Publications by authors named "Alan J Stewart"

89 Publications

Correction.

Am Nat 2021 Sep 24;198(3):438-439. Epub 2021 Jun 24.

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http://dx.doi.org/10.1086/715499DOI Listing
September 2021

Prognostic features of the tumour microenvironment in oesophageal adenocarcinoma.

Biochim Biophys Acta Rev Cancer 2021 Jul 29;1876(2):188598. Epub 2021 Jul 29.

Department of Surgery, Trinity Translational Medicine Institute, St. James's Hospital, Dublin 8, Ireland; Trinity St James's Cancer Institute, St James's Hospital, Dublin 8, Ireland. Electronic address:

Oesophageal adenocarcinoma (OAC) is a disease with an incredibly poor survival rate and a complex makeup. The growth and spread of OAC tumours are profoundly influenced by their surrounding microenvironment and the properties of the tumour itself. Constant crosstalk between the tumour and its microenvironment is key to the survival of the tumour and ultimately the death of the patient. The tumour microenvironment (TME) is composed of a complex milieu of cell types including cancer associated fibroblasts (CAFs) which make up the tumour stroma, endothelial cells which line blood and lymphatic vessels and infiltrating immune cell populations. These various cell types and the tumour constantly communicate through environmental cues including fluctuations in pH, hypoxia and the release of mitogens such as cytokines, chemokines and growth factors, many of which help promote malignant progression. Eventually clusters of tumour cells such as tumour buds break away and spread through the lymphatic system to nearby lymph nodes or enter the circulation forming secondary metastasis. Collectively, these factors need to be considered when assessing and treating patients clinically. This review aims to summarise the ways in which these various factors are currently assessed and how they relate to patient treatment and outcome at an individual level.
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http://dx.doi.org/10.1016/j.bbcan.2021.188598DOI Listing
July 2021

Albumin-mediated alteration of plasma zinc speciation by fatty acids modulates blood clotting in type-2 diabetes.

Chem Sci 2021 Feb 1;12(11):4079-4093. Epub 2021 Feb 1.

School of Medicine, University of St Andrews Fife KY16 9TF St Andrews UK +44 (0)1334 463482 +44 (0)1334 463546.

Zn is an essential regulator of coagulation and is released from activated platelets. In plasma, the free Zn concentration is fine-tuned through buffering by human serum albumin (HSA). Importantly, the ability of HSA to bind/buffer Zn is compromised by co-transported non-esterified fatty acids (NEFAs). Given the role of Zn in blood clot formation, we hypothesise that Zn displacement from HSA by NEFAs in certain conditions (such as type 2 diabetes mellitus, T2DM) impacts on the cellular and protein arms of coagulation. To test this hypothesis, we assessed the extent to which increasing concentrations of a range of medium- and long-chain NEFAs reduced Zn-binding ability of HSA. Amongst the NEFAs tested, palmitate (16 : 0) and stearate (18 : 0) were the most effective at suppressing zinc-binding, whilst the mono-unsaturated palmitoleate (16 : 1c9) was markedly less effective. Assessment of platelet aggregation and fibrin clotting parameters in purified systems and in pooled plasma suggested that the HSA-mediated impact of the model NEFA myristate on zinc speciation intensified the effects of Zn alone. The effects of elevated Zn alone on fibrin clot density and fibre thickness in a purified protein system were mirrored in samples from T2DM patients, who have derranged NEFA metabolism. Crucially, T2DM individuals had increased total plasma NEFAs compared to controls, with the concentrations of key saturated (myristate, palmitate, stearate) and mono-unsaturated (oleate, -vaccenate) NEFAs positively correlating with clot density. Collectively, these data strongly support the concept that elevated NEFA levels contribute to altered coagulation in T2DM through dysregulation of plasma zinc speciation.
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http://dx.doi.org/10.1039/d0sc06605bDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8179462PMC
February 2021

Mapping evidence on integrated conservation and health projects worldwide: an appeal for help in identifying past and ongoing interventions.

Lancet Planet Health 2021 06;5(6):e335

Evolution, behaviour and environment, School of Life Sciences, University of Sussex, Falmer, UK.

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http://dx.doi.org/10.1016/S2542-5196(21)00133-9DOI Listing
June 2021

Leptin and Obesity: Role and Clinical Implication.

Front Endocrinol (Lausanne) 2021 18;12:585887. Epub 2021 May 18.

Department of Radiobiology and Molecular Genetics, "VINČA" Institute of Nuclear Sciences - National Institute of the Republic of Serbia, University of Belgrade, Belgrade, Serbia.

The peptide hormone leptin regulates food intake, body mass, and reproductive function and plays a role in fetal growth, proinflammatory immune responses, angiogenesis and lipolysis. Leptin is a product of the obese () gene and, following synthesis and secretion from fat cells in white adipose tissue, binds to and activates its cognate receptor, the leptin receptor (LEP-R). LEP-R distribution facilitates leptin's pleiotropic effects, playing a crucial role in regulating body mass a negative feedback mechanism between adipose tissue and the hypothalamus. Leptin resistance is characterized by reduced satiety, over-consumption of nutrients, and increased total body mass. Often this leads to obesity, which reduces the effectiveness of using exogenous leptin as a therapeutic agent. Thus, combining leptin therapies with leptin sensitizers may help overcome such resistance and, consequently, obesity. This review examines recent data obtained from human and animal studies related to leptin, its role in obesity, and its usefulness in obesity treatment.
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http://dx.doi.org/10.3389/fendo.2021.585887DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8167040PMC
May 2021

Spatial scaling of plant and bird diversity from 50 to 10,000 ha in a lowland tropical rainforest.

Oecologia 2021 May 5;196(1):101-113. Epub 2021 May 5.

Biology Centre, Institute of Entomology, Czech Academy of Sciences, Branišovská 1760, 370 05, Ceske Budejovice, Czech Republic.

While there are numerous studies of diversity patterns both within local communities and at regional scales, the intermediate scale of tens to thousands of km is often neglected. Here we present detailed local data on plant communities (using 20 × 20 m plots) and bird communities (using point counts) for a 50 ha ForestGEO plot in lowland rainforest at Wanang, Papua New Guinea. We compare these local diversity patterns with those documented in the surrounding 10,000 ha of lowland rainforest. Woody plant species richness was lower within 50 ha (88% of 10,000 ha richness), even when both were surveyed with identical sampling effort. In contrast, bird communities exhibited identical species accumulation patterns at both spatial scales. Similarity in species composition (Chao-Jaccard) remained constant while similarity in dominance structure (Bray-Curtis) decreased with increased distance between samples across the range from < 1 to 13.8 km for both plant and bird communities. The similarity decay was more rapid in plants, but in both cases was slow. The results indicate low to zero beta-diversity at the spatial scale represented here, particularly for birds but also for woody plants. A 50 ha plot provided a highly accurate representation of broader-scale diversity and community composition within 10,000 ha for birds, and a relatively good representation for woody plants. This suggests potential for wider generalization of data from ForestGEO plots which are almost always locally unreplicated, at least for those in lowland tropical forest.
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http://dx.doi.org/10.1007/s00442-021-04925-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8139934PMC
May 2021

Differential outcomes of novel plant-herbivore associations between an invading planthopper and native and invasive Spartina cordgrass species.

Oecologia 2021 Apr 31;195(4):983-994. Epub 2021 Mar 31.

School of Life Sciences, University of Sussex, Brighton, BN1 9QG, UK.

Non-native plants may benefit, briefly or permanently, from natural enemy release in their invaded range, or may form novel interactions with native enemy species. Likewise, newly arrived herbivores may develop novel associations with native plants or, where their hosts have arrived ahead of them, re-establish interactions that existed previously in their ancestral ranges. Predicting outcomes from this diversity of novel and re-established interactions between plants and their herbivores presents a major challenge for invasion biology. We report on interactions between the recently arrived invasive planthopper Prokelisia marginata, and the multi-ploidy Spartina complex of four native and introduced species in Britain, each representing a different level of shared evolutionary history with the herbivore. As predicted, S. alterniflora, the ancestral host, was least impacted by planthopper herbivory, with the previously unexposed native S. maritima, a nationally threatened species, suffering the greatest impacts on leaf length gain, new leaf growth and relative water content. Contrary to expectations, glasshouse trials showed P. marginata to preferentially oviposit on the invasive allododecaploid S. anglica, on which it achieved earlier egg hatch, faster nymphal development, larger female body size and greatest final population size. We suggest P. marginata is in the process of rapid adaptation to maximise its performance on what is now the most abundant and widespread host in Britain. The diversity of novel and re-established interactions of the herbivore with this multi-ploidy complex makes this a highly valuable system for the study of the evolutionary ecology of plant-insect interactions and their influence on invasion dynamics.
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http://dx.doi.org/10.1007/s00442-021-04898-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8052223PMC
April 2021

Pulse Dipolar EPR Reveals Double-Histidine Motif Cu-NTA Spin-Labeling Robustness against Competitor Ions.

J Phys Chem Lett 2021 Mar 13;12(11):2815-2819. Epub 2021 Mar 13.

School of Medicine, University of St. Andrews, North Haugh, St. Andrews, KY16 9TF, U.K.

Pulse-dipolar EPR is an appealing strategy for structural characterization of complex systems in solution that complements other biophysical techniques. Significantly, the emergence of genetically encoded self-assembling spin labels exploiting exogenously introduced double-histidine motifs in conjunction with Cu-chelates offers high precision distance determination in systems nonpermissive to thiol-directed spin labeling. However, the noncovalency of this interaction exposes potential vulnerabilities to competition from adventitious divalent metal ions, and pH sensitivity. Herein, a combination of room-temperature isothermal titration calorimetry (ITC) and cryogenic relaxation-induced dipolar modulation enhancement (RIDME) measurements are applied to the model protein group G. protein G, B1 domain (GB1). Results demonstrate double-histidine motif spin labeling using Cu-nitrilotriacetic acid (Cu-NTA) is robust against the competitor ligand Zn-NTA at >1000-fold molar excess, and high nM binding affinity is surprisingly retained under acidic and basic conditions even though room temperature affinity shows a stronger pH dependence. This indicates the strategy is well-suited for diverse biological applications, with the requirement of other metal ion cofactors or slightly acidic pH not necessarily being prohibitive.
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http://dx.doi.org/10.1021/acs.jpclett.1c00211DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8006131PMC
March 2021

Ablation of Results in Transient Bone Hypomineralization.

JBMR Plus 2021 Feb 8;5(2):e10439. Epub 2020 Dec 8.

The Roslin Institute and Royal (Dick) School of Veterinary Studies University of Edinburgh Midlothian UK.

Biomineralization is a fundamental process key to the development of the skeleton. The phosphatase orphan phosphatase 1 (PHOSPHO1), which likely functions within extracellular matrix vesicles, has emerged as a critical regulator of biomineralization. However, the biochemical pathways that generate intravesicular PHOSPHO1 substrates are currently unknown. We hypothesized that the enzyme ectonucleotide pyrophosphatase/phosphodiesterase 6 (ENPP6) is an upstream source of the PHOSPHO1 substrate. To test this, we characterized skeletal phenotypes of mice homozygous for a targeted deletion of ( ). Micro-computed tomography of the trabecular compartment revealed transient hypomineralization in tibias ( < 0.05) that normalized by 12 weeks of age. Whole-bone cortical analysis also revealed significantly hypomineralized proximal bone in 4- but not 12-week-old mice ( < 0.05) compared with WT animals. Back-scattered SEM revealed a failure in 4-week-old trabecular bone of mineralization foci to propagate. Static histomorphometry revealed increased osteoid volume ( > 0.01) and osteoid surface ( < 0.05), which recovered by 12 weeks but was not accompanied by changes in osteoblast or osteoclast number. This study is the first to characterize the skeletal phenotype of mice, revealing transient hypomineralization in young animals compared with WT controls. These data suggest that ENPP6 is important for bone mineralization and may function upstream of PHOSPHO1 as a novel means of generating its substrates inside matrix vesicles. © 2020 The Authors. published by Wiley Periodicals LLC. on behalf of American Society for Bone and Mineral Research.
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http://dx.doi.org/10.1002/jbm4.10439DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7872340PMC
February 2021

Species Separation within, and Preliminary Phylogeny for, the Leafhopper Genus with Particular Reference to the Putative British Endemic (Hemiptera: Cicadellidae).

Insects 2020 Nov 13;11(11). Epub 2020 Nov 13.

Cardiff School of Biosciences, Sir Martin Evans Building, Museum Avenue, Cardiff CF10 3AX, UK.

The subfamily Aphrodinae (Hemiptera: Cicadellidae) contains ~33 species in Europe within four genera. Species in two genera in particular, and , have proved to be difficult to distinguish morphologically. Our aim was to determine the status of the putative species , found only on the RSPB Nature Reserve at Dungeness, Kent, a possible rare UK endemic. DNA from samples of all seven UK species (plus from the Czech Republic) were sequenced using parts of the mitochondrial cytochrome oxidase I and 16S rRNA genes. Bayesian inference phylogenies were created. Specimens of each species clustered into monophyletic groups, except for , and . Two specimens grouped with repeatedly with strong support, and the remaining clustered within . Genetic distances suggest that and are a single interbreeding population (0% divergence), while and diverged by only 0.28%. Shared haplotypes between , and strongly suggest interbreeding, although misidentification may also explain these topologies. However, all clustered together in the trees. A conservative approach might be to treat , until other evidence is forthcoming, as a possible endemic subspecies.
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http://dx.doi.org/10.3390/insects11110799DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7697789PMC
November 2020

Health service needs and perspectives of remote forest communities in Papua New Guinea: study protocol for combined clinical and rapid anthropological assessments with parallel treatment of urgent cases.

BMJ Open 2020 10 31;10(10):e041784. Epub 2020 Oct 31.

Primary Care and Public Health, and NIHR Global Health Research Unit on Neglected Tropical Diseases, Brighton and Sussex Medical School, Falmer, UK.

Introduction: Our project follows community requests for health service incorporation into conservation collaborations in the rainforests of Papua New Guinea (PNG). This protocol is for health needs assessments, our first step in coplanning medical provision in communities with no existing health data.

Methods And Analysis: The study includes clinical assessments and rapid anthropological assessment procedures (RAP) exploring the health needs and perspectives of partner communities in two areas, conducted over 6 weeks fieldwork. First, in Wanang village (population c.200), which is set in lowland rainforest. Second, in six communities (population c.3000) along an altitudinal transect up the highest mountain in PNG, Mount Wilhelm. Individual primary care assessments incorporate physical examinations and questioning (providing qualitative and quantitative data) while RAP includes focus groups, interviews and field observations (providing qualitative data). Given absence of in-community primary care, treatments are offered alongside research activity but will not form part of the study. Data are collected by a research fellow, primary care clinician and two PNG research technicians. After quantitative and qualitative analyses, we will report: ethnoclassifications of disease, causes, symptoms and perceived appropriate treatment; community rankings of disease importance and service needs; attitudes regarding health service provision; disease burdens and associations with altitudinal-related variables and cultural practices. To aid wider use study tools are in online supplemental file, and paper and ODK versions are available free from the corresponding author.

Ethics And Dissemination: Challenges include supporting informed consent in communities with low literacy and diverse cultures, moral duties to provide treatment alongside research in medically underserved areas while minimising risks of therapeutic misconception and inappropriate inducement, and PNG research capacity building. Brighton and Sussex Medical School (UK), PNG Institute of Medical Research and PNG Medical Research Advisory Committee have approved the study. Dissemination will be via journals, village meetings and plain language summaries.
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http://dx.doi.org/10.1136/bmjopen-2020-041784DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7733180PMC
October 2020

Atherosclerosis Linked to Aberrant Amino Acid Metabolism and Immunosuppressive Amino Acid Catabolizing Enzymes.

Front Immunol 2020 28;11:551758. Epub 2020 Sep 28.

Department of Radiobiology and Molecular Genetics, "VINČA" Institute of Nuclear Sciences - National Institute of the Republic of Serbia, University of Belgrade, Belgrade, Serbia.

Cardiovascular disease is the leading global health concern and responsible for more deaths worldwide than any other type of disorder. Atherosclerosis is a chronic inflammatory disease in the arterial wall, which underpins several types of cardiovascular disease. It has emerged that a strong relationship exists between alterations in amino acid (AA) metabolism and the development of atherosclerosis. Recent studies have reported positive correlations between levels of branched-chain amino acids (BCAAs) such as leucine, valine, and isoleucine in plasma and the occurrence of metabolic disturbances. Elevated serum levels of BCAAs indicate a high cardiometabolic risk. Thus, BCAAs may also impact atherosclerosis prevention and offer a novel therapeutic strategy for specific individuals at risk of coronary events. The metabolism of AAs, such as L-arginine, homoarginine, and L-tryptophan, is recognized as a critical regulator of vascular homeostasis. Dietary intake of homoarginine, taurine, and glycine can improve atherosclerosis by endothelium remodeling. Available data also suggest that the regulation of AA metabolism by indoleamine 2,3-dioxygenase (IDO) and arginases 1 and 2 are mediated through various immunological signals and that immunosuppressive AA metabolizing enzymes are promising therapeutic targets against atherosclerosis. Further clinical studies and basic studies that make use of animal models are required. Here we review recent data examining links between AA metabolism and the development of atherosclerosis.
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http://dx.doi.org/10.3389/fimmu.2020.551758DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7549398PMC
April 2021

Lipidomic profiling of plasma free fatty acids in type-1 diabetes highlights specific changes in lipid metabolism.

Biochim Biophys Acta Mol Cell Biol Lipids 2021 01 1;1866(1):158823. Epub 2020 Oct 1.

School of Medicine, University of St Andrews, Medical and Biological Sciences Building, St Andrews, Fife KY16 9TF, United Kingdom. Electronic address:

Type-1 diabetes mellitus (T1DM) is associated with metabolic changes leading to alterations in glucose and lipid handling. While T1DM-associated effects on many major plasma lipids have been characterised, such effects on plasma free fatty acids (FFA) have not been fully examined. Using gas chromatography-mass spectrometry, we measured the plasma concentrations of FFA species in individuals with T1DM (n = 44) and age/sex-matched healthy controls (n = 44). Relationships between FFA species and various parameters were evaluated. Plasma concentrations of myristate (14:0), palmitoleate (16:1), palmitate (16:0), linoleate (18:2), oleate (18:1c9), cis-vaccenate (18:1c11), eicosapentaenoate (20:5), arachidonate (20:4) and docosahexanoate (22:6) were reduced in the T1DM group (p < 0.0001 for all, except p = 0.0020 for eicosapentaenoate and p = 0.0068 for arachidonate); α-linolenate (18:3) and dihomo-γ-linolenate (20:3) concentrations were unchanged. The saturated/unsaturated FFA ratio, n-3/n-6 ratio, de novo lipogenesis index (palmitate (main lipogenesis product)/linoleate (only found in diet)) and elongase index (oleate/palmitoleate) were increased in the T1DM group (p = 0.0166, p = 0.0089, p < 0.0001 and p = 0.0008 respectively). The stearoyl-CoA desaturase 1 (SCD1) index 1 (palmitoleate/palmitate) and index 2 (oleate/stearate) were reduced in T1DM (p < 0.0001 for both). The delta-(5)-desaturase (D5D) index (arachidonate/dihomo-γ-linolenate) was unchanged. Age and sex had no effect on plasma FFA concentrations in T1DM, while SCD1 index 1 was positively correlated (p = 0.098) and elongase index negatively correlated with age (p = 0.0363). HbA1c was negatively correlated with all plasma FFA concentrations measured except α-linolenate and dihomo-γ-linolenate. Correlations were observed between plasma FFA concentrations and cholesterol and HDL concentrations, but not LDL concentration or diabetes duration. Collectively, these results aid our understanding of T1DM and its effects on lipid metabolism.
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http://dx.doi.org/10.1016/j.bbalip.2020.158823DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7695620PMC
January 2021

Artificial lighting impairs mate attraction in a nocturnal capital breeder.

J Exp Biol 2020 09 30;223(Pt 19). Epub 2020 Sep 30.

School of Life Sciences, University of Sussex, Falmer, Brighton BN1 9QG, UK

Artificial lighting at night (ALAN) is increasingly recognised as having negative effects on many organisms, though the exact mechanisms remain unclear. Glow worms are likely susceptible to ALAN because females use bioluminescence to signal to attract males. We quantified the impact of ALAN by comparing the efficacy of traps that mimicked females to attract males in the presence or absence of a white artificial light source (ALS). Illuminated traps attracted fewer males than did traps in the dark. Illuminated traps closer to the ALS attracted fewer males than those further away, whereas traps in the dark attracted similar numbers of males up to 40 m from the ALS. Thus, ALAN impedes females' ability to attract males, the effect increasing with light intensity. Consequently, ALAN potentially affects glow worms' fecundity and long-term population survival. More broadly, this study emphasises the potentially severe deleterious effects of ALAN upon nocturnal insect populations.
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http://dx.doi.org/10.1242/jeb.229146DOI Listing
September 2020

Reduced Plasma Magnesium Levels in Type-1 Diabetes Associate with Prothrombotic Changes in Fibrin Clotting and Fibrinolysis.

Thromb Haemost 2020 Feb 3;120(2):243-252. Epub 2020 Jan 3.

School of Medicine, University of St Andrews, St Andrews, United Kingdom.

Individuals with type-1 diabetes mellitus (T1DM) have a higher risk of thrombosis and low plasma magnesium concentrations. As magnesium is a known regulator of fibrin network formation, we investigated potential associations between fibrin clot properties and plasma magnesium concentrations in 45 individuals with T1DM and 47 age- and sex-matched controls without diabetes. Fibrin clot characteristics were assessed using a validated turbidimetric assay and associations with plasma magnesium concentration were examined. Plasma concentrations of fibrinogen, plasminogen activator inhibitor-1 (PAI-1), and lipids were measured and fibrin fiber diameters assessed using scanning electron microscopy. Fibrin clot maximum absorbance was unchanged in subjects with T1DM compared with controls, while lysis time was prolonged ( = 0.0273). No differences in fibrin fiber diameters or in lipid profile were observed between T1DM and controls. PAI-1 concentration was lower in the T1DM group compared with the controls ( = 0.0232) and positively correlated with lysis time ( = 0.0023). Plasma magnesium concentration was lower in the T1DM group compared with controls ( < 0.0001). Magnesium concentration negatively correlated with clot maximum absorbance ( = 0.0215) and lysis time ( = 0.0464). A turbidimetric fibrin clot lysis assay performed in a purified system that included PAI-1 and 0 to 3.2 mM Mg showed a shortening of lysis time with increasing Mg concentrations ( = 0.0004). Our findings reveal that plasma magnesium concentration is associated with changes in fibrin clot and lysis parameters.
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http://dx.doi.org/10.1055/s-0039-3402808DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6997031PMC
February 2020

Coagulatory Defects in Type-1 and Type-2 Diabetes.

Int J Mol Sci 2019 Dec 16;20(24). Epub 2019 Dec 16.

Medical and Biological Sciences Building, School of Medicine, University of St Andrews, St Andrews KY16 9TF, UK.

Diabetes (both type-1 and type-2) affects millions of individuals worldwide. A major cause of death for individuals with diabetes is cardiovascular diseases, in part since both types of diabetes lead to physiological changes that affect haemostasis. Those changes include altered concentrations of coagulatory proteins, hyper-activation of platelets, changes in metal ion homeostasis, alterations in lipid metabolism (leading to lipotoxicity in the heart and atherosclerosis), the presence of pro-coagulatory microparticles and endothelial dysfunction. In this review, we explore the different mechanisms by which diabetes leads to an increased risk of developing coagulatory disorders and how this differs between type-1 and type-2 diabetes.
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http://dx.doi.org/10.3390/ijms20246345DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6940903PMC
December 2019

A metalloproteomic analysis of interactions between plasma proteins and zinc: elevated fatty acid levels affect zinc distribution.

Metallomics 2019 11 15;11(11):1805-1819. Epub 2019 Oct 15.

Department of Chemistry, University of Warwick, Coventry, CV4 7AL, UK.

Serum albumin is a highly abundant plasma protein associated with the transport of metal ions, pharmaceuticals, fatty acids and a variety of small molecules in the blood. Once thought of as a molecular 'sponge', mounting evidence suggests that the albumin-facilitated transport of chemically diverse entities is not independent. One such example is the transport of Zn ions and non-esterified 'free' fatty acids (FFAs) by albumin, both of which bind at high affinity sites located in close proximity. Our previous research suggests that their transport in blood plasma is linked via an allosteric mechanism on serum albumin. In direct competition, albumin-bound FFAs significantly decrease the binding capacity of albumin for Zn, with one of the predicted consequences being a change in plasma/serum zinc speciation. Using liquid chromatography (LC), ICP-MS and fluorescence assays, our work provides a quantitative assessment of this phenomenon, and finds that in the presence of high FFA concentrations encountered in various physiological conditions, a significant proportion of albumin-bound Zn is re-distributed amongst plasma/serum proteins. Using peptide mass fingerprinting and immunodetection, we identify candidate acceptor proteins for Zn liberated from albumin. These include histidine-rich glycoprotein (HRG), a multifunctional protein associated with the regulation of blood coagulation, and members of the complement system involved in the innate immune response. Our findings highlight how FFA-mediated changes in extracellular metal speciation might contribute to the progression of certain pathological conditions.
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http://dx.doi.org/10.1039/c9mt00177hDOI Listing
November 2019

Changes in Plasma Free Fatty Acids Associated with Type-2 Diabetes.

Nutrients 2019 Aug 28;11(9). Epub 2019 Aug 28.

School of Medicine, University of St Andrews, St Andrews KY16 9TF, UK.

Type 2 diabetes mellitus (T2DM) is associated with increased total plasma free fatty acid (FFA) concentrations and an elevated risk of cardiovascular disease. The exact mechanisms by which the plasma FFA profile of subjects with T2DM changes is unclear, but it is thought that dietary fats and changes to lipid metabolism are likely to contribute. Therefore, establishing the changes in concentrations of specific FFAs in an individual's plasma is important. Each type of FFA has different effects on physiological processes, including the regulation of lipolysis and lipogenesis in adipose tissue, inflammation, endocrine signalling and the composition and properties of cellular membranes. Alterations in such processes due to altered plasma FFA concentrations/profiles can potentially result in the development of insulin resistance and coagulatory defects. Finally, fibrates and statins, lipid-regulating drugs prescribed to subjects with T2DM, are also thought to exert part of their beneficial effects by impacting on plasma FFA concentrations. Thus, it is also interesting to consider their effects on the concentration of FFAs in plasma. Collectively, we review how FFAs are altered in T2DM and explore the likely downstream physiological and pathological implications of such changes.
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http://dx.doi.org/10.3390/nu11092022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6770316PMC
August 2019

Sub-Micromolar Pulse Dipolar EPR Spectroscopy Reveals Increasing Cu -labelling of Double-Histidine Motifs with Lower Temperature.

Angew Chem Int Ed Engl 2019 08 18;58(34):11681-11685. Epub 2019 Jul 18.

EaStCHEM School of Chemistry, Biomedical Sciences Research Complex, and Centre of Magnetic Resonance, University of St Andrews, North Haugh, St Andrews, KY16 9ST, UK.

Electron paramagnetic resonance (EPR) distance measurements are making increasingly important contributions to the studies of biomolecules by providing highly accurate geometric constraints. Combining double-histidine motifs with Cu spin labels can further increase the precision of distance measurements. It is also useful for proteins containing essential cysteines that can interfere with thiol-specific labelling. However, the non-covalent Cu coordination approach is vulnerable to low binding-affinity. Herein, dissociation constants (K ) are investigated directly from the modulation depths of relaxation-induced dipolar modulation enhancement (RIDME) EPR experiments. This reveals low- to sub-μm Cu K s under EPR distance measurement conditions at cryogenic temperatures. We show the feasibility of exploiting the double-histidine motif for EPR applications even at sub-μm protein concentrations in orthogonally labelled Cu -nitroxide systems using a commercial Q-band EPR instrument.
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http://dx.doi.org/10.1002/anie.201904848DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6771633PMC
August 2019

Quantitative proteomic changes in LPS-activated monocyte-derived dendritic cells: A SWATH-MS study.

Sci Rep 2019 03 13;9(1):4343. Epub 2019 Mar 13.

School of Medicine, University of St Andrews, St Andrews, KY16 9TF, UK.

Dendritic cells are key immune cells that respond to pathogens and co-ordinate many innate and adaptive immune responses. Quantitative mass spectrometry using Sequential Window Acquisition of all THeoretical fragment-ion spectra-Mass Spectrometry (SWATH-MS) was performed here to determine the global alterations in monocyte-derived dendritic cells (moDCs) in response to stimulation with lipopolysaccharide (LPS). A moDC library of 4,666 proteins was generated and proteins were quantified at 0, 6 and 24 h post-LPS stimulation using SWATH-MS. At 6 h and 24 h post-LPS exposure, the relative abundance of 227 and 282 proteins was statistically significantly altered (p-value ≤ 0.05), respectively. Functional annotation of proteins exhibiting significant changes in expression between the various time points led to the identification of clusters of proteins implicated in distinct cellular processes including interferon and interleukin signalling, endocytosis, the ER-phagosome pathway and antigen-presentation. In SWATH-MS major histocompatibility complex (MHC) class I proteins were highly upregulated at 24 h, whilst MHC class II proteins exhibited comparatively fewer changes over this period. This study provides new detailed insight into the global proteomic changes that occur in moDCs during antigen processing and presentation and further demonstrates the potential of SWATH-MS for the quantitative study of proteins involved in cellular processes.
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http://dx.doi.org/10.1038/s41598-019-40773-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6416353PMC
March 2019

Total plasma magnesium, zinc, copper and selenium concentrations in type-I and type-II diabetes.

Biometals 2019 02 22;32(1):123-138. Epub 2019 Jan 22.

School of Medicine, University of St Andrews, Medical and Biological Sciences Building, St Andrews, Fife, KY16 9TF, UK.

Glycemia and insulin resistance are important regulators of multiple physiological processes and their dysregulation has wide-ranging consequences, including alterations in plasma concentrations of metal micronutrients. Here, magnesium, zinc, copper, selenium and glycated albumin (HbA1c) concentrations and quartile differences were examined in 45 subjects with type-I diabetes (T1DM), 54 subjects with type-II diabetes (T2DM) and 62 control subjects in order to assess potential differences between sexes and between T1DM and T2DM. Plasma magnesium concentration was decreased in T1DM subjects, with the second, third and fourth quartiles of magnesium concentrations associated with the absence of T1DM. This effect was observed in females but not males. In T2DM, the highest quartile of selenium concentrations and the third quartile of copper concentrations associated with the absence of diabetes in males. The highest quartile of magnesium concentrations was associated with the absence of T2DM in males but not females. HbA1c correlated with plasma concentrations of magnesium (negatively, in both sexes together in T1DM and T1DM males), copper (positively, in T1DM males and in both sexes together in T2DM), selenium (positively, in both sexes together in T1DM and T2DM, and T2DM females) and with zinc/copper ratio (negatively, in both sexes together in T1DM and T2DM). This study shows that plasma magnesium concentration is altered to the highest degree in T1DM, while in T2DM, plasma selenium and copper concentrations are significantly affected. This work increases our understanding of how T1DM and T2DM affects plasma metal concentrations and may have future implications for diabetes management.
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http://dx.doi.org/10.1007/s10534-018-00167-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7004817PMC
February 2019

On the origin of proteins in human drusen: The meet, greet and stick hypothesis.

Prog Retin Eye Res 2019 05 17;70:55-84. Epub 2018 Dec 17.

UCL Institute of Ophthalmology, University College London, London, UK; Centre for Experimental Medicine, School of Medicine, Dentistry and Biomedical Sciences, Queen's University, Belfast, Northern Ireland, UK.

Retinal drusen formation is not only a clinical hallmark for the development of age-related macular degeneration (AMD) but also for other disorders, such as Alzheimer's disease and renal diseases. The initiation and growth of drusen is poorly understood. Attention has focused on lipids and minerals, but relatively little is known about the origin of drusen-associated proteins and how they are retained in the space between the basal lamina of the retinal pigment epithelium and the inner collagenous layer space (sub-RPE-BL space). While some authors suggested that drusen proteins are mainly derived from cellular debris from processed photoreceptor outer segments and the RPE, others suggest a choroidal cell or blood origin. Here, we reviewed and supplemented the existing literature on the molecular composition of the retina/choroid complex, to gain a more complete understanding of the sources of proteins in drusen. These "drusenomics" studies showed that a considerable proportion of currently identified drusen proteins is uniquely originating from the blood. A smaller, but still large fraction of drusen proteins comes from both blood and/or RPE. Only a small proportion of drusen proteins is uniquely derived from the photoreceptors or choroid. We next evaluated how drusen components may "meet, greet and stick" to each other and/or to structures like hydroxyapatite spherules to form macroscopic deposits in the sub-RPE-BL space. Finally, we discuss implications of our findings with respect to the previously proposed homology between drusenogenesis in AMD and plaque formation in atherosclerosis.
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http://dx.doi.org/10.1016/j.preteyeres.2018.12.003DOI Listing
May 2019

Developing a list of invasive alien species likely to threaten biodiversity and ecosystems in the European Union.

Glob Chang Biol 2019 03 12;25(3):1032-1048. Epub 2018 Dec 12.

Institut National de la Recherche Agronomique, Zoologie Forestière, UR 0633, Ardon Orleans Cedex 2, France.

The European Union (EU) has recently published its first list of invasive alien species (IAS) of EU concern to which current legislation must apply. The list comprises species known to pose great threats to biodiversity and needs to be maintained and updated. Horizon scanning is seen as critical to identify the most threatening potential IAS that do not yet occur in Europe to be subsequently risk assessed for future listing. Accordingly, we present a systematic consensus horizon scanning procedure to derive a ranked list of potential IAS likely to arrive, establish, spread and have an impact on biodiversity in the region over the next decade. The approach is unique in the continental scale examined, the breadth of taxonomic groups and environments considered, and the methods and data sources used. International experts were brought together to address five broad thematic groups of potential IAS. For each thematic group the experts first independently assembled lists of potential IAS not yet established in the EU but potentially threatening biodiversity if introduced. Experts were asked to score the species within their thematic group for their separate likelihoods of i) arrival, ii) establishment, iii) spread, and iv) magnitude of the potential negative impact on biodiversity within the EU. Experts then convened for a 2-day workshop applying consensus methods to compile a ranked list of potential IAS. From an initial working list of 329 species, a list of 66 species not yet established in the EU that were considered to be very high (8 species), high (40 species) or medium (18 species) risk species was derived. Here, we present these species highlighting the potential negative impacts and the most likely biogeographic regions to be affected by these potential IAS.
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http://dx.doi.org/10.1111/gcb.14527DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7380041PMC
March 2019

Crosstalk between zinc and free fatty acids in plasma.

Biochim Biophys Acta Mol Cell Biol Lipids 2019 04 25;1864(4):532-542. Epub 2018 Sep 25.

Department of Chemistry, University of Warwick, Coventry CV4 7AL, UK. Electronic address:

In mammalian blood plasma, serum albumin acts as a transport protein for free fatty acids, other lipids and hydrophobic molecules including neurodegenerative peptides, and essential metal ions such as zinc to allow their systemic distribution. Importantly, binding of these chemically extremely diverse entities is not independent, but linked allosterically. One particularly intriguing allosteric link exists between free fatty acid and zinc binding. Albumin thus mediates crosstalk between energy status/metabolism and organismal zinc handling. In recognition of the fact that even small changes in extracellular zinc concentration and speciation modulate the function of many cell types, the albumin-mediated impact of free fatty acid concentration on zinc distribution may be significant for both normal physiological processes including energy metabolism, insulin activity, heparin neutralisation, blood coagulation, and zinc signalling, and a range of disease states, including metabolic syndrome, cardiovascular disease, myocardial ischemia, diabetes, and thrombosis.
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http://dx.doi.org/10.1016/j.bbalip.2018.09.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6372834PMC
April 2019

Influence of zinc on glycosaminoglycan neutralisation during coagulation.

Metallomics 2018 09;10(9):1180-1190

School of Medicine, University of St Andrews, Medical and Biological Sciences Building, St Andrews, Fife, UK.

Heparan sulfate (HS), dermatan sulfate (DS) and heparin are glycosaminoglycans (GAGs) that serve as key natural and pharmacological anticoagulants. During normal clotting such agents require to be inactivated or neutralised. Several proteins have been reported to facilitate their neutralisation, which reside in platelet α-granules and are released following platelet activation. These include histidine-rich-glycoprotein (HRG), fibrinogen and high-molecular-weight kininogen (HMWK). Zinc ions (Zn2+) are also present in α-granules at a high concentration and participate in the propagation of coagulation by influencing the binding of neutralising proteins to GAGs. Zn2+ in many cases increases the affinity of these proteins to GAGs, and is thus an important regulator of GAG neutralisation and haemostasis. Binding of Zn2+ to HRG, HMWK and fibrinogen is mediated predominantly through coordination to histidine residues but the mechanisms by which Zn2+ increases the affinity of the proteins for GAGs are not yet completely clear. Here we will review current knowledge of how Zn2+ binds to and influences the neutralisation of GAGs and describe the importance of this process in both normal and pathogenic clotting.
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http://dx.doi.org/10.1039/c8mt00159fDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6148461PMC
September 2018

Ischemia-modified albumin: Crosstalk between fatty acid and cobalt binding.

Prostaglandins Leukot Essent Fatty Acids 2018 08 20;135:147-157. Epub 2018 Jul 20.

School of Medicine, University of St Andrews, St Andrews, United Kingdom. Electronic address:

Myocardial ischemia is difficult to diagnose effectively with still few well-defined biochemical markers for identification in advance, or in the absence of myocardial necrosis. "Ischemia-modified albumin" (IMA), a form of albumin displaying reduced cobalt-binding affinity, is significantly elevated in ischemic patients, and the albumin cobalt-binding (ACB) assay can measure its level indirectly. Elucidating the molecular mechanism underlying the identity of IMA and the ACB assay hinges on understanding metal-binding properties of albumin. Albumin binds most metal ions and harbours four primary metal binding sites: site A, site B, the N-terminal site (NTS), and the free thiol at Cys34. Previous efforts to clarify the identity of IMA and the causes for its reduced cobalt-binding capacity were focused on the NTS site, but the degree of N-terminal modification could not be correlated to the presence of ischemia. More recent work suggested that Co ions as used in the ACB assay bind preferentially to site B, then to site A, and finally to the NTS. This insight paved the way for a new consistent molecular basis of the ACB assay: albumin is also the main plasma carrier for free fatty acids (FFAs), and binding of a fatty acid to the high-affinity site FA2 results in conformational changes in albumin which prevent metal binding at site A and partially at site B. Thus, this review advances the hypothesis that high IMA levels in myocardial ischemia and many other conditions originate from high plasma FFA levels hampering the binding of Co to sites A and/or B. This is supported by biophysical studies and the co-association of a range of pathological conditions with positive ACB assays and high plasma FFA levels.
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http://dx.doi.org/10.1016/j.plefa.2018.07.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6109191PMC
August 2018

Native electrospray mass spectrometry approaches to probe the interaction between zinc and an anti-angiogenic peptide from histidine-rich glycoprotein.

Sci Rep 2018 06 5;8(1):8646. Epub 2018 Jun 5.

Department of Chemistry, University of Warwick, Coventry, UK.

Zinc modulates the biological function of histidine-rich glycoprotein (HRG) through binding to its His-rich region (HRR). The Zn-binding properties of a 35 amino-acid biologically-active peptide mimic of the HRR, HRGP330, were investigated using dissociative mass spectrometry approaches in addition to travelling-wave ion mobility mass spectrometry (TWIM-MS). Native mass spectrometry confirmed zinc binding to HRGP330; however, broadening of the H NMR resonances upon addition of Zn ions precluded the attainment of structural information. A complementary approach employing TWIM-MS indicated that HRGP330 has a more compact structure in the presence of Zn ions. Top-down MS/MS data supported a metal-binding-induced conformational change, as fewer fragments were observed for Zn-bound HRGP330. Zn-bound fragments of both N-terminal and C-terminal ends of the peptide were identified from collision-induced dissociation (CID) and electron transfer dissociation/proton transfer reaction (ETD/PTR) experiments, suggesting that multiple binding sites exist within this region of HRG. The combination of mass spectrometry and NMR approaches provides new insight into the highly dynamic interaction between zinc and this His-rich peptide.
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http://dx.doi.org/10.1038/s41598-018-26924-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5988744PMC
June 2018

Glycosaminoglycan Neutralization in Coagulation Control.

Arterioscler Thromb Vasc Biol 2018 06 19;38(6):1258-1270. Epub 2018 Apr 19.

From the School of Medicine, University of St Andrews, Fife, United Kingdom.

The glycosaminoglycans (GAGs) heparan sulfate, dermatan sulfate, and heparin are important anticoagulants that inhibit clot formation through interactions with antithrombin and heparin cofactor II. Unfractionated heparin, low-molecular-weight heparin, and heparin-derived drugs are often the main treatments used clinically to handle coagulatory disorders. A wide range of proteins have been reported to bind and neutralize these GAGs to promote clot formation. Such neutralizing proteins are involved in a variety of other physiological processes, including inflammation, transport, and signaling. It is clear that these interactions are important for the control of normal coagulation and influence the efficacy of heparin and heparin-based therapeutics. In addition to neutralization, the anticoagulant activities of GAGs may also be regulated through reduced synthesis or by degradation. In this review, we describe GAG neutralization, the proteins involved, and the molecular processes that contribute to the regulation of anticoagulant GAG activity.
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http://dx.doi.org/10.1161/ATVBAHA.118.311102DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5965931PMC
June 2018

Quantitative analysis of hydroxyapatite-binding plasma proteins in genotyped individuals with late-stage age-related macular degeneration.

Exp Eye Res 2018 07 24;172:21-29. Epub 2018 Mar 24.

School of Medicine, University of St Andrews, St Andrews, KY16 9TF, UK. Electronic address:

Age-related macular degeneration (AMD) is associated with the formation of sub-retinal pigment epithelial (RPE) deposits that block circulatory exchange with the retina. The factors that contribute to deposit formation are not well understood. Recently, we identified the presence of spherular hydroxyapatite (HAP) structures within sub-RPE deposits to which several AMD-associated proteins were bound. This suggested that protein binding to HAP represents a potential mechanism for the retention of proteins in the sub-RPE space. Here we performed quantitative proteomics using Sequential Window Acquisition of all THeoretical fragment-ion spectra-Mass Spectrometry (SWATH-MS) on plasma samples from 23 patients with late-stage neovascular AMD following HAP-binding. Individuals were genotyped for the high risk CFH variant (T1277C) and binding to HAP was compared between wild type and risk variants. From a library of 242 HAP binding plasma proteins (1% false discovery rate), SWATH-MS revealed significant quantitative differences in the abundance of 32 HAP-binding proteins (p < 0.05) between the two homozygous groups. The concentrations of six proteins (FHR1, FHR3, APOC4, C4A, C4B and PZP) in the HAP eluted fractions and whole plasma were further analysed using ELISA and their presence in sections from human cadaver eyes was examined using immunofluorescence. All six proteins were found to be present in the RPE/choroid interface, and four of these (FHR1, FHR3, APOC4 and PZP) were associated with spherules in sub-RPE space. This study provides qualitative and quantitative information relating to the degree by which plasma proteins may contribute to sub-RPE deposit formation through binding to HAP spherules and how genetic differences might contribute to deposit formation.
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http://dx.doi.org/10.1016/j.exer.2018.03.023DOI Listing
July 2018

Erythritol Attenuates Postprandial Blood Glucose by Inhibiting α-Glucosidase.

J Agric Food Chem 2018 Feb 5;66(6):1401-1407. Epub 2018 Feb 5.

Key Laboratory of Tibetan Medicine Research, Northwest Plateau Institute of Biology, Chinese Academy of Sciences , 23 Xinning Road, Xining, Qinghai 810001, People's Republic of China.

Diabetes mellitus (DM) is a serious metabolic disorder, where impaired postprandial blood glucose regulation often leads to severe health complications. The natural chemical erythritol is a C4 polyol approved by the U.S. Food and Drug Administration for use as a sweetener. Here, we examined a potential role for erythritol in the control of postprandial blood glucose levels in DM. An anti-postprandial hyperglycemia effect upon erythritol administration (500 mg kg) was demonstrated in alloxan-induced DM model mice by monitoring changes in blood glucose after intragastric administration of drugs and starch. We also found that erythritol most likely exerts its anti-postprandial hyperglycemic activities by inhibiting α-glucosidase in a competitive manner. This was supported by enzyme activity assays and molecular modeling experiments. In the latter experiments, it was possible to successfully dock erythritol into the catalytic pocket of α-glucosidase, with the resultant interaction likely driven by electrostatic interactions involving Asp215, Asp69, and Arg446 residues. This study suggests that erythritol may not only serve as a glucose substitute but also be a useful agent in the treatment of DM to help manage postprandial blood glucose levels.
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http://dx.doi.org/10.1021/acs.jafc.7b05033DOI Listing
February 2018
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