Redox Ratio Research Articles

Inhibition of B-cell lymphoma 2 family proteins alters optical redox ratio, mitochondrial polarization, and cell energetics independent of cell state.

.SignificanceThe optical redox ratio (ORR) [autofluorescence intensity of the reduced form of nicotinamide adenine dinucleotide (phosphate) (NAD(P)H)/flavin adenine dinucleotide (FAD)] provides a label-free method to quantify cellular metabolism. However, it is unclear whether changes in the ORR with B-cell lymphoma 2 (Bcl-2) family protein inhibition are due to metabolic stress alone or compromised cell viability.AimDetermine whether ABT-263 (navitoclax, Bcl-2 family inhibitor) changes the ORR due to changes in mitochondrial function that are independent of changes in cell viability.ApproachSW48 colon cancer cells were used to investigate changes in ORR, mitochondrial membrane potential, oxygen consumption rates, and cell state (cell growth, viability, proliferation, apoptosis, autophagy, and senescence) with ABT-263, TAK-228 [sapanisertib, mammalian target of rapamycin complex 1/2 (mTORC 1/2) inhibitor], and their combination at 24 h.ResultsChanges in the ORR with Bcl-2 inhibition are driven by increases in both NAD(P)H and FAD autofluorescence, corresponding with increased basal metabolic rate and increased mitochondrial polarization. ABT-263 treatment does not change cell viability or induce autophagy but does induce a senescent phenotype. The metabolic changes seen with ABT-263 treatment are mitigated by combination with mTORC1/2 inhibition.ConclusionsThe ORR is sensitive to increases in mitochondrial polarization, energetic state, and cell senescence, which can change independently from cell viability.

Different Effects of RNAi-Mediated Downregulation or Chemical Inhibition of NAMPT in an Isogenic IDH Mutant and Wild-Type Glioma Cell Model.

The IDH1R132H mutation in glioma results in the neoenzymatic function of IDH1, leading to the production of the oncometabolite 2-hydroxyglutarate (2-HG), alterations in energy metabolism and changes in the cellular redox household. Although shifts in the redox ratio NADPH/NADP+ were described, the consequences for the NAD+ synthesis pathways and potential therapeutic interventions were largely unexplored. Here, we describe the effects of heterozygous IDH1R132H on the redox system in a CRISPR/Cas edited glioblastoma model and compare them with IDH1 wild-type (IDH1wt) cells. Besides an increase in 2-HG and decrease in NADPH, we observed an increase in NAD+ in IDH1R132H glioblastoma cells. RT-qPCR analysis revealed the upregulation of the expression of the NAD+ synthesis enzyme nicotinamide phosphoribosyltransferase (NAMPT). Knockdown of NAMPT resulted in significantly reduced viability in IDH1R132H glioblastoma cells. Given this dependence of IDH1R132H cells on NAMPT expression, we explored the effects of the NAMPT inhibitors FK866, GMX1778 and GNE-617. Surprisingly, these agents were equally cytotoxic to IDH1R132H and IDH1wt cells. Altogether, our results indicate that targeting the NAD+ synthesis pathway is a promising therapeutic strategy in IDH mutant gliomas; however, the agent should be carefully considered since three small-molecule inhibitors of NAMPT tested in this study were not suitable for this purpose.

Pyridine nucleotide redox potential in coronary smooth muscle couples myocardial blood flow to cardiac metabolism

Adequate oxygen delivery to the heart during stress is essential for sustaining cardiac function. Acute increases in myocardial oxygen demand evoke coronary vasodilation and enhance perfusion via functional upregulation of smooth muscle voltage-gated K+ (Kv) channels. Because this response is controlled by Kv1 accessory subunits (i.e., Kvβ), which are NAD(P)(H)-dependent aldo-keto reductases, we tested the hypothesis that oxygen demand modifies arterial [NAD(H)]i, and that resultant cytosolic pyridine nucleotide redox state influences Kv1 activity. High-resolution imaging mass spectrometry and live-cell imaging reveal cardiac workload-dependent increases in NADH:NAD+ in intramyocardial arterial myocytes. Intracellular NAD(P)(H) redox ratios reflecting elevated oxygen demand potentiate native coronary Kv1 activity in a Kvβ2-dependent manner. Ablation of Kvβ2 catalysis suppresses redox-dependent increases in Kv1 activity, vasodilation, and the relationship between cardiac workload and myocardial blood flow. Collectively, this work suggests that the pyridine nucleotide sensitivity and enzymatic activity of Kvβ2 controls coronary vasoreactivity and myocardial blood flow during metabolic stress.

Insight into redox regulation of apoptosis in cancer cells with multiparametric live-cell microscopy

Cellular redox status and the level of reactive oxygen species (ROS) are important regulators of apoptotic potential, playing a crucial role in the growth of cancer cell and their resistance to apoptosis. However, the relationships between the redox status and ROS production during apoptosis remain poorly explored. In this study, we present an investigation on the correlations between the production of ROS, the redox ratio FAD/NAD(P)H, the proportions of the reduced nicotinamide cofactors NADH and NADPH, and caspase-3 activity in cancer cells at the level of individual cells. Two-photon excitation fluorescence lifetime imaging microscopy (FLIM) was applied to monitor simultaneously apoptosis using the genetically encoded sensor of caspase-3, mKate2-DEVD-iRFP, and the autofluorescence of redox cofactors in colorectal cancer cells upon stimulation of apoptosis with staurosporine, cisplatin or hydrogen peroxide. We found that, irrespective of the apoptotic stimulus used, ROS accumulation correlated well with both the elevated pool of mitochondrial, enzyme-bound NADH and caspase-3 activation. Meanwhile, a shift in the contribution of bound NADH could develop independently of the apoptosis, and this was observed in the case of cisplatin. An increase in the proportion of bound NADPH was detected only in staurosporine-treated cells, this likely being associated with a high level of ROS production and their resulting detoxification. The results of the study favor the discovery of new therapeutic strategies based on manipulation of the cellular redox balance, which could help improve the anti-tumor activity of drugs and overcome apoptotic resistance.

Effects of Orally Administered Nicotinamide Riboside on Bioenergetic Metabolism, Oxidative Stress and Cognition in Mild Cognitive Impairment and Mild Alzheimer's Disease

<h3>Introduction</h3> Alzheimer's Disease (AD) is a neurodegenerative disease affecting millions of people worldwide, representing significant health and economic burdens. There has been a recent drive to develop more effective AD treatments. A potential strategy involves enhancing brain bioenergetics, or the metabolism of fuel molecules to create and use energy through glycolysis and oxidative phosphorylation (OxPhos). The regulation of mitochondrial turnover and function in the brain declines as we age, and it has become increasingly accepted that these metabolic alterations precede hallmark AD pathology, including amyloid-beta (Aβ) plaques and neurofibrillary tangles. An overlapping set of mechanisms involving redox dysregulation, oxidative stress, and mitochondrial dysfunction may be related to mild cognitive impairment (MCI) and AD. Nicotinamide adenine dinucleotide (NAD) has been identified as a promising molecular target for bioenergetic treatments as it is requisite for many redox reactions. The redox ratio (RR, i.e., NAD+/NADH) is a mark of the system's ability to carry out ATP synthesis. NAD+ levels decrease as we age, which compromises energy production in the mitochondria, impairs clearance of damaged mitochondria, and impacts our brain's capacity to react to oxidative stress. Since mitochondrial dysfunction and oxidative stress are believed to play a role in the pathophysiology of age-related diseases, boosting NAD levels could be an effective AD treatment. Nicotinamide Riboside (NR) is a naturally occurring, over the counter, commercially available, and safe vitamin B3-based NAD+ precursor supplement. Several lines of evidence suggest that NR is a mitochondrially-favored NAD+ precursor. Data supports the safety and efficacy of using NR to increase NAD+ levels in humans, but these studies have not measured the RR at critical tissue sites, such as the brain, to determine the therapeutic potential of oral NR supplementation. This study proposes a novel intervention strategy for patients with MCI or mild AD that targets redox dysregulation and oxidative stress in the brain using NR. Our primary objective is to validate the effect of NR supplementation on NAD+ levels and RR in the brain of patients with MCI or mild AD. Our secondary aim is to quantify the impact of NR supplementation on ensuing markers of mitochondrial function and oxidative stress in MCI and mild AD patients. Our third, exploratory aim, is to correlate neuroimaging measurements with clinical assessments. <h3>Methods</h3> A 12-week open-label, clinical trial of NR supported by the National Institute on Aging (R01AG066670, Du/Forester), we plan to enroll a total of 50 participants - 25 with MCI and 25 with mild AD, all with at least one copy of the APOE ε4 allele. Participants will be asked to take 1,000 mg NR daily over 12-weeks, and to come in for three scanning visits. We will use novel neuroimaging methods —spectroscopy on a 4T scanner to enable in vivo measurement of NAD+, NADH and RR, along with markers of mitochondrial function tied to NAD, including creatine kinase (CK), ATPase, and glutathione (GSH)—a molecule essential for cellular repair. Our primary outcome measures are NAD+ levels and RR. Our secondary outcome measures are CK/ATPase activity and GSH levels. Our exploratory outcome measures include the Repeatable Battery for the Assessment of Neuropsychological Status (RBANS) for cognition and the Alzheimer's Disease Cooperative Study-Activities of Daily Living (ADCS-ADL) for daily functioning. <h3>Results</h3> Previous clinical trials of NR pharmacokinetics in humans support the safety and efficacy of using NR to increase NAD+ levels in the plasma. Emerging data also highlights important health benefits this supplement may have. We recently received preliminary spectroscopic data following a course of NR supplementation in healthy volunteers (N=4) which observed elevations in NAD+ and RR after two weeks. Only a small difference of downstream effects, CK/ATPase activity and GSH levels, were observed, which could be due to the short period of NR treatment. We hypothesize that oral NR will increase NAD+ and RR in the medial prefrontal cortex and will increase GSH levels and CK/ATPase activity in the brain. For our exploratory aim, we hypothesize that oral NR will improve cognitive status and functional abilities, as measured by the RBANS and ADCS-ADL. <h3>Conclusions</h3> We are investigating NR to target redox dysregulation and oxidative stress in the brain in those with MCI or mild AD. We aim to document and quantify the neurobiological and clinical effects of NR supplementation in MCI and mild AD using in vivo brain imaging. This novel intervention strategy of brain bioenergetics has thus far received minute empirical attention and may represent an alternative strategy for treating MCI and mild AD. Early detection and repair of neural dysfunction at this critical stage could significantly alter disease course and improve functional outcomes. <h3>This research was funded by</h3> National Institutes of Health

S-nitrosoglutathione alleviates hyperglycemia-induced neurobehavioral deficits involving nitro-oxidative stress and aberrant monaminergic system

The present study evaluated the protective role of S-nitrosoglutathione (GSNO) in preventing hyperglycemia-induced nitro-oxidative stress and alterations in monoaminergic system associated with neurobehavioral deficits in mice. Mice were subjected to diabetes by intraperitoneal injection of streptozotocin (40 mg/kg body weight) for 5 days, whereas GSNO (100 μg/kg body weight) was administered daily via oral route for 8 weeks. Diabetic mice showed deficits in neurobehavioral functions associated with memory, learning, anxiety and motor coordination. These neurobehavioral deficits observed in diabetic mice may be attributed to decrease in norepinephrine (NE), dopamine (DA), serotonin (5-HT) and increased monoamine oxidase (MAO) activity in cortex and hippocampus. Further, a significant increase in reactive oxygen species (ROS), protein carbonyls, nitrotyrosine (NT) and lipid peroxidation were observed in brain regions of diabetic animals suggesting increased nitro-oxidative stress. Hyperglycemia induced nitro-oxidative stress appears to involve reduction in redox ratio (GSH/GSSG) and enzymatic antioxidants; catalase (CAT) and superoxide dismutase (SOD) in cortex and hippocampus. However, GSNO supplementation was able to ameliorate alterations in monoaminergic system and nitro-oxidative stress in the brain regions thereby restoring neurobehavioural functions. These findings suggest GSNO as potential therapeutic molecule to prevent diabetic encephalopathy.

Effect of Normobaric Hypoxia on Alterations in Redox Homeostasis, Nitrosative Stress, Inflammation, and Lysosomal Function following Acute Physical Exercise.

Hypoxia is a recognized inducer of oxidative stress during prolonged physical activity. Nevertheless, previous studies have not systematically examined the effects of normoxia and hypoxia during acute physical exercise. The study is aimed at evaluating the relationship between enzymatic and nonenzymatic antioxidant barrier, total antioxidant/oxidant status, oxidative and nitrosative damage, inflammation, and lysosomal function in different acute exercise protocols under normoxia and hypoxia. Fifteen competitive athletes were recruited for the study. They were subjected to two types of acute cycling exercise with different intensities and durations: graded exercise until exhaustion (GE) and simulated 30 km individual time trial (TT). Both exercise protocols were performed under normoxic and hypoxic (FiO2 = 16.5%) conditions. The number of subjects was determined based on our previous experiment, assuming the test power = 0.8 and α = 0.05. We demonstrated enhanced enzymatic antioxidant systems during hypoxic exercise (GE: ↑ catalase (CAT), ↑ superoxide dismutase; TT: ↑ CAT) with a concomitant decrease in plasma reduced glutathione. In athletes exercising in hypoxia, redox status was shifted in favor of oxidation reactions (GE: ↑ total oxidant status, ↓ redox ratio), leading to increased oxidation/nitration of proteins (GE: ↑ advanced oxidation protein products (AOPP), ↑ ischemia-modified albumin, ↑ 3-nitrotyrosine, ↑ S-nitrosothiols; TT: ↑ AOPP) and lipids (GE: ↑ malondialdehyde). Concentrations of nitric oxide and its metabolites (peroxynitrite) were significantly higher in the plasma of hypoxic exercisers with an associated increase in inflammatory mediators (GE: ↑ myeloperoxidase, ↑ tumor necrosis factor-alpha) and lysosomal exoglycosidase activity (GE: ↑ N-acetyl-β-hexosaminidase, ↑ β-glucuronidase). Our study indicates that even a single intensive exercise session disrupts the antioxidant barrier and leads to increased oxidative and nitrosative damage at the systemic level. High-intensity exercise until exhaustion (GE) alters redox homeostasis more than the less intense exercise (TT, near the anaerobic threshold) of longer duration (20.2 ± 1.9 min vs. 61.1 ± 5.4 min—normoxia; 18.0 ± 1.9 min vs. 63.7 ± 3.0 min—hypoxia), while hypoxia significantly exacerbates oxidative stress, inflammation, and lysosomal dysfunction in athletic subjects.

Multispectral autofluorescence characteristics of reproductive aging in old and young mouse oocytes

Increasing age has a major detrimental impact on female fertility, which, with an ageing population, has major sociological implications. This impact is primarily mediated through deteriorating quality of the oocyte. Deteriorating oocyte quality with biological age is the greatest rate-limiting factor to female fertility. Here we have used label-free, non-invasive multi-spectral imaging to identify unique autofluorescence profiles of oocytes from young and aged animals. Discriminant analysis demonstrated that young oocytes have a distinct autofluorescent profile which accurately distinguishes them from aged oocytes. We recently showed that treatment with the nicotinamide adenine dinucleotide (NAD+) precursor nicotinamide mononucleotide (NMN) restored oocyte quality and fertility in aged animals, and when our analysis was applied to oocytes from aged animals treated with NMN, 85% of these oocytes were classified as having the autofluorescent signature of young animals. Spectral unmixing using the Robust Dependent Component Analysis (RoDECA) algorithm demonstrated that NMN treatment altered the metabolic profile of oocytes, increasing free NAD(P)H, protein bound NAD(P)H, redox ratio and the ratio of bound to free NAD(P)H. The frequency of oocytes with simultaneously high NAD(P)H and flavin content was also significantly increased in mice treated with NMN. Young and Aged + NMN oocytes had a smoother spectral distribution, with the distribution of NAD(P)H in young oocytes specifically differing from that of aged oocytes. Identifying the multispectral profile of oocyte autofluorescence during aging could have utility as a non-invasive and sensitive measure of oocyte quality.

In vivo fluorescence lifetime imaging of macrophage intracellular metabolism during wound responses in zebrafish.

The function of macrophages in vitro is linked to their metabolic rewiring. However, macrophage metabolism remains poorly characterized in situ. Here, we used two-photon intensity and lifetime imaging of autofluorescent metabolic coenzymes, nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) and flavin adenine dinucleotide (FAD), to assess the metabolism of macrophages in the wound microenvironment. Inhibiting glycolysis reduced NAD(P)H mean lifetime and made the intracellular redox state of macrophages more oxidized, as indicated by reduced optical redox ratio. We found that TNFα+ macrophages had lower NAD(P)H mean lifetime and were more oxidized compared to TNFα- macrophages. Both infection and thermal injury induced a macrophage population with a more oxidized redox state in wounded tissues. Kinetic analysis detected temporal changes in the optical redox ratio during tissue repair, revealing a shift toward a more reduced redox state over time. Metformin reduced TNFα+ wound macrophages, made intracellular redox state more reduced and improved tissue repair. By contrast, depletion of STAT6 increased TNFα+ wound macrophages, made redox state more oxidized and impaired regeneration. Our findings suggest that autofluorescence of NAD(P)H and FAD is sensitive to dynamic changes in intracellular metabolism in tissues and can be used to probe the temporal and spatial regulation of macrophage metabolism during tissue damage and repair.

Exercise decreases PP2A-specific reversible thiol oxidation in human erythrocytes: Implications for redox biomarkers

New readily accessible systemic redox biomarkers are needed to understand the biological roles reactive oxygen species (ROS) play in humans because overtly flawed, technically fraught, and unspecific assays severely hamper translational progress. The antibody-linked oxi-state assay (ALISA) makes it possible to develop valid ROS-sensitive target-specific protein thiol redox state biomarkers in a readily accessible microplate format. Here, we used a maximal exercise bout to disrupt redox homeostasis in a physiologically meaningful way to determine whether the catalytic core of the serine/threonine protein phosphatase PP2A is a candidate systemic redox biomarker in human erythrocytes. We reasoned that: constitutive oxidative stress (e.g., haemoglobin autoxidation) would sensitise erythrocytes to disrupted ion homeostasis as manifested by increased oxidation of the ion regulatory phosphatase PP2A. Unexpectedly, an acute bout of maximal exercise lasting ~16 min decreased PP2A-specific reversible thiol oxidation (redox ratio, rest: 0.46; exercise: 0.33) without changing PP2A content (rest: 193 pg/ml; exercise: 191 pg/ml). The need for only 3–4 μl of sample to perform ALISA means PP2A-specific reversible thiol oxidation is a capillary—fingertip blood—compatible candidate redox biomarker. Consistent with biologically meaningful redox regulation, thiol reductant-inducible PP2A activity was significantly greater (+10%) at rest compared to exercise. We establish a route to developing new readily measurable protein thiol redox biomarkers for understanding the biological roles ROS play in humans.

Quantitative analysis of the interaction of ethanol metabolism with gluconeogenesis and fatty acid oxidation in the perfused liver of fasted rats

Ethanol is known to significantly affect gluconeogenesis and lipid metabolism in the liver, primarily by altering the redox ratio in both cytosol and mitochondria. The effect of ethanol was analyzed using a comprehensive, dynamic model of liver metabolism that takes into account sub-cellular compartmentation, detailed kinetics for the citric acid cycle, ethanol and acetaldehyde oxidation, and gluconeogenesis, and inter-compartmental transport of metabolites, including the malate-aspartate shuttle. The kinetic expression for alcohol dehydrogenase takes into account inhibition by ethanol and NADH. Simulations of perfusions of the rat liver were performed with various combinations of substrates (lactate, pyruvate, and fatty acids), with subsequent addition of ethanol to the perfusate. The model successfully predicts NADH/NAD+, in both cytosol and mitochondria, the expected directional flux of reducing equivalents between the two compartments during perfusion with different gluconeogenic precursors, and the effect of ethanol on glucose and ketone body production. This model can serve as a platform for in silico experiments investigating the effects of ethanol on the many dehydrogenases, and thus the major carbohydrate and lipid metabolic pathways in the liver, as well as potential effects of various drugs that may interact with ethanol.

Fluorescence lifetime imaging microscopy (FLIM) detects differences in metabolic signatures between euploid and aneuploid human blastocysts.

Can non-invasive imaging with fluorescence lifetime imaging microscopy (FLIM) detect metabolic differences in euploid versus aneuploid human blastocysts? FLIM has identified significant metabolic differences between euploid and aneuploid blastocysts. Prior studies have demonstrated that FLIM can detect metabolic differences in mouse oocytes and embryos and in discarded human blastocysts. This was a prospective observational study from August 2019 to February 2020. Embryo metabolic state was assessed using FLIM to measure the autofluorescence metabolic factors nicotinamide adenine dinucleotide dehydrogenase together with nicotinamide adenine phosphate dinucleotide dehydrogenase (NAD(P)H) and flavin adenine dinucleotide (FAD). Eight metabolic FLIM parameters were obtained from each blastocyst (four for NAD(P)H and four for FAD): short (T1) and long (T2) fluorescence lifetime, fluorescence intensity (I) and fraction of the molecules engaged with enzymes (F). The redox ratio (NAD(P)H-I)/(FAD-I) was also calculated for each image. This study was performed at a single academically affiliated centre where there were 156 discarded frozen blastocysts (n = 17 euploids; 139 aneuploids) included. Ploidy status was determined by pre-implantation genetic testing for aneuploidy (PGT-A). Discarded human blastocysts were compared using single FLIM parameters. Additionally, inner cell mass (ICM) and trophectoderm (TE) were also evaluated. Multilevel models were used for analysis. A post-hoc correction used Benjamini-Hochberg's false discovery rate, at a q-value of 0.05. Comparing euploid (n = 17) versus aneuploid (n = 139) embryos, a significant difference was seen in NAD(P)H-F (P < 0.04), FAD-I (P < 0.04) and redox ratio (P < 0.05). Euploid ICM (n = 15) versus aneuploid ICM (n = 119) also demonstrated significantly different signatures in NAD(P)H-F (P < 0.009), FAD-I (P < 0.03) and redox ratio (P < 0.03). Similarly, euploid TE (n = 15) versus aneuploid TE (n = 119) had significant differences in NAD(P)H-F (P < 0.0001) and FAD-I (P < 0.04). This study utilized discarded human blastocysts, and these embryos may differ metabolically from non-discarded human embryos. The blastocysts analysed were vitrified after PGT-A biopsy and it is unclear how the vitrification process may affect the metabolic profile of blastocysts. Our study was also limited by the small number of rare donated euploid embryos available for analysis. Euploid embryos are very rarely discarded due to their value to patients trying to conceive, which limits their use for research purposes. However, we controlled for the imbalance with the bootstrap resampling analysis. These findings provide preliminary evidence that FLIM may be a useful non-invasive clinical tool to assist in identifying the ploidy status of embryos. The study was supported by the Blavatnik Biomedical Accelerator Grant at Harvard University. Becker and Hickl GmbH and Boston Electronics sponsored research with the loaning of equipment for FLIM. D.J.N. is an inventor on patent US20170039415A1. There are no other conflicts of interest to declare. N/A.

Development of an optical fiber-based redox monitoring system for tissue metabolism.

An optical redox ratio can potentially be used to report on the dynamics of cell and tissue metabolism and define altered metabolic conditions for different pathologies. While there are methods to measure the optical redox ratio, they are not particularly suited to real-time in situ or in vivo analysis. Here, we have developed a fiber-optic system to measure redox ratios in cells and tissues and two mathematical models to enable real-time, in vivo redox measurements. The optical redox ratios in tissue explants are correlated directly with endogenous NADH/FAD fluorescence emissions. We apply the mathematical models to the two-photon microscopy data and show consistent results. We also used our fiber-optic system to measure redox in different tissues and show consistent results between the two models, hence demonstrating proof-of-principle. This innovative redox monitoring system will have practical applications for defining different metabolic disease states.

Structural, redox and isotopic behaviors of iron in geological silicate glasses: A NRIXS study of Lamb-Mössbauer factors and force constants

We report a combined conventional Mössbauer and synchrotron Nuclear Resonant Inelastic X-ray Scattering (NRIXS) study of a series of basalt, andesite, dacite, and rhyolite glasses at temperatures ranging from 5 to 1223 K. These glasses were synthesized under controlled oxygen fugacities and span a wide range of Fe3+/Fetot ratios. As expected from theory, we find that in these glasses, the inverse of the Lamb-Mössbauer factor (the recoil-free fraction) correlates linearly with the fraction of the conventional Mössbauer spectrum that can be ascribed to Fe3+. Extrapolating the linear relationships to pure Fe3+ and Fe2+ endmembers yields a ratio for the Lamb-Mössbauer factors C of Fe3+ and Fe2+ of 1.203 ± 0.017 (1σ), with no clear dependence on the chemical composition of the glass. We show that the materials studied follow the harmonic approximation up to 1223 K for olivine and 773 K for basaltic glass, allowing us to extrapolate C in temperature. The temperature-dependence of C is well approximated by the formula C=1.203±0.033T/300 over the temperature range 50–750 K. This calibration is used to correct previous redox ratio determinations. We investigate how chemical composition and iron redox state influence the mean force constant of iron in glasses, a key driver of Fe isotopic fractionation during magma generation and differentiation. As previously documented by Dauphas et al. (2014), we find clear correlation between bond strength and iron redox ratio that is secondarily modulated by coordination effects in more alkali-rich felsic magmas. New data on simplified glass compositions reveal that network modifiers, notably K and Na, seem to exert an important control on Fe2+ bond strengths. The refined ratio of Mössbauer factors for Fe2+ and Fe3+ is used to improve on calibrations of iron redox state in geological glasses using XANES spectroscopy. We reevaluate the Fe3+/Fetot ratio of MORBs based on previously published XANES and Mössbauer data. 0.087±0.005

Information entropy of quantitative chemometric endogenous fluorescence improves photonic lung cancer diagnosis.

Quantitative chemometric widefield endogenous fluorescence microscopy (CFM) maps the endogenous absolute chromophore concentration and spatial distribution in cells and tissue sections label-free from fluorescence color images under broadband excitation and detection. By quantifying the endogenous chromophores, including tryptophan, elastin, reduced nicotinamide adenine dinucleotide [NAD(P)H], and flavin adenine dinucleotide (FAD), CFM reveals the biochemical environment and subcellular structure. Here we show that the chromophore information entropy, marking its spatial distribution pattern of quantitative chemometric endogenous fluorescence at the microscopic scale, improves photonic lung cancer diagnosis with independent diagnostic power to the cellular metabolism biomarker. NAD(P)H and FAD's information entropy is found to decrease from normal to perilesional to cancerous tissue, whereas the information entropy for the redox ratios [FAD/tryptophan and FAD/NAD(P)H] is smaller for the normal tissue than both perilesional and cancerous tissue. CFM imaging of the specimen's inherent biochemical and structural properties eliminates the dependence on measurement details and facilitates robust, accurate diagnosis. The synergy of quantifying absolute chromophore concentration and information entropy achieves high accuracies for a three-class classification of lung tissue into normal, perilesional, and cancerous ones and a three-class classification of lung cancers into grade 1, grade 2, and grade 3 using a support vector machine, outperforming the chromophore concentration biomarkers.

Synthesis, characterization and density Functional theory (DFT) study of LaCeO3

Nanosized perovskite oxide of LaCeO3 was synthesized using the EDTA-citrate method and calcined at 600 °C for 5 h. The white powder obtained from synthesis was characterized using X-ray Diffraction (XRD), Scanning Electron Microscopy (SEM), Raman Spectroscopy, and UV–Vis Diffuse Reflectance Spectroscopy (UV–Vis DRS) Analysis. The developed perovskite can be used as catalyst in soot oxidation, VOC oxidation, and photo thermal reactions, and materials in solar cells, sensors and fuel cell. XRD analysis confirms the crystalline nature of the perovskite with a predominantly cubic fluorite structure. Morphological studies concluded that the LaCeO3 perovskite has a crystalline microstructure with a large number of honeycomb-like macropores with a minor crystalline phase. UV–Vis DRS tells us that the absorbance acquired can be attributed to the variations in the composition of Ce. Tauc plot allotted based on the data obtained from UV–vis Spectroscopy gave an energy band gap of 3.15 eV, signifying that the material may be classified as a semiconductor. Raman Spectroscopy tells us that the sharp peak obtained at 446 cm−1 corresponds to the stretching of the Ce-O bond. XPS analysis corresponds to La 3d with binding energy gap of 3.3 eV, Ce 3d with redox ratio 49.88% and O 1 s peak with surface oxygen vacancy ratio of 65.5%. DFT analysis of the sample result in direct band gap of 0.802 eV and indirect band gap of 1.716 eV.

Feasibility of Non-invasive Measurement of Tumour NAD(H) by In Vivo Phosphorus-31 Magnetic Resonance Spectroscopy.

Importance of the redox status of nicotinamide adenine dinucleotide (NAD), including its oxidized (NAD+) and reduced (NADH) forms, has been shown in many biological processes. However, NAD(H) redox status assessment is traditionally limited to biochemical assays in vitro or optical redox imaging (ORI) for superficial tissues in vivo and for deep tissues ex vivo. In recent years, phosphorous-31 magnetic resonance spectroscopy (31P-MRS) was utilized to quantify NAD+, NADH, and the redox ratio NAD+/NADH in normal tissues in vivo. The quantification is based on the spectral fitting of the upfield shoulder of the αATP peak that contains signals of NAD+ (a quartet) and NADH (a singlet), assuming pH-independence of peak positions. To evaluate the feasibility of measuring tumour NAD(H) redox status in vivo, we fitted single voxel 31P-MR spectra of subcutaneous mouse xenografts of human breast cancer cell lines acquired on a 9.4-T horizontal bore preclinical MR scanner. We found larger variations in the chemical shift offsets of NAD+ and NADH from αATP in these tumours than the literature values of normal tissues. Furthermore, our 31P-MR spectra of αATP, NAD+, and NADH solution phantoms indicated that the chemical shift of αATP and thus the offsets between NAD(H) and αATP were pH dependent. Therefore, whether tumour pH should be incorporated into the spectral fitting model should be further evaluated. Additionally, spectral resolution and signal-to-noise ratio should be improved by optimising 31P-MRS protocols, increasing data acquisition time, and using a more sensitive coil for signal detection.

Physicochemical properties and structure of titania-containing metallurgical slags: a review

The titanium industry can hardly bypass the titania-containing slags, and the slag physicochemical properties are essential in the metallurgical reactor design and process control. The TiO2–FeO-based slags and TiO2–SiO2–CaO-based slags are the main metallurgical slag systems in the titanium resource utilization processes. To elaborate the role of TiO2 in the physicochemical properties of titania-containing metallurgical slags, the physicochemical properties including titanium redox ratio, liquidus temperature, viscosity, electrical conductivity, density, surface tension, thermal conductivity, and sulfide capacity were critically reviewed. Moreover, the property prediction models were briefly introduced with regards to the limitations of the existing models. The property prediction models are still required to evolve since not all properties of titania-containing slags can be well modeled. As the slag structure has an intimate relationship with slag properties, the structural details of the titania-containing slag were investigated by using a combination of spectroscopic technologies, but the knowledge of the slag structure was not fully ascertained. The potential research fields related to the physicochemical properties and structure of the titania-containing slags were also suggested.

Maturation- and degeneration-dependent articular cartilage metabolism via optical redox ratio imaging.

From the two metabolic processes in healthy cartilage, glycolysis has been associated with proliferation and oxidative phosphorylation (oxphos) with matrix synthesis. Recently, metabolic dysregulation was significantly correlated with cartilage degradation and osteoarthritis progression. While these findings suggest maturation predisposes cartilage to metabolic instability with consequences for tissue maintenance, these links have not been shown. Therefore, this study sought to address three hypotheses (a) chondrocytes exhibit differential metabolic activity between immaturity (0-4 months), adolescence (5-18 months), and maturity (>18 months); (b) perturbation of metabolic activity has consequences on expression of genes pertinent to cartilage tissue maintenance; and (c) severity of cartilage damage is positively correlated with glycolysis and oxphos activity as well as optical redox ratio in postadolescent cartilage. Porcine femoral cartilage samples from pigs (3 days to 6 years) underwent optical redox ratio imaging, which measures autofluorescence of NAD(P)H and FAD. Gene expression analysis and histological scoring was conducted for comparison against imaging metrics. NAD(P)H and FAD autofluorescence both demonstrated increasing intensity with age, while optical redox ratio was lowest in adolescent samples compared to immature or mature samples. Inhibition of glycolysis suppressed expression of Col2, Col1, ADAMTS4, and ADAMTS5, while oxphos inhibition had no effect. FAD fluorescence and optical redox ratio were positively correlated with histological degeneration. This study demonstrates maturation- and degeneration-dependent metabolic activity in cartilage and explores the consequences of this differential activity on gene expression. This study aids our basic understanding of cartilage biology and highlights opportunity for potential diagnostic applications.

Simultaneous NAD(P)H and FAD fluorescence lifetime microscopy of long UVA\u2013induced metabolic stress in reconstructed human skin

Solar ultraviolet longwave UVA1 exposure of human skin has short-term consequences at cellular and molecular level, leading at long-term to photoaging. Following exposure, reactive oxygen species (ROS) are generated, inducing oxidative stress that might impair cellular metabolic activity. However, the dynamic of UVA1 impact on cellular metabolism remains unknown because of lacking adequate live imaging techniques. Here we assess the UVA1-induced metabolic stress response in reconstructed human skin with multicolor two-photon fluorescence lifetime microscopy (FLIM). Simultaneous imaging of nicotinamide adenine dinucleotide (NAD(P)H) and flavin adenine dinucleotide (FAD) by wavelength mixing allows quantifying cellular metabolism in function of NAD(P)+/NAD(P)H and FAD/FADH2 redox ratios. After UVA1 exposure, we observe an increase of fraction of bound NAD(P)H and decrease of fraction of bound FAD indicating a metabolic switch from glycolysis to oxidative phosphorylation or oxidative stress possibly correlated to ROS generation. NAD(P)H and FAD biomarkers have unique temporal dynamic and sensitivity to skin cell types and UVA1 dose. While the FAD biomarker is UVA1 dose-dependent in keratinocytes, the NAD(P)H biomarker shows no dose dependence in keratinocytes, but is directly affected after exposure in fibroblasts, thus reflecting different skin cells sensitivities to oxidative stress. Finally, we show that a sunscreen including a UVA1 filter prevents UVA1 metabolic stress response from occurring.