Counteract Lipid Peroxidation Research Articles

Catalase, Glutathione Peroxidase, and Peroxiredoxin 2 in Erythrocyte Cytosol and Membrane in Hereditary Spherocytosis, Sickle Cell Disease, and β-Thalassemia.

Catalase (CAT), glutathione peroxidase (GPx), and peroxiredoxin 2 (Prx2) can counteract the deleterious effects of oxidative stress (OS). Their binding to the red blood cell (RBC) membrane has been reported in non-immune hemolytic anemias (NIHAs). Our aim was to evaluate the relationships between CAT, GPx, and Prx2, focusing on their role at the RBC membrane, in hereditary spherocytosis (HS), sickle cell disease (SCD), β-thalassemia (β-thal), and healthy individuals. The studies were performed in plasma and in the RBC cytosol and membrane, evaluating OS biomarkers and the enzymatic activities and/or the amounts of CAT, GPx, and Prx2. The binding of the enzymes to the membrane appears to be the primary protective mechanism against oxidative membrane injuries in healthy RBCs. In HS (unsplenectomized) and β-thal, translocation from the cytosol to the membrane of CAT and Prx2, respectively, was observed, probably to counteract lipid peroxidation. RBCs from splenectomized HS patients showed the highest membrane-bound hemoglobin, CAT, and GPx amounts in the membrane. SCD patients presented the lowest amount of enzyme linkage, possibly due to structural changes induced by sickle hemoglobin. The OS-induced changes and antioxidant response were different between the studied NIHAs and may contribute to the different clinical patterns in these patients.

Elovanoids Suppress Lipid Peroxidation after Experimental Stroke: An LC-MS-based Untargeted Lipidomics Analysis

At the onset and progression of neurodegenerative diseases, enhanced reactive oxygen species (ROS) and an overall uncompensated oxidative stress environment evolve. Oxidized phospholipids (OxPLs) cause detrimental perturbations to synaptic membranes and other parts of the nervous system. We tested if the lipid mediators Elovanoids (ELVs) that exert neuroprotective signaling also counteract lipid peroxidation using an experimental ischemic stroke model. Male Sprague-Dawley rats after 2 hours of middle cerebral artery occlusion (MCAo) were used. ELV-N34 was administered intranasally at 1 hour, 24 hours and 48 hours after, and rats were sacrificed on day 3; the brains were removed and sampled into the right and left hemispheres and the cortex, subcortex. Untargeted lipidomics were performed using a Waters Xevo G2-XS QTof system under MSE mode, and collected MS/MS spectra for the OxPLs was processed and searched against a reference spectral library by MS-DIAL software. ELV-N34 reduced oxidized phosphatidylcholine (OxPC) and phosphatidylethanolamine (OxPE) molecular species. We suggest that ELVs target lipid peroxidation as part of their neuroprotective bioactivity.

Targeting ferroptosis by poly(acrylic) acid coated Mn3O4 nanoparticles alleviates acute liver injury

Ferroptosis, a newly characterized form of regulated cell death, is induced by excessive accumulation of lipid peroxidation catalyzed by intracellular bioactive iron. Increasing evidence has suggested that ferroptosis is involved in the pathogenesis of several human diseases, including acute liver injury. Targeted inhibition of ferroptosis holds great promise for the clinical treatment of these diseases. Herein, we report a simple and one-pot synthesis of ultrasmall poly(acrylic) acid coated Mn3O4 nanoparticles (PAA@Mn3O4-NPs, PMO), which perform multiple antioxidant enzyme-mimicking activities and can scavenge broad-spectrum reactive oxygen species. PMO could potently suppress ferroptosis. Mechanistically, after being absorbed mainly through macropinocytosis, PMO are largely enriched in lysosomes, where PMO detoxify ROS, inhibit ferritinophagy-mediated iron mobilization and preserve mTOR activation, which collectively confer the prominent inhibition of ferroptosis. Additionally, PMO injection potently counteracts lipid peroxidation and alleviates acetaminophen- and ischaemia/reperfusion-induced acute liver injury in mice. Collectively, our results reveal that biocompatible PMO act as potent ferroptosis inhibitors through multifaceted mechanisms, which ensures that PMO have great translational potential for the clinical treatment of ferroptosis-related acute liver injury.

Abstract 6144: Polyploidization enables initial survival of chemotherapy but a progressively increased susceptibility to ferroptosis

Abstract Once cancer has metastasized it remains incurable owing to evolved resistance to nearly all systemic therapies. Classical views of therapeutic resistance include tumor heterogeneity and genetic instability, but our group and others have identified an unappreciated cancer cell phenotype, the polyaneuploid cancer cell (PACC) state, that provides an alternative model. The PACC state is induced upon exposure to a variety of applied stressors, including multiple different classes of chemotherapy, and is characterized by polyploidization of the aneuploid cancer genome. We have shown that this polyploidization is accomplished through endocycling that leads to an enlarged cell size with increased transcriptomic and protein content, higher oxidative buffer capacity, and increased autophagy. We have demonstrated that PACCs are present in a variety of cell lines, arise in response to multiple stressors in vitro, and are resistant to subsequent treatment with several classes of chemotherapy. Thus, we hypothesize that the PACC state is driving therapeutic resistance to current treatment regimens. Elimination of this overlooked phenotype is critical for sustained anticancer therapy response. Cancer cells, including those in the PACC state, control canonical cell cycle and cell death programs to evade apoptosis from external stressors including chemotherapy, highlighting an opportunity to leverage cell cycle-independent death pathways as an alternative therapeutic strategy1. Ferroptosis is a cell cycle agnostic immunogenic form of cell death that is an ideal pathway for elimination of cells in the PACC state. Characterization of ROS levels, glutathione content, labile iron homeostasis, and lipid content indicate that cells in the PACC state have a mounting susceptibility to ferroptosis. Our data suggests that cells in the PACC state keep ferroptosis in check through elevated levels of reduced glutathione and proficient redox activity, including turn over of GPX4, the sole enzyme counteracting lipid peroxidation. Direct inhibition of GPX4 is highly toxic to cells in the PACC state, particularly at later timepoints post therapy removal when the immediate stress response is largely resolved. These studies will directly lead to development of a therapeutic strategy specifically targeting the PACC state with the intention of eliminating cancer resistance. 1Loftus, L.V.; Amend, S.R.; Pienta, K.J. Interplay between Cell Death and Cell Proliferation Reveals New Strategies for Cancer Therapy. Int. J. Mol. Sci. 2022, 23, 4723. https://doi.org/10.3390/ijms23094723 Citation Format: Luke Loftus. Polyploidization enables initial survival of chemotherapy but a progressively increased susceptibility to ferroptosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6144.

Effects of carnosine on the embryonic development and TiO2 nanoparticles-induced oxidative stress on Zebrafish.

Oxidative stress is due to an unbalance between pro-oxidants, such as reactive oxygen (ROS) and nitrogen (RNS) species, and antioxidants/antioxidant system. Under physiological conditions these species are involved in different cellular processes such as cellular homeostasis and immune response, while an excessive production of ROS/RNS has been linked to the development of various diseases such as cancer, diabetes, and Alzheimer's disease. In this context, the naturally occurring dipeptide carnosine has shown the ability to scavenge ROS, counteract lipid peroxidation, and inhibit proteins oxidation. Titanium dioxide nanoparticles (TiO2-NPs) have been widely used to produce cosmetics, in wastewater treatment, in food industry, and in healthcare product. As consequence, these NPs are often released into aquatic environments. The Danio rerio (commonly called zebrafish) embryos exposure to TiO2-NPs did not affect the hatching rate, but induced oxidative stress. According to this scenario, in the present study, we first investigated the effects of carnosine exposure and of a sub-toxic administration of TiO2-NPs on the development and survival of zebrafish embryos/larvae measured through the acute embryo toxicity test (FET-Test). Zebrafish larvae represent a useful model to study oxidative stress-linked disorders and to test antioxidant molecules, while carnosine was selected based on its well-known multimodal mechanism of action that includes a strong antioxidant activity. Once the basal effects of carnosine were assessed, we then evaluated its effects on TiO2-NPs-induced oxidative stress in zebrafish larvae, measured in terms of total ROS production (measured with 2,7-dichlorodihydrofluorescein diacetate probe) and protein expression by immunohistochemistry of two cellular stress markers, 70 kDa-heat shock protein (Hsp70) and metallothioneins (MTs). We demonstrated that carnosine did not alter the phenotypes of both embryos and larvae of zebrafish at different hours post fertilization. Carnosine was instead able to significantly decrease the enhancement of ROS levels in zebrafish larvae exposed to TiO2-NPs and its antioxidant effect was paralleled by the rescue of the protein expression levels of Hsp70 and MTs. Our results suggest a therapeutic potential of carnosine as a new pharmacological tool in the context of pathologies characterized by oxidative stress such as neurodegenerative disorders.

Natural Astaxanthin Is a Green Antioxidant Able to Counteract Lipid Peroxidation and Ferroptotic Cell Death

Astaxanthin is a red orange xanthophyll carotenoid produced mainly by microalgae but which can also be chemically synthesized. As demonstrated by several studies, this lipophilic molecule is endowed with potent antioxidant properties and is able to modulate biological functions. Unlike synthetic astaxanthin, natural astaxanthin (NAst) is considered safe for human nutrition, and its production is considered eco-friendly. The antioxidant activity of astaxanthin depends on its bioavailability, which, in turn, is related to its hydrophobicity. In this study, we analyzed the water-solubility of NAst and assessed its protective effect against oxidative stress by means of different approaches using a neuroblastoma cell model. Moreover, due to its highly lipophilic nature, astaxanthin is particularly protective against lipid peroxidation; therefore, the role of NAst in counteracting ferroptosis was investigated. This recently discovered process of programmed cell death is indeed characterized by iron-dependent lipid peroxidation and seems to be linked to the onset and development of oxidative-stress-related diseases. The promising results of this study, together with the "green sources" from which astaxanthin could derive, suggest a potential role for NAst in the prevention and co-treatment of chronic degenerative diseases by means of a sustainable approach.

1,8-cineole and ginger extract (Zingiber officinale Rosc) as stress mitigator for transportation of largemouth bass (Micropterus salmoides L.)

Transportation stress is considered the major cause of production and economic losses in aquaculture. Here, we examined the effects of dried ginger extract (GE) and 1,8-cineole (CIN) on juvenile largemouth bass (Micropterus salmoides) subjected to simulated transport by quantifying serum metabolites, antioxidant enzyme activity, and haematological features in conjunction with whole transcriptome analysis. Under transportation stress, the addition of dried ginger extract (20 μg/L) or 1,8-cineole (30 μg/L) significantly reduced serum cortisol levels in juvenile largemouth bass. Additionally, CIN administration was associated with a significant increase in total protein and glucose levels in serum, and reduced alkaline phosphatase (AKP) and alanine aminotransferase (ALT) enzyme activity. GE and CIN significantly increased serum lysozyme activity, glutathione peroxidase (GSH-Px) activity and total antioxidant capacity (T-AOC), thereby counteracting lipid peroxidation. Transportation stress-induced malondialdehyde (MDA) production was reduced under exposure to GE and CIN, while white blood cell (WBC) counts were increased in the treatment group compared with controls. Whole transcriptome analysis of M. salmoides liver tissue identified 533 significantly differentially expressed genes (DEGs) between the CIN and transport-only control treatment groups. DEGs were mainly enriched in endocrine system, immune system and metabolic pathways. This study demonstrates that dried ginger extract or 1,8-cineole can potentially regulate M. salmoides response to transportation stress, thereby providing novel insights into the mechanisms underlying immune response and antioxidant pathways during sedation, supporting 1,8-cineole as an effective potential antioxidant agent in aquaculture.

Persistent ferroptosis promotes cervical squamous intraepithelial lesion development and oncogenesis by regulating KRAS expression in patients with high risk-HPV infection

Cervical squamous cell carcinoma (CSCC) is a type of female cancer that affects millions of families worldwide. Human papillomavirus (HPV) infection is the main reason for CSCC formation, and squamous intraepithelial lesions (SILs) induced by high-risk HPV (HR-HPV) infection are considered precancerous lesions. A previous study reported that HPV-infected cancer cells were able to counteract lipid peroxidation for survival. Recent research has reported that ferroptosis acts in an iron-dependent lipid peroxidation manner to kill cancer cells, and it is proposed as a new approach for female cancer therapy. Here, we investigated the role of ferroptosis throughout SIL development into CSCC. We found that ferroptosis occurred in SIL, but anti-ferroptosis emerged in CSCC. Our data further indicated that an antiferroptotic effect was formed in response to persistent ferroptosis and then promoted oncogenesis. Altogether, we provide novel insight into ferroptosis in cervical SIL development and suggest a potential therapeutic target for the treatment of CSCC.

Nutraceutical and therapeutic potential of Phycocyanobilin for treating Alzheimer’s disease

Alzheimer's disease (AD) is a devastating neurodegenerative condition provoking the loss of cognitive and memory performances. Despite huge efforts to develop effective AD therapies, there is still no cure for this neurological condition. Here, we review the main biological properties of Phycocyanobilin (PCB), accounting for its potential uses against AD. PCB, given individually or released in vivo from C-Phycocyanin (C-PC), acts as a bioactive-molecule-mediating antioxidant, is anti-inflammatory and has immunomodulatory activities. PCB/C-PC are able to scavenge reactive oxygen and nitrogen species, to counteract lipid peroxidation and to inhibit enzymes such as NADPH oxidase and COX-2. In animal models of multiple sclerosis and ischemic stroke, these compounds induce remyelination as demonstrated by electron microscopy and the expression of genes such as Mal up-regulation of and Lingo-1 down-regulation. These treatments also reduce pro-inflammatory cytokines levels and induce immune suppressive genes. PCB/C-PC protects isolated rat brain mitochondria and inactivate microglia, astrocytes and neuronal apoptosis mediators. Such processes are all involved in the pathogenic cascade of AD, and thus PCB may effectively mitigate the injury in this condition. Furthermore, PCB can be administered safely by oral or parenteral routes and therefore, could be commercially offered as a nutraceutical supplement or as a pharmaceutical drug.

Global warming overrides physiological anti-predatory mechanisms in intertidal rock pool fish Gobius paganellus

In nature, a multitude of factors influences the fitness of an organism at a given time, which makes single stressor assessments far from ecologically relevant scenarios. This study focused on the effects of water temperature and predation stress on the metabolism and body mass gain of a common intertidal rock pool fish, Gobius paganellus, addressing the following hypotheses: (1) the energy metabolism of G. paganellus under predation stress is reduced; (2) G. paganellus shows thermal compensation under heat stress; and (3) thermal stress is the dominant stressor that may override predation stress responses. Individuals were exposed to simulated predation stress and temperature increase from 20 °C to 29 °C, and both stressors combined. Physiological effects were addressed using biochemical biomarkers related with energy metabolism (isocitrate dehydrogenase, lactate dehydrogenase, energy available, energy consumption rates), oxidative stress (superoxide dismutase, catalase, DNA damage, lipid peroxidation), and biotransformation (glutathione-S-transferase).The results of this study revealed that predation stress reduced the cellular metabolism of G. paganellus, and enhanced storage of protein reserves. As hypothesized, hyperthermia decreased the aerobic mitochondrial metabolism, indicating thermal compensation mechanisms to resist against unfavourable temperatures. Hyperthermia was the dominant stressor overriding the physiological responses to predation stress. Both stressors combined might further have synergistically activated detoxification pathways, even though not strong enough to counteract lipid peroxidation and DNA damage completely. The synergistic effect of combined thermal and predation stress thus may not only increase the risk of being preyed upon, but also may indicate extra energy trade-off for the basal metabolism, which in turn may have ecologically relevant consequences for general body functions such as somatic growth and reproduction. The present findings clearly underline the ecological importance of multi-stressor assessments to provide a better and holistic picture of physiological responses towards more realistic evaluations of climate change consequences for intertidal populations.

Phytochemical and Toxicological Effect of Ethanol Extract of Heliotropium Indicum on Liver of Male Albino Rats

Heliotropium indicum is a plant of the Boraginaceae family reported to have multiple beneficial health effects, including analgesic, anti-malaria, anti-microbial, anti-tumor, anti-inflammatory, amongst others. 42 Male Wistar rats were used for this study. Group A served as control and received distilled water only. Groups B, C, D, E, and F were fed with the extract of Heliotropium Indicum in different concentrations. (500 mg/kg, 1000 mg/kg, 1500 mg/kg 2000 mg/kg and 3000 mg/kg) respectively. This study showed that Heliotropium indicum contains phytochemicals such as saponins tannins, cardiac glycosides, and terpenoids. Our results revealed that Heliotropium indicum decreased the ALT, AST, ALP, and bilirubin levels, especially at doses from 1500 mg/kg B.W. Also, inflammatory cytokine expressions were significantly reduced at similar doses. However, the level of antioxidants (GSH, CAT, GST, SOD, GPx) were significantly increased by Heliotropium indicum administration at doses ranging from 1500 mg/kg to 3000 mg/kg. The antioxidative potentials of Heliotropium indicum are further complemented by its ability to reduce inflammatory cytokine expression, including TNF-α, IL-1β, IL-6, COX-2 as well as NO and LPO. Ethanol extract of Heliotropium indicum at doses between 1500 mg/kg and 3000 mg/kg has antioxidative, anti-inflammatory effects and can counteract lipid peroxidation.

First characterization of Pompia intrea candied fruit: The headspace chemical profile, polar extract composition and its biological activities

Extracts obtained from Pompia intrea (PI), a traditional candied fruit whose main ingredient is the pompia fruit (Citrus monstruosa L.) were evaluated for the first time. Volatile compounds were evaluated by headspace solid-phase microextraction (HS-SPME) followed by GC-FID/MS analyses. Polar compounds were identified by high-performance liquid chromatography coupled with electrospray mass spectrometry LC-ESI-Orbitrap-MS/MS and quantified by liquid chromatography coupled with ultraviolet/visible detection (LC-DAD). The antioxidant activity of these extracts was tested using the FRAP, CUPRAC, ABTS•+ and DPPH● assays. Moreover, their ability to protect intestinal cells against lipid peroxidation was studied. The HS-SPME GC-FID/MS confirmed the presence of typical molecules originating from the fruit (mainly terpenes, but particularly limonene). The LC-DAD and LC-ESI-(HR)MSn profiles showed high levels of neohesperidin (45.7 ± 11.1 mg/L) and 5-(hydroxymethyl)furfural (40.8 ± 23.5 mg/L). The results showed that the PI extracts contained high levels of total phenols and exhibited considerable antioxidant activity, which was significantly correlated to the presence of specific compounds such as neoeriocitrin and neohesperidin. Furthermore, pretreatments with different concentrations of PI extracts preserved enterocytes from oxidative damage by scavenging reactive oxygen species, thus counteracting lipid peroxidation. This study suggests that consumption of PI could provide intake of compounds with ascertained biological activity.

Taurine: A Regulator of Cellular Redox Homeostasis and Skeletal Muscle Function.

Taurine is a nonproteinogenic ß-aminosulfonic acid. Important dietary sources of taurine are fish and seafood. Taurine interacts with ion channels, stabilizes membranes, and regulates the cell volume. These actions confirm its high concentrations in excitable tissues like retina, neurons, and muscles. Retinal degeneration, cardiomyopathy, as well as skeletal muscle malfunction are evident in taurine-deficient phenotypes. There is evidence that taurine counteracts lipid peroxidation and increases cellular antioxidant defense in response to inflammation. In activated neutrophils, taurine reacts with hypochloric acid to form taurine chloramine, which triggers the Kelch-like ECH-associated protein 1-nuclear factor E2-related factor 1 (Keap1-Nrf2) pathway. Consequently, Nrf2 target genes, such as heme oxygenase-1 and catalase, are induced. Furthermore, taurine may prevent an overload of reactive oxygen species (ROS) directly by an inhibition of ROS generation within the respiratory chain. Taurine affects mitochondrial bioenergetics and taurine-deficient mice exhibit an impaired exercise performance. Moreover, some studies demonstrate that taurine enhances the glycogen repletion in the postexercise recovery phase. In the case of taurine deficiency, many studies observed a phenotype known in muscle senescence and skeletal muscle disorders. Overall, taurine plays an important role in cellular redox homeostasis and skeletal muscle function.

Age related distribution of 4-hydroxy 2-nonenal immunoreactivity in human retina

The retina is prone to be damaged by oxidative stress (OS), owing to its constant exposure to light, high rate of oxygen consumption and high membrane lipid content. Lipid peroxidation in aging human retina has been shown by biochemical means. However, information on the cellular sites of OS and antioxidant responses in aging human retina remains limited. Here, we show distribution of immunoreactivity (IR) to a marker of lipid peroxidation (4-hydroxy 2-nonenal [HNE] and antioxidant enzymes involved in counteracting lipid peroxidation (glutathione S-transferase-π1 and glutarexoxin-1) in donor human retinas at different ages (35–91 years; N = 24). Initially, HNE-IR was present in few macular cone outer segments (COS, sixth decade). With aging, IR appeared in many COS and peaked at ninth decade (14 vs 62 per 3850 μm2 area between 6 and 9 decade; p < 0.001) in the parafovea then seen elsewhere (perifoveal, mid-peripheral and nasal). IR was seen in the parafovea of all retinas, whereas it was present in 8/24 of perifoveal and 6/24 of mid-peripheral retinas, indicating that the parafovea is susceptible to undergo lipid peroxidation. Foveolar COS were immunonegative until 81 years, which developed IR later (>83 years). IR to glutathione S-transferase-π1 was moderate until eight decade and then showed a decrease in photoreceptor cells between ninth and tenth decade, while glutaredoxin-1 maintained a steady expression with aging. Damaged COS were present in aged retinas, and inner segments and photoreceptor nuclei also showed some degree of alterations. Although there was increased lipid peroxidation with aging, cone death was minimal in those retinas. The two antioxidant enzymes studied here, may play a role in protecting photoreceptors against OS with advanced aging.

Effect of Vitamin E and Polyunsaturated Fatty Acids on Cryopreserved Sperm Quality in Bos taurus Bulls Under Testicular Heat Stress

ABSTRACTTaurine bulls are highly susceptible to heat stress, leading to increased oxidative stress (OS) and impaired sperm viability. Polyunsaturated fatty acids (PUFAs) supplementation can be an alternative to improve semen quality, which also results in more sperm susceptibility to lipid peroxidation. Moreover, this deleterious effect can be exacerbated in animals affected by heat stress. Vitamin E is a key antioxidant that counteracts lipid peroxidation of sperm membrane caused by OS. Thus, combining PUFAs with vitamin E may improve sperm quality. In this context, this study aimed to evaluate the effect of interaction between PUFAs and vitamin E on sperm quality in Bos taurus bulls under testicular heat stress. Sixteen taurine bulls under testicular heat stress were randomly assigned in four groups: Control, Vitamin E, PUFA, and PUFA + Vitamin E. All groups lasted for 60 days. Samples were cryopreserved/thawed and analyzed for motility variables (CASA), membrane and acrosome integrity, mitochondrial activity, susceptibility to oxidative stress, DNA integrity, and sperm-binding capacity. Results showed that vitamin E had a beneficial effect on some sperm characteristics, whereas PUFA supplementation had an adverse effect when the two treatments were evaluated separately. Finally, the association between PUFAs and vitamin E did not improve sperm quality.

Tu1558 - Portal Triad Clamping in Rats, and Reperfusion. BPC 157 Acutely Counteracts Lipid Peroxidation in Ischemia and in Reperfusion in Liver

Tu1558 - Portal Triad Clamping in Rats, and Reperfusion. BPC 157 Acutely Counteracts Lipid Peroxidation in Ischemia and in Reperfusion in Liver

Novel antioxidant capacity assay for lipophilic compounds using electron paramagnetic resonance spectroscopy.

A novel antioxidant capacity assay for lipophilic compounds was developed using electron paramagnetic resonance (EPR) spectroscopy. The assay is based on antioxidant’s scavenging ability against the tert-butoxyl radical generated photolytically from di-tert-butyl peroxide in ethyl acetate, and named the tert-butoxyl-based antioxidant capacity (BAC) assay. The radical was trapped by spin trap, 5,5-dimethyl-1-pyrroline-N-oxide, and EPR signal intensity of the spin adduct was used as a quantitative marker of radical levels. Signal intensity decreased in a dose-dependent manner in the presence of an antioxidant that competitively reacts with the radical, which was utilized to evaluate BAC values.The BAC method enabled the accurate estimation of antioxidant capacity for lipophilic materials that may counteract lipid peroxidation in biological membranes. The BAC values for quercetin and caffeic acid are 0.639 ± 0.020 and 0.118 ± 0.012 trolox equivalents, respectively, which are much smaller than values obtained by other aqueous methods such as H-ORAC and ORAC-EPR. Thus, antioxidants present in a non-aqueous environment should be evaluated using a non-aqueous system. In combination with in situ ascorbate reduction, the BAC method was capable of accurately determining the antioxidant capacity of water-insoluble materials that may be reduced in living cells.

The use of Pomegranate Juice for Counteract Lipid Peroxidation that Naturally occurred During Liquid Storage of Roosters’ Semen

Materials and Methods: In an attempt to find a suitable in vitro storage method for roosters’ semen, an experiment was conducted to study the influence of inclusion pomegranate juice (PJ) into semen diluent on semen quality during liquid storage for up to 36 h. A total of 60 White Leghorn roosters, 40 weeks of age, randomly divided into 6 treatment groups (10 males each) were used in this study. Treatment groups were as follows: T1 = fresh semen, T2 = semen diluted 1:2 with Al-Daraji 2 diluent (AD2D) alone, T3 – T6 = semen diluted 1:2 with AD2D supplemented with 2 ml, 4 ml, 6 ml or 8 ml of PJ / 100 ml of diluent, respectively. Semen samples were assessed after in vitro storage at 4 – 6˚C for 12 h, 24 h or 36 h as regards mass motility, individual motility and percentages of dead spermatozoa, abnormal spermatozoa and acrosomal abnormalities. Results: Results revealed that supplementing the diluent of roosters semen with PJ (T3, T4, T5 and T6) and then storing it for different storage periods (12 h, 24 h or 36 h) resulted in significant (p < 0.05) improvement in spermatozoa motility, viability, morphology and acrosomal integrity in comparison with the control group (T1). Moreover, T5 and T6 surpassed other treatments with respect to these semen characteristics, while there were no significant differences between T2, T3 and T4 concerning all semen traits included in this study. Conclusion: In conclusion, the substitution of AD2D diluent composition with PJ significantly improves the quality of roosters semen that is in vitro stored for up to 36 h. Furthermore, the positive effect of PJ observed in this study may be due to enhanced sperm resistance to lipid peroxidation that naturally occurred during in vitro storage of avian semen.

Morphofunctional properties of red blood cells and erythropoiesis intensity during 105-day isolation

Throughout the 105-day experiment with isolation and confinement, the energy-producing system of red blood cells did not exhibit substantial changes. A compensatory increase in G6PD activity intensified reparative processes in the cell and inhibited lipid peroxidation (LPO). The possibility of LPO activation in the erythrocyte plasma membrane can be inferred from changes in lipid and phospholipid fractions, which testifies to an increase in the membrane microviscosity on days 70 and 105 of the experiment and after its completion. On days 70 and 105, the content of erythropoietin in all test subjects decreased, which points to a reduced erythropoietic activity and a decrease in the content of hemoglobin, erythrocyte count, hematocrit value, and hemoglobin content per erythrocyte. These shifts persisted after the experiment although erythropoietin level was slightly increased on these days. The shifts were adaptive by nature while changes in biochemical values were compensatory reactions aimed at counteracting lipid peroxidation activation by emotional stresses of the experiment.

Breaking the chain at the membrane: paraoxonase 2 counteracts lipid peroxidation at the plasma membrane

Lipid peroxidation through electrophilic molecules of extracellular origin is involved in the pathogenesis of many inflammatory conditions. To counteract free radical actions at the plasma membrane, cells host a variety of antioxidative enzymes. Here we analyzed localization, membrane topology, and trafficking of PON2 a member of the paraoxonase family of 3 enzymatically active proteins (PON1-3) found to have antiatherogenic properties. Immunohistochemistry localized PON2 to the villous tip of human intestinal epithelial cells. Employing membrane preparations, surface biotinylation experiments, and mutational analyses in HEK 293T and HeLa cells, we demonstrate that PON2 is a type II transmembrane protein. A hydrophobic stretch in the N terminus was identified as single transmembrane domain of PON2. The enzymatically active domain faced the extracellular compartment, where it suppressed lipid peroxidation (P<0.05) and regulated the glucosylceramide content, as demonstrated by mass spectrometry (P<0.05). PON2 translocation to the plasma membrane was dependent on intracellular calcium responses and could be induced to >10-fold as compared to baseline (P=0.0001) by oxidative stress. Taken together, these data identify the paraoxonase protein PON2 as a type II transmembrane protein, which is dynamically translocated to the plasma membrane in response to oxidative stress to counteract lipid peroxidation.