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BACKGROUND AND PURPOSE: Incidental findings on brain MR and variation of the circle of Willis (CoW) are relatively common among the general population. Ethnic differences have been described before, but few studies have explored the prevalence of incidental intracranial cerebrovascular findings and CoW variants in the setting of a single multi-ethnic cohort. The purpose of this investigation is to describe both incidental cerebrovascular findings and the morphology of the CoW on high-resolution 3T time-of-flight MR angiography (ToF MRA) in a UK tri-ethnic population-based cohort and to present updated prevalence estimates and morphologic reference values. MATERIALS AND METHODS: We studied participants from the UK Southall and Brent Revisited (SABRE) study who underwent 3T brain MRI between 2014-2018. ToF MRA images were assessed for the presence of incidental cerebrovascular imaging findings and used to determine CoW anatomy. RESULTS: 750 participants (mean age: 71.28 ± 6.46 years, range [46-90], 337 female), 322 White Europeans, 253 South Asians, and 175 African Caribbeans, were included. Incidental cerebrovascular findings were observed in 84 subjects (11.2%, 95% CI [9.0-13.7]; 38 women, 45.24%, 95% CI [34.34-56.48]), cerebral aneurysms being the most frequent, followed by intracranial arterial stenoses (ICAS) with highest prevalence among South Asians compared to White European (OR: 2.72, 95% CI [1.22-6.08], p = .015) and African Caribbean subjects (OR: 2.79, 95% CI [1.00-7.82], p = .051). Other findings included arteriovenous malformations and infundibula. The CoW was found to be more often complete in women than in men (25.22% compared to 18.41%, p = .024), and in African Caribbean (34.86%), compared to White European (19.19%), and South Asian (14.23%) subjects (p <0.001 each). CONCLUSIONS: ICAS were independently associated with ethnicity after adjusting for vascular risk factors, having the highest prevalence among South Asians. The prevalence of aneurysms was higher than in previous population-based studies. We observed anatomical differences in the CoW configuration between women, men, and ethnicities. ABBREVIATIONS: BP = Blood pressure; ICAS = Intracranial arterial stenoses; CoW = Circle of Willis; CVM = Cerebral vascular malformations; OR = Odds ratio; ToF MRA = Time-of-flight MR angiography.
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A method for structure refinement of molecules based on residual dipolar coupling (RDC) data is proposed. It calculates RDC values using magnetic-field rotational sampling of the rotational degrees of freedom of a molecule in conjunction with molecule-internal configurational sampling. By applying rotational sampling, as is occurring in the experiment, leading to observable RDCs, the method stays close to the experiment. It avoids the use of an alignment tensor and, therefore, the assumptions that the overall rotation of the molecule is decoupled from its internal motions and that the molecule is rigid. Two simple molecules, a relatively rigid and a very flexible cyclo-octane molecule with eight aliphatic side chains containing 24 united atoms, serve as so-called "toy model" test systems. The method demonstrates the influence of molecular flexibility, force-field dominance, and the number of RDC restraints available on the outcome of structure refinement based on RDCs. Magnetic-field rotational sampling is basically equivalent but more efficient than explicitly sampling the rotational degrees of freedom of the molecule. In addition, the performance of the method is less dependent on the number NRDC of measured RDC-values available. The restraining forces bias the overall orientation distribution of the molecule correctly. This study suggests that the information content of RDCs with respect to molecular structure is limited.
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In NMR experiments, residual dipolar couplings (RDCs) in a molecule can be measured by averaging the dipolar couplings (DCs) over the rotational motion of a molecule in an environment that induces a slight anisotropic orientation distribution of the molecule. Since the shape of the anisotropic distribution cannot be measured, it is standard practice to use a particular orientation distribution of the molecule with respect to the magnetic field, in the form of a so-called alignment tensor (AT), to calculate RDC-values for the molecule. Since the same alignment tensor is commonly used to calculate the different RDCs of a molecule, this approach rests on the assumption that the rotational motion of the molecule is decoupled from its internal motions and that the molecule is rigid. The validity of these two assumptions is investigated for a small, simple molecule, using a relatively rigid atomic interaction function or force field and a more flexible one. By simulating the molecule using an orientation-biasing force an anisotropic rotational distribution can be generated, for which RDCs can be obtained. Using these RDCs as target RDCs when applying one of the two approaches of structure refinement based on RDCs, it can be investigated how well the target RDCs are approximated in the RDC restraining and whether the corresponding nonuniform orientation distribution is reproduced. For the relatively rigid version of the molecule, the AT approach reproduces the target RDC-values, although the nonuniform orientation distribution of the angle θab,H between the vector râab connecting two atoms a and b in the molecule and the vector representing the direction of the magnetic field Hâ as generated in the orientation-biasing simulation cannot be reproduced in the AT RDC-restraining simulation. For the relatively flexible version of the molecule, the AT approach fails to reproduce both the target RDC values and the nonuniform orientation distribution. For biomolecules with flexible parts, the application of the AT approach is thus not recommended. Instead, a method based on sampling of the rotational and internal degrees of freedom of the molecule should be applied in molecular structure determination or refinement based on measured RDCs.
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Modelos Moleculares , Rotación , Anisotropía , Espectroscopía de Resonancia Magnética/métodosRESUMEN
During pregnancy the maternal pancreatic islets of Langerhans undergo adaptive changes to compensate for gestational insulin resistance. The lactogenic hormones are well established to play a key role in regulating the islet adaptation to pregnancy, and one of the mechanisms through which they act is through upregulating ß-cell serotonin production. During pregnancy islet serotonin levels are significantly elevated, where it is released from the ß-cells to drive the adaptive response through paracrine and autocrine effects. We have previously shown that placental kisspeptin (KP) also plays a role in promoting the elevated insulin secretion and ß-cell proliferation observed during pregnancy, although the precise mechanisms involved are unclear. In the present study we investigated the effects of KP on expression of pro-proliferative genes and serotonin biosynthesis within rodent islets. Whilst KP had limited effect on pro-proliferative gene expression at the time points tested, KP did significantly stimulate expression of the serotonin biosynthesis enzyme Tph-1. Furthermore, the islets of pregnant ß-cell-specific GPR54 knockdown mice were found to contain significantly fewer serotonin-positive ß-cells when compared to pregnant controls. Our previous studies suggested that reduced placental kisspeptin production, with consequent impaired kisspeptin-dependent ß-cell compensation, may be a factor in the development of GDM in humans. These current data suggest that, similar to the lactogenic hormones, KP may also contribute to serotonin biosynthesis and subsequent islet signalling during pregnancy. Furthermore, upregulation of serotonin biosynthesis may represent a common mechanism through which multiple signals might influence the islet adaptation to pregnancy.
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Células Secretoras de Insulina , Islotes Pancreáticos , Humanos , Embarazo , Ratones , Femenino , Animales , Kisspeptinas/metabolismo , Insulina/metabolismo , Serotonina/metabolismo , Placenta/metabolismo , Células Secretoras de Insulina/metabolismo , Islotes Pancreáticos/metabolismo , Prolactina/metabolismoRESUMEN
Imaging markers of cerebral small vessel disease provide valuable information on brain health, but their manual assessment is time-consuming and hampered by substantial intra- and interrater variability. Automated rating may benefit biomedical research, as well as clinical assessment, but diagnostic reliability of existing algorithms is unknown. Here, we present the results of the VAscular Lesions DetectiOn and Segmentation (Where is VALDO?) challenge that was run as a satellite event at the international conference on Medical Image Computing and Computer Aided Intervention (MICCAI) 2021. This challenge aimed to promote the development of methods for automated detection and segmentation of small and sparse imaging markers of cerebral small vessel disease, namely enlarged perivascular spaces (EPVS) (Task 1), cerebral microbleeds (Task 2) and lacunes of presumed vascular origin (Task 3) while leveraging weak and noisy labels. Overall, 12 teams participated in the challenge proposing solutions for one or more tasks (4 for Task 1-EPVS, 9 for Task 2-Microbleeds and 6 for Task 3-Lacunes). Multi-cohort data was used in both training and evaluation. Results showed a large variability in performance both across teams and across tasks, with promising results notably for Task 1-EPVS and Task 2-Microbleeds and not practically useful results yet for Task 3-Lacunes. It also highlighted the performance inconsistency across cases that may deter use at an individual level, while still proving useful at a population level.
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Enfermedades de los Pequeños Vasos Cerebrales , Imagen por Resonancia Magnética , Humanos , Imagen por Resonancia Magnética/métodos , Reproducibilidad de los Resultados , Enfermedades de los Pequeños Vasos Cerebrales/diagnóstico por imagen , Hemorragia Cerebral , ComputadoresRESUMEN
Phaeochromocytomas (PCC) and paragangliomas (PGL), cumulatively referred to as PPGLs, are neuroendocrine tumours arising from neural crest-derived cells in the sympathetic and parasympathetic nervous systems. Predicting future tumour behaviour and the likelihood of metastatic disease remains problematic as genotype-phenotype correlations are limited, the disease has variable penetrance and, to date, no reliable molecular, cellular or histological markers have emerged. Tumour metabolism quantification can be considered as a method to delineating tumour aggressiveness by utilising hyperpolarised 13 C-MR (HP-MR). The technique may provide an opportunity to non-invasively characterise disease behaviour. Here, we present the first instance of the analysis of PPGL metabolism via HP-MR in a single case.
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In adverse pregnancy a perturbed redox environment is associated with abnormal early-life cardiovascular development and function. Previous studies have noted alterations in the expression and/or activity of Nuclear Factor E2 Related Factor 2 (NRF2) and its antioxidant targets during human gestational diabetic (GDM) pregnancy, however to our knowledge the functional role of NRF2 in fetal 'priming' of cardiovascular dysfunction in obese and GDM pregnancy has not been investigated. Using a murine model of obesity-induced glucose dysregulated pregnancy, we demonstrate that NRF2 activation by maternal sulforaphane (SFN) supplementation normalizes NRF2-linked NQO1, GCL and CuZnSOD expression in maternal and fetal liver placental and fetal heart tissue by gestational day 17.5. Activation of NRF2 in utero in wild type but not NRF2 deficient mice improved markers of placental efficiency and partially restored fetal growth. SFN supplementation was associated with reduced markers of fetal cardiac oxidative stress, including Nox2 and 3-nitrotyrosine, as well as attenuation of cardiac mass and cardiomyocyte area in male offspring by postnatal day 52 and improved vascular function in male and female offspring by postnatal day 98. Our findings are the first to highlight the functional consequences of NRF2 modulation in utero on early-life cardiovascular function in offspring, demonstrating that activation of NRF2 affords cardiovascular protection in offspring of pregnancies affected by redox dysregulation.
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Factor 2 Relacionado con NF-E2 , Placenta , Humanos , Ratones , Masculino , Femenino , Embarazo , Animales , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Placenta/metabolismo , Oxidación-Reducción , Isotiocianatos/farmacología , Obesidad/metabolismo , Estrés Oxidativo , Miocitos Cardíacos/metabolismoRESUMEN
BACKGROUND: Different dementia support roles exist but evidence is lacking on which aspects are best, for whom, and in what circumstances, and on their associated costs and benefits. Phase 1 of the Dementia PersonAlised Care Team programme (D-PACT) developed a post-diagnostic primary care-based intervention for people with dementia and their carers and assessed the feasibility of a trial. AIM: Phase 2 of the programme aims to 1) refine the programme theory on how, when, and for whom the intervention works; and 2) evaluate its value and impact. DESIGN & SETTING: A realist longitudinal mixed-methods evaluation will be conducted in urban, rural, and coastal areas across South West and North West England where low-income or ethnic minority populations (for example, South Asian) are represented. Design was informed by patient, public, and professional stakeholder input and phase 1 findings. METHOD: High-volume qualitative and quantitative data will be collected longitudinally from people with dementia, carers, and practitioners. Analyses will comprise the following: 1) realist longitudinal case studies; 2) conversation analysis of recorded interactions; 3) statistical analyses of outcome and experience questionnaires; 4a) health economic analysis examining costs of delivery; and 4b) realist economic analysis of high-cost events and 'near misses'. All findings will be synthesised using a joint display table, evidence appraisal tool, triangulation, and stakeholder co-analysis. CONCLUSION: The realist evaluation will describe how, why, and for whom the intervention does or does not lead to change over time. It will also demonstrate how a non-randomised design can be more appropriate for complex interventions with similar questions or populations.
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BACKGROUND: Three-dimensional (3D) multiecho balanced steady-state free precession (ME-bSSFP) has previously been demonstrated in preclinical hyperpolarized (HP) 13 C-MRI in vivo experiments, and it may be suitable for clinical metabolic imaging of prostate cancer (PCa). PURPOSE: To validate a signal simulation framework for the use of sequence parameter optimization. To demonstrate the feasibility of ME-bSSFP for HP 13 C-MRI in patients. To evaluate the metabolism in PCa measured by ME-bSSFP. STUDY TYPE: Retrospective single-center cohort study. PHANTOMS/POPULATION: Phantoms containing aqueous solutions of [1-13 C] lactate (2.3 M) and [13 C] urea (8 M). Eight patients (mean age 67 ± 6 years) with biopsy-confirmed Gleason 3 + 4 (n = 7) and 4 + 3 (n = 1) PCa. FIELD STRENGTH/SEQUENCES: 1 H MRI at 3 T with T2 -weighted turbo spin-echo sequence used for spatial localization and spoiled dual gradient-echo sequence used for B0 -field measurement. ME-bSSFP sequence for 13 C MR spectroscopic imaging with retrospective multipoint IDEAL metabolite separation. ASSESSMENT: The primary endpoint was the analysis of pyruvate-to-lactate conversion in PCa and healthy prostate regions of interest (ROIs) using model-free area under the curve (AUC) ratios and a one-directional kinetic model (kP ). The secondary objectives were to investigate the correlation between simulated and experimental ME-bSSFP metabolite signals for HP 13 C-MRI parameter optimization. STATISTICAL TESTS: Pearson correlation coefficients with 95% confidence intervals and paired t-tests. The level of statistical significance was set at P < 0.05. RESULTS: Strong correlations between simulated and empirical ME-bSSFP signals were found (r > 0.96). Therefore, the simulation framework was used for sequence optimization. Whole prostate metabolic HP 13 C-MRI, observing the conversion of pyruvate into lactate, with a temporal resolution of 6 seconds was demonstrated using ME-bSSFP. Both assessed metrics resulted in significant differences between PCa (mean ± SD) (AUC = 0.33 ± 012, kP = 0.038 ± 0.014) and healthy (AUC = 0.15 ± 0.10, kP = 0.011 ± 0.007) ROIs. DATA CONCLUSION: Metabolic HP 13 C-MRI in the prostate using ME-bSSFP allows for differentiation between aggressive PCa and healthy tissue. EVIDENCE LEVEL: 2 TECHNICAL EFFICACY: Stage 1.
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Neoplasias de la Próstata , Ácido Pirúvico , Masculino , Humanos , Persona de Mediana Edad , Anciano , Ácido Pirúvico/química , Ácido Pirúvico/metabolismo , Estudios Retrospectivos , Estudios de Cohortes , Neoplasias de la Próstata/diagnóstico por imagen , Imagen por Resonancia Magnética/métodos , Ácido LácticoRESUMEN
Background and aims: Risk of stroke and dementia is markedly higher in people of South Asian and African Caribbean descent than white Europeans in the UK. This is unexplained by cardiovascular risk factors (CVRF). We hypothesized this might indicate accelerated early vascular aging (EVA) and that EVA might account for stronger associations between cerebral large artery characteristics and markers of small vessel disease. Methods: 360 participants in a tri-ethnic population-based study (120 per ethnic group) underwent cerebral and vertebral MRI. Length and median diameter of the basilar artery (BA) were derived from Time of Flight images, while white matter hyperintensities (WMH) volumes were obtained from T1 and FLAIR images. Associations between BA characteristics and CVRF were assessed using multivariable linear regression. Partial correlation coefficients between WMH load and BA characteristics were calculated after adjustment for CVRF and other potential confounders. Results: BA diameter was strongly associated with age in South Asians (+11.3 µm/year 95% CI = [3.05; 19.62]; p = 0.008), with unconvincing relationships in African Caribbeans (3.4 µm/year [-5.26, 12.12]; p = 0.436) or Europeans (2.6 µm/year [-5.75, 10.87]; p = 0.543). BA length was associated with age in South Asians (+0.34 mm/year [0.02; 0.65]; p = 0.037) and African Caribbeans (+0.39 mm/year [0.12; 0.65]; p = 0.005) but not Europeans (+0.08 mm/year [-0.26; 0.41]; p = 0.653). BA diameter (rho = 0.210; p = 0.022) and length (rho = 0.261; p = 0.004) were associated with frontal WMH load in South Asians (persisting after multivariable adjustment for CVRF). Conclusions: Compared with Europeans, the basilar artery undergoes more accelerated EVA in South Asians and in African Caribbeans, albeit to a lesser extent. Such EVA may contribute to the higher burden of CSVD observed in South Asians and excess risk of stroke, vascular cognitive impairment and dementia observed in these ethnic groups.
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Molecular dynamics (MD) simulations have been used to characterize the effects of backbone N-amination of residues in a model ß-hairpin peptide. This modification is of considerable interest as N-aminated peptides have been shown to inhibit amyloid-type aggregation. Six derivatives of the ß-hairpin peptide, which contain one, two, or four N-aminated residues, have been studied. For each peptide 100 ns MD simulations starting from the folded ß-hairpin structure were performed. The effects of the N-amination prove to be very sequence dependent. N-Amination of a residue involved in interstrand hydrogen bonding (Val3) leads to unfolding of the ß-hairpin, whereas N-amination of a residue toward the C-terminus (Leu11) gives fraying at the termini of the peptide. In the other derivatives the peptide remains folded, with increasing levels of N-amination reducing the right-handed twist of the ß-hairpin and favoring population of a type II' rather than a type I' ß-turn. MD simulations (100 ns) have also been run for each peptide starting from an unfolded extended chain. Here, the peptide with four N-aminated residues shows the most folding into the ß-hairpin (34%). Analysis of the simulations shows that N-amination favors the population of ß (φ, ψ) conformations by the preceding residue due to, at least in part, a network of weak NH2(i)-CO(i) and NH2(i)-CO(i-2) hydrogen bonds. It also leads to a reduction of misfolding because of changes in the hydrogen-bonding potential. Both of these features help funnel the peptide to the folded ß-hairpin structure. The conformational insights provided through this work give a firm foundation for the design of N-aminated peptide inhibitors for modulating protein-protein interactions and aggregation.
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Péptidos , Pliegue de Proteína , Enlace de Hidrógeno , Secuencia de Aminoácidos , Aminación , Estructura Secundaria de Proteína , Péptidos/químicaRESUMEN
A method for structure refinement of molecules based on residual dipolar coupling (RDC) data is proposed. It calculates RDC values using rotational and molecule-internal configurational sampling instead of the common refinement procedure that is based on the approximation of the nonuniform rotational distribution of the molecule by a single alignment tensor representing the average nonuniformity of this distribution. Using rotational sampling, as is occurring in the experiment leading to observable RDCs, the method stays close to the experiment. It avoids the use of an alignment tensor and thus the assumption that the overall rotation of the molecule is decoupled from its internal motions and that the molecule be rigid. Two simple molecules, two-united-atomic ethane and a cyclooctane molecule with eight side chains, containing 24 united atoms, serve as the so-called "toy model" test systems. The method demonstrates the influence of molecular flexibility and force-field deficiencies on the outcome of structure refinement based on RDCs. For a molecule of a given size (number of atoms Nat), there must be a sufficiently large number NRDC of measured RDC values available to allow the restraining forces to bias the overall orientation distribution of the molecule. If the ratio NRDC/Nat gets too small, the RDC-restraining forces will either not be strong enough to change the overall rotational direction of the molecule such that the target RDC values are approximated well or will be so strong that they induce a local deformation of the molecule. In the latter case, the size or inertia of the molecule hinders a restraining-induced overall rotation and the internal structure of the molecule is not strong enough to avoid local deformation due to the restraining forces.
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Resonancia Magnética Nuclear Biomolecular , Espectroscopía de Resonancia Magnética , Conformación Molecular , Resonancia Magnética Nuclear Biomolecular/métodosRESUMEN
INTRODUCTION: Multiparametric MRI (mpMRI) is now widely used to risk stratify men with a suspicion of prostate cancer and identify suspicious regions for biopsy. However, the technique has modest specificity and a high false-positive rate, especially in men with mpMRI scored as indeterminate (3/5) or likely (4/5) to have clinically significant cancer (csPCa) (Gleason ≥3+4). Advanced MRI techniques have emerged which seek to improve this characterisation and could predict biopsy results non-invasively. Before these techniques are translated clinically, robust histological and clinical validation is required. METHODS AND ANALYSIS: This study aims to clinically validate two advanced MRI techniques in a prospectively recruited cohort of men suspected of prostate cancer. Histological analysis of men undergoing biopsy or prostatectomy will be used for biological validation of biomarkers derived from Vascular and Extracellular Restricted Diffusion for Cytometry in Tumours and Luminal Water imaging. In particular, prostatectomy specimens will be processed using three-dimension printed patient-specific moulds to allow for accurate MRI and histology mapping. The index tests will be compared with the histological reference standard to derive false positive rate and true positive rate for men with mpMRI scores which are indeterminate (3/5) or likely (4/5) to have clinically significant prostate cancer (csPCa). Histopathological validation from both biopsy and prostatectomy samples will provide the best ground truth in validating promising MRI techniques which could predict biopsy results and help avoid unnecessary biopsies in men suspected of prostate cancer. ETHICS AND DISSEMINATION: Ethical approval was granted by the London-Queen Square Research Ethics Committee (19/LO/1803) on 23 January 2020. Results from the study will be presented at conferences and submitted to peer-reviewed journals for publication. Results will also be available on ClinicalTrials.gov. TRIAL REGISTRATION NUMBER: NCT04792138.
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Imágenes de Resonancia Magnética Multiparamétrica , Neoplasias de la Próstata , Biomarcadores , Humanos , Biopsia Guiada por Imagen , Imagen por Resonancia Magnética , Masculino , Estudios Prospectivos , Neoplasias de la Próstata/diagnóstico por imagen , Neoplasias de la Próstata/patologíaRESUMEN
In protein simulation or structure refinement based on values of observable quantities measured in (aqueous) solution, solvent (water) molecules may be explicitly treated, omitted, or represented by a potential of mean-solvation-force term, depending on protein coordinates only, in the force field used. These three approaches are compared for hen egg white lysozyme (HEWL). This 129-residue non-spherical protein contains a variety of secondary-structure elements, and ample experimental data are available: 1630 atom-atom Nuclear Overhauser Enhancement (NOE) upper distance bounds, 213 3 J-couplings and 200 S2 order parameters. These data are used to compare the performance of the three approaches. It is found that a molecular dynamics (MD) simulation in explicit water approximates the experimental data much better than stochastic dynamics (SD) simulation in vacuo without or with a solvent-accessible-surface-area (SASA) implicit-solvation term added to the force field. This is due to the missing energetic and entropic contributions and hydrogen-bonding capacities of the water molecules and the missing dielectric screening effect of this high-permittivity solvent. Omission of explicit water molecules leads to compaction of the protein, an increased internal strain, distortion of exposed loop and turn regions and excessive intra-protein hydrogen bonding. As a consequence, the conformation and dynamics of groups on the surface of the protein, which may play a key role in protein-protein interactions or ligand or substrate binding, may be incorrectly modelled. It is thus recommended to include water molecules explicitly in structure refinement of proteins in aqueous solution based on nuclear magnetic resonance (NMR) or other experimentally measured data.
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Simulación de Dinámica Molecular , Muramidasa , Simulación por Computador , Muramidasa/química , Proteínas/química , Solventes/química , AguaRESUMEN
OBJECTIVE: To develop a phantom system which can be integrated with an automated injection system, eliminating the experimental variability that arises with manual injection; for the purposes of pulse sequence testing and metric derivation in hyperpolarised 13C-MR. METHODS: The custom dynamic phantom was machined from Ultem and filled with a nicotinamide adenine dinucleotide and lactate dehydrogenase mixture dissolved in phosphate buffered saline. Hyperpolarised [1-13C]-pyruvate was then injected into the phantom (n = 8) via an automated syringe pump and the conversion of pyruvate to lactate monitored through a 13C imaging sequence. RESULTS: The phantom showed low coefficient of variation for the lactate to pyruvate peak signal heights (11.6%) and dynamic area-under curve ratios (11.0%). The variance for the lactate dehydrogenase enzyme rate constant (kP) was also seen to be low at 15.6%. CONCLUSION: The dynamic phantom demonstrates high reproducibility for quantification of 13C-hyperpolarised MR-derived metrics. Establishing such a phantom is needed to facilitate development of hyperpolarsed 13C-MR pulse sequenced; and moreover, to enable multisite hyperpolarised 13C-MR clinical trials where assessment of metric variability across sites is critical. ADVANCES IN KNOWLEDGE: The dynamic phantom developed during the course of this study will be a useful tool in testing new pulse sequences and standardisation in future hyperpolarised work.
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Imagen por Resonancia Magnética , Ácido Pirúvico , Isótopos de Carbono , Humanos , Lactato Deshidrogenasas , Ácido Láctico , Imagen por Resonancia Magnética/métodos , Espectroscopía de Resonancia Magnética/métodos , Reproducibilidad de los ResultadosRESUMEN
Sustained and localized delivery of chemotherapeutics in postoperative cancer treatment leads to a radical improvement in prognosis and a much decreased risk of tumor recurrence. In this work, polydopamine (PDA)-coated superparamagnetic iron oxide nanoparticle (SPION)-loaded polycaprolactone and poly(lactic-co-glycolic acid) fibers were developed as a potential implant to ensure safe and sustained release of the chemotherapeutic drug methotrexate (MTX), as well as provide local contrast for magnetic resonance imaging (MRI). Fibres were prepared by co-axial electrospinning and loaded with MTX-layered double hydroxide (LDH) nanocomposites in the core, yielding organic-inorganic hybrids ranging from 1.23 to 1.48 µm in diameter. After surface coating with PDA, SPIONs were subsequently loaded on the fibre surface and found to be evenly distributed, providing high MRI contrast. In vitro drug release studies showed the PDA coated fibres gave sustained release of MTX over 18 days, and the release profile is responsive to conditions representative of the tumor microenvironment such as slightly acidic pH values or elevated concentrations of the reducing agent glutathione (GSH). In vitro studies with Caco-2 and A549 cells showed highly effective killing with the PDA coated formulations, which was further enhanced at higher levels of GSH. The fibres hence have the potential to act as an implantable drug-eluting platform for the sustained release of cytotoxic agents within a tumor site, providing a novel treatment option for post-operative cancer patients.
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Nanocompuestos , Medicina de Precisión , Células CACO-2 , Humanos , Indoles , Imagen por Resonancia Magnética , Polímeros , Nanomedicina TeranósticaRESUMEN
Diabetes mellitus is characterised by hyperglycaemia, which results from an absolute or relative lack of insulin. Chronic and acute hyperglycaemia are associated with a range of health complications and an overall increased risk of mortality. Mouse models are vital in understanding the pathogenesis of this disease and its complications, as well as for developing new diabetes therapeutics. However, for experimental questions to be suitably tested, it is critical that factors inherent to the animal model are considered, as these can have profound impacts on experimental outcome, data reproducibility and robustness. In this review, we discuss key considerations relating to model choice, physiological characteristics (such as age, sex and genetic background) and husbandry practices and explore the impact of these on common experimental readouts used in preclinical diabetes research.
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Investigación Biomédica/métodos , Diabetes Mellitus Experimental/terapia , Diabetes Mellitus Tipo 2/terapia , Manejo de la Enfermedad , Resistencia a la Insulina/fisiología , Animales , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Tipo 2/genética , RatonesRESUMEN
Multiple crystal structures of the homo-trimeric protein disulphide isomerase PmScsC reveal that the peptide which links the trimerization stalk and catalytic domain can adopt helical, ß-strand and loop conformations. This region has been called a 'shape-shifter' peptide. Characterisation of this peptide using NMR experiments and MD simulations has shown that it is essentially disordered in solution. Analysis of the PmScsC crystal structures identifies the role of intermolecular contacts, within an assembly of protein molecules, in stabilising the different linker peptide conformations. These context-dependent conformational properties may be important functionally, allowing for the binding and disulphide shuffling of a variety of protein substrates to PmScsC. They also have a relevance for our understanding of protein aggregation and misfolding showing how intermolecular quaternary interactions can lead to ß-sheet formation by a sequence that in other contexts adopts a helical structure. This 'shape-shifting' peptide region within PmScsC is reminiscent of one-to-many molecular recognition features (MoRFs) found in intrinsically disordered proteins which are able to adopt different conformations when they fold upon binding to their protein partners.
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Proteína Disulfuro Isomerasas/química , Proteína Disulfuro Isomerasas/ultraestructura , Secuencia de Aminoácidos , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/ultraestructura , Dominio Catalítico , Proteínas Intrínsecamente Desordenadas/química , Proteínas Intrínsecamente Desordenadas/ultraestructura , Espectroscopía de Resonancia Magnética , Simulación de Dinámica Molecular , Péptidos , Proteína Disulfuro Isomerasas/metabolismo , Dominios Proteicos , Proteus mirabilis/enzimología , Proteus mirabilis/metabolismoRESUMEN
E. coli nitroreductase NfsB (also called NfnB) has been studied extensively, largely due to its potential for cancer gene therapy. A homodimeric flavoprotein of 216 residues, it catalyses the reduction of nitroaromatics to cytotoxic hydroxylamines by NADH and NADPH and also the reduction of quinones to hydroxyquinones. Its role in vivo is not known but it is postulated to be involved in reducing oxidative stress. The crystal structures of the wild type protein and several homologues have been determined in the absence and presence of ligands, including nicotinate as a mimic of the headpiece of the nicotinamide cofactors. There is little effect on the overall structure of the protein on binding ligands, but, from the B factors, there appears to be a decrease in mobility of 2 helices near the active site. As a first step towards examining the dynamics of the protein in solution with and without ligand, we have assigned the backbone 13C, 15N, and 1HN resonances of NfsB and examined the effect of the binding of nicotinate on the amide 15N, and 1HN shifts.
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Proteínas de Escherichia coli , Resonancia Magnética Nuclear Biomolecular , Dominio Catalítico , NitrorreductasasRESUMEN
Values of 3J-couplings as obtained from NMR experiments on proteins cannot easily be used to determine protein structure due to the difficulty of accounting for the high sensitivity of intermediate 3J-coupling values (4-8 Hz) to the averaging period that must cover the conformational variability of the torsional angle related to the 3J-coupling, and due to the difficulty of handling the multiple-valued character of the inverse Karplus relation between torsional angle and 3J-coupling. Both problems can be solved by using 3J-coupling time-averaging local-elevation restraining MD simulation. Application to the protein hen egg white lysozyme using 213 backbone and side-chain 3J-coupling restraints shows that a conformational ensemble compatible with the experimental data can be obtained using this technique, and that accounting for averaging and the ability of the algorithm to escape from local minima for the torsional angle induced by the Karplus relation, are essential for a comprehensive use of 3J-coupling data in protein structure determination.