 11, 13, 14, 16, 17, 19 and 20) in biological systems. In this study, HpDoHE were synthesized by photooxidation, and the corresponding hydroxides were obtained by reduction with NaBH4. The isolated isomers were characterized by LC-MS/MS, and unique and specific fragment ions were chosen to construct a selected reaction monitoring (SRM) method for the targeted quantitative analysis of each HpDoHE and HDoHE isomer. The detection limits for the LC-MS/MS-SRM assay were 1−670 pg for HpDoHE and 0.5−8.5 pg for HDoHE injected onto a column. Using this method, it was possible to detect the basal levels of HDoHE isomers in both rat plasma and brain samples. Therefore, the developed LC-MS/MS-SRM can be used as an important tool to identify and quantify the hydro(pero)xy derivatives of DHA in biological system and may be helpful for the oxidative lipidomic studies.
Mitochondrial cholesterol has been reported to be increased under specific pathological conditions associated with enhanced oxidative stress parameters. In this scenario, cholesterol oxidation would be increased, leading to the production of reactive aldehydes, including cholesterol carboxyaldehyde (ChAld). By using SDS micelles as a mitochondrial mimetic model, we have demonstrated that ChAld covalently modifies cytochrome c (cytc), a protein known to participate in electron transport and apoptosis signaling. This mimetic model induces changes in cytc structure in the same way as mitochondrial membranes do. Tryptic digestion of the cytc-ChAld adduct followed by MALDI-TOF/TOF analyses revealed that modifications occur at Lys residues (K22) localized at cytc site L, a site involved in protein-protein and protein-membrane interactions. Interestingly, ChAld ligation prevented cytc detachment from liposomes even under high ionic strength conditions. Overall, it can be concluded that ChAld ligation to Lys residues at site L creates a hydrophobic tail at cytc, which promotes cytc anchoring to the membrane. Although not investigated in detail in this study, cytc adduction to cholesterol derived aldehydes could have implications in cytc release from mitochondria under apoptotic stimuli.
Lipid peroxidation is a well-known process that has been implicated in many diseases. Recent evidence has shown that mitochondrial cholesterol levels are increased under specific conditions, making it an important target for peroxidation inside the mitochondria. Cholesterol peroxidation generates, as primary products, several hydroperoxides (ChOOH), which can react with transition metals and metalloproteins. In this sense, cytochrome c (CYTC), a heme protein largely found in the mitochondria, becomes a candidate to react with ChOOH. Using CYTC associated with SDS micelles to mimic mitochondrial conditions, we show that ChOOH induces dose-dependent CYTC Soret band bleaching, indicating that it is using ChOOH as a substrate. This reaction leads to protein oligomerization, suggesting the formation of a protein radical that, subsequently, recombines, giving dimers, trimers, and tetramers. EPR experiments confirmed the production of carbon-centered radicals from both protein and lipid in the presence of ChOOH. Similar results were obtained with linoleic acid hydroperoxides (LAOOH). In addition, replacing SDS micelles by cardiolipin-containing liposomes as the mitochondrial mimetic led to similar results with either ChOOH or LAOOH. Importantly, kinetic experiments show that CYTC bleaching is faster with ChOOH than with H2O2, suggesting that these hydroperoxides could be relevant substrates for CYTC peroxidase-like activity in biological media. Altogether, these results show that CYTC induces homolytic cleavage of lipid-derived hydroperoxides, producing lipid and protein radicals.
Recebido em 16/8/10; aceito em 20/4/11; publicado na web em 8/7/11 METABOLISM, OXIDATION AND BIOLOGICAL IMPLICATIONS OF DOCOSAHEXAENOIC ACID IN NEURODEGENERATIVE DISEASES. Docosahexaenoic acid (C22:6, n-3, DHA) is a polyunsaturated fatty acid (PUFA) present in large concentrations in the brain and, due to the presence of six double bonds in its structure, is highly susceptible to oxidation by enzymes and reactive oxygen/nitrogen species. The peroxidation of PUFAs has been implicated in an increasing number of human disorders, including neurodegenerative diseases. Hence, a better understanding of the metabolism pathways of DHA should provide new insights about its role in neurodegenerative diseases. Here we review the main aspects related to DHA metabolism, as well as, the recent findings showing its association with neurodegenerative diseases.Keywords: docosahexaenoic acid; metabolism; neurodegenerative diseases. INTRODUÇÃOOs lipídios constituem um grupo heterogêneo de compostos que têm papel crucial na célula, cuja característica comum é a solubilidade em solventes orgânicos e insolubilidade na água. As funções biológicas dos mesmos são tão diversas quanto a sua química. Sua importância pode ser notada pelo número crescente de estudos apontando o envolvimento de seus produtos de oxidação e de disfunções no metabolismo de lipídios em doenças crônico-degenerativas. 1-4Os ácidos graxos são ácidos carboxílicos com cadeias de hidrocarbonetos contendo de 4 a 36 carbonos, sendo encontrados principalmente esterificados a triglicerídios, colesterol e fosfolipídios constituintes das membranas celulares. Eles podem ser classificados em saturados (contendo apenas ligações simples), monoinsaturados (uma ligação dupla) ou poli-insaturados (duas ou mais ligações duplas). Os ácidos graxos poli-insaturados podem ser representados usando-se a abreviação estrutural que os identifica única e exclusivamente através do número de carbonos e de duplas ligações. Também podem ser denominados a partir da posição das ligações duplas, em relação ao grupamento metila terminal, utilizando-se para tanto a denominação ω ou n, 5 sendo a denominação n a recomendada pela IUPAC. 6Estudos mostram que os ácidos graxos poli-insaturados n-3 (ômega-3) e n-6 (ômega-6) têm efeitos distintos nas células, sendo o primeiro associado a efeitos benéficos e o segundo, a efeitos deletérios. 7,8 Os ácidos linoleico (18:2n-6) e linolênico (18:3n-3) são os precursores dos ácidos graxos n-6 e n-3, respectivamente. São considerados essenciais por não serem sintetizados de novo pelos tecidos de vertebrados, sendo necessária a ingestão destes pela dieta. 9O ácido docosahexaenoico (4,7,10,13,16,19-22:6, DHA) é um ácido graxo ômega-3 de 22 carbonos e 6 insaturações. Está enriquecido nas membranas neurais do córtex cerebral e na retina, 10,11 onde 25-35% se encontram esterificados em aminofosfolipídios como a fosfatidilserina, fosfatidiletanolamina e em plasmalogênios.12,13 O DHA constitui > 17% do peso total de ácidos graxos no cérebro de ratos adultos e > 33% do total ...
Docosahexaenoic acid (C22:6, n-3, DHA) is a polyunsaturated fatty acid highly enriched in the brain. This fatty acid can be easily oxidized yielding hydroperoxides as primary products. Cu, Zn-Superoxide dismutase (SOD1) aggregation is a common hallmark of Amyotrophic Lateral Sclerosis (ALS) and the molecular mechanisms behind their formation are not completely understood. Here we investigated the effect of DHA and its hydroperoxides (DHAOOH) on human SOD1 oligomerization in vitro. DHA induced the formation of high-molecular-weight (HMW) SOD1 species (>700 kDa). Aggregation was dependent on free thiols and occurred primarily with the protein in its apo-form. SOD1 incubation with DHA was accompanied by changes in protein structure leading to exposure of protein hydrophobic patches and formation of non-amyloid aggregates. Site-directed mutagenesis studies demonstrated that Cys 6 and Cys 111 in wild-type and Cys 6 in ALS-linked G93A mutant are required for aggregation. In contrast, DHAOOH did not induce HMW species formation but promoted abnormal covalent dimerization of apo-SOD1 that was resistant to SDS and thiol reductants. Overall, our data demonstrate that DHA and DHAOOH induce distinct types of apo-SOD1 oligomerization leading to the formation of HMW and low-molecular-weight species, respectively.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.