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Molecular & Cellular Proteomics : MCP Nov 2020Small admixtures in water, of metal ions, often act as cell growth regulators. Here we report that enrichment of deuterium content in water, normally found at 8 mm...
Small admixtures in water, of metal ions, often act as cell growth regulators. Here we report that enrichment of deuterium content in water, normally found at 8 mm concentration, two-three folds increases cell proliferation and lowers the oxidative stress level as well. Acting as an anti-oxidant, deuterium-enriched water prevents the toxic effect of such oxidative agents as hydrogen peroxide and auranofin. This action is opposite to that of deuterium depletion that is known to suppress cell growth and induce oxidative stress in mitochondria. We thus hypothesize that deuterium may be a natural cell growth regulator that controls mitochondrial oxidation-reduction balance. Because growth acceleration is reduced approximately by half by addition to water a minute amount (0.15%) of O isotope, at least part of the deuterium effect on cell growth can be explained by the isotopic resonance phenomenon. A slight (≈2-fold) enrichment of deuterium in water accelerates human cell growth. Quantitative MS based proteomics determined changes in protein abundances and redox states and found that deuterium-enriched water acts mainly through decreasing ROS production in mitochondria. This action is opposite to that of deuterium depletion that suppresses cell growth by inducing oxidative stress. Thus deuterium may be a natural cell growth regulator that controls mitochondrial oxidation-reduction balance. The role of isotopic resonance in this effect was validated by further experiments on bacteria.
Topics: Antioxidants; Cell Line, Tumor; Cell Proliferation; Cell Survival; Deuterium; Escherichia coli; Humans; Metabolic Networks and Pathways; Mitochondria; Oxidation-Reduction; Oxidative Stress; Principal Component Analysis; Protein Interaction Maps; Proteome; Proteomics; Reactive Oxygen Species; Tandem Mass Spectrometry; Water
PubMed: 32769093
DOI: 10.1074/mcp.RA120.002231 -
The American Journal of Clinical... Jul 2022Most authorities define childhood overweight/obesity as a BMI exceeding the same high centile cutoff at all ages, but it seems unlikely that true obesity prevalence...
BACKGROUND
Most authorities define childhood overweight/obesity as a BMI exceeding the same high centile cutoff at all ages, but it seems unlikely that true obesity prevalence (excess body fat) is constant throughout childhood.
OBJECTIVES
We investigated how fat mass (FM) and lean mass (LM), adjusted for height, relate to BMI and each other across childhood, using a uniquely large database of body composition measures, estimated using gold standard methods.
METHODS
Cross-sectional and cohort data were collated from representative samples of healthy children aged 6 wk to 20 y and children attending obesity clinics aged 7-16 y. Body composition was measured by deuterium dilution up to age 4 y, and by either deuterium or the criterion 4-component model from 4 to 20 y. FM and LM were expressed, respectively, as fat mass index (FMI; FM/height2) and lean mass index (LMI; LM/height2).
RESULTS
There were 2367 measurements of weight, height, and body composition from 1953 individuals. Before age 6 y, the variability in FMI, LMI, and BMI was much less than after; FMI was low (mainly <8 kg/m2) and FMI and LMI were weakly negatively correlated. From mid-childhood, upper limits for both BMI and FMI rose, but FMI in children with BMI <91st centile still rarely exceeded 8. With increasing age, the correlation of FMI with LMI rose to 0.5-0.7, driven mainly by children with a high FMI also having a high LMI.
CONCLUSIONS
Raised fat levels are much less common at younger than older ages, and young children with a high BMI centile have lower FMI than older children with the same BMI centile. Current BMI centile cutoffs thus overdiagnose obesity in younger groups. More stringent cutoffs are required for children aged <6 y, matching the WHO recommendation for 0-5 y.
Topics: Adolescent; Body Composition; Body Mass Index; Child; Child, Preschool; Cross-Sectional Studies; Deuterium; Humans; Overdiagnosis; Pediatric Obesity
PubMed: 34967839
DOI: 10.1093/ajcn/nqab421 -
Molecular & Cellular Proteomics : MCP Oct 2022The assembly of proteins and peptides into amyloid fibrils is causally linked to serious disorders such as Alzheimer's disease. Multiple proteins have been shown to...
The assembly of proteins and peptides into amyloid fibrils is causally linked to serious disorders such as Alzheimer's disease. Multiple proteins have been shown to prevent amyloid formation in vitro and in vivo, ranging from highly specific chaperone-client pairs to completely nonspecific binding of aggregation-prone peptides. The underlying interactions remain elusive. Here, we turn to the machine learning-based structure prediction algorithm AlphaFold2 to obtain models for the nonspecific interactions of β-lactoglobulin, transthyretin, or thioredoxin 80 with the model amyloid peptide amyloid β and the highly specific complex between the BRICHOS chaperone domain of C-terminal region of lung surfactant protein C and its polyvaline target. Using a combination of native mass spectrometry (MS) and ion mobility MS, we show that nonspecific chaperoning is driven predominantly by hydrophobic interactions of amyloid β with hydrophobic surfaces in β-lactoglobulin, transthyretin, and thioredoxin 80, and in part regulated by oligomer stability. For C-terminal region of lung surfactant protein C, native MS and hydrogen-deuterium exchange MS reveal that a disordered region recognizes the polyvaline target by forming a complementary β-strand. Hence, we show that AlphaFold2 and MS can yield atomistic models of hard-to-capture protein interactions that reveal different chaperoning mechanisms based on separate ligand properties and may provide possible clues for specific therapeutic intervention.
Topics: Humans; Amyloid; Amyloid beta-Peptides; Prealbumin; Deuterium; Ligands; Molecular Chaperones; Mass Spectrometry; Machine Learning; Thioredoxins; Lactoglobulins; Pulmonary Surfactant-Associated Proteins
PubMed: 36115577
DOI: 10.1016/j.mcpro.2022.100413 -
The American Journal of Clinical... Apr 2021Milk is an important high-quality animal protein source in low- and middle-income countries (LMICs). Although the true ileal digestibility and absorption of milk has...
BACKGROUND
Milk is an important high-quality animal protein source in low- and middle-income countries (LMICs). Although the true ileal digestibility and absorption of milk has been shown to be high in French adults, this may be lower in individuals from LMICs who are at risk of environmental enteropathy.
OBJECTIVE
To determine the true ileal indispensable amino acid (IAA) digestibility of intrinsically labeled goat milk protein in South Indian women of reproductive age (WRA), using the dual-isotope tracer technique, and to measure intestinal absorption of amino acid and inert sugar in the same participants using L-allo-isoleucine and a dual-sugar assay.
METHODS
Milk with 2H-labeled protein collected from a lactating goat fed intrinsically 2H-labeled fodder (maize and cowpea) was spray dried. Labeled milk protein was administered in a plateau feeding protocol to WRA with normal BMI, in whom urinary lactulose and mannitol recovery and the lactulose/mannitol ratio (LMR) were measured, to determine its true ileal IAA digestibility by the dual-isotope tracer technique with a reference U-13C-amino acid mixture. A phenylalanine absorption index was calculated from the plasma to meal ratio of 13C9 phenylalanine within the digestibility protocol. On a separate day, the allo-isoleucine absorption index was estimated from the ratio of plasma allo-isoleucine enrichments after oral 13C6-15N-L- and intravenous 2H10-L-allo-isoleucine administration.
RESULTS
The means ± SDs of true ileal IAA digestibility of goat milk protein, lactulose and mannitol recovery, LMR, allo-isoleucine and phenylalanine absorption index were 94.0 ± 2.9%, 0.09 ± 0.03%, 7.9 ± 2.3%, 0.012 ± 0.004, 88.4 ± 3.8% and 24.5 ± 1.6%, respectively. The LMR correlated with the allo-isoleucine absorption index (rs = -0.93, P = 0.008).
CONCLUSION
The true ileal digestibility of goat milk protein in South Indian WRA with normal intestinal absorptive function and integrity was comparable to earlier estimates in healthy French adults.
Topics: Adult; Amino Acids; Animal Feed; Animals; Deuterium; Digestion; Female; Goats; Humans; Ileum; Lactulose; Mannitol; Milk; Milk Proteins; Young Adult
PubMed: 33677496
DOI: 10.1093/ajcn/nqaa400 -
Magnetic Resonance in Medicine Jul 2023To monitor the metabolic turnover of β-hydroxybutyrate (BHB) oxidation using H-MRS in conjunction with intravenous administration of H labeled BHB.
PURPOSE
To monitor the metabolic turnover of β-hydroxybutyrate (BHB) oxidation using H-MRS in conjunction with intravenous administration of H labeled BHB.
METHODS
Nine-month-old mice were infused with [3,4,4,4]- H -BHB (d -BHB; 3.11 g/kg) through the tail vein using a bolus variable infusion rate for a period of 90 min. The labeling of downstream cerebral metabolites from the oxidative metabolism of d -BHB was monitored using H-MRS spectra acquired with a home-built H surface coil on a 9.4T preclinical MR scanner with a temporal resolution of 6.25 min. An exponential model was fit to the BHB and glutamate/glutamine (Glx) turnover curves to determine rate constants of metabolite turnover and to aid in the visualization of metabolite time courses.
RESULTS
Deuterium label was incorporated into Glx from BHB metabolism through the tricarboxylic acid (TCA) cycle, with an increase in the level of [4,4]- H -Glx (d -Glx) over time and reaching a quasi-steady state concentration of ∼0.6 ± 0.1 mM following 30 min of infusion. Complete oxidative metabolic breakdown of d -BHB also resulted in the formation of semi-heavy water (HDO), with a four-fold (10.1 to ∼42.1 ± 7.3 mM) linear (R = 0.998) increase in its concentration by the end of infusion. The rate constant of Glx turnover from d -BHB metabolism was determined to be 0.034 ± 0.004 min .
CONCLUSION
H-MRS can be used to monitor the cerebral metabolism of BHB with its deuterated form by measuring the downstream labeling of Glx. The integration of H-MRS with deuterated BHB substrate provides an alternative and clinically promising MRS tool to detect neurometabolic fluxes in healthy and disease conditions.
Topics: Mice; Animals; 3-Hydroxybutyric Acid; Deuterium; Oxidation-Reduction; Brain
PubMed: 36971349
DOI: 10.1002/mrm.29648 -
Current Research in Structural Biology 2022Myelin is a natural and dynamic multilamellar membrane structure that continues to be of significant biological and neurological interest, especially with respect to its...
Myelin is a natural and dynamic multilamellar membrane structure that continues to be of significant biological and neurological interest, especially with respect to its biosynthesis and assembly during its normal formation, maintenance, and pathological breakdown. To explore the usefulness of neutron diffraction in the structural analysis of myelin, we investigated the use of labeling by metabolically incorporating non-toxic levels of deuterium (H; D) via drinking water into a pregnant dam (D-dam) and her developing embryos. All of the mice were sacrificed when the pups (D-pups) were 55 days old. Myelinated sciatic nerves were dissected, fixed in glutaraldehyde and examined by neutron diffraction. Parallel samples that were unfixed (trigeminal nerves) were frozen for mass spectrometry (MS). The diffraction patterns of the nerves from deuterium-fed mice (D-mice) vs. the controls (H-mice) had major differences in the intensities of the Bragg peaks but no appreciable differences in myelin periodicity. Neutron scattering density profiles showed an appreciable increase in density at the center of the lipid-rich membrane bilayer. This increase was greater in D-pups than in D-dam, and its localization was consistent with deuteration of lipid hydrocarbon, which predominates over transmembrane protein in myelin. MS analysis of the lipids isolated from the trigeminal nerves demonstrated that in the pups the percentage of lipids that had one or more deuterium atoms was uniformly high across lipid species (97.6% ± 2.0%), whereas in the mother the lipids were substantially less deuterated (60.6% ± 26.4%) with levels varying among lipid species and subspecies. The mass distribution pattern of deuterium-containing isotopologues indicated the fraction (in %) of each lipid (sub-)species having one or more deuteriums incorporated: in the D-pups, the pattern was always bell-shaped, and the average number of D atoms ranged from a low of ∼4 in fatty acid to a high of ∼9 in cerebroside. By contrast, in D-dam most lipids had more complex, overlapping distributions that were weighted toward a lower average number of deuteriums, which ranged from a low of ∼3-4 in fatty acid and in one species of sulfatide to a high of 6-7 in cerebroside and sphingomyelin. The consistently high level of deuteration in D-pups can be attributed to their lipogenesis during gestation and rapid, postnatal myelination. The widely varying levels of deuteration in D-dam, by contrast, likely depends on the relative metabolic stability of the particular lipid species during myelin maintenance. Our current findings demonstrate that stably-incorporated D label can be detected and localized using neutron diffraction in a complex tissue such as myelin; and moreover, that MS can be used to screen a broad range of deuterated lipid species to monitor differential rates of lipid turnover. In addition to helping to develop a comprehensive understanding of the synthesis and turnover of specific lipids in normal and abnormal myelin, our results also suggest application to studies on myelin proteins (which constitute only 20-30% by dry mass of the myelin, vs. 70-80% for lipid), as well as more broadly to the molecular constituents of other biological tissues.
PubMed: 35941866
DOI: 10.1016/j.crstbi.2022.06.003 -
Journal of the American Society For... Jul 2022Antimicrobial resistance is a serious challenge facing human and veterinary health. Current methods of detecting resistance are limited in turn-around time or universal...
Antimicrobial resistance is a serious challenge facing human and veterinary health. Current methods of detecting resistance are limited in turn-around time or universal detection. In this work, a new antimicrobial susceptibility test is developed and validated, which utilizes deuterium labeling of membrane lipids to track the growth of bacterial cells. We hypothesize that deuterium uptake and subsequent labeling of lipids can be detected using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Additionally, bacteria growth is performed on the MALDI target, minimizing sample preparation materials and time. When two strains are grown in the presence of deuterium oxide, labeling can be detected in as little as 30 min to 2 h. The labeling efficiency, or the ratio of labeled to unlabeled lipid peaks, provides information about the growth rate of bacteria. This growth ratio can differentiate between resistant and susceptible strains of bacteria as a resistant strain will maintain ∼50% labeling efficiency between untreated and treated cultures. In comparison, a susceptible strain will see a decrease in fractional abundance of deuterium from ∼50% in the untreated to ∼10% in the treated. This approach is applied to measure the minimum inhibitory concentration (MIC) of the resistant and susceptible strains from on-target microdroplet culture in a range of antibiotic concentrations. The first antibiotic concentration with a significant decrease in fractional abundance of deuterium correlates well with a traditionally obtained MIC using broth dilution, indicating the clinical relevance of the results.
Topics: Anti-Bacterial Agents; Bacteria; Deuterium; Drug Resistance, Bacterial; Escherichia coli; Humans; Lipids; Microbial Sensitivity Tests; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
PubMed: 35623100
DOI: 10.1021/jasms.2c00056 -
Life (Basel, Switzerland) Nov 2020Hydrogen/Deuterium eXchange Mass Spectrometry (HDX-MS) is a rapidly evolving technique for analyzing structural features and dynamic properties of proteins. It may stand... (Review)
Review
Hydrogen/Deuterium eXchange Mass Spectrometry (HDX-MS) is a rapidly evolving technique for analyzing structural features and dynamic properties of proteins. It may stand alone or serve as a complementary method to cryo-electron-microscopy (EM) or other structural biology approaches. HDX-MS is capable of providing information on individual proteins as well as large protein complexes. Owing to recent methodological advancements and improving availability of instrumentation, HDX-MS is becoming a routine technique for some applications. When dealing with samples of low to medium complexity and sizes of less than 150 kDa, conformation and ligand interaction analyses by HDX-MS are already almost routine applications. This is also well supported by the rapid evolution of the computational (software) background that facilitates the analysis of the obtained experimental data. HDX-MS can cope at times with analytes that are difficult to tackle by any other approach. Large complexes like viral capsids as well as disordered proteins can also be analyzed by this method. HDX-MS has recently become an established tool in the drug discovery process and biopharmaceutical development, as it is now also capable of dissecting post-translational modifications and membrane proteins. This mini review provides the reader with an introduction to the technique and a brief overview of the most common applications. Furthermore, the most challenging likely applications, the analyses of glycosylated and membrane proteins, are also highlighted.
PubMed: 33203161
DOI: 10.3390/life10110286 -
Cell Proliferation Jan 2023Elimination of brain tumour initiating cells (BTICs) is important for the good prognosis of malignant brain tumour treatment. To develop a novel strategy targeting...
OBJECTIVES
Elimination of brain tumour initiating cells (BTICs) is important for the good prognosis of malignant brain tumour treatment. To develop a novel strategy targeting BTICs, we studied NR2E1(TLX) involved self-renewal mechanism of BTICs and explored the intervention means.
MATERIALS AND METHODS
NR2E1 and its interacting protein-LSD1 in BTICs were studied by gene interference combined with cell growth, tumour sphere formation, co-immunoprecipitation and chromatin immunoprecipitation assays. NR2E1 interacting peptide of LSD1 was identified by Amide Hydrogen/Deuterium Exchange and Mass Spectrometry (HDX-MS) and analysed by in vitro functional assays. The in vivo function of the peptide was examined with intracranial mouse model by transplanting patient-derived BTICs.
RESULTS
We found NR2E1 recruits LSD1, a lysine demethylase, to demethylate mono- and di-methylated histone 3 Lys4 (H3K4me/me2) at the Pten promoter and repress its expression, thereby promoting BTIC proliferation. Using Amide Hydrogen/Deuterium Exchange and Mass Spectrometry (HDX-MS) method, we identified four LSD1 peptides that may interact with NR2E1. One of the peptides, LSD1-197-211 that locates at the LSD1 SWIRM domain, strongly inhibited BTIC proliferation by promoting Pten expression through interfering NR2E1 and LSD1 function. Furthermore, overexpression of this peptide in human BTICs can inhibit intracranial tumour formation.
CONCLUSION
Peptide LSD1-197-211 can repress BTICs by interfering the synergistic function of NR2E1 and LSD1 and may be a promising lead peptide for brain tumour therapy in future.
Topics: Animals; Humans; Mice; Amides; Brain; Cell Proliferation; Deuterium; Histone Demethylases; Neoplastic Stem Cells; Orphan Nuclear Receptors; Peptides; Receptors, Cytoplasmic and Nuclear
PubMed: 36321378
DOI: 10.1111/cpr.13350 -
IEEE Transactions on Medical Imaging Dec 2021Deuterium magnetic resonance spectroscopic imaging (DMRSI) has recently been recognized as a potentially powerful tool for noninvasive imaging of brain energy metabolism...
Deuterium magnetic resonance spectroscopic imaging (DMRSI) has recently been recognized as a potentially powerful tool for noninvasive imaging of brain energy metabolism and tumor. However, the low sensitivity of DMRSI has significantly limited its utility for both research and clinical applications. This work presents a novel machine learning-based method to address this limitation. The proposed method synergistically integrates physics-based subspace modeling and data-driven deep learning for effective denoising, making high-resolution dynamic DMRSI possible. Specifically, a novel subspace model was used to represent the dynamic DMRSI signals; deep neural networks were trained to capture the low-dimensional manifolds of the spectral and temporal distributions of practical dynamic DMRSI data. The learned subspace and manifold structures were integrated via a regularization formulation to remove measurement noise. Theoretical analysis, computer simulations, and in vivo experiments have been conducted to demonstrate the denoising efficacy of the proposed method which enabled high-resolution imaging capability. The translational potential was demonstrated in tumor-bearing rats, where the Warburg effect associated with cancer metabolism and tumor heterogeneity were successfully captured. The new method may not only provide an effective tool to enhance the sensitivity of DMRSI for basic research and clinical applications but also provide a framework for denoising other spatiospectral data.
Topics: Algorithms; Animals; Deuterium; Machine Learning; Magnetic Resonance Imaging; Magnetic Resonance Spectroscopy; Rats
PubMed: 34319872
DOI: 10.1109/TMI.2021.3101149