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Among the causes of male infertility, enough attention is paid to oxidative stress, which in turn is a pathogenetic link in the inflammatory process. However, there is practically no information on the content of oxidized modified proteins in the semen, which makes it difficult to study the pathogenesis of diseases of the male reproductive system. In part, protein oxidation may be due to the production of reactive oxygen species by microorganisms, both directly and indirectly through the activation of immune system cells. The aim of the research was to study the level of oxidized modified proteins and changes in immunoglobulin concentrations in the semen under bacteriospermia. A study was made of the ejaculate of 48 men who applied to the clinic for infertility in marriage. The comparison group consisted of 32 practically healthy men who had no growth of microorganisms in the ejaculate samples. When conducting bacteriological analysis, the studied samples were diluted 10 times and used the generally accepted method. The concentration of albumin, immunoglobulins A, M, G, E was determined in the spermatic fluid. The oxidative modification of proteins was evaluated in the reaction with 2,4-dinitrophenylhydrazine. The concentration of oxidized proteins was expressed in nmol/mg of the total protein of the studied biological fluid. The biuret method was used to determine the protein concentration. Statistical analysis of the results was performed using descriptive statistics and Student's t-test for paired data. The concentration of protein in the seminal fluid did not differ significantly among the studied groups. The albumin concentration (16.96±1.28 mg/mL) was statistically significantly lower in the absence of microorganism growth than in bacteriospermia. With bacteriospermia, a decrease in the concentration of IgM and IgA and an increase in the level of IgG were noted. The degree of protein oxidation is maximum when enterobacteria are isolated from seminal fluid. Thus, during the studies it was found that, despite the absence of a clinic, with asymptomatic bacteriospermia, the secretion of immunoglobulins G into the semen is observed. The accumulation of oxidized proteins in the seminal fluid in bacteriospermia has been shown.

Oxidized protein overloading caused by diabetes is one accelerating pathological pathway in diabetic encephalopathy development. To determine whether the PA28-regulated function of the proteasome plays a role in diabetes-induced oxidative damaged protein degradation, brain PA28α and PA28β interference experiments were performed in a high-fat diet (HFD) and streptozotocin (STZ)-induced rat model. The present results showed that proteasome activity was changed in the brains of diabetic rats, but the constitutive subunits were not. In vivo PA28α and PA28β inhibition via adeno-associated virus (AAV) shRNA infection successfully decreased PA28 protein levels and further exacerbated oxidized proteins load by regulating proteasome catalytic activity. These findings suggest that the proteasome plays a role in the elimination of oxidized proteins and that PA28 is functionally involved in the regulation of proteasome activity in vivo. This study suggests that abnormal protein turbulence occurring in the diabetic brain could be explained by the proteasome-mediated degradation pathway. Changes in proteasome activity regulator PA28 could be a reason to induce oxidative aggregation in diabetic brain. Proteasome regulator PA28 inhibition in vivo by AAV vector injection could aggravate oxidized proteins abundance in brain of HFD-STZ diabetic rat model. [ABSTRACT FROM AUTHOR]

Trigonelline (TRIG) is a natural compound in an alkaloid family found in diverse plants. This compound exerts anti-inflammatory, anti-allergic, anti-oxidative and anti-fibrotic activities in several disease models. However, its beneficial role in endothelial injury, especially induced by diabetes, is unclear. We, therefore, evaluated the effects of TRIG on the cellular proteome of human endothelial (EA.hy926) cells followed by functional validation in high-glucose (HG)-induced endothelial deteriorations. Label-free quantification using nanoLC-ESI-Qq-TOF MS/MS revealed 40 downregulated and 29 upregulated proteins induced by TRIG. Functional enrichment analysis using DAVID and REVIGO tools suggested the involvement of these altered proteins in several biological processes and molecular functions, particularly cell-cell adhesion, ATP metabolic process, cell redox homeostasis, cadherin binding, and ATP hydrolysis activity. Experimental validation showed that HG triggered endothelial-to-mesenchymal transition (EndMT) (as demonstrated by increased spindle index and mesenchymal markers, i.e., fibronectin and vimentin, and decreased endothelial markers, i.e., PECAM-1 and VE-cadherin), increased oxidized proteins, and reduced intracellular ATP, active mitochondria, endothelial tube/mesh formation and VEGF secretion. However, TRIG successfully abolished all these defects induced by HG. These data indicate that TRIG prevents HG-induced EndMT, oxidative stress, mitochondrial dysfunction, and impaired angiogenic activity in human endothelial cells. [Display omitted] • Trigonelline (TRIG) significantly altered levels of 69 proteins in EA.hy926 cells. • The altered proteins were involved in many biological processes/molecular functions. • High-glucose (HG) triggered endothelial-to-mesenchymal transition (EndMT). • HG induced oxidative stress, mitochondrial dysfunction and impaired angiogenesis. • TRIG could prevent all these HG-induced defects in human endothelial cells. [ABSTRACT FROM AUTHOR]

This study aimed to assess the antioxidant activity of golden chlorella (GoC) and grape pomace (GrP) extracts both in vitro and in pea protein-based extrudates. We hypothesized that GoC/GrP would limit oxidation of proteins in the extrudates compared with commercial antioxidants. The results showed that GoC extract was effective in metal chelation and GrP extract possessed excellent radical scavenging activity and reducing power. Protein oxidation inevitably occurred after low-moisture extrusion in terms of elevated level of protein carbonyls and the gradual loss of thiols. LC-MS/MS revealed that the monoxidation and 4-hydroxynonenal adduction were the major oxidative modifications, and legumin was the most susceptible globulin for oxidation. The GoC/GrP extracts effectively retarded the oxidation progress in extrudates by lower intensity of oxidized peptides, whereas protein electrophoretic profiles remained unaffected. This study highlighted the great potential of GoC/GrP as natural antioxidants in plant-based foods., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Palmitic acid (PA) leads to lipotoxicity in type 2 diabetes and induces oxidative stress in podocytes. Oxidized cellular proteins are degraded by proteasomes. The role of proteasomes in PA- or oxidative stress-induced podocyte injury and pathogenesis of diabetic nephropathy (DN) is unknown. We investigated the effects of PA on expression of 20S and 26S proteasomes, proteasome activator 28 (PA28) regulators, and the immunoproteasome in cultured podocytes and renal cortical tissues of db/db and db/m mice using Western blot analysis. Glomerular areas and glomerular basement membrane (GBM) widths of db/db and db/m mice were examined using morphometry. Short-term incubation of PA or low levels of H2O2 upregulated only the immunoproteasome in cultured podocytes. Long-term exposure of podocytes to PA ultimately downregulated the immunoproteasome as with other proteasomes, whereas oleic acid (OA) or eicosapentaenoic acid (EPA) restored the PA-induced decreased protein levels. In db/db mice, renal cortical immunoproteasome expression with PA28α was significantly decreased compared with db/m mice, and glomerular areas and GBM widths were significantly increased compared with db/m mice. Feeding of an OA-rich olive oil or EPA-rich fish oil protected db/db mice against the reduced renal cortical immunoproteasome expression, glomerular enlargement, and GBM thickening. These results demonstrate that lipotoxicity downregulates the immunoproteasome in podocytes and kidneys in type 2 diabetes and that OA and EPA protected type 2 diabetic mice against decreased renal cortical immunoproteasome expression and the progression of DN. Given this, lipotoxicity-induced podocyte injury with impaired immunoproteasome expression appears to play an important role in the pathogenesis of DN. [ABSTRACT FROM AUTHOR]

Protein oxidation readily occurs in postmortem muscle during storage and processing. Over the past decade new analytical methods have been developed and new aspects of protein oxidation in meat have been studied, such as the reaction mechanism, and impacts on eating quality and nutritional value. It is now evident that amino acid side chains in myofibrillar proteins undergoes modifications due to oxidative stress. In turn this will lead to formation of new protein-protein cross-links in structural proteins, however, also the overall level of fixed-charge groups attached to the peptide backbones is modified. Meat texture and water-holding are important quality attributes and they are affected by the oxidation of structural proteins. Different mechanisms have been suggested to explain the oxidation-induced quality changes, focusing mainly on reduced proteolysis and formation of cross-links. This review explores the current understanding of protein oxidation in fresh meat in relation to texture and water-holding. The consequences of protein oxidation at molecular level in relation to oxidation-induced cross-linking and changes in net charges of myofibrillar proteins, and the impacts on texture and water-holding are discussed. [ABSTRACT FROM AUTHOR]

Abstract The skeletal muscle is a crucial tissue for maintaining whole body homeostasis. Aging seems to have a disruptive effect on skeletal muscle homeostasis including proteostasis. However, how aging specifically impacts slow and fast twitch fiber types remains elusive. Muscle proteostasis is largely maintained by the proteasomal system. Here we characterized the proteasomal system in two different fiber types, using a non-sarcopenic aging model. By analyzing the proteasomal activity and amount, as well as the polyubiquitinated proteins and the level of protein oxidation in Musculus soleus (Sol) and Musculus extensor digitorum longus (EDL), we found that the slow twitch Sol muscle shows an overall higher respiratory and proteasomal activity in young and old animals. However, especially during aging the fast twitch EDL muscle reduces protein oxidation by an increase of antioxidant capacity. Thus, under adaptive non-sarcopenic conditions, the two fibers types seem to have different strategies to avoid age-related changes. [ABSTRACT FROM AUTHOR]

The proteasome is a multimeric and multicatalytic intracellular protease responsible for the degradation of proteins involved in cell cycle control, various signaling processes, antigen presentation, and control of protein synthesis. The central catalytic complex of the proteasome is called the 20S core particle. The majority of these are flanked on one or both sides by regulatory units. Most common among these units is the 19S regulatory unit. When coupled to the 19S unit, the complex is termed the asymmetric or symmetric 26S proteasome depending on whether one or both sides are coupled to the 19S unit, respectively. The 26S proteasome recognizes poly-ubiquitinylated substrates targeted for proteolysis. Targeted proteins interact with the 19S unit where they are deubiquitinylated, unfolded, and translocated to the 20S catalytic chamber for degradation. The 26S proteasome is responsible for the degradation of major proteins involved in the regulation of the cellular cycle, antigen presentation and control of protein synthesis. Alternatively, the proteasome is also active when dissociated from regulatory units. This free pool of 20S proteasome is described in yeast to mammalian cells. The free 20S proteasome degrades proteins by a process independent of poly-ubiquitinylation and ATP consumption. Oxidatively modified proteins and other substrates are degraded in this manner. The 20S proteasome comprises two central heptamers (β-rings) where the catalytic sites are located and two external heptamers (α-rings) that are responsible for proteasomal gating. Because the 20S proteasome lacks regulatory units, it is unclear what mechanisms regulate the gating of α-rings between open and closed forms. In the present review, we discuss 20S proteasomal gating modulation through a redox mechanism, namely, S-glutathionylation of cysteine residues located in the α-rings, and the consequence of this post-translational modification on 20S proteasomal function.

High neutrophil (PMN, Polymorphonuclear neutrophil) counts in the endometrium of cows affected by endometritis, suggests the involvement of oxidative stress (OS) among the causes of impaired fertility. Protein oxidation, in particular, advanced oxidation protein products (AOPP), are OS biomarkers linked to PMN activity. To test this hypothesis, the relationship between protein oxidation and uterus health was studied in thirty-eight dairy cows during the puerperium. The animals were found to be cycling, without any signs of disease and pharmacological treatments. PMN count was performed either through a cytobrush or a uterine horn lavage (UHL). Cows were classified into four groups, based on the uterine ultrasonographic characteristics and the PMN percentage in the uterine horns with a higher percentage of high neutrophil horn (HNH). They were classified as: Healthy (H); Subclinical Endometritis (SCE); Grade 1 Endometritis (EM1); and Grade 2 Endometritis (EM2). AOPP and carbonyls were measured in plasma and UHL. UHL samples underwent Western blot analysis to visualize the carbonyl and dityrosine formation. Plasma AOPP were higher (p < 0.05) in EM2. AOPP and carbonyl group concentrations were higher in the HNH samples (p < 0.05). Protein concentration in the UHL was higher in the EM2 (p < 0.05). Carbonyl and dityrosine formation was more intense in EM1 and EM2. Protein oxidation observed in the EM2 suggests the presence of an inflammatory status in the uterus which, if not adequately hindered, could result in low fertility.

Protein oxidation has become a topic of great scientific interest in the field of food science and nutrition. Food proteins are known to be preferential targets of radical species, and protein oxidation has relevant consequences on protein functionality and food quality. Current trends in this field call attention to the nutritional and health dimensions of oxidized foods. Both lipid and protein oxidation products are accumulated in food during processing and storage and also upon food intake during the subsequent digestion phases. The gastrointestinal tract and internal organs are exposed to the cytotoxic and mutagenic potential of these species. While the molecular basis of the pathogenesis of particular dietary lipid oxidation products is well known, the impact of dietary oxidized proteins on human health has been largely ignored. The well-established association between in vivo protein oxidation and aging and age-related diseases urges scientists to investigate the contribution of dietary protein oxidation to particular pathological conditions. Recent reports indicate the involvement of dietary protein oxidation species on particular health disorders, which emphasizes the link between dietary and in vivo protein oxidation. [ABSTRACT FROM AUTHOR]

Reactive oxygen species (ROS) are engaged in several processes essential for normal cell functioning, such as differentiation, anti-microbial defense, stimulus sensing and signaling. Interestingly, recent studies imply that cellular signal transduction and gene regulation are mediated not only directly by ROS but also by the molecules derived from ROS-mediated oxidation. Lipid peroxidation leads to non-enzymatic formation of oxylipins. These molecules were shown to modulate expression of signaling associated genes including genes encoding phosphatases, kinases and transcription factors. Oxidized peptides derived from protein oxidation might be engaged in organelle-specific ROS signaling. In turn, oxidation of particular mRNAs leads to decrease in the level of encoded proteins and thus, contributes to the post-transcriptional regulation of gene expression. Present mini review summarizes latest findings concerning involvement of products of lipid, protein and RNA oxidation in signal transduction and gene regulation.

ABSTRACT The nonsteroidal anti-inflammatory drug ketorolac is extensively used in the treatment of acute postoperative pain. This pharmaceutical has been found at concentrations of 0.2-60 µg/L in diverse water bodies around the world; however, its effects on aquatic organisms remain unknown. The present study, evaluated the oxidative stress and genotoxicity induced by sublethal concentrations of ketorolac (1 and 60 µg/L) on liver, brain, and blood of the common carp Cyprinus carpio. This toxicant induced oxidative damage (increased lipid peroxidation, hydroperoxide content, and protein carbonyl content) as well as changes in antioxidant status (superoxide dismutase, catalase, and glutathione peroxidase activity) in liver and brain of carp. In blood, ketorolac increased the frequency of micronuclei and is therefore genotoxic for the test species. The effects observed were time and concentration dependent. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1035-1043, 2016. [ABSTRACT FROM AUTHOR]

Disinfection by-products (DBPs) are generated through the reaction of chlorine with organic and inorganic matter in indoor swimming pools. Different DBPs are present in indoor swimming pools. This study evaluated the effects of different chlorinated formations in oxidative stress and lung damage in 20 swimmers after 40 min of aerobic swimming in 3 indoor pools with different characteristics. Biological samples were collected to measure lung damage (serum-surfactantassociated proteins A and B), oxidative stress parameters (plasma protein carbonylation and malondialdehyde, and wholeblood glutathione oxidation), and swimming exertion values (blood lactate) before and after exercise. Free chlorine and combined chlorine in water, and chlorine in air samples were determined in all the swimming pools. Chlorination as disinfection treatment led to the formation of chloramines in water samples, mainly mono- and dichloramine. However, free chlorine was the predominate species in ultraviolet-treated swimming pool. Levels of total chlorine increased as a function of the swimming activity in chlorinated swimming pools. The lower quality of the installation resulted in a higher content of total chlorine, especially in air samples, and therefore a higher exposure of the swimmer to DBPs. However, the concentration level of chlorinated DBPs did not result in significant variation in serum-surfactant-associated proteins A and oxidative stress parameters in swimmers. In conclusion, the quality of the installation affected the DBPs concentration; however, it did not lead to lung epithelial damage and oxidative stress parameters in swimmers. [ABSTRACT FROM AUTHOR]

The activities of antioxidant enzymes and the contents of the oxidized forms of the proteins in the blood of the Black Sea round goby Neogobius melanostomus, which were sampled in the waters of Sevastopol in 2003 and 2012, were studied. An increase in the values of the tested biomarkers in fish catches of 2012 compared to those of 2003 was recorded. The causes for the observed changes are discussed. [ABSTRACT FROM AUTHOR]

Se estudiaron 24 hombres y 28 mujeres con edades comprendidas entre 60 y 91 años, con el objetivo de evaluar el estado antioxidante y los indicadores de daño oxidativo de una población de ancianos de Las Tunas. En el suero sanguíneo se determinó la concentración de los antioxidantes no enzimáticos: vitamina A, b caroteno, vitamina E y vitamina C, así como del zinc, la actividad de las enzimas antioxidantes superóxido dismutasa y catalasa y la concentración de los indicadores de daño oxidativo, malondialdehído y proteínas oxidadas. Con excepción de la vitamina C y del zinc para la población total y las mujeres, los demás indicadores antioxidantes se encuentran en el rango reportado como normal. El estado de la vitamina C y del zinc es deficiente en un elevado porcentaje de los individuos; en tanto que la actividad de las enzimas antioxidantes resulta ser superior en relación con adultos mayores, sin embargo la concentración de los indicadores de daño oxidativo se encuentra dentro de los límites normales, lo que pudiera ser indicativo de un estrés oxidativo compensadoTwenty four men and twenty eight females aged 60 to 91 years were studied to evaluate the antioxidant status and the oxidative damage indicators of an elderly population in Las Tunas province. Concentration in blood serum of non enzymatic antioxidants such as vitamin A, b carotene, vitamin E, vitamin C, and zinc; the activity of antioxidant enzymes superoxide dismutase and catalase as well as concentration of indicators of oxidative damage, malondialdehyde and oxidized proteins were determined. Except for vitamin C and zinc for the whole population and the females respectively, the rest of antioxidant indicators are within the reported normal ranges. The states of Vitamin C and zinc are deficient in a high percentage of individuals whereas the activity of antioxidant enzymes is higher in older adults. However, the concentration of oxidative damage indicators are within the normal limits, which may indicate compensated oxidative stress

Reactive oxygen species (ROS) are engaged in several processes essential for normal cell functioning, such as differentiation, anti-microbial defense, stimulus sensing and signaling. Interestingly, recent studies imply that cellular signal transduction and gene regulation are mediated not only directly by ROS but also by the molecules derived from ROS-mediated oxidation. Lipid peroxidation leads to non-enzymatic formation of oxylipins. These molecules were shown to modulate expression of signaling associated genes including genes encoding phosphatases, kinases and transcription factors. Oxidized peptides derived from protein oxidation might be engaged in organelle-specific ROS signaling. In turn, oxidation of particular mRNAs leads to decrease in the level of encoded proteins and thus, contributes to the post-transcriptional regulation of gene expression. Present mini review summarizes latest findings concerning involvement of products of lipid, protein and RNA oxidation in signal transduction and gene regulation. [ABSTRACT FROM AUTHOR]

The nonsteroidal anti-inflammatory naproxen sodium is among the most commonly used pharmaceuticals in Mexico. Nevertheless, there is no adequate regulation to control its sale, use, and disposal. This agent can enter water bodies by diverse pathways, attaining significant concentrations, especially in sediments since they are the final reservoir of many organic contaminants, and may induce damage on hydrobionts. Naproxen sodium is water-soluble, shows stability in aquatic ecosystems, and can form reactive products when oxidized. The aim of this study was to evaluate the oxidative stress and consequent damage to genetic material induced by naproxen sodium in sediments on Hyalella azteca. After 48 h of exposure to artificial sediments enriched with 76.6 and 339.2 mg kg (equivalent to 1/10 of the LC and the NOAEL obtained in an earlier 48-h acute toxicity assay), the following biomarkers were evaluated: lipid peroxidation level, protein carbonyl content, activity of the antioxidant enzymes, and DNA damage. Results show that naproxen sodium induces oxidative stress (increased lipid and protein oxidation, as well as superoxide dismutase and catalase activity and decrease of glutathione peroxidase activity) and genotoxicity (increased oxidative damage of DNA) at sublethal concentrations on H. azteca, being the damage, concentration dependent. [ABSTRACT FROM AUTHOR]

Protein S-glutathionylation is a post-translational modification that controls many cellular pathways. Recently, we demonstrated that the α5-subunit of the 20S proteasome is S-glutathionylated in yeast cells grown to the stationary phase in rich medium containing glucose, stimulating 20S core gate opening and increasing the degradation of oxidized proteins. In the present study, we evaluated the correlation between proteasomal S-glutathionylation and the intracellular redox status. The redox status was controlled by growing yeast cells in distinct carbon sources which induced respiratory (glycerol/ethanol) or fermentative (glucose) metabolism. Cells grown under glycerol/ethanol displayed higher reductive power when compared to cells grown under glucose. When purified from cells grown in glucose, 20S proteasome α5-subunit exhibited an intense anti-glutathione labeling. A higher frequency of the open catalytic chamber gate was observed in the S-glutathionylated preparations as demonstrated by transmission electron microscopy. Therefore, cells that had been grown in glucose displayed an increased ability to degrade oxidized proteins. The results of the present study suggest that 20S proteasomal S-glutathionylation is a relevant adaptive response to oxidative stress that is capable to sense the intracellular redox environment, leading to the removal of oxidized proteins via a process that is not dependent upon ubiquitylation and ATP consumption. [ABSTRACT FROM AUTHOR]

Pharmaceutical agents, like diclofenac and acetaminophen, are sold without prescription leading to excessive use. These agents may reach water bodies through various routes and attain significant concentrations, posing a risk to hydrobiont health. Diverse studies have shown that during the biotransformation of these compounds, reactive metabolites and reactive oxygen species are produced which induce oxidative stress and damage to diverse biomolecules. However, toxicity studies that assess the effects of a mixture of contaminants are scarce, being very important as this is how they are actually in the environment. The present study aimed to evaluate the oxidative stress induced by mixture of diclofenac and acetaminophen on Cyprinus carpio and compare with the effect produced by these pharmaceuticals in isolation. A 96-h sublethal toxicity assay of the tested pharmaceuticals (isolated and in mixture) was performed and the following biomarkers were evaluated: lipid peroxidation, protein carbonyl content, and activity of the antioxidant enzymes superoxide dismutase, catalase, and glutathione peroxidase. The pharmaceuticals evaluated induce oxidative stress on C. carpio in isolated form and as a mixture, but the level of damage being dependent on the organ evaluated as well as the type of toxicant and form of exposure (in isolation or as a mixture). [ABSTRACT FROM AUTHOR]

The strictly regulated removal of oxidized structures is a universal stress response of eukaryotic cells that targets damaged or toxic components for vacuolar or lysosomal degradation. Autophagy stands at the crossroad between cell survival and death. It promotes survival by degrading proteins and organelles damaged during oxidative stress, but it is also activated as a part of death programs, when the damage cannot be overcome. Evidence is accumulating that the cellular sites of ROS production and signaling may be primary targets of autophagy. Therefore, autophagosomal targeting of mitochondria (mitophagy) is of particular importance. Mitophagy is a selective process that can specifically target dysfunctional mitochondria, but also mitophagy may play a role in controlling the number and quality of mitochondria during stress. Here we review the mechanisms of both non-specific autophagy and mitochondrial targeting in plants, drawing analogies and emphasizing differences with yeast and mammalian systems. [ABSTRACT FROM AUTHOR]

Despite the widely accepted belief that selenium toxicity in plants is manifested by the misincorporation of selenocysteine into selenoproteins, there is a lack of data suggesting that selenoproteins are malformed or misfolded. Plant mechanisms to prevent the formation of selenoproteins are associated with increased selenium tolerance, yet there is no evidence to suggest that selenoproteins are malformed or potentially misfolded. We reasoned that if selenoproteins are malformed, then they might be degraded by the ubiquitin-proteasome pathway. The data demonstrate that selenate treatment induced the accumulation of both oxidized and ubiquitinated proteins, thus implicating both the 20S and 26S proteasome of Stanleya pinnata, a selenium-hyperaccumulating plant, in a selenate response. Inhibition of the proteasome increases the amount of selenium incorporated into protein, but not other elements. Furthermore, a higher percentage of selenium was found in a ubiquitinated protein fraction compared with other elements, suggesting that malformed selenoproteins are preferentially ubiquitinated and removed by the proteasome. Additionally, levels of the 20S and 26S proteasome and two heat shock proteins increase upon selenate treatment. Arabidopsis mutants with defects in the 26S proteasome have decreased selenium tolerance, which further supports the hypothesis that the 26S proteasome probably prevents selenium toxicity by removing selenoproteins. [ABSTRACT FROM AUTHOR]

The principal function of the proteasome is targeted degradation of intracellular proteins. Proteasome dysfunction has been observed in experimental cardiomyopathies and implicated in human congestive heart failure. Measures to enhance proteasome proteolytic function are currently lacking but would be beneficial in testing the pathogenic role of proteasome dysfunction and could have significant therapeutic potential. The association of proteasome activator 28 (PA28) with the 20S proteasome may play a role in antigen processing. It is unclear, however, whether the PA28 plays any important role outside of antigen presentation, although up-regulation of PA28 has been observed in certain types of cardiomyopathy. Here, we show that PA28α expression (PA28αOE) stabilized PA28β, increased 11S proteasomes, and enhanced the degradation of a previously validated proteasome surrogate substrate (GFPu) in cultured neonatal rat cardiomyocytes. PA28αOE significantly attenuated H2O2-induced increases in the protein carbonyls and markedly suppressed apoptosis in cultured cardiomyocytes under basal conditions or when stressed by H2O2-. We conclude that PA28αOE is sufficient to up-regulate 11S proteasomes, enhance proteasome-mediated removal of misfolded and oxidized proteins, and protect against oxidative stress in cardiomyocytes, providing a highly sought means to increase proteasomal degradation of abnormal cellular proteins. [ABSTRACT FROM AUTHOR]

AtFtsH4 is one of four inner membrane-bound mitochondrial ATP-dependent metalloproteases in Arabidopsis thaliana, called AAA proteases, whose catalytic site is exposed to the intermembrane space. In the present study, we used a reverse-genetic approach to investigate the physiological role of the AtFtsH4 protease. We found that loss of AtFtsH4 did not significantly affect Arabidopsis growth under optimal conditions (long days); however, severe morphological and developmental abnormalities in late rosette development occurred under short-day conditions. The asymmetric shape and irregular serration of expanding leaf blades were the most striking features of the ftsh4 mutant phenotype. The severe abnormal morphology of the leaf blades was accompanied by ultrastructural changes in mitochondria and chloroplasts. These abnormalities correlated with elevated levels of reactive oxygen species and carbonylated mitochondrial proteins. We found that two classes of molecular chaperones, Hsp70 and prohibitins, were over-expressed in ftsh4 mutants during late vegetative growth under both short- and long-day conditions. Taken together, our data indicate that lack of AtFtsH4 results in impairment of organelle development and Arabidopsis leaf morphology under short-day conditions. [ABSTRACT FROM AUTHOR]

Abstract: Reactions of tryptophan residues in proteins with radical and other oxidative species frequently lead to cleavage of the indole ring, modifying tryptophan residues into N-formylkynurenine (NFK) and kynurenine. Tryptophan modification has been detected in physiologically important proteins and has been associated with a number of human disease conditions. Modified residues have been identified through various combinations of proteomic analyses, tryptic digestion, HPLC, and mass spectrometry. Here we present a novel, immunological approach using polyclonal antiserum for detection of NFK. The specificity of our antiserum is confirmed using photooxidation and radical-mediated oxidation of proteins with and without tryptophan residues. The sensitivity of our antiserum is validated through detection of NFK in photooxidized myoglobin (two tryptophan residues) and in carbonate radical-oxidized human SOD1, which contains a single tryptophan residue. Analysis of photooxidized milk also shows that our antiserum can detect NFK residues in a mixture of proteins. Results from mass spectrometric analysis of photooxidized myoglobin samples corroborate the immunological data, detecting an increase in NFK content as the extent of photooxidation increases. [Copyright &y& Elsevier]

Elevated levels of oxidized proteins are reported in diseased tissue from age-related pathologies such as atherosclerosis, neurodegenerative disorders, and cataract. Unlike the precise mechanisms that exist for the repair of nucleic acids, lipids, and carbohydrates, the primary pathway for the repair of oxidized proteins is complete catabolism to their constitutive amino acids. This process can be inefficient as is evidenced by their accumulation. It is generally considered that damaged proteins are degraded by the proteasome; however, this is only true for mildly oxidized proteins, because substrates must be unfolded to enter the narrow catalytic core. Rather, evidence suggests that moderately or heavily oxidized proteins are endocytosed and enter the endosomal/lysosomal system, indicating co-operation between the proteasomes and the lysosomes. Heavily modified substrates are incompletely degraded and accumulate within the lysosomal compartments resulting in the formation of lipofuscin-like, autofluorescent aggregates. Accumulation eventually results in impaired turnover of large organelles such as proteasomes and mitochondria, lysosomal destablization, leakage of proteases into the cytosol and apoptosis. In this review, we summarize reports published since our last assessments of the field of oxidized protein degradation including a role for modified proteins in the induction of apoptosis. © 2009 IUBMB IUBMB Life, 61(5): 522–527, 2009 [ABSTRACT FROM AUTHOR]

Abstract: Prior work has shown that d-amphetamine (AMPH) treatment or voluntary exercise improves cognitive functions after traumatic brain injury (TBI). In addition, voluntary exercise increases levels of brain-derived neurotrophic factor (BDNF). The current study was conducted to determine how AMPH and exercise treatments, either alone or in combination, affect molecular events that may underlie recovery following controlled cortical impact (CCI) injury in rats. We also determined if these treatments reduced injury-induced oxidative stress. Following a CCI or sham injury, rats received AMPH (1 mg/kg/day) or saline treatment via an ALZET® pump and were housed with or without access to a running wheel for 7 days. CCI rats ran significantly less than sham controls, but exercise level was not altered by drug treatment. On day 7 the hippocampus ipsilateral to injury was harvested and BDNF, synapsin I and phosphorylated (P) -synapsin I proteins were quantified. Exercise or AMPH alone significantly increased BDNF protein in sham and CCI rats, but this effect was lost with the combined treatment. In sham-injured rats synapsin I increased significantly after AMPH or exercise, but did not increase after combined treatment. Synapsin levels, including the P-synapsin/total synapsin ratio, were reduced from sham controls in the saline-treated CCI groups, with or without exercise. AMPH treatment significantly increased the P-synapsin/total synapsin ratio after CCI, an effect that was attenuated by combining AMPH with exercise. Exercise or AMPH treatment alone significantly decreased hippocampal carbonyl groups on oxidized proteins in the CCI rats, compared with saline-treated sedentary counterparts, but this reduction in a marker of oxidative stress was not found with the combination of exercise and AMPH treatment. These results indicate that, whereas exercise or AMPH treatment alone may induce plasticity and reduce oxidative stress after TBI, combining these treatments may cancel each other''s therapeutic effects. [Copyright &y& Elsevier]

Abstract: Aging has been associated with zinc deficiency, leading to chronic inflammation and subsequent oxidative stress, especially in the immune system. The increased oxidative stress provokes the accumulation of oxidized proteins, raising the problem of the efficacy of intracellular protein maintenance systems responsible for the elimination of oxidatively modified proteins. Our objective was to analyse the effect of zinc supplementation in the elderly on protein maintenance in peripheral blood lymphocytes. The status of the proteasome, which is in charge of oxidized protein degradation and the repair enzymes peptide methionine sulfoxide reductases, which can reverse methionine oxidation in proteins, were analysed on peripheral blood lymphocytes collected from 20 elderly subjects (age range between 59 and 85 years old) before and after zinc supplementation (10mg of zinc per day for 48±2 days). A decrease of oxidized protein content in zinc supplemented subjects was observed and was associated with an increase of expression levels and/or activities of proteasome and methionine sulfoxide reductases. Our results indicate that zinc treatment could enhance the anti-oxidative defences of peripheral blood lymphocytes by increasing the activities of protein maintenance systems responsible for the elimination of oxidatively modified proteins. [Copyright &y& Elsevier]

Nitric oxide (NO) is a small, ubiquitous molecule, whose physiological function in plants has recently been widely investigated. It seems that one of its pivotal properties is the antioxidant capacity, enabling plants to alleviate the effects of the oxidized stress. In this work we investigated the role of NO in soybean ( Glycine max L. Cv. Navico) cell suspension treated with cadmium. Sodium nitroprusside (SNP), nitric oxide donor, markedly decreased the negative influence of Cd2+ on cell growth. It was also found to stimulate superoxide dismutase (SOD, EC 1.15.1.1). Using specific fluorochromes — dihydroethidine (DHE) and 2′,7′- dichlorofluorescein (DCFH-DA) it was shown that NO was very effective in reducing the level of superoxide anion (O) and hydrogen peroxide, respectively. Furthermore, as evaluated by means of NO specific fluorochrome 4,5-diaminofluorescein diacetate (DAF-2DA), increased production of NO was found in Cd-treated cells. In cadmium-stressed cells SNP lowered the level of oxidized proteins. Our results suggest that the antioxidant properties of nitric oxide in Cd-treated soybean cells rely mainly on its ability to direct scavenging of ROS and stimulation of the antioxidant system. [ABSTRACT FROM AUTHOR]

We recently reported that the transient expression of polyglutamine tracts of various size in exon 1 of the huntingtin polypeptide (httEx1) generated abnormally high levels of intracellular reactive oxygen species that directly contributed to cell death. Here, we compared the protection generated by heat shock proteins to that provided by the antioxidant agent N-acetyl-l-cysteine. In cells expressing httEx1 with 72 glutamine repeats (httEx1-72Q), the overexpression of Hsp27 or Hsp70 plus Hdj-1(Hsp40) or treatment of the cells with N-acetyl-l-cysteine inhibited not only mitochondrial membrane potential disruption but also the increase in reactive oxygen species, nitric oxide and protein oxidation. However, only heat shock proteins and not N-acetyl-l-cysteine reduced the size of the inclusion bodies formed by httEx1-72Q. In cells expressing httEx1 polypeptide with 103 glutamine repeats (httEx1-103Q), heat shock proteins neither decreased oxidative damage nor reduced the size of the inclusions. In contrast, N-acetyl-l-cysteine still efficiently decreased the oxidative damage induced by httEx1-103Q polypeptide without altering the inclusions. N-Acetyl-l-cysteine was inactive with regard to proteasome inhibition, whereas heat shock proteins partially restored the caspase-like activity of this protease. These observations suggest some relationships between the presence of inclusion bodies and the oxidative damage induced by httEx1-polyQ. [ABSTRACT FROM AUTHOR]

Abstract: Proteins accumulate during aging and form insoluble protein aggregates. Microglia are responsible for their removal from the brain. During aging, changes within the microglia might play a crucial role in the malfunctioning of these cells. Therefore, we isolated primary microglial cells from adult rats and compared their activation status and their ability to degrade proteins to that of microglial cells isolated from newborn animals. The ability of adult microglial cells to degrade proteins is substantially decreased. However, the preincubation of microglial cells with vitamin E improves significantly the degradation of such modified proteins. The degradation of proteins from apoptotic vesicles is decreased in microglia isolated from adult rats. This might be the result of a suppression of the CD36 receptor due to vitamin E treatment. We concluded that microglial cells isolated from adult organisms have different metabolic properties and seem to be a more valuable model to study age-related diseases. [Copyright &y& Elsevier]

Proteins are major target for radicals and other oxidants when these are formed in both intra- and extracellular environments in vivo. Formation of lesions on proteins may be highly sensitive protein-based biomarkers for oxidative damage in mammalian systems. Oxidized proteins are often functionally inactive and their unfolding is associated with enhanced susceptibility to proteinases. ROS scavenging activities of intact proteins are weaker than those of misfolded proteins or equivalent concentrations of their constituent amino acids. Protein oxidation and enhanced proteolytic degradation, therefore, have been suggested to cause a net increase in ROS scavenging capacity. However, certain oxidized proteins are poorly handled by cells, and together with possible alterations in the rate of production of oxidized proteins, may contribute to the observed accumulation and damaging actions of oxidized proteins during ageing and in pathologies such as diabetes, arteriosclerosis and neurodegenerative diseases. Protein oxidation may play a controlling role in cellular remodelling and cell growth. There is some evidence that antioxidant supplementation may protect against protein oxidation, but additional controlled studies of antioxidant intake to evaluate the significance of dietary/pharmacological antioxidants in preventing physiological/pathological oxidative changes are needed. [ABSTRACT FROM AUTHOR]

Summary: When the tea (Camellia sinensis) leaf water potential was −1.1MPa (Moderate water deficit), there was 58% inhibition of photosynthesis accompanied by increased zeaxanthin, malondialdehyde, oxidized proteins and superoxide dismutase activity. When the leaf water potential was −2MPa (severe water deficit), there was nearly complete inhibition of photosynthesis apart from a decrease in chlorophylls, β-carotene, neoxanthin and lutein. Water deficit at this level caused further conversion of violaxanthin to zeaxanthin, suggesting damage to the photosynthetic apparatus. There were consistent decreases in antioxidants and pyridine nucleotides, and accumulation of catalytic Fe, malondialdehyde and oxidized proteins. It is inferred that, in tea plants, the increase in catalytic Fe and the decrease in antioxidant protection may be involved in the oxidative damage caused by severe water deficit, but not necessarily in the incipient stress induced by moderate water deficit. [Copyright &y& Elsevier]

Modification of proteins by reactive oxygen species is implicated in different disorders. The proteasome is a multicatalytic proteinase in charge of intracellular protein turnover and of oxidized proteins degradation. Consequently, proteasome function is very important in controlling the level of altered proteins in eukaryotic cells. Evidence for a decline in proteasome activity during skin photo-aging has been provided in Bulteau et al. in 2002. The ability of a lipid algae extract (Phaeodactylum tricornutum) to stimulate 20S proteasome peptidase activities was described by Nizard et al. in 2001. Furthermore, keratinocytes treated with Phaeodactylum tricornutum extract and then UVA and UVB irradiated, exhibited a sustained level of proteasome activity comparable to the one of nonirradiated cells. The level of modified proteins can be quantified by measurement of protein carbonyl content (Oxyblot technique), which has been shown to increase with aging and other disorders. In this paper, it is described that, in the presence of this lipid algae extract, the level of oxidized proteins is reduced, as assessed by the Oxyblot technique. These results are obtained both with culture of human keratinocytes and stratum corneum skin cells (obtained by stripping) from human volunteers. Altogether, these results argue for the presence of compounds in this algae extract that have a stimulating and/or protective effect on proteasome activity, resulting in a decreased level of protein oxidation. [ABSTRACT FROM AUTHOR]

In order to study the role of Helicobacter pylori infection in gastric carcinogenesis, we have measured oxidized (carbonyls) and nitrated (nitrotyrosine-containing) proteins as markers for oxidative and nitrative stress in 216 human gastric biopsies using dot and western immunoblots and correlated the results with H. pylori, cagA status, expression of interleukin-8 and inducible nitric oxide synthase (iNOS) mRNAs, and gastric pathology. Higher levels of both oxidized and nitrated proteins were found in patients with either chronic gastritis or duodenal ulcer than in those with normal mucosa. The levels of modified proteins were significantly higher in inflamed samples infected with H. pylori, especially cagA+ strains, and in those with expression of interleukin-8 and iNOS mRNAs than in those negative for these parameters. These results indicate that infection with cagA+ H. pylori induces significant oxidative and nitrative stress in stomach mucosa, contributing to the pathogenesis of H. pylori-associated gastroduodenal diseases. [ABSTRACT FROM AUTHOR]

The intragastric alcohol infusion rat model (IAIRM) of alcoholic liver disease (ALD) has been utilized in various laboratories to study various aspects of ALD pathogenesis including oxidative stress, cytokine upregulation, hypoxic damage, apoptosis, ubiquitin-proteasome pathway and CYP2E1 induction. The basic value of the model is that it produces pathologic changes which resemble ALD including microvesicular and macrovesicular fat, megamitochondria, apoptosis, central lobular and pericellular fibrosis, portal fibrosis, bridging fibrosis, central necrosis, and mixed inflammatory infiltrate including PMNs and lymphocytes. The model is valuable because the diet and ethanol intake are totally under the control of the investigator. A steady state can be maintained with high or low blood alcohol levels for long periods. The cycling of the blood alcohol levels, when a constant infusion rate of alcohol is maintained, simulates binge drinking. Using this model the importance of dietary fat, especially the degree of saturation of the fatty acids on the induction of liver pathology, has been documented. The role of endotoxin, the Kupffer cell, TNFα, and NADPH oxidase have been demonstrated. The importance of 2E1 in oxidative stress induction has been shown using inhibitors of the isozyme. The importance of dietary iron in the pathogenesis of cirrhosis has been documented. Acetaldehyde has been shown to play a role in preventing liver pathology by preventing NFκB activation. Using the model, to maintain high blood alcohol levels is found to be necessary to demonstrate proteasomal peptidase inhibition. Ubiquitin synthesis is also inhibited at high blood alcohol levels in the IAIRM model. Oxidized proteins accumulate in the liver at high blood alcohol levels. Neoantigens derived from protein adducts formed with products of oxidation induce autoimmune mechanisms of liver injury. Thus, in many ways the model has revolutionized our understanding of the pathogenesis of ALD.Copyright © 2001 National Science Council, ROC and S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Mitochondria actively participate in oxygenic metabolism and are one of the major sources of reactive oxygen species (ROS) production in plant cells. However, instead of measuring ROS concentrations in organelles it is more worthwhile to observe active ROS generation or downstream oxidation products, because the steady state level of ROS is easily buffered. Here, we describe how to measure the in vitro production of superoxide anion radicals (O 2 · - ) by mitochondria and the release of O 2 · - into the cytosol. A method to determine glutathione, which is the most abundant mitochondrial low-mass antioxidant, is presented since changes in the redox state of glutathione can be indicative of the oxidative action of ROS. The identification of oxidative damage to mitochondrial components is the ultimate symptom that ROS homeostasis is not under control. We present how to determine the extent of oxidation of membrane lipids and the carbonylation of mitochondrial proteins. In summary, oxidative stress symptoms have to be analyzed at different levels, including ROS production, scavenging capacity, and signs of destruction, which only together can be considered markers of mitochondrial ROS status., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

Caffeine is an active ingredient found in coffee and energy beverages. Its hepatoprotective effects against liver fibrosis are well-documented. Nonetheless, its renoprotective effects against renal fibrogenesis and epithelial-mesenchymal transition (EMT) processes remain unclear and under-investigated. In this study, the protective effects of caffeine against oxalate-induced EMT in renal tubular cells were evaluated by various assays to measure expression levels of epithelial and mesenchymal markers, cell migrating activity, level of oxidized proteins, and expression of Nrf2 and Snail1. Oxalate at sublethal dose significantly suppressed cell proliferation but increased cell elongation, spindle index and migration. Oxalate also decreased expression of epithelial markers (zonula occludens-1 (ZO-1) and E-cadherin) but increased expression of mesenchymal markers (fibronectin, vimentin and α-smooth muscle actin (α-SMA)). All of these EMT-inducing effects of oxalate could be prevented by pretreatment with caffeine. While oxalate increased oxidized proteins and Snail1 levels, it decreased Nrf2 expression. Caffeine could preserve all these molecules to their basal (control) levels. Finally, silencing of Nrf2 expression by small interfering RNA (siRNA) could abolish such protective effects of caffeine on oxalate-induced EMT. Our data indicate that the renoprotective effects of caffeine against oxalate-induced EMT is mediated, at least in part, by its anti-oxidative property through activation of Nrf2 signaling and suppression of Snail1 transcription factor. [Display omitted] • Oxalate decreases cell proliferation but increases cell elongation and spindle index. • Oxalate reduces epithelial phenotype but enhances mesenchymal phenotype of cells. • Oxalate enhances cell migrating activity. • Oxalate decreases Nrf2 expression but increases Snail1 expression. • All of the oxalate-induced EMT features can be prevented by caffeine pretreatment. [ABSTRACT FROM AUTHOR]

The interaction of reactive oxygen species (ROS) with lipids, proteins, nucleic acids and hydrocarbonates promotes acute and chronic tissue damage, mediates immunomodulation and triggers autoimmunity in systemic lupus erythematous (SLE) patients. The aim of the study was to determine the pathophysiological mechanisms of the oxidative stress-related damage and molecular mechanisms to counteract oxidative stimuli in lupus nephritis. Our study included 38 SLE patients with lupus nephritis (LN group), 44 SLE patients without renal impairment (non-LN group) and 40 healthy volunteers as control group. In the present paper, we evaluated serum lipid peroxidation, DNA oxidation, oxidized proteins, carbohydrate oxidation, and endogenous protective systems. We detected defective DNA repair mechanisms via 8-oxoguanine-DNA-glycosylase (OGG1), the reduced regulatory effect of soluble receptor for advanced glycation end products (sRAGE) in the activation of AGE-RAGE axis, low levels of thiols, disulphide bonds formation and high nitrotyrosination in lupus nephritis. All these data help us to identify more molecular mechanisms to counteract oxidative stress in LN that could permit a more precise assessment of disease prognosis, as well as developing new therapeutic targets. [ABSTRACT FROM AUTHOR]

Mounting clinical and experimental evidence indicates that free radicals play important roles in many physiological and pathological conditions. The wider application of free radical measurement has increased awareness of functional implications of radical-induced impairment of the oxidative/antioxidative balance. In the following review, the role of oxygen free radicals in some human and experimental pathological conditions is described, with particular emphasis on the mechanisms by which they produce oxidative damage to lipids, proteins, and nucleic bases. The role of free radicals and the activation of the antioxidant systems in arteriosclerosis and ageing, diabetes, ischemia/reperfusion injury, ethanol intoxication, and liver steatosis is discussed. Therapeutic approaches to the use of antioxidants have been described and prospects for clinical use have been considered. [ABSTRACT FROM AUTHOR]

While the posttranslational N-terminal arginylation of proteins has been demonstrated in a variety of eukaryotic cells including neurons and their axons, the targets of the reaction are poorly understood. Several lines of evidence suggest that arginylation may be a cytoprotective mechanism used by cells to target oxidatively damaged (and thus potentially toxic) proteins for degradation. In the present experiments, we have begun to test this hypothesis by incubating oxidized test proteins in a rat brain extract capable of arginylating endogenous proteins. Bovine serum albumin, pancreatic ribonuclease-A and the A-chain of insulin were chosen as test proteins and either oxidized by metal catalyzed oxidation or purchased in their oxidized forms and incubated with the extract and [3H]Arg. SDS PAGE of the incubation product showed [3H]Arg migrating with the oxidized forms of BSA and RNase but not with the un-oxidized form of BSA. Following incubation with the oxidized A-chain of insulin, analysis of the [3H]product by SDS PAGE and HPLC showed co-migration of [3H]Arg with A-chain standards and amino acid sequencing showed [3H]Arg at the N-terminus of the A-chain of insulin. The data suggest that oxidative damage to a protein may be a signal for its N-terminal arginylation. [ABSTRACT FROM AUTHOR]

Spinal muscular atrophy (SMA) type 1 is a severe infantile autosomal-recessive neuromuscular disorder caused by a survival motor neuron 1 gene (SMN1) mutation and characterized by progressive muscle weakness. Without supportive care, SMA type 1 is rapidly fatal. The antisense oligonucleotide nusinersen has recently improved the natural course of this disease. Here, we investigated, with a functional proteomic approach, cerebrospinal fluid (CSF) protein profiles from SMA type 1 patients who underwent nusinersen administration to clarify the biochemical response to the treatment and to monitor disease progression based on therapy. Six months after starting treatment (12 mg/5 mL × four doses of loading regimen administered at days 0, 14, 28, and 63), we observed a generalized reversion trend of the CSF protein pattern from our patient cohort to that of control donors. Notably, a marked up-regulation of apolipoprotein A1 and apolipoprotein E and a consistent variation in transthyretin proteoform occurrence were detected. Since these multifunctional proteins are critically active in biomolecular processes aberrant in SMA, i.e., synaptogenesis and neurite growth, neuronal survival and plasticity, inflammation, and oxidative stress control, their nusinersen induced modulation may support SMN improved-expression effects. Hence, these lipoproteins and transthyretin could represent valuable biomarkers to assess patient responsiveness and disease progression. [ABSTRACT FROM AUTHOR]

Palmitic acid (PA) leads to lipotoxicity in type 2 diabetes and induces oxidative stress in podocytes. Oxidized cellular proteins are degraded by proteasomes. The role of proteasomes in PA- or oxidative stress-induced podocyte injury and pathogenesis of diabetic nephropathy (DN) is unknown. We investigated the effects of PA on expression of 20S and 26S proteasomes, proteasome activator 28 (PA28) regulators, and the immunoproteasome in cultured podocytes and renal cortical tissues of db/db and db/m mice using Western blot analysis. Glomerular areas and glomerular basement membrane (GBM) widths of db/db and db/m mice were examined using morphometry. Short-term incubation of PA or low levels of H 2 O 2 upregulated only the immunoproteasome in cultured podocytes. Long-term exposure of podocytes to PA ultimately downregulated the immunoproteasome as with other proteasomes, whereas oleic acid (OA) or eicosapentaenoic acid (EPA) restored the PA-induced decreased protein levels. In db/db mice, renal cortical immunoproteasome expression with PA28α was significantly decreased compared with db/m mice, and glomerular areas and GBM widths were significantly increased compared with db/m mice. Feeding of an OA-rich olive oil or EPA-rich fish oil protected db/db mice against the reduced renal cortical immunoproteasome expression, glomerular enlargement, and GBM thickening. These results demonstrate that lipotoxicity downregulates the immunoproteasome in podocytes and kidneys in type 2 diabetes and that OA and EPA protected type 2 diabetic mice against decreased renal cortical immunoproteasome expression and the progression of DN. Given this, lipotoxicity-induced podocyte injury with impaired immunoproteasome expression appears to play an important role in the pathogenesis of DN. NEW & NOTEWORTHY In podocytes, PA rapidly induced immunoproteasome expression but ultimately decreased it, while OA and EPA restored the decreased immunoproteasome levels. In the renal cortex of type 2 diabetic mice, immunoproteasome expression was significantly decreased, whereas feeding of OA-rich olive oil or EPA-rich fish oil diets protected them against the reduced immunoproteasome expression and progression of diabetic nephropathy. Thus, lipotoxicity-induced podocyte injury with impaired immunoproteasome expression may be related to the pathogenesis of diabetic nephropathy.