Noninvasive quantification of ascorbate and glutathione concentration in the elderly human brain.
ABSTRACT: In this study, ascorbate (Asc) and glutathione (GSH) concentrations were quantified noninvasively using double-edited (1)H MRS at 4 T in the occipital cortex of healthy young [age (mean ± standard deviation) = 20.4 ± 1.4 years] and elderly (age = 76.6 ± 6.1 years) human subjects. Elderly subjects had a lower GSH concentration than younger subjects (p < 0.05). The Asc concentration was not significantly associated with age. Furthermore, the lactate (Lac) concentration was higher in elderly than young subjects. Lower GSH and higher Lac concentrations are indications of defective protection against oxidative damage and impaired mitochondrial respiration. The extent to which the observed concentration differences could be associated with physiological differences and methodological artifacts is discussed. In conclusion, GSH and Asc concentrations were compared noninvasively for the first time in young vs elderly subjects.
Project description:The transverse relaxation times (T(2)) and concentrations of Ascorbate (Asc) and glutathione (GSH) were measured from a single dataset of double-edited spectra that were acquired at several TEs at 4 T in the human brain. Six TEs between 102 and 152 ms were utilized to calculate T(2) for the group of 12 subjects scanned five times each. Spectra measured at all six TEs were summed to quantify the concentration in each individual scan. LCModel fitting was optimized for the quantification of the Asc and GSH double-edited spectra. When the fitted baseline was constrained to be flat, T(2) was found to be 67 ms (95% confidence interval, 50-83 ms) for GSH and ≤115 ms for Asc using the sum of spectra measured over 60 scans. The Asc and GSH concentrations quantified in each of the 60 scans were 0.62 ± 0.08 and 0.81 ± 0.11 µmol/g [mean ± standard deviation (SD), n = 60], respectively, using 10 µmol/g N-acetylaspartate as an internal reference and assuming a constant influence of N-acetylaspartate and antioxidant T(2) relaxation in the reference solution and in vivo. The T(2) value of GSH was measured for the first time in the human brain. The data are consistent with short T(2) for both antioxidants. These T(2) values are essential for the absolute quantification of Asc and GSH concentrations measured at long TE, and provide a critical step towards addressing assumptions about T(2), and therefore towards the quantification of concentrations without the possibility of systematic bias.
Project description:N-aryl maleimides can undergo a 1,4-Michael-type addition reaction with reduced glutathione (GSH), leading to a decreased concentration of GSH and an increased concentration of free radicals (FRs) in cells. GSH is a critical scavenging molecule responsible for protecting cells from oxidation and for maintaining redox homeostasis. N-aryl maleimides disturb redox homeostasis in cells because they scavenge thiol-containing molecules, especially GSH. This study aimed at measuring the concentrations of GSH and FRs by electronic paramagnetic resonance (EPR), in the brain and liver tissue of male Wistar rats (ex vivo) at different ages and after treatment with 3,5-dimaleimylbenzoic acid (3,5-DMB). Our results showed a relationship between age and the concentrations of GSH and FRs in cells. In young rats, the concentration of GSH was higher than in old rats, while the concentration of FRs was higher in adult rats than in young rats, suggesting an inverse relationship between GSH and FRs. On the other hand, the reaction of 3,5-DMB (an electrophilic maleimide) with cellular GSH increased the FR content. The results of this study contribute to the awareness that the process of aging implies not only a loss of tissue function but also essential changes in the molecular contents of cells, especially the concentrations of FRs and GSH.
Project description:Ascorbate (Asc, vitamin C) was quantified in the human brain noninvasively using two different (1)H NMR spectroscopy methods: short-echo time STEAM and MEGA-PRESS homonuclear editing. Taking advantage of increased sensitivity and chemical shift dispersion at 7 T, Asc was quantified with increased reliability relative to our previous study accomplished at 4 T. Asc concentration quantified from short-echo time spectra measured from the occipital lobe of eight healthy subjects ([Asc] = 1.1 +/- 0.3 micromol/g, mean +/- SD) was in excellent agreement with Asc concentration quantified from the same volume of interest using homonuclear editing ([Asc] = 1.2 +/- 0.2 micromol/g). This agreement indicates that at 7 T, Asc can be reliably quantified in the human brain simultaneously with 15 other metabolites. Additional advantages of the short-echo time approach were: shorter measurement time than homonuclear editing and minimal effect of T(2) relaxation on Asc quantification. High magnetic field was also beneficial for Asc quantification with MEGA-PRESS because increased chemical shift dispersion enabled editing with full efficiency, which resulted in a supra-linear gain in signal-to-noise ratio relative to 4 T.
Project description:Two spectral editing techniques for the simultaneous detection of glutathione (GSH) and lactate (Lac) in the human brain at 3 T are described and evaluated. These methods, 'sMEGA' (sinc-MEscher and GArwood) and 'DEW' (Double Editing With), were optimized to detect GSH and Lac simultaneously at 3 T using density-matrix simulations and validation in phantoms. Simulations to test for co-edited metabolites within the detected GSH region of the spectrum were also performed. In vivo data were acquired in the midline parietal region of seven subjects using both methods, and compared with conventional MEGA-PRESS (MEscher and GArwood-Point RESolved Spectroscopy) acquisitions of GSH and Lac. Simulations and phantom experiments showed that sMEGA and DEW had a high editing efficiency for both GSH and Lac. In the phantom, the editing efficiency of GSH was >88% relative to a conventional GSH MEGA-PRESS acquisition, whereas, for Lac, the editing efficiency was >95% relative to a conventional Lac MEGA-PRESS acquisition. Simulations also showed that the editing efficiency of both methods was comparable with separate MEGA-PRESS acquisitions of the same metabolites. In addition, simulations and in vivo spectra showed that, at a TE of 140 ms, there was a partial overlap between creatine (Cr) and GSH peaks, and that N-acetyl aspartate/N-acetyl aspartyl glutamate (NAA/NAAG) were sufficiently resolved from GSH. In vivo measurements showed that both sMEGA and DEW edited GSH and Lac reliably with the same editing efficiency as conventional MEGA-PRESS acquisitions of the same metabolites, with measured GSH integrals of 2.23 ± 0.51, 2.31 ± 0.38, 2.38 ± 0.53 and measured Lac integrals of 1.72 ± 0.67, 1.55 ± 0.35 and 1.53 ± 0.54 for MEGA-PRESS, DEW and sMEGA, respectively. Simultaneous detection of GSH and Lac using sMEGA and DEW is possible at 3 T with high editing efficiency.
Project description:More than 2.5 million patients in the U.S. require treatment for pressure ulcers annually, and the elderly are at particularly high risk for pressure ulcer development. Current therapy for pressure ulcers consists of conservative medical management for shallow lesions and aggressive debridement and surgery for deeper lesions. The current study uses a murine model to address the hypothesis that adipose-derived stromal/stem cell (ASC) treatment would accelerate and enhance pressure ulcer repair. The dorsal skin of both young (2 months old [mo]) and old (20 mo) C57BL/6J female mice was sandwiched between external magnets for 12 hours over 2 consecutive days to initiate a pressure ulcer. One day following the induction, mice were injected with ASCs isolated from congenic mice transgenic for the green fluorescent protein under a ubiquitous promoter. Relative to phosphate-buffered saline-treated controls, ASC-treated mice displayed a cell concentration-dependent acceleration of wound closure, improved epidermal/dermal architecture, increased adipogenesis, and reduced inflammatory cell infiltration. The ASC-induced improvements occurred in both young and elderly recipients, although the expression profile of angiogenic, immunomodulatory, and reparative mRNAs differed as a function of age. The results are consistent with clinical reports that fat grafting improved skin architecture in thermal injuries; the authors of this published study have invoked ASC-based mechanisms to account for their clinical outcomes. Thus, the current proof-of-principle study sets the stage for clinical translation of autologous and/or allogeneic ASC treatment of pressure ulcers.Adipose-derived stromal/stem cells (ASCs) promote the healing of pressure ulcer wounds in both young and old mice. ASCs enhance wound healing rates through adipogenic differentiation and regeneration of the underlying architecture of the skin.
Project description:BACKGROUND:Dotinurad is a novel selective urate reabsorption inhibitor (SURI) that selectively inhibits the reabsorption of uric acid in renal tubules and promotes the excretion of uric acid into urine. In this study, the effects of age and gender on the pharmacokinetics (PK), pharmacodynamics (PD), and safety of dotinurad were evaluated in healthy subjects. METHODS:An open-label study of a single oral administration of dotinurad 1 mg was conducted in elderly (??65 years) Japanese males and females, and young (20-35 years) males and females (six patients each). RESULTS:Following a single-dose administration of dotinurad, the change in dotinurad plasma concentration showed a similar profile across groups. Regarding the PK parameters, there was no significant difference between elderly and young subjects. On comparing males and females, significant differences were observed in some parameters in elderly subjects. However, these differences in some parameters could not be detected by adjust for body weight. When PD parameters in elderly and young subjects were compared, significant differences were observed in some parameters in male subjects. On comparing males and females, significant differences were observed in some parameters in young subjects; however, the percent change in serum uric acid concentration decreased over time was relatively close for both groups. There were no clinically relevant safety problems. CONCLUSION:Age and gender had no clinically meaningful effect on the PK, PD, and safety of dotinurad. CLINICAL TRIALS:ClinicalTrials.gov identifier: NCT02344875.
Project description:Age-associated accumulation of somatic mutations in mitochondrial DNA (mtDNA) has been proposed to be responsible for the age-associated mitochondrial respiration defects found in elderly human subjects. In contrasts, our previous studies proposed that the age-associated respiration defects found in human fibroblasts are caused not by mtDNA mutations. To addressed these controversial issues, we carried out microarray analysis of two young (TIG3S and TIG121) and two elderly (TIG107 and TIG102) fibroblasts. Overall design: Young (TIG3S and TIG121) and elderly (TIG107 and TIG102) fibroblasts are analyzed.
Project description:The shift from marine to plant-based ingredients in fish feeds affects the dietary concentrations and bioavailability of micronutrients, amino acids and lipids and consequently warrants a re-evaluation of dietary nutrient recommendations. In the present study, an Atlantic salmon diet high in plant ingredients was supplemented with graded levels of nutrient premix (NP), containing selected amino acids, taurine, cholesterol, vitamins and minerals. This article presents the results on the antioxidant nutrients vitamin C, E and selenium (Se), and effects on tissue redox status. The feed ingredients appeared to contain sufficient levels of vitamin E and Se to cover the requirements to prevent clinical deficiency symptoms. The body levels of ?-tocopherol (TOH) in parr and that of Se in parr and post-smolt showed a linear relationship with dietary concentration, while ?-TOH in post-smolt seemed to be saturable with a breakpoint near 140 mg kg-1. Ascorbic acid (Asc) concentration in the basal feed was below the expected minimum requirement, but the experimental period was probably too short for the fish to develop visible deficiency symptoms. Asc was saturable in both parr and post-smolt whole body at dietary concentrations of 190 and 63-89 mg kg-1, respectively. Maximum whole body Asc concentration was approximately 40 mg kg-1 in parr and 14 mg kg-1 in post-smolt. Retention ranged from 41 to 10% in parr and from -206 to 12% in post-smolt with increasing NP supplementation. This indicates that the post-smolts had an extraordinarily high consumption of Asc. Analyses of glutathione (GSH) and glutathione disulphide (GSSG) concentrations and the calculated GSH based redox potentials in liver and muscle tissue, indicated only minor effects of diets on redox regulation. However, the post-smolt were more oxidized than the parr. This was supported by the high consumption of Asc and high expression of gpx1 and gpx3 in liver. Based on the present trials, the recommendations for supplementation of vitamin C and E in diets for Atlantic salmon are similar to current practices, e.g. 150 mg kg-1 of ?-TOH and 190 mg kg-1 Asc which was the saturating concentration in parr. Higher concentrations than what would prevent clinical deficiency symptoms are necessary to protect fish against incidents of oxidative stress and to improve immune and stress responses. There were no indications that the Se requirement exceeded the current recommendation of 0.3 mg kg-1.
Project description:Weaning usually leads to stress in livestock, which has a negative impact on their growth and development. Research on oxidative stress and inflammation induced by weaning has not been reported in goats. Here, we focused on oxidative stress profile and inflammation status of the lower gut (jejunum, ileum, and colon) of goats. First, we illustrated the status of antioxidant activity and inflammation in the intestine of young goats on pre-(2 weeks postnatal, 2 wkpn) or post-(11 wkpn, weaning at day 45 postnatal)-weaned period of young goats. Malondialdehyde (MDA) was higher (p < 0.0001) in jejunum and ileum of the young goats in 11 wkpn than that in 2 wkpn, whereas superoxide dismutase (SOD) activity was lower (p = 0.012) in the lower gut of the young goats with 11 wkpn than that in 2 wkpn. Furthermore, we intended to explore the protective influence of a probiotic additive (Lactobacillus plantarum (LAC) P-8, 10 g/d) and a prebiotic additive (Sangrovit®, Macleaya cordata (MAC) extract 3.75% w/w premix, 0.3 g/d) on intestinal oxidative stress and inflammation status of early-weaned young goats (average weights of 5.63 ± 0.30 kg, weaned on d 45 postnatal). We observed that LAC reduced MDA in jejunum and ileum (p < 0.0001), increased SOD activity in ileum (p < 0.01), and increased glutathione peroxidase (GSH-Px) activity in jejunum (p < 0.05). Similarly, MAC reduced MDA contents (p < 0.0001), increased SOD activities (p < 0.01) in both of ileum and jejunum, and increased GSH-Px activity (p < 0.05) in jejunum. However, there were no differences in feed intake, average daily gain, inflammation parameters (interleukin 2 and interleukin 6), and colon oxidative stress profile (MDA, SOD, or GSH-Px) among treatments. These results provide evidence that weaning induces oxidative damage in the lower gut of young goats, and the oxidative damage in the small intestine can be reduced by adding the addition of LAC or MAC in diets depending on the region of the lower gut.
Project description:A decline in brain function is a characteristic feature of healthy aging; however, little is known about the biologic basis of this phenomenon. To determine whether there are alterations in brain mitochondrial metabolism associated with healthy aging, we combined (13)C/(1)H magnetic resonance spectroscopy with infusions of [1-(13)C]glucose and [2-(13)C]acetate to quantitatively characterize rates of neuronal and astroglial tricarboxylic acid cycles, as well as neuroglial glutamate-glutamine cycling, in healthy elderly and young volunteers. Compared with young subjects, neuronal mitochondrial metabolism and glutamate-glutamine cycle flux was approximately 30% lower in elderly subjects. The reduction in individual subjects correlated strongly with reductions in N-acetylaspartate and glutamate concentrations consistent with chronic reductions in brain mitochondrial function. In elderly subjects infused with [2-(13)C]acetate labeling of glutamine, C4 and C3 differed from that of the young subjects, indicating age-related changes in glial mitochondrial metabolism. Taken together, these studies show that healthy aging is associated with reduced neuronal mitochondrial metabolism and altered glial mitochondrial metabolism, which may in part be responsible for declines in brain function.