The Effect of Fat Intake with Increased Omega-6-to-Omega-3 Polyunsaturated Fatty Acid Ratio in Animal Models of Early and Late Alzheimer's Disease-like Pathogenesis.

This work aims to clarify the effect of dietary polyunsaturated fatty acid (PUFA) intake on the adult brain affected by amyloid pathology. McGill-R-Thy1-APP transgenic (Tg) rat and 5xFAD Tg mouse models that represent earlier or later disease stages were employed. The animals were exposed to a control diet (CD) or an HFD based on corn oil, from young (rats) or adult (mice) ages for 24 or 10 weeks, respectively. In rats and mice, the HFD impaired reference memory in wild-type (WT) animals but did not worsen it in Tg, did not cause obesity, and did not increase triglycerides or glucose levels. Conversely, the HFD promoted stronger microglial activation in Tg vs. WT rats but had no effect on cerebral amyloid deposition. IFN-γ, IL-1ß, and IL-6 plasma levels were increased in Tg rats, regardless of diet, while CXCL1 chemokine levels were increased in HFD-fed mice, regardless of genotype. Hippocampal 3-nitrotyrosine levels tended to increase in HFD-fed Tg rats but not in mice. Overall, an HFD with an elevated omega-6-to-omega-3 ratio as compared to the CD (25:1 vs. 8.4:1) did not aggravate the outcome of AD regardless of the stage of amyloid pathology, suggesting that many neurobiological processes relevant to AD are not directly dependent on PUFA intake.

4-Hydroxynonenal impairs miRNA maturation in heart failure via Dicer post-translational modification.

BACKGROUND AND AIMS: Developing novel therapies to battle the global public health burden of heart failure remains challenging. This study investigates the underlying mechanisms and potential treatment for 4-hydroxynonenal (4-HNE) deleterious effects in heart failure. METHODS: Biochemical, functional, and histochemical measurements were applied to identify 4-HNE adducts in rat and human failing hearts. In vitro studies were performed to validate 4-HNE targets. RESULTS: 4-HNE, a reactive aldehyde by-product of mitochondrial dysfunction in heart failure, covalently inhibits Dicer, an RNase III endonuclease essential for microRNA (miRNA) biogenesis. 4-HNE inhibition of Dicer impairs miRNA processing. Mechanistically, 4-HNE binds to recombinant human Dicer through an intermolecular interaction that disrupts both activity and stability of Dicer in a concentration- and time-dependent manner. Dithiothreitol neutralization of 4-HNE or replacing 4-HNE-targeted residues in Dicer prevents 4-HNE inhibition of Dicer in vitro. Interestingly, end-stage human failing hearts from three different heart failure aetiologies display defective 4-HNE clearance, decreased Dicer activity, and miRNA biogenesis impairment. Notably, boosting 4-HNE clearance through pharmacological re-activation of mitochondrial aldehyde dehydrogenase 2 (ALDH2) using Alda-1 or its improved orally bioavailable derivative AD-9308 restores Dicer activity. ALDH2 is a major enzyme responsible for 4-HNE removal. Importantly, this response is accompanied by improved miRNA maturation and cardiac function/remodelling in a pre-clinical model of heart failure. CONCLUSIONS: 4-HNE inhibition of Dicer directly impairs miRNA biogenesis in heart failure. Strikingly, decreasing cardiac 4-HNE levels through pharmacological ALDH2 activation is sufficient to re-establish Dicer activity and miRNA biogenesis; thereby representing potential treatment for patients with heart failure.

Photochemically-induced protein tyrosine nitration in vitro and in cellula by 5-methyl-1,4-dinitro-1H-imidazole (DNI): synthesis and biochemical characterization.

The photochemical nitrating agent 5-methyl-1,4-dinitro-1H-imidazole (DNI) has been recently described as an effective tool for nitrating tyrosine residues in proteins under 390 nm irradiation (Long T. et al., 2021). Herein, we describe the one-step synthesis of DNI from the precursor 4-methyl-5-nitro-1H-imidazole with good yield (66%) and high purity (>99%). Spectral analysis of DNI reveals two maximum peaks (228 and 290 nm) with maximum nitration yields and kinetics occurring at 290 nm. Electron paramagnetic resonance (EPR)- and mass spectrometry (MS)- spin trapping analysis evidenced the formation of nitrogen dioxide (•NO2) upon irradiation of DNI, implying the homolysis of the N-N bond in the DNI molecule. Irradiation of DNI at 290, 390 nm, or UVA light (315-400 nm), produced tyrosine nitration, with yields approaching ca. 30% with respect to DNI at 290 nm exposure. Indeed, using alpha-synuclein as a model protein, the main protein post-translational modification triggered by DNI was the generation of 3-nitrotyrosine as shown by MS analysis. Additionally, the formation of di-tyrosine was also observed. Finally, intracellular •NO2 production upon DNI photolysis in bovine aortic endothelial cells was evidenced by the nitration of the tyrosine analog probe p-hydroxyphenylacetic acid (PHPA) and cellular protein tyrosine nitration.

Disulfides form persulfides at alkaline pH leading to potential overestimations in the cold cyanolysis method.

It is well established that proteins and peptides can release sulfur under alkaline treatment, mainly through the ß-elimination of disulfides with the concomitant formation of persulfides and dehydroalanine derivatives. In this study, we evaluated the formation of glutathione persulfide (GSSH/GSS-) by exposure of glutathione disulfide (GSSG) to alkaline conditions. The kinetics of the reaction between GSSG and HO- was investigated by UV-Vis absorbance, reaction with 5,5'-dithio-bis-(2-nitrobenzoic acid) (DTNB), and cold cyanolysis, obtaining an apparent second-order rate constant of ∼10-3 M-1 s-1 at 25 °C. The formation of GSSH and the dehydroalanine derivative was confirmed by HPLC and/or mass spectrometry. However, the mixtures did not equilibrate in a timescale of hours, and additional species, including thiol and diverse sulfane sulfur compounds were also formed, probably through further reactions of the persulfide. Cold cyanolysis is frequently used to quantify persulfides, since it measures sulfane sulfur. This method involves a step in which the sample to be analyzed is incubated with cyanide at alkaline pH. When cold cyanolysis was applied to samples containing GSSG, sulfane sulfur products that were not present in the original sample were measured. Thus, our results reveal the risk of overestimating the amount of sulfane sulfur compounds in samples that contain disulfides due to their decay to persulfides and other sulfane sulfur compounds at alkaline pH. Overall, our study highlights that the ß-elimination of disulfides is a potential source of persulfides, although we do not recommend the preparation of GSSH from incubation of GSSG in alkali. Our study also highlights the importance of being cautious when doing and interpreting cold cyanolysis experiments.

Mitochondrial Peroxiredoxin 3 Is Rapidly Oxidized and Hyperoxidized by Fatty Acid Hydroperoxides.

Human peroxiredoxin 3 (HsPrx3) is a thiol-based peroxidase responsible for the reduction of most hydrogen peroxide and peroxynitrite formed in mitochondria. Mitochondrial disfunction can lead to membrane lipoperoxidation, resulting in the formation of lipid-bound fatty acid hydroperoxides (LFA-OOHs) which can be released to become free fatty acid hydroperoxides (fFA-OOHs). Herein, we report that HsPrx3 is oxidized and hyperoxidized by fFA-OOHs including those derived from arachidonic acid and eicosapentaenoic acid peroxidation at position 15 with remarkably high rate constants of oxidation (>3.5 × 107 M-1s-1) and hyperoxidation (~2 × 107 M-1s-1). The endoperoxide-hydroperoxide PGG2, an intermediate in prostanoid synthesis, oxidized HsPrx3 with a similar rate constant, but was less effective in causing hyperoxidation. Biophysical methodologies suggest that HsPrx3 can bind hydrophobic structures. Indeed, molecular dynamic simulations allowed the identification of a hydrophobic patch near the enzyme active site that can allocate the hydroperoxide group of fFA-OOHs in close proximity to the thiolate in the peroxidatic cysteine. Simulations performed using available and herein reported kinetic data indicate that HsPrx3 should be considered a main target for mitochondrial fFA-OOHs. Finally, kinetic simulation analysis support that mitochondrial fFA-OOHs formation fluxes in the range of nM/s are expected to contribute to HsPrx3 hyperoxidation, a modification that has been detected in vivo under physiological and pathological conditions.

Multiple oxidative post-translational modifications of human glutamine synthetase mediate peroxynitrite-dependent enzyme inactivation and aggregation.

Glutamine synthetase (GS), which catalyzes the ATP-dependent synthesis of L-glutamine from L-glutamate and ammonia, is a ubiquitous and conserved enzyme that plays a pivotal role in nitrogen metabolism across all life domains. In vertebrates, GS is highly expressed in astrocytes, where its activity sustains the glutamate-glutamine cycle at glutamatergic synapses and is thus essential for maintaining brain homeostasis. In fact, decreased GS levels or activity have been associated with neurodegenerative diseases, with these alterations attributed to oxidative post-translational modifications of the protein, in particular tyrosine nitration. In this study, we expressed and purified human GS (HsGS) and performed an in-depth analysis of its oxidative inactivation by peroxynitrite (ONOO-) in vitro. We found that ONOO- exposure led to a dose-dependent loss of HsGS activity, the oxidation of cysteine, methionine, and tyrosine residues and also the nitration of tryptophan and tyrosine residues. Peptide mapping by LC-MS/MS through combined H216O/H218O trypsin digestion identified up to 10 tyrosine nitration sites and five types of dityrosine cross-links; these modifications were further scrutinized by structural analysis. Tyrosine residues 171, 185, 269, 283, and 336 were the main nitration targets; however, tyrosine-to-phenylalanine HsGS mutants revealed that their sole nitration was not responsible for enzyme inactivation. In addition, we observed that ONOO- induced HsGS aggregation and activity loss. Thiol oxidation was a key modification to elicit aggregation, as it was also induced by hydrogen peroxide treatment. Taken together, our results indicate that multiple oxidative events at various sites are responsible for the inactivation and aggregation of human GS.

Energy, protein and redox metabolism underlying adaptive responses in New Zealand versus North American Holstein cows in pasture-based dairy systems.

This study explored the metabolic adaptions to grazing conditions of two Holstein genetic strains (GS; North American, NAH; New Zealand, NZH) in two feeding strategies (FS; restricted, P30, vs. maximised, PMAX, grazing). Four groups (NAH-P30, NZH-P30, NAH-PMAX and NZH-PMAX; n = 10 cows each) were compared between -45 and 180 days in milk (DIM). NZH cows had lower (p = 0.02) fat and protein corrected milk (FPCM) yield and a tendency for lower (p = 0.09) body condition score concomitantly with a trend (p < 0.07) for higher average plasma insulin and lower (p = 0.01) 3-methylhistidine (3MH) at -45 DIM than NAH. Plasma glucose tended to be affected by the triple interaction GS × FS × DIM (p = 0.06) as it was similar between NAH-P30 and NZH-P30, but higher (p ≤ 0.02) for NZH-PMAX than NAH-PMAX except at 21 DIM. The physiological imbalance index was affected by the GS × FS interaction (p < 0.01) as it was lower (p < 0.01) only for NZH-PMAX versus NAH-PMAX. NZH cows had higher (p = 0.01) plasma thiobarbituric acid reactive substances at -45 DIM and tended to have higher protein carbonyls (p = 0.10) and superoxide dismutase (SOD) activity (p = 0.06) on average, and had higher (p < 0.01) α-tocopherol during mid-lactation than NAH Regarding the FS, FPCM was similar (p = 0.12) among them, but PMAX cows had higher (p < 0.01) plasma non-esterified fatty acids and 3MH, and lower insulin (p < 0.01) than P30 at 100 DIM. PMAX cows showed higher average SOD activity (p = 0.01) and plasma α-tocopherol at 100 and 180 DIM (p < 0.01). Under grazing, NZH cows can have a better energy status and lower muscle mobilisation but a higher redox reactivity. Maximising grazing can worsen energy status and muscle mobilisation while improving antioxidant response with no effect on FPCM.

Fast and biphasic 8-nitroguanine production from guanine and peroxynitrite.

Guanine (Gua), among purines, is a preferred oxidation/nitration target because of its low one-electron redox potential. The reactive oxygen/nitrogen species peroxynitrite (ONOO-), produced in vivo by the reaction between nitric oxide (•NO) and superoxide radical (O2•‒), is responsible for several oxidative modifications in biomolecules, including nitration, nitrosation, oxidation, and peroxidation. In particular, the nitration of Gua, although detected, as well as its reaction kinetics have been seldom investigated. Thus, we studied the concentration- and temperature-dependent formation of 8-nitroguanine (8-NitroGua) in phosphate buffer (pH 7.40) using stopped-flow spectrophotometry. Traces showed a biexponential behavior, with best-fit rate constants: kfast = 4.4 s-1 and kslow = 0.41 s-1 (30 °C, 400 µM both Gua and ONOO-). kfast increased linearly with the concentration of both reactants whereas kslow was concentration-independent. Linear regression analysis of kfast as a function of Gua and ONOO- concentration yielded values of 2.5-6.3 × 103 M-1s-1 and 1.5-3.5 s-1 for the second-order (slope) and first-order (ordinate) rate constants, respectively (30 °C). Since ONOO- is a short-lived species, its decay kinetics was also taken into account for this analysis. The 8-NitroGua product was stable for at least 4 h, so no spontaneous denitration was observed. Stopped-flow assays using antioxidants and free-radical scavengers suggested a mixed direct/indirect reaction mechanism for 8-NitroGua formation. Gua nitration by ONOO- was also observed in the presence of physiologically relevant CO2 concentrations. The reaction product identity, its yield (∼4.2%, with 400 µM ONOO- and 200 µM Gua), and the reaction mechanism were unequivocally determined by HPLC-MS/MS experiments. In conclusion, 8-NitroGua production at physiologic pH reached significant levels in a few hundred milliseconds, suggesting that the process might be kinetically relevant in vivo and can likely cause permanent nitrative damage to DNA bases.

HPLC-MS/MS Oxylipin Analysis of Plasma from Amyotrophic Lateral Sclerosis Patients.

Oxylipins play a critical role in regulating the onset and resolution phase of inflammation. Despite inflammation is a pathological hallmark in amyotrophic lateral sclerosis (ALS), the plasma oxylipin profile of ALS patients has not been assessed yet. Herein, we develop an oxylipin profile-targeted analysis of plasma from 74 ALS patients and controls. We found a significant decrease in linoleic acid-derived oxylipins in ALS patients, including 9-hydroxy-octadecadienoic acid (9-HODE) and 13-HODE. These derivatives have been reported as important regulators of inflammation on different cell systems. In addition, some 5-lipoxygenase metabolites, such as 5-hydroxy- eicosatetraenoic acid also showed a significant decrease in ALS plasma samples. Isoprostanes of the F2α family were detected only in ALS patients but not in control samples, while the hydroxylated metabolite 11-HETE significantly decreased. Despite our effort to analyze specialized pro-resolving mediators, they were not detected in plasma samples. However, we found the levels of 14-hydroxy-docosahexaenoic acid, a marker pathway of the Maresin biosynthesis, were also reduced in ALS patients, suggesting a defective activation in the resolution programs of inflammation in ALS. We further analyze oxylipin concentration levels in plasma from ALS patients to detect correlations between these metabolites and some clinical parameters. Interestingly, we found that plasmatic levels of 13-HODE and 9-HODE positively correlate with disease duration, expressed as days since onset. In summary, we developed a method to analyze "(oxy)lipidomics" in ALS human plasma and found new profiles of metabolites and novel lipid derivatives with unknown biological activities as potential footprints of disease onset.

Administration of Maresin-1 ameliorates the physiopathology of experimental autoimmune encephalomyelitis.

BACKGROUND: Resolution of inflammation is an active and regulated process that leads to the clearance of cell debris and immune cells from the challenged tissue, facilitating the recovery of homeostasis. This physiological response is coordinated by endogenous bioactive lipids known as specialized pro-resolving mediators (SPMs). When resolution fails, inflammation becomes uncontrolled leading chronic inflammation and tissue damage, as occurs in multiple sclerosis (MS). METHODS: SPMs and the key biosynthetic enzymes involved in SPM production were analysed by metabololipidomics and qPCR in active brain lesions, serum and peripheral blood mononuclear cells (PBMC) of MS patients as well as in the spinal cord of mice with experimental autoimmune encephalomyelitis (EAE). We also tested the therapeutic actions of the SPM coined Maresin-1 (MaR1) in EAE mice and studied its impact on inflammation by doing luminex and flow cytometry analysis. RESULTS: We show that levels of MaR1 and other SPMs were below the limit of detection or not increased in the spinal cord of EAE mice, whereas the production of pro-inflammatory eicosanoids was induced during disease progression. Similarly, we reveal that SPMs were undetected in serum and active brain lesion samples of MS patients, which was linked to impaired expression of the enzymes involved in the biosynthetic pathways of SPMs. We demonstrate that exogenous administration of MaR1 in EAE mice suppressed the protein levels of various pro-inflammatory cytokines and reduced immune cells counts in the spinal cord and blood. MaR1 also decreased the numbers of Th1 cells but increased the accumulation of regulatory T cells and drove macrophage polarization towards an anti-inflammatory phenotype. Importantly, we provide clear evidence that administration of MaR1 in mice with clinical signs of EAE enhanced neurological outcomes and protected from demyelination. CONCLUSIONS: This study reveals that there is an imbalance in the production of SPMs in MS patients and in EAE mice, and that increasing the bioavailability of SPMs, such as MaR1, minimizes inflammation and mediates therapeutic actions. Thus, these data suggest that immunoresolvent therapies, such as MaR1, could be a novel avenue for the treatment of MS.

Decreased proteasomal cleavage at nitrotyrosine sites in proteins and peptides.

Removal of moderately oxidized proteins is mainly carried out by the proteasome, while highly modified proteins are no longer degradable. However, in the case of proteins modified by nitration of tyrosine residues to 3-nitrotyrosine (NO2Y), the role of the proteasome remains to be established. For this purpose, degradation assays and mass spectrometry analyses were performed using isolated proteasome and purified fractions of native cytochrome c (Cyt c) and tyrosine nitrated proteoforms (NO2Y74-Cyt c and NO2Y97-Cyt c). While Cyt c treated under mild conditions with hydrogen peroxide was preferentially degraded by the proteasome, NO2Y74- and NO2Y97-Cyt c species did not show an increased degradation rate with respect to native Cyt c. Peptide mapping analysis confirmed a decreased chymotrypsin-like cleavage at C-terminal of NO2Y sites within the protein, with respect to unmodified Y residues. Additionally, studies with the proteasome substrate suc-LLVY-AMC (Y-AMC) and its NO2Y-containing analog, suc-LLVNO2Y-AMC (NO2Y-AMC) were performed, both using isolated 20S-proteasome and astrocytoma cell lysates as the proteasomal source. Comparisons of both substrates showed a significantly decreased proteasome activity towards NO2Y-AMC. Moreover, NO2Y-AMC, but not Y-AMC degradation rates, were largely diminished by increasing the reaction pH, suggesting an inhibitory influence of the additional negative charge contained in NO2Y-AMC secondary to nitration. The mechanism of slowing of proteasome activity in NO2Y-contaning peptides was further substantiated in studies using the phenylalanine and nitro-phenylalanine peptide analog substrates. Finally, degradation rates of Y-AMC and NO2Y-AMC with proteinase K were the same, demonstrating the selective inability of the proteasome to readily cleave at nitrotyrosine sites. Altogether, data indicate that the proteasome has a decreased capability to cleave at C-terminal of NO2Y residues in proteins with respect to the unmodified residues, making this a possible factor that decreases the turnover of oxidized proteins, if they are not unfolded, and facilitating the accumulation of nitrated proteins.

Antiplatelet activity and chemical analysis of leaf and fruit extracts from Aristotelia chilensis.

Aristotelia chilensis (Mol.) Stuntz, also known as maqui, is a plant native to Chile without chemical characterization and quantification of the bioactive compounds present in it. HPLC-UV and HPLC-MS/MS studies have shown the presence, at different concentrations, of phenolic and anthocyanin compounds in fruit and leave extracts of the domesticated maqui clones Luna Nueva, Morena, and Perla Negra. The extracts from leaves and unripe fruits of Luna Nueva and Morena clones significantly inhibit platelet aggregation induced by several agonists; the extracts inhibit platelet granule secretion by decreasing the exposure of P-selectin and CD63 at the platelet membrane. Reactive oxygen species formation in platelets is lower in the presence of maqui extracts. Statistical Pearson analysis supports the levels of phenolic and anthocyanin compounds being responsible for the antiaggregant maqui effects. This work is the first evidence of antiplatelet activity from Aristotelia chilensis giving added value to the use of leaves and unripe fruits from this species.

Olive oil-derived nitro-fatty acids: protection of mitochondrial function in non-alcoholic fatty liver disease.

Non-alcoholic fatty liver disease (NAFLD) is characterized by excessive liver fat deposition in the absence of significant alcohol intake. Since extra virgin olive oil (EVOO) reduces fat accumulation, we analyzed the involvement of nitro-fatty acids (NO2-FA) on the beneficial effects of EVOO consumption on NAFLD. Nitro-fatty acids formation was observed during digestion in mice supplemented with EVOO and nitrite. Mice fed with a high-fat diet (HF) presented lower plasma NO2-FA levels than normal chow, and circulating concentrations recovered when the HF diet was supplemented with 10% EVOO plus nitrite. Under NO2-FA formation conditions, liver hemoxygenase-1 expression significantly increased while decreased body weight and fat liver accumulation. Mitochondrial dysfunction plays a central role in the pathogenesis of NAFLD while NO2-FA has been shown to protect from mitochondrial oxidative damage. Accordingly, an improvement of respiratory indexes was observed when mice were supplemented with both EVOO plus nitrite. Liver mitochondrial complexes II and V activities were greater in mice with EVOO supplementation and further improved in the presence of nitrite. Overall, our results strongly suggest a positive correlation between NO2-OA formation from EVOO and the observed improvement of mitochondrial function in NAFLD. The formation of NO2-FA can account for the health benefits associated with EVOO consumption.

Acidity and nucleophilic reactivity of glutathione persulfide.

Persulfides (RSSH/RSS-) participate in sulfur trafficking and metabolic processes, and are proposed to mediate the signaling effects of hydrogen sulfide (H2S). Despite their growing relevance, their chemical properties are poorly understood. Herein, we studied experimentally and computationally the formation, acidity, and nucleophilicity of glutathione persulfide (GSSH/GSS-), the derivative of the abundant cellular thiol glutathione (GSH). We characterized the kinetics and equilibrium of GSSH formation from glutathione disulfide and H2S. A pKa of 5.45 for GSSH was determined, which is 3.49 units below that of GSH. The reactions of GSSH with the physiologically relevant electrophiles peroxynitrite and hydrogen peroxide, and with the probe monobromobimane, were studied and compared with those of thiols. These reactions occurred through SN2 mechanisms. At neutral pH, GSSH reacted faster than GSH because of increased availability of the anion and, depending on the electrophile, increased reactivity. In addition, GSS- presented higher nucleophilicity with respect to a thiolate with similar basicity. This can be interpreted in terms of the so-called α effect, i.e. the increased reactivity of a nucleophile when the atom adjacent to the nucleophilic atom has high electron density. The magnitude of the α effect correlated with the Brønsted nucleophilic factor, ßnuc, for the reactions with thiolates and with the ability of the leaving group. Our study constitutes the first determination of the pKa of a biological persulfide and the first examination of the α effect in sulfur nucleophiles, and sheds light on the chemical basis of the biological properties of persulfides.

Data of detection and characterization of nitrated conjugated-linoleic acid (NO2-cLA) in LDL.

Under physiological and pathophysiological conditions, lipid nitration occurs generating nitro-fatty acids (NFA) with pleiotropic activities as modulation of inflammatory cell responses. Foam cell formation and atherosclerotic lesion development have been extensively related to low-density lipoprotein (LDL) oxidation. Considering our manuscript "Fatty acid nitration in human low-density lipoprotein" (https://doi.org/10.1016/j.abb.2019.108190), herein we report the oxidation versus nitration of human LDL protein and lipid fractions. Data is shown on LDL fatty acid nitration, in particular, formation and quantitation of nitro-conjugated linoleic acid (NO2-cLA) under mild nitration conditions. In parallel to NO2-cLA formation, depletion of endogenous antioxidants, protein tyrosine nitration, and carbonyl formation is observed. Overall, our data propose the formation of a potential anti-atherogenic form of LDL carrying NFA.

Fatty acid nitration in human low-density lipoprotein.

Lipid nitration occurs during physiological and pathophysiological conditions, generating a variety of biomolecules capable to modulate inflammatory cell responses. Low-density lipoprotein (LDL) oxidation has been extensively related to atherosclerotic lesion development while oxidative modifications confer the particle pro-atherogenic features. Herein, we reviewed the oxidation versus nitration of human LDL protein and lipid fractions. We propose that unsaturated fatty acids present in LDL can be nitrated under mild nitration conditions, suggesting an anti-atherogenic role for LDL carrying nitro-fatty acids (NFA).

Kinetics of formation and reactivity of the persulfide in the one-cysteine peroxiredoxin from Mycobacterium tuberculosis.

Hydrogen sulfide (H2S) participates in prokaryotic metabolism and is associated with several physiological functions in mammals. H2S reacts with oxidized thiol derivatives (i.e. disulfides and sulfenic acids) and thereby forms persulfides, which are plausible transducers of the H2S-mediated signaling effects. The one-cysteine peroxiredoxin alkyl hydroperoxide reductase E from Mycobacterium tuberculosis (MtAhpE-SH) reacts fast with hydroperoxides, forming a stable sulfenic acid (MtAhpE-SOH), which we chose here as a model to study the interactions between H2S and peroxiredoxins (Prx). MtAhpE-SOH reacted with H2S, forming a persulfide (MtAhpE-SSH) detectable by mass spectrometry. The rate constant for this reaction was (1.4 ± 0.2) × 103 m-1 s-1 (pH 7.4, 25 °C), six times higher than that reported for the reaction with the main low-molecular-weight thiol in M. tuberculosis, mycothiol. H2S was able to complete the catalytic cycle of MtAhpE and, according to kinetic considerations, it could represent an alternative substrate in M. tuberculosis. MtAhpE-SSH reacted 43 times faster than did MtAhpE-SH with the unspecific electrophile 4,4'-dithiodipyridine, a disulfide that exhibits no preferential reactivity with peroxidatic cysteines, but MtAhpE-SSH was less reactive toward specific Prx substrates such as hydrogen peroxide and peroxynitrite. According to molecular dynamics simulations, this loss of specific reactivity could be explained by alterations in the MtAhpE active site. MtAhpE-SSH could transfer its sulfane sulfur to a low-molecular-weight thiol, a process likely facilitated by the low pKa of the leaving thiol MtAhpE-SH, highlighting the possibility that Prx participates in transpersulfidation. The findings of our study contribute to the understanding of persulfide formation and reactivity.

Impaired hepatic mitochondrial function during early lactation in dairy cows: Association with protein lysine acetylation.

Early lactation is an energy-deming period for dairy cows which may lead to negative energy balance, threatening animal health and consequently productivity. Herein we studied hepatic mitochondrial function in Holstein-Friesian multiparous dairy cows during lactation, under two different feeding strategies. During the first 180 days postpartum the cows were fed a total mixed ration (70% forage: 30% concentrate) ad libitum (non-grazing group, G0) or grazed Festuca arundinacea or Mendicago sativa plus supplementation (grazing group, G1). From 180 to 250 days postpartum, all cows grazed Festuca arundinacea were supplemented with total mixed ration. Mitochondrial function was assessed measuring oxygen consumption rate in liver biopsies revealed that maximum respiratory rate decreased significantly in grazing cows during early lactation, yet was unchanged in non-grazing cows during the lactation curve. While no differences could be found in mitochondrial content or oxidative stress markers, a significant increase in protein lysine acetylation was found in grazing cows during early lactation but not in cows from the non-grazing group. Mitochondrial acetylation positively correlated with liver triglycerides ß-hydroxybutyrate plasma levels, well-known markers of negative energy balance, while a negative correlation was found with the maximum respiratory rate sirtuin 3 levels. To our knowledge this is the first report of mitochondrial function in liver biopsies of dairy cows during lactation. On the whole our results indicate that mitochondrial function is impaired during early lactation in grazing cows that acetylation may account for changes in mitochondrial function in this period. Additionally, our results suggest that feeding total mixed ration during early lactation may be an efficient protective strategy.

Corrigendum to "Protective Effects of a Polyphenol-Rich Extract from Syzygium cumini (L.) Skeels Leaf on Oxidative Stress-Induced Diabetic Rats".

[This corrects the article DOI: 10.1155/2018/5386079.].