Oxidized Glutathione Ratio Research Articles

Hepatoprotective Effect of Nicotinamide Versus Lead-Motivated Hepatotoxicity in Rats via Correcting Effect on Nuclear Factor-kβ Pathway and Glutathione Metabolism.

Lead (Pb) poisoning is one of the pivotal environmental issues and prompts liver dysfunction by elevating oxidative stress and inflammation. Nicotinamide (NA) deficiency enhances sensitivity to Pb toxicity. So, we investigated the effect of nicotinamide (NA) on the rat's liver histopathological and biochemical profiles in a rat model of Pb toxicity. Thirty-six rats were divided into four groups (nine rats at each): normal (N), lead toxicity (Pbt), and NA-treated N and Pbt groups. Treated groups took NA (180 mg/L in drinking water for one month). Pb intoxication was motivated in rats by acquiring 50 mg/L lead acetate in drinking water. Oxidative stress markers (advanced oxidation protein products and malondialdehyde), antioxidant markers (total glutathione, reduced glutathione to oxidized glutathione ratio, ferric ion reducing power, catalase, and paraoxonase-1), and inflammatory markers (hepatic nuclear factor-kβ expression, interleukin 1β level, and myeloperoxidase activity) in sera and liver homogenates were determined. In addition, the biochemical parameters of the liver function were measured. Finally, the liver of rats was evaluated by histopathological observation. NA corrected lead-persuaded biochemical and histopathological changes in the rat's liver. In addition, treatment decreased Pb, oxidative stress, and inflammatory markers in the sera and liver homogenates of N and Pbt groups. In addition, it elevated antioxidant markers (p < 0.001). NA prevented Pb-induced liver histopathological alternations and reduced liver dysfunction by reducing Pb, oxidative stress, and inflammation. Moreover, raising GSH/GSSG and diminishing the hepatic NF-kβ pathway are cardinal mechanisms of the treatment against Pb-motivated hepatotoxicity in rats.

Ndufs4 KO mice: A model to study comorbid mood disorders associated with mitochondrial dysfunction

The Ndufs4 knockout (KO) mouse is a validated and robust preclinical model of mitochondrial diseases (specifically Leigh syndrome), that displays a narrow window of relative phenotypical normality, despite its inherent mitochondrial complex I dysfunction and severe phenotype. Preclinical observations related to psychiatric comorbidities that arise in patients with mitochondrial diseases and indeed in Leigh syndrome are, however, yet to be investigated in this model. Strengthening this narrative is the fact that major depression and bipolar disorder are known to present with deficits in mitochondrial function. We therefore screened the behavioural profile of male and female Ndufs4 KO mice (relative to heterozygous; HET and wildtype; WT mice) between postnatal days 28 and 35 for locomotor, depressive- and anxiety-like alterations and linked it with selected brain biomarkers, viz. serotonin, kynurenine, and redox status in brain areas relevant to psychiatric pathologies (i.e., prefrontal cortex, hippocampus, and striatum). The Ndufs4 KO mice initially displayed depressive-like behaviour in the tail suspension test on PND31 but not on PND35 in the forced swim test. In the mirror box test, increased risk resilience was observed. Serotonin levels of KO mice, compared to HET controls, were increased on PND36, together with increased tryptophan to serotonin and kynurenine turnover. Kynurenine to kynurenic acid turnover was however decreased, while reduced versus oxidized glutathione ratio (GSH/GSSG) was increased. When considering the comorbid psychiatric traits of patients with mitochondrial disorders, this work elaborates on the neuropsychiatric profile of the Ndufs KO mouse. Secondly, despite locomotor differences, Ndufs4 KO mice present with a behavioural profile not unlike rodent models of bipolar disorder, namely variable mood states and risk-taking behaviour. The model may elucidate the bio-energetic mechanisms underlying mood disorders, especially in the presence of mitochondrial disease. Studies are however required to further validate the model's translational relevance.

Piezoeletric cold atmospheric plasma induces apoptosis and autophagy in human hepatocellular carcinoma cells through blocking glycolysis and AKT/mTOR/HIF-1α pathway

Hepatocellular carcinoma (HCC) is the sixth most prevalent cancer and the fourth leading cause of cancer-related death worldwide. Advanced or metastatic HCC is currently managed using systemic drug therapy with unsatisfactory patient survival. Cold atmospheric plasma has emerged as a promising, physicochemical, and broad-spectrum oncotherapy. In this preclinical study, we investigated the anti-neoplastic functions and mechanism of piezoelectric direct discharge technology-based CAP, Piezo-CAP, on HCC in vitro and in vivo. Various HCC cells lines, such as SMMC7721, HepG2 and LM3, were used as in vitro cancer model for the phenotypic and mechanistic studies. Specifically, the cell counting Kit-8 and colony formation assay, flow cytometry, Transwell assay, Western blot, reactive oxygen species (ROS) assay, and glutathione to oxidized glutathione ratio (GSH/GSSG) assay were used to demonstrate plasma-induced changes in HCC cell proliferation, cell cycle progression, migration and invasion, epithelial-to-mesenchymal transition, intracellular ROS, and antioxidant capacity, respectively. In addition, the Acridine orange and ethidium bromide (AO/EB) staining and transmission electron microscopy were performed for cellular and subcellular assessment of HCC cell apoptosis. The Ad-mCherry-RFP-LC3B fluorescent double-labeled lentiviral system was used to detect autophagic flux. On the other hand, RNA-sequencing, quantitative real-time PCR, and Western blot were used to demonstrate plasma-induced metabolic and molecular disruption of tumor glycolysis and oncogenic proliferation, respectively. In vivo experiments using a human cell-line-derived xenograft model and immunohistochemistry (IHC) were utilized to investigate the mechanism. Piezo-CAP exerted anti-neoplastic functions through inhibiting cell proliferation, migration and invasion, and promote cell apoptosis and autophagy. Treatment of Piezo-CAP could suppress proliferation and induce autophagy of HCC cells through simultaneously disrupts cancer survival pathways of redox deregulation, glycolytic pathway, and PI3K/AKT/mTOR/HIF1α pathway signaling. Moreover, upon translation of these in vitro results into the tissue level, Piezo-CAP significantly suppressed in situ tumor growth. These findings collectively suggest that Piezo-CAP-induced apoptosis and autophagy of HCC cells though a multitargeted blockade of major cancer survival pathways of deregulated redox balance, glycolysis, and PI3K/AKT/mTOR/HIF-1α signaling.

RRx-001: a chimeric triple action NLRP3 inhibitor, Nrf2 inducer, and nitric oxide superagonist.

RRx-001 is a shape shifting small molecule with Fast Track designation for the prevention/amelioration of chemoradiation-induced severe oral mucositis (SOM) in newly diagnosed Head and Neck cancer. It has been intentionally developed or "engineered" as a chimeric single molecular entity that targets multiple redox-based mechanisms. Like an antibody drug conjugate (ADC), RRx-001 contains, at one end a "targeting" moiety, which binds to the NLRP3 inflammasome and inhibits it as well as Kelch-like ECH-associated protein 1 (KEAP1), the negative regulator of Nrf2, and, at the other end, a conformationally constrained, dinitro containing 4 membered ring, which fragments under conditions of hypoxia and reduction to release therapeutically active metabolites i.e., the payload. This "payload", which is delivered specifically to hypoperfused and inflamed areas, includes nitric oxide, nitric oxide related species and carbon-centered radicals. As observed with ADCs, RRx-001 contains a backbone amide "linker" attached to a binding site, which correlates with the Fab region of an antibody, and to the dinitroazetidine payload, which is microenvironmentally activated. However, unlike ADCs, whose large size impacts their pharmacokinetic properties, RRx-001 is a nonpolar small molecule that easily crosses cell membranes and the blood brain barrier (BBB) and distributes systemically. This short review is organized around the de novo design and in vivo pro-oxidant/pro-inflammatory and antioxidant/anti-inflammatory activity of RRx-001, which, in turn, depends on the reduced to oxidized glutathione ratio and the oxygenation status of tissues.

Altered acylcarnitine metabolism and inflexible mitochondrial fuel utilization characterize the loss of neonatal myocardial regeneration capacity

Myocardial regeneration capacity declines during the first week after birth, and this decline is linked to adaptation to oxidative metabolism. Utilizing this regenerative window, we characterized the metabolic changes in myocardial injury in 1-day-old regeneration-competent and 7-day-old regeneration-compromised mice. The mice were either sham-operated or received left anterior descending coronary artery ligation to induce myocardial infarction (MI) and acute ischemic heart failure. Myocardial samples were collected 21 days after operations for metabolomic, transcriptomic and proteomic analyses. Phenotypic characterizations were carried out using echocardiography, histology and mitochondrial structural and functional assessments. In both groups, MI induced an early decline in cardiac function that persisted in the regeneration-compromised mice over time. By integrating the findings from metabolomic, transcriptomic and proteomic examinations, we linked regeneration failure to the accumulation of long-chain acylcarnitines and insufficient metabolic capacity for fatty acid beta-oxidation. Decreased expression of the redox-sensitive mitochondrial Slc25a20 carnitine-acylcarnitine translocase together with a decreased reduced:oxidized glutathione ratio in the myocardium in the regeneration-compromised mice pointed to a defect in the redox-sensitive acylcarnitine transport to the mitochondrial matrix. Rather than a forced shift from the preferred adult myocardial oxidative fuel source, our results suggest the facilitation of mitochondrial fatty acid transport and improvement of the beta-oxidation pathway as a means to overcome the metabolic barrier for repair and regeneration in adult mammals after MI and heart failure.

How does phosphorus influence Cd tolerance strategy in arbuscular mycorrhizal - Phragmites australis symbiotic system?

To clarify how phosphorus (P) influences arbuscular mycorrhizal fungi (AMF) interactions with host plants, we measured the effects of variation in environmental P levels and AMF colonization on photosynthesis, element absorption, ultrastructure, antioxidant capacity, and transcription mechanisms in Phragmites australis (P. australis) under cadmium (Cd) stress. AMF maintained photosynthetic stability, element balance, subcellular integrity and enhanced antioxidant capacity by upregulating antioxidant gene expression. Specifically, AMF overcame Cd-induced stomatal limitation, and mycorrhizal dependence peaked in the high Cd-moderate P treatment (156.08%). Antioxidants and compatible solutes responded to P-level changes: the primary driving forces of removing reactive oxygen species (ROS) and maintaining osmotic balance were superoxide dismutase, catalase, and sugars at limited P levels and total polyphenol, flavonoid, peroxidase, and proline at abundant P levels, we refer to this phenomenon as “functional link.” AMF and phosphorus enhanced Cd tolerance in P. australis, but the regulation of AMF was P-dependent. Phosphorus prevented increases in total glutathione content and AMF-induced GSH/GSSG ratio (reduced to oxidized glutathione ratio) by inhibiting the expression of assimilatory sulfate reduction and glutathione reductase genes. The AMF-induced flavonoid synthesis pathway was regulated by P, and AMF activated Cd-tolerance mechanisms by inducing P-dependent signaling.

Oxidative stress responses in Puntius sarana collected from some environmentally contaminated areas of River Mahananda, Malda, West Bengal.

Environmental contaminant-induced aquatic pollution threatens the aquatic biota and causes oxidative stress in the aquatic organisms. Mahananda River, the lifeline of Malda district, West Bengal, India is presently experiencing environmental crisis due to increasing level of contaminants which deteriorate the water quality along with its biota. Puntius sarana is an economically important edible fish species in the River Mahananda that seems to be declining, owing to environmental pollution. Therefore, the present study aimed to investigate the health status of Puntius sarana by evaluating different oxidative stress biomarkers. The sampling sites were selected according to population density and possible anthropogenic activities around the River Mahananda. Significantly higher concentration of Zn2+ (p < 0.01), Cu2+ (p < 0.05) in different sampling sites has been observed, compared to the control site. Results of the water quality index and water pollution index confirmed the pollution status of different sampling sites. Significant decreases were observed in superoxide dismutase and catalase activity, compared to the control. The decreased values of reduced and oxidized glutathione ratio also indicate the increased oxidative stress of Puntius sarana. The principal component analysis showed that total hardness, calcium hardness, nitrate, manganese, zinc, copper, iron and arsenic were the main descriptors of the aquatic pollution. Therefore, findings of the present study demonstrate the ecotoxicological impacts of various anthropogenic activities which necessitates the need for continuous monitoring of the fresh water resources and its ecosystem from further contamination.

Quantitation of Glutathione and Oxidized Glutathione Ratios from Biological Matrices Using LC-MS/MS.

Oxidation of glutathione (GSH) to its disulfide dimer (GSSG) is the major mechanism by which cells balance reactive oxygen species (ROS) and mitigate oxidative stress. Thus, measuring the ratio of GSH/GSSG is an ideal way to assess oxidative stress within a cell. Quantitative mass spectrometry offers an ideal method to measure the GSH/GSSG ratio and can be applied to a variety of biological matrices and disease models. The following chapter details the design, optimization, and execution of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay to measure the GSH/GSSG ratio.

N-Acetylcysteine effects on glutathione and glutamate in schizophrenia: A preliminary MRS study

N-acetylcysteine (NAC) is a commonly used antioxidant that may have beneficial effects for schizophrenia. In this double-blind, randomized, placebo-controlled preliminary study, 40 patients with schizophrenia or schizoaffective disorder were randomized to receive 2400 mg NAC daily or placebo over eight weeks to examine the effects of NAC on prefrontal magnetic resonance spectroscopy levels of glutathione and glutamate. Secondary outcomes included negative symptoms, cognition, and plasma glutathione levels. We found that NAC treatment was associated with increased glutathione (statistically significant) and decreased glutamate (trend-level) compared with placebo in medial prefrontal cortex but not dorsolateral prefrontal cortex. We also observed a baseline association between medial prefrontal cortex levels of glutathione and plasma reduced / oxidized glutathione ratios. No treatment effects on symptoms or cognition were observed. Taken together, these findings indicate that treatment with N-acetylcysteine may increase medial prefrontal cortical levels of glutathione after eight weeks of treatment. These changes in cortical levels of glutathione may serve as an early biomarker of later clinical change and may underlie the cognitive and symptomatic improvements reported in longer-term treatment studies.

Changes in the concentration, distribution, and speciation of arsenic in the hyperaccumulator Pteris vittata at different growth stages

The arsenic (As) hyperaccumulator has become a model plant for the study of the interaction between plants and trace elements. However, the change in As concentration, distribution and speciation of hyperaccumulator Pteris vittata at different growth stages, especially with the aging process remains unknown. We collected P. vittata at different growth ages and analyzed As concentration, distribution, and speciation. Furthermore, metabolic profiling was conducted for P. vittata at different growth stages. With aging, the reduced glutathione/ oxidized glutathione ratio decreased while the malondialdehyde content increased, accompanied by the change in the main As speciation in P. vittata from arsenite to arsenate. Metabolic profiling also indicated significant difference in the compositions of metabolites during different growth stages. Specifically, flavonoid compounds were found to be positively correlated with As concentration. Results indicated that with the aging of P. vittata, the redox potential increased in the pinnae, leading to the oxidation of As, which may have impacted the distribution of As in this fern. Furthermore, the correlation between As concentration and flavonoid compounds implied the essential role of flavonoid metabolism in the accumulation and transport of As in this plant.

The ataxia-telangiectasia mutated gene product regulates the cellular acid-labile sulfide fraction

The ataxia-telangiectasia mutated (ATM) protein regulates cell cycle checkpoints, the cellular redox state, and double-stranded DNA break repair. ATM loss causes the disorder ataxia-telangiectasia (A-T), distinguished by ataxia, telangiectasias, dysregulated cellular redox and iron responses, and an increased cancer risk. We examined the sulfur pool in A-T cells, with and without an ATM expression vector. While free and bound sulfide levels were not changed with ATM expression, the acid-labile sulfide faction was significantly increased. ATM expression also increased cysteine desulfurase (NFS1), NFU1 iron-sulfur cluster scaffold homolog protein, and several mitochondrial complex I proteins’ expression. Additionally, ATM expression suppressed cystathionine β-synthase and cystathionine γ-synthase protein expression, cystathionine γ-synthase enzymatic activity, and increased the reduced to oxidized glutathione ratio. This last observation is interesting, as dysregulated glutathione is implicated in A-T pathology. As ATM expression increases the expression of proteins central in initiating 2Fe-2S and 4Fe-4S cluster formation (NFS1 and NFU1, respectively), and the acid-labile sulfide faction is composed of sulfur incorporated into Fe-S clusters, our data indicates that ATM regulates aspects of Fe-S cluster biosynthesis, the transsulfuration pathway, and glutathione redox cycling. Thus, our data may explain some of the redox- and iron-related pathologies seen in A-T.

Effects of sulfur amino acid supplementation on broiler chickens exposed to acute and chronic cyclic heat stress

Chronic heat stress can result in oxidative damage from increased reactive oxygen species. One proposed method to alleviate the chronic effects of HS is the supplementation of sulfur amino acids (SAA) which can be metabolized to glutathione, an important antioxidant. Therefore, the objective of this experiment was to determine the effects of dietary SAA content on broiler chickens exposed to HS from 28 to 35 d on broiler performance, body temperature, intestinal permeability, and oxidative status. Four experimental treatments were arranged as a 2 × 2 factorial consisting of HS (6 h at 33.3°C followed by 18 h at 27.8°C from 28 to 35 d of age) and Thermoneutral (TN- 22.2°C continuously from 28 to 35 d) and 2 dietary concentrations of SAA formulated at 100% (0.95, 0.87, and 0.80% for starter, grower, and finisher diets) or 130% SAA (1.24, 1.13, and 1.04% for starter, grower, and finisher diets). A total of 648-day-old, male Ross 708 chicks were placed in 36 pens with 18 chicks/pen and 9 replicates per treatment. Data were analyzed as a 2 × 2 factorial in JMP 14 (P ≤ 0.05). No interaction effects were observed on broiler live performance (P > 0.05). As expected, HS reduced BWG by 92 g and increased FCR by 11 points from 28 to 35 d of age compared to TN, respectively (P ≤ 0.05). The supplementation of SAA had no effect on live performance (P > 0.05). Cloacal temperatures were increased by 1.7, 1.4, and 1.2°C with HS at 28, 31, and 35 d compared to TN, respectively (P ≤ 0.05) and dietary SAA did not alter cloacal temperatures. At 28 d of age, supplementation of SAA to birds exposed to HS interacted as serum FITC-dextran (an indicator of intestinal permeability) was reduced to that of the TN group (P ≤ 0.05). The interaction was lost at 31 d, but HS still increased intestinal permeability (P ≤ 0.05). By 35 d, broilers were able to adapt to the HS conditions and intestinal permeability was unaffected (P > 0.05). Potential oxidative damage was reduced by increased SAA supplementation as indicated by an improvement in the reduced glutathione to oxidized glutathione ratio of 5 and 45 % at 28 (P = 0.08) and 35 d (P ≤ 0.05). These data suggest that intestinal permeability is compromised initially and to at least three d of heat exposure before the bird can adjust. However, oxidative damage in the liver of broilers exposed to HS is more chronic, building over the entire 7 d HS period and increased dietary SAA might have some protective effects on both broiler intestinal permeability and oxidative stress responses to HS.

Polyamines protect boar sperm from oxidative stress in vitro.

Sperm are susceptible to excessive reactive oxygen species (ROS). Spermine and spermidine are secreted in large amounts by the prostate and potent natural free radical scavengers and protect cells against redox disorder. Thus, we used boar sperm as a model to study the polyamines uptake and elucidate whether polyamines protected sperm from ROS stress. Seven mature and fertile Duroc boars (aged 15 to 30 mo) were used in this study. In experiment 1, spermine and spermidine (3.6±0.3 and 3.3±0.2 mmol/L, respectively) were abundant in seminal plasma, and the content of polyamine decreased (P < 0.05) after preservation at 17 °C for 7 d or incubation at 37 °C for 6h. In experiment 2, using labeling of spermine or spermidine by conjugation with fluorescein isothiocyanate and ultra-high-performance liquid chromatography, we found that the accumulation of spermine or spermidine in sperm was inhibited by quinidine and dl-tetrahydropalmatine (THP, organic cation transporters [OCT] inhibitors, P < 0.05), but not mildronate and l-carnitine (organic cation/carnitine transporter [OCTN] inhibitors, P > 0.05). In experiment 3, the addition of spermine or spermidine (0.5 mmol/L) in the extender resulted in higher motility, plasma membrane and acrosome integrity, and lower ROS level after preservation in vitro at 17 °C for 7 d (P < 0.05). In experiment 4, in the condition of oxidative stress (treatment with H2O2 at 37 °C for 2h), the addition of spermine (1 mmol/L) or spermidine (0.5 mmol/L) in extender increased activities of glutathione peroxidase, glutathione reductase, and glutathione S-transferase; reduced glutathione and oxidized glutathione ratio (P < 0.05); and alleviate oxidative stress-induced lipid peroxidation, DNA damage, mitochondrial membrane potential (ΔΨm) decline, adenosine triphosphate depletion, and intracellular calcium concentration ([Ca2+]i) overload (P < 0.05), thereby improving boar sperm motility, the integrity of plasma membrane and acrosome (P < 0.05) in vitro. These data suggest that spermine and spermidine alleviate oxidative stress via the antioxidant capacity, thereby improving the efficacy of boar semen preservation.

Dimethyl Fumarate Ameliorates Nucleus Pulposus Cell Dysfunction through Activating the Nrf2/HO-1 Pathway in Intervertebral Disc Degeneration.

Background Oxidative stress, inflammation, and nucleus pulposus cells (NPCs) apoptosis are involved in pathogenesis of intervertebral disc (IVD) degeneration (IVDD). Dimethyl fumarate (DMF) has been found to effectively depress oxidative stress and inflammation via the Nrf2 pathway. Hence, this project was designed to explore the underlying mechanisms of how DMF protects NPCs from damage by LPS challenge. Methods and Results CCK8 assay and flow cytometry of apoptosis indicated that DMF treatment attenuated LPS-induced NPC damage. Western blot analysis demonstrated that DMF enhanced the expressions of nuclear factor-erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) in LPS-challenged NPCs. DMF treatment significantly decreased the accumulation of ROS, downregulated inflammatory cytokines (p-NF-κB, IL-1β, and TNF-α), and ER stress-associated apoptosis proteins (Bip, calpain-1, caspase-12, caspase-3, and Bax) in LPS-challenged NPCs. The level of antiapoptotic protein Bcl-2 was promoted by DMF treatment in LPS-challenged NPCs. Glutathione (GSH) assay showed that DMF treatment improved reduced to oxidized glutathione ratio in LPS-challenged NPCs. Furthermore, the results of western blot analysis indicated that in LPS-challenged NPCs, DMF treatment ameliorated the elevated levels of matrix degradation enzymes (MMP-13, aggrecanase 1) and type I collagen and the reduced levels of matrix composition (type II collagen and ACAN). However, Nrf2 knockdown abolished these protective effects of DMF. Conclusion Our data suggested that treatment with DMF mitigated LPS-induced oxidative stress, inflammation, and ER stress-associated apoptosis in NPCs via the Nrf2/HO-1 signaling pathway, thus reliving LPS-induced dysfunction of NPCs, which offered a novel potential pharmacological treatment strategy for IVDD.

Bone mineral density and oxidative stress in adolescent girls with anorexia nervosa

Oxidative stress appears to be involved in the pathogenesis of osteoporosis-a serious complication of anorexia nervosa (AN). We evaluated the oxidative status in adolescent girls with AN and its potential relationship with bone mineral density (BMD). Girls with AN (n = 43) and age-matched healthy controls (n = 20) underwent anthropometric and BMD examination. Markers of bone turnover, oxidative stress, and antioxidant status were measured. Participants with AN and controls did not differ in BMD at the lumbar spine (p = 0.17) and total body less head BMD (p = 0.08). BMD at the total hip was lower (p < 0.001) in the AN group compared with the controls. Levels of antioxidant status markers-ferric reduction antioxidant power, total antioxidant capacity, and reduced and oxidized glutathione ratio (all p < 0.001)-were significantly lower, whereas those of advanced oxidation protein products (AOPP), fructosamines, and advanced glycation end products (AGEs) (all p < 0.001) were higher in AN patients than in healthy controls. BMD and bone turnover markers were positively correlated with antioxidant status markers, while they were negatively correlated with AOPP, fructosamines, and AGEs levels. Conclusion: This is the first study to assess a potential association between oxidative status and BMD in adolescents with AN. We demonstrated that in young girls, the imbalance of oxidative status and reduced BMD are concurrently manifested at the time of the diagnosis of AN. Disturbance of oxidative status could play a pathogenetic role in AN-associated decreased BMD. What is Known: • Osteoporosis is a serious complication of AN, and in affected adolescents may result in a permanent deficit in bone mass. • Oxidative and carbonyl stress may be involved in the development of bone loss. What is New: • Adolescents girls with AN have impaired antioxidant defense and increased oxidative damage to biomolecules. • Disturbance of oxidative status could affect bone loss and could contribute to decreased BMD in adolescent females with AN.

Nanoceria Prevents Glucose-Induced Protein Glycation in Eye Lens Cells.

Cerium oxide nanoparticles (nanoceria) are generally known for their recyclable antioxidative properties making them an appealing biomaterial for protecting against physiological and pathological age-related changes that are caused by reactive oxygen species (ROS). Cataract is one such pathology that has been associated with oxidation and glycation of the lens proteins (crystallins) leading to aggregation and opacification. A novel coated nanoceria formulation has been previously shown to enter the human lens epithelial cells (HLECs) and protect them from oxidative stress induced by hydrogen peroxide (H2O2). In this work, the mechanism of nanoceria uptake in HLECs is studied and multiple anti-cataractogenic properties are assessed in vitro. Our results show that the nanoceria provide multiple beneficial actions to delay cataract progression by (1) acting as a catalase mimetic in cells with inhibited catalase, (2) improving reduced to oxidised glutathione ratio (GSH/GSSG) in HLECs, and (3) inhibiting the non-enzymatic glucose-induced glycation of the chaperone lens protein α-crystallin. Given the multifactorial nature of cataract progression, the varied actions of nanoceria render them promising candidates for potential non-surgical therapeutic treatment.

Mutant Nmnat1 leads to a retina-specific decrease of NAD+ accompanied by increased poly(ADP-ribose) in a mouse model of NMNAT1-associated retinal degeneration.

Nicotinamide mononucleotide adenylyltransferase 1 (NMNAT1) is required for nuclear nicotinamide adenine mononucleotide (NAD+) biosynthesis in all nucleated cells, and despite its functional ubiquity, mutations in this gene lead to an isolated retinal degeneration. The mechanisms underlying how mutant NMNAT1 causes disease are not well understood, nor is the reason why the pathology is confined to the retina. Using a mouse model of NMNAT1-associated retinal degeneration that harbors the p.Val9Met mutation, we tested the hypothesis that decreased function of mutant NMNAT1 has a greater effect on the levels of NAD+ in the retina than elsewhere in the body. Measurements by liquid chromatography with tandem mass spectrometry showed an early and sustained decrease of NAD+ in mutant retinas that was not observed in other tissues. To understand how consumers of nuclear NAD+ are affected by the reduced availability of NAD+ in mutant retinas, poly(ADP-ribose) polymerase (PARP) and nuclear sirtuin activity were evaluated. PARP activity was elevated during disease progression, as evidenced by overproduction of poly(ADP-ribose) (PAR) in photoreceptors, whereas histone deacetylation activity of nuclear sirtuins was not altered. We hypothesized that PARP could be activated because of elevated levels of oxidative stress; however, we did not observe oxidative DNA damage, lipid peroxidation, or a low glutathione to oxidized glutathione ratio. Terminal deoxynucleotidyl transferase dUTP nick end labeling staining revealed that photoreceptors appear to ultimately die by apoptosis, although the low NAD+ levels and overproduction of PAR suggest that cell death may include aspects of the parthanatos cell death pathway.

Can ocean warming alter sub-lethal effects of antiepileptic and antihistaminic pharmaceuticals in marine bivalves?

The negative effects induced in marine organisms by Climate Change related abiotic factors consequences, namely ocean warming, are well-known. However, few works studied the combined impacts of ocean warming and contaminants, as pharmaceutical drugs. Carbamazepine (CBZ) and cetirizine (CTZ) occur in the marine environment, showing negative effects in marine organisms. This study aimed to evaluate the impacts of ocean warming on the effects of CBZ and CTZ, when acting individually and combined (drug vs drug), in the edible clam Ruditapes philippinarum. For that, drugs concentration, bioconcentration factors and biochemical parameters, related with clam’s metabolic capacity and oxidative stress, were evaluated after 28 days exposure to environmentally relevant scenarios of these stressors. The results showed limited impacts of the drugs (single and combined) at control and warming condition. Indeed, it appeared that warming improved the oxidative status of contaminated clams (higher reduced to oxidized glutathione ratio, lower lipid peroxidation and protein carbonylation levels), especially when both drugs were combined. This may result from clam’s defence mechanisms activation and reduced metabolic capacity that, respectively, increased elimination and limited production of reactive oxygen species. At low stress levels, defence mechanisms were not activated which resulted into oxidative stress. The present findings highlighted that under higher stress levels clams may be able to activate defence strategies that were sufficient to avoid cellular damages and loss of redox homeostasis. Nevertheless, low concentrations were tested in the present study and the observed responses may greatly change under increased pollution levels or temperatures. Further research on this topic is needed since marine heat waves are increasing in frequency and intensity and pollution levels of some pharmaceuticals are also increasing in coastal systems.

Repeated exposure to transient obstructive sleep apnea–related conditions causes an atrial fibrillation substrate in a chronic rat model

High night-to-night variability in obstructive sleep apnea (OSA) is associated with atrial fibrillation (AF). Obstructive apneas are characterized by intermittent deoxygenation-reoxygenation and intrathoracic pressure swings during ineffective inspiration against occluded upper airways. We elucidated the effect of repeated exposure to transient OSA conditions simulated by intermittent negative upper airway pressure (INAP) on the development of an AF substrate. INAP (48 events/4 h; apnea-hypopnea index 12 events/h) was applied in sedated spontaneously breathing rats (2% isoflurane) to simulate mild-to-moderate OSA. Rats without INAP served as a control group (CTR). In an acute test series (ATS), rats were either killed immediately (n = 9 per group) or after 24 hours of recovery (ATS-REC: n = 5 per group). To simulate high night-to-night variability in OSA, INAP applications (n = 10; 24 events/4 h; apnea-hypopnea index 6/h) were repeated every second day for 3 weeks in a chronic test series (CTS). INAP increased atrial oxidative stress acutely, represented in decreases of reduced to oxidized glutathione ratio (ATS: INAP: 0.33 ± 0.05 vs CTR: 1 ± 0.26; P = .016), which was reversible after 24 hours (ATS-REC: INAP vs CTR; P = .274). Although atrial oxidative stress did not accumulate in the CTS, atrial histological analysis revealed increased cardiomyocyte diameters, reduced connexin 43 expression, and increased interstitial fibrosis formation (CTS: INAP 7.0% ± 0.5% vs CTR 5.1% ± 0.3%; P = .013), which were associated with longer inducible AF episodes (CTS: INAP: 11.65 ± 4.43 seconds vs CTR: 0.7 ± 0.33 seconds; P = .033). Acute simulation of OSA was associated with reversible atrial oxidative stress. Cumulative exposure to these transient OSA-related conditions resulted in AF substrates and was associated with increased AF susceptibility. Mild-to-moderate OSA with high night-to-night variability may deserve intensive management to prevent atrial substrate development.

Perfusion reduces bispecific antibody aggregation via mitigating mitochondrial dysfunction-induced glutathione oxidation and ER stress in CHO cells

One major challenge observed for the expression of therapeutic bispecific antibodies (BisAbs) is high product aggregates. Aggregates increase the risk of immune responses in patients and therefore must be removed at the expense of purification yields. BisAbs contain engineered disulfide bonds, which have been demonstrated to form product aggregates, if mispaired. However, the underlying intracellular mechanisms leading to product aggregate formation remain unknown. We demonstrate that impaired glutathione regulation underlies BisAb aggregation formation in a CHO cell process. Aggregate formation was evaluated for the same clonal CHO cell line producing a BisAb using fed-batch and perfusion processes. The perfusion process produced significantly lower BisAb aggregates compared to the fed-batch process. Perfusion bioreactors attenuated mitochondrial dysfunction and ER stress resulting in a favorable intracellular redox environment as indicated by improved reduced to oxidized glutathione ratio. Conversely, mitochondrial dysfunction-induced glutathione oxidation and ER stress disrupted the intracellular redox homeostasis, leading to product aggregation in the fed-batch process. Combined, our results demonstrate that mitochondrial dysfunction and ER stress impaired glutathione regulation leading to higher product aggregates in the fed-batch process. This is the first study to utilize perfusion bioreactors as a tool to demonstrate the intracellular mechanisms underlying product aggregation formation.