Effect of ferritin-containing fractions with different iron loading on lipid peroxidation.
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ABSTRACT:
SUBMITTER: Gutteridge JM
PROVIDER: S-EPMC1154126 | biostudies-other | 1983 Feb
REPOSITORIES: biostudies-other
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Project description:Cultured chick embryo hepatocytes were iron-loaded with ferric nitrilotriacetate. Iron-loading was confirmed by both quantitative cellular iron determinations and ultrastructural studies. With iron-loading, lipid peroxidation, as detected by malonaldehyde released into the medium, occurred at a linear rate for 12h, after which time the rate of malonaldehyde production decreased. No cell toxicity, as detected by lactate dehydrogenase release, was noted. The amount of malonaldehyde recovered in the medium after 18h of exposure to iron represented 24-33% of the total malonaldehyde that could be produced by incubating lysed cells with iron and ascorbate. Cellular glutathione was not affected by iron-stimulated lipid peroxidation, but was increased by allylisopropylacetamide. Although iron-loading by itself had no effect on activity of 5-aminolaevulinate synthase, the first and rate-limiting step in haem synthesis, iron-loading in the presence of the porphyrogenic drug allylisopropylacetamide increased levels of 5-aminolaevulinate synthase 6-fold over levels induced by the drug alone. The antioxidant, butylated hydroxytoluene, totally inhibited iron-stimulated lipid peroxidation, but did not interfere with the effect of iron-loading to potentiate an increase in 5-aminolaevulinate synthase. After 18h of exposure to iron, followed by a change to fresh medium, the iron remaining within the cells did not stimulate further lipid peroxidation over the following 18h, but did potentiate an increase in 5-aminolaevulinate synthase on exposure to allylisopropylacetamide. It therefore appears that lipid peroxidation is not the mechanism by which iron potentiates induction of hepatic 5-aminolaevulinate synthase.
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Project description:Ferritin and haemosiderin were shown, by the measurement of malondialdehyde production and loss of polyunsaturated fatty acids, to stimulate lipid peroxidation in liposomes. At pH 7.4 ascorbate was additionally required to achieve peroxidation; however, peroxidation occurred at pH 4.5 in the presence of iron-proteins alone. The damage was completely inhibited by the incorporation of chain-breaking antioxidants (alpha-tocopherol and butylated hydroxytoluene) into the liposomes. Metal chelators (desferrioxamine and EDTA) also completely inhibited lipid peroxidation. These and further results indicate that, at pH 4.5, even in the absence of a reducing agent, iron is released from haemosiderin and can mediate oxidative damage to a lipid membrane.
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Project description:Hereditary ferritinopathy (HF) is a neurodegenerative disease characterized by intracellular ferritin inclusion bodies (IBs) and iron accumulation throughout the central nervous system. Ferritin IBs are composed of mutant ferritin light chain as well as wild-type light (Wt-FTL) and heavy chain (FTH1) polypeptides. In vitro studies have shown that the mutant light chain polypeptide p.Phe167SerfsX26 (Mt-FTL) forms soluble ferritin 24-mer homopolymers having a specific structural disruption that explains its functional problems of reduced ability to incorporate iron and aggregation during iron loading. However, because ferritins are usually 24-mer heteropolymers and all three polypeptides are found in IBs, we investigated the properties of Mt-FTL/FTH1 and Mt-FTL/Wt-FTL heteropolymeric ferritins. We show here the facile assembly of Mt-FTL and FTH1 subunits into soluble ferritin heteropolymers, but their ability to incorporate iron was significantly reduced relative to Wt-FTL/FTH1 heteropolymers. In addition, Mt-FTL/FTH1 heteropolymers formed aggregates during iron loading, contrasting Wt-FTL/FTH1 heteropolymers and similar to what was seen for Mt-FTL homopolymers. The resulting precipitate contained both Mt-FTL and FTH1 polypeptides as do ferritin IBs in patients with HF. The presence of Mt-FTL subunits in Mt-FTL/Wt-FTL heteropolymers also caused iron loading-induced aggregation relative to Wt-FTL homopolymers, with the precipitate containing Mt- and Wt-FTL polypeptides again paralleling HF. Our data demonstrate that co-assembly with wild-type subunits does not circumvent the functional problems caused by mutant subunits. Furthermore, the functional problems characterized here in heteropolymers that contain mutant subunits parallel those problems previously reported in homopolymers composed exclusively of mutant subunits, which strongly suggests that the structural disruption characterized previously in Mt-FTL homopolymers occurs in a similar manner and to a significant extent in both Mt-FTL/FTH1 and Mt-FTL/Wt-FTL heteropolymers.
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Project description:Traditional concepts of life sciences consider oxidative stress as a fundamental process of aging and various diseases including cancer, whereas traditional medicine recommends dietary intake of iron to support physiological functions of the organism. However, due to its strong pro-oxidative capacity, if not controlled well, iron can trigger harmful oxidative stress manifested eventually by toxic chain reactions of lipid peroxidation. Such effects of iron are considered to be major disadvantages of uncontrolled iron usage, although ferroptosis seems to be an important defense mechanism attenuating cancer development. Therefore, a variety of iron-containing nanoparticles were developed for experimental radio-, chemo-, and photodynamic as well as magnetic dynamic nanosystems that alter redox homeostasis in cancer cells. Moreover, studies carried over recent decades have revealed that even the end products of lipid peroxidation, represented by 4-hydroxynonenal (4-HNE), could have desirable effects even acting as kinds of selective anticancer substances produced by non-malignant cells for defense again invading cancer. Therefore, advanced nanotechnologies should be developed for using iron to trigger targeted lipid peroxidation as an anticancer option of integrative biomedicine.
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Project description:Ferritin is a protein that stores and releases iron to prevent diseases associated with iron dysregulation in plants, animals, and bacteria. The conversion between iron-loaded holo-ferritin and empty apo-ferritin is an important process for iron regulation. To date, studies of ferritin have used either ensemble measurements to quantify the characteristics of a large number of proteins or single-molecule approaches to interrogate labeled or modified proteins. Here we demonstrate the first real-time study of the dynamics of iron ion loading and biomineralization within a single, unlabeled ferritin protein. Using optical nanotweezers, we trapped single apo- and holo-ferritins indefinitely, distinguished one from the other, and monitored their structural dynamics in real time. The study presented here deepens the understanding of the iron uptake mechanism of ferritin proteins, which may lead to new therapeutics for iron-related diseases.
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Project description:The aim of this study was to evaluate metal binding and antioxidant activities of hydrolyzed oat bran proteins followed by the determination of peptide sequences. Protamex oat bran protein hydrolysates (OBPH) were separated by reverse-phase HPLC into eight peptide fractions (F1-F8) and their abilities to either chelate metals (Fe2+, Ca2+) or prevent the oxidation of lipids were investigated. In the Fe2+ chelation assay, OBPH had significantly (p < 0.05) higher activity (39.7 %) than the best performed fraction F7 (22.8 %). The second most active was F5 with 12.1 % chelating activity and this was higher than the activity of the tripeptide glutathione (5.8 %) used as control. The two most Fe2+ chelating fractions (F5, F7) however had weak calcium binding (0.6-1.0 %) properties at peptide concentration ranging from 0.2 to 1.0 mg/mL. In the lipid peroxidation assay, OBPH and all HPLC fractions prevented the oxidation of linoleic acid. More than 60 peptides mainly derived from globulin and avenin proteins were identified using tandem mass spectrometry.
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Project description:Taurine is a nonproteinogenic amino sulfonic acid in mammals. Interestingly, skeletal muscle is unable to synthesize taurine endogenously, and the processing of muscular taurine changes throughout ageing and under specific pathophysiological conditions, such as muscular dystrophy. Ageing and disease are also associated with altered iron metabolism, especially when there is an excess of labile iron. The present study addresses the question of whether taurine connects cytoprotective effects and redox homeostasis in a previously unknown iron-dependent manner. Using cultured differentiated C2C12 myotubes, the impact of taurine on markers of lipid peroxidation, redox-sensitive enzymes and iron-related proteins was studied. Significant increases in the heme protein myoglobin and the iron storage protein ferritin were observed in response to taurine treatment. Taurine supplementation reduced lipid peroxidation and BODIPY oxidation by ~60 and 25%, respectively. Furthermore, the mRNA levels of redox-sensitive heme oxygenase (Hmox1), catalase (Cat) and glutamate-cysteine ligase (Gclc) and the total cellular glutathione content were lower in taurine-supplemented cells than they were in the control cells. We suggest that taurine may inhibit the initiation and propagation of lipid peroxidation by lowering basal levels of cellular stress, perhaps through reduction of the cellular labile iron pool.
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Project description:Protein aggregation and abnormal lipid homeostasis are both implicated in neurodegeneration through unknown mechanisms. Here we demonstrate that aggregate-membrane interaction is critical to induce a form of cell death called ferroptosis. Importantly, the aggregate-membrane interaction that drives ferroptosis depends both on the conformational structure of the aggregate, as well as the oxidation state of the lipid membrane. We generated human stem cell-derived models of synucleinopathy, characterized by the intracellular formation of α-synuclein aggregates that bind to membranes. In human iPSC-derived neurons with SNCA triplication, physiological concentrations of glutamate and dopamine induce abnormal calcium signaling owing to the incorporation of excess α-synuclein oligomers into membranes, leading to altered membrane conductance and abnormal calcium influx. α-synuclein oligomers further induce lipid peroxidation. Targeted inhibition of lipid peroxidation prevents the aggregate-membrane interaction, abolishes aberrant calcium fluxes, and restores physiological calcium signaling. Inhibition of lipid peroxidation, and reduction of iron-dependent accumulation of free radicals, further prevents oligomer-induced toxicity in human neurons. In summary, we report that peroxidation of polyunsaturated fatty acids underlies the incorporation of β-sheet-rich aggregates into the membranes, and that additionally induces neuronal death. This suggests a role for ferroptosis in Parkinson's disease, and highlights a new mechanism by which lipid peroxidation causes cell death.
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Project description:One approach to magnetogenetics uses radiofrequency (RF) waves to activate transient receptor potential channels (TRPV1 and TRPV4) that are coupled to cellular ferritins. The mechanisms underlying this effect are unclear and controversial. Theoretical calculations suggest that the heat produced by RF fields is likely orders of magnitude weaker than needed for channel activation. Using the FeRIC (Ferritin iron Redistribution to Ion Channels) system, we have uncovered a mechanism of activation of ferritin-tagged channels via a biochemical pathway initiated by RF disturbance of ferritin and mediated by ferritin-associated iron. We show that, in cells expressing TRPVFeRIC channels, RF increases the levels of the labile iron pool in a ferritin-dependent manner. Free iron participates in chemical reactions, producing reactive oxygen species and oxidized lipids that ultimately activate the TRPVFeRIC channels. This biochemical pathway predicts a similar RF-induced activation of other lipid-sensitive TRP channels and may guide future magnetogenetic designs.
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