Background: Accurate definition of the extent and severity of left-ventricular assist device (LVAD) infection may facilitate therapeutic decision making and targeted surgical intervention. Here, we explore the value of ¹⁸F-fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) for guidance of patient management. Methods: Fifty-seven LVAD-carrying patients received 85 whole-body ¹⁸F-FDG PET/CT scans for the work-up of device infection. Clinical follow-up was obtained over a period of up to two years. Results: PET/CT showed various patterns of infectious involvement of the 4 LVAD components: driveline entry point (77% of cases), subcutaneous driveline path (87%), pump pocket (49%) and outflow tract (58%). Driveline smears revealed staphylococcus or pseudomonas strains as the underlying pathogen in a majority of cases (48 and 34%, respectively). At receiver-operating characteristics analysis, an ¹⁸F-FDG standardized uptake value (SUV) >2.5 was most accurate to identify smear-positive driveline infection. Infection of 3 or all 4 LVAD components showed a trend towards lower survival vs infection of 2 or less components (P = 0.089), while involvement of thoracic lymph nodes was significantly associated with adverse outcome (P = 0.001 for nodal SUV above vs below median). Finally, patients that underwent early surgical revision within 3 months after PET/CT (n = 21) required significantly less inpatient hospital care during follow-up when compared to those receiving delayed surgical revision (n = 11; p<0.05). Conclusion: Whole-body ¹⁸F-FDG PET/CT identifies the extent of LVAD infection and predicts adverse outcome. Initial experience suggests that early image-guided surgical intervention may facilitate a less complicated subsequent course.

2-Deoxy-2-[18F]fluoro-D-glucose (2-FDG) with positron emission tomography (2-FDG-PET) is undeniably useful in the clinic, among other uses, to monitor change over time using the 2-FDG standardized uptake values (SUV) metric. This report suggests some potentially serious caveats for this and related roles for 2-FDG PET. Most critical is the assumption that there is an exact proportionality between glucose metabolism and 2-FDG metabolism, called the lumped constant, LC. This report describes that LC is not constant for a specific tissue and may be variable before and after disease treatment. The purpose of this work is not to deny the clinical value of 2-FDG PET; it is a reminder that when one extends the use of an appropriately qualified imaging method, new observations may arise and further validation would be necessary. Current understanding of glucose-based energetics in vivo is based on the quantification of glucose metabolic rates with 2-FDG PET, a method that permits the non-invasive assessment in various human disorders. However, 2-FDG is only a good substrate for facilitated-glucose transporters (GLUTs) but not for sodium-dependent glucose co-transporters (SGLTs), which have recently been shown to be distributed in multiple human tissues. Thus, the GLUT-mediated in vivo glucose utilization measured by 2-FDG PET would be blinded to the potentially substantial role of functional SGLTs in glucose transport and utilization. Therefore, in these circumstances the 2-FDG LC used to quantify in vivo glucose utilization should not be expected to remain constant. 2-FDG LC variations have been especially significant in tumors, particularly at different stages of cancer development, affecting the accuracy of quantitative glucose measures and potentially limiting the prognostic value of 2-FDG, as well as its accuracy in monitoring treatments. SGLT-mediated glucose transport can be estimated using α-methyl-4-deoxy-4-[18F]fluoro-D-glucopyranoside (Me-4FDG). Utilizing both 2-FDG and Me-4FDG should provide a more complete picture of glucose utilization via both GLUT and SGLT transporters in health and disease stages. Given the widespread use of 2-FDG PET to infer glucose metabolism, appreciating the potential limitations of 2-FDG as a surrogate for glucose metabolic rate and the potential reasons for variability in LC is relevant. Even when the readout for the 2-FDG PET study is only an SUV parameter, variability in LC is important, particularly if it changes over the course of disease progression (e.g., an evolving tumor).

Objectives: Esthesioneuroblastoma (ENB) is rare with limited therapeutic options when unresectable or metastatic; however, expression of somatostatin receptors qualifies it for peptide receptor radionuclide therapy (PRRT). We report outcomes of PRRT in ENB from two referral centers. Methods: Using PRRT databases at two European Neuroendocrine Tumour Society Centers of Excellence, case finding was undertaken between 2004-2018 for patients who had PRRT with recurrent/metastatic ENB deemed unsuitable for further conventional therapies. Evaluations of response using a composite reference standard and for survival were performed. Results: Of seven patients, four had partial response, two had disease stabilization and one had early progression. Possible side effects include worsening CSF-leaks. Median progression-free survival was 17 months (range, 0-30), and median overall survival was 32 months (range, 4–53). Conclusion: PRRT shows promising efficacy and moderate survival duration in unresectable locally advanced or metastatic ENB warranting larger cohort studies incorporating measures of quality of life.

Purpose: To determine the effect of smooth filter and partial volume correction (PVC) method on measured prostate-specific membrane antigen (PSMA) activity in small metastatic lesions and to determine the impact of these changes on the molecular imaging (mi) PSMA scoring. Materials & Methods: Men with biochemical recurrence of prostate cancer with negative CT and bone scintigraphy were referred for ¹⁸F-DCFPyL PET/CT. Examinations were performed on one of 2 PET/CT scanners (GE Discovery 610 or Siemens mCT40). All suspected tumor sites were manually contoured on co-registered CT and PET images, and each was assigned a miPSMA score as per the PROMISE criteria. The PVC factors were calculated for every lesion using the anatomical CT and then applied to the unsmoothed PET images. The miPSMA scores, with and without the corrections, were compared, and a simplified "rule of thumb" (RoT) correction factor (CF) was derived for lesions at various sizes (<4mm, 4-7mm, 7-9mm, 9-12mm). This was then applied to the original dataset and miPSMA scores obtained using the RoT CF were compared to those found using the actual corrections. Results: There were 75 men (median age, 69 years; median serum PSA of 3.69 ug/L) with 232 metastatic nodes < 12 mm in diameter (mean lesion volume of 313.5 ± 309.6 mm³). Mean SUV_max before and after correction was 11.0 ± 9.3 and 28.5 ± 22.8, respectively (p<0.00001). The mean CF for lesions <4mm (n = 22), 4-7mm (n = 140), 7-9mm (n = 50), 9-12 mm (n = 20) was 4 (range: 2.5-6.4), 2.8 (range: 1.6-4.9), 2.3 (range: 1.6-3.3) and 1.8 (range 1.4-2.4), respectively. Overall miPSMA scores were concordant between the corrected dataset and RoT in 205/232 lesions (88.4%). Conclusion: There is a significant effect of smooth filter and partial volume correction on measured PSMA activity in small nodal metastases, impacting the miPSMA score.

Latest digital whole-body PET scanners provide a combination of higher sensitivity and improved spatial and timing resolution. We performed a lesion detectability study on two generations of Siemens Biograph PET/CT scanners, the mCT and Vision, to study the impact of improved physical performance on clinical performance. Our hypothesis is that the improved performance of the Vision will result in improved lesion detectability, allowing shorter imaging times or equivalently, lower injected dose. Methods: Data were acquired with the Society of Nuclear Medicine and Molecular Imaging Clinical Trials Network torso phantom combined with a 20-cm diameter cylindrical phantom. Spherical lesions were emulated by acquiring spheres-in-air data, and combining it with the phantom data to generate combined datasets with embedded lesions of known contrast. Two sphere sizes and uptakes were used: 9.89 mm diameter spheres with 6:1 (lung) and 3:1 (cylinder) and 4.95 mm diameter spheres with 9.6:1 (lung) and 4.5:1 (cylinder) local activity concentration uptakes. Standard image reconstruction was performed: ordinary Poisson ordered subsets expectation maximization algorithm with point spread function and time-of-flight modeling and post-reconstruction smoothing with a 5 mm Gaussian filter. The Vision images were also generated without any post-reconstruction smoothing. Generalized scan statistics methodology was used to estimate the area under the localization receiver operating characteristic curve (ALROC). Results: Higher sensitivity and improved TOF performance of Vision leads to reduced contrast in the background noise nodule distribution. Measured lesion contrast is also higher on the Vision due to its improved spatial resolution. Hence, the ALROC values are noticeably higher for the Vision relative to the mCT. Conclusion: Improved overall performance of the Vision provides a factor of 4-6 reduction in imaging time (or injected dose) over the mCT when using the ALROC metric for lesions >9.89 mm in diameter. Smaller lesions are barely detected in the mCT, leading to even higher ALROC gains with the Vision. Improved spatial resolution of the Vision also leads to a higher measured contrast that is closer to the real uptake, implying improved quantification. Post-reconstruction smoothing, however, reduces this improvement in measured contrast, thereby reducing the ALROC values for small, high uptake lesions.

The purpose of this study was to establish the dose-response relationship of selective internal radiation therapy (SIRT) in patients with metastatic colorectal cancer (mCRC), when informed by radiobiological sensitivity parameters derived from mCRC cell lines exposed to yttrium-90 (⁹⁰Y). Methods: 23 mCRC patients with liver metastases refractory to chemotherapy were included. ⁹⁰Y bremsstrahlung SPECT images were transformed into dose maps assuming the local dose deposition method. Baseline and follow-up CT scans were segmented to derive liver and tumor volumes. Mean, median, and D₇₀ (minimum dose to 70% of tumor volume) values determined from dose maps were correlated with change in tumor volume and vRECIST response using linear and logistic regression, respectively. Radiosensitivity parameters determined by clonogenic assays of mCRC cell lines HT-29 and DLD-1 after exposure to ⁹⁰Y or external beam radiotherapy (EBRT; 6MV photons) were used in biological effective dose (BED) calculations. Results: Mean administered radioactivity was 1469±428 MBq (847-2185 MBq), achieving a mean radiation absorbed tumor dose of 35.5±9.4 Gy and mean normal liver dose of 26.4±6.8 Gy. A 1.0 Gy increase in mean, median, and D₇₀ absorbed dose was associated with reduction in tumor volume of 1.8%, 1.8%, and 1.5%, respectively, and increased probability of vRECIST response (odds ratio: 1.09, 1.09, and 1.10 respectively). Threshold mean, median and D₇₀ doses for response were 48.3, 48.8, and 41.8 Gy respectively. EBRT-equivalent BEDs for ⁹⁰Y are up to 50% smaller than those calculated by applying protraction-corrected radiobiological parameters derived from EBRT alone. Conclusion: Dosimetric studies have assumed equivalence between ⁹⁰Y SIRT and EBRT, leading to inflation of BED for SIRT and possible under-treatment. Radiobiological parameters for ⁹⁰Y were applied to a BED model, providing a calculation method that has the potential to improve assessment of tumor control.

It was hypothesized that the brain β-amyloid buildup curve plateaus at an early symptomatic Alzheimer's disease (AD) stage. Atrophy-related partial volume effects (PVEs) degrade signal in hot-spot imaging techniques, such as amyloid positron emission tomography (PET). This longitudinal analysis of amyloid-sensitive PET data investigated the shape of the β-amyloid curve in AD applying PVE correction (PVEC). We analyzed baseline and 2-year follow-up data of 216 symptomatic individuals on the AD continuum (positive amyloid status) enrolled in Alzheimer's Disease Neuroimaging Initiative (17 AD dementia, 199 mild cognitive impairment), including 18F-florbetapir PET, magnetic resonance imaging and mini mental state examination (MMSE) scores. For PVEC, the modified Müller-Gärtner method was performed. Compared to non-PVE-corrected data, PVE-corrected data yielded significantly higher regional and composite standardized uptake value ratio (SUVR) changes over time (P=0.0002 for composite SUVRs). Longitudinal SUVR changes in relation to MMSE decreases showed a significantly higher slope of the regression line in the PVE-corrected as compared to the non-PVE-corrected PET data (F=7.1, P=0.008). These PVEC results indicate that the β-amyloid buildup curve does not plateau at an early symptomatic disease stage. A further evaluation of the impact of PVEC on the in-vivo characterization of time-dependent AD pathology, including the reliable assessment and comparison of other amyloid tracers, is warranted.

The prostate-specific membrane antigen (PSMA) is an excellent target for theranostic applications in prostate cancer (PCa). However, PSMA-targeted radioligand therapy can cause undesirable effects due to high accumulation of PSMA radiotracers in salivary glands and kidneys. This study assessed orally administered monosodium glutamate (MSG) as a potential means of reducing kidney and salivary gland radiation exposure using a PSMA targeting radiotracer. Methods: This prospective, double-blind, placebo-controlled study enrolled 10 biochemically recurrent PCa patients. Each subject served as his own control. [¹⁸F]DCFPyl PET/CT imaging sessions were performed 3 – 7 days apart, following oral administration of either 12.7 g of MSG or placebo. Data from the two sets of images were analyzed by placing regions of interest on lacrimal, parotid and submandibular glands, left ventricle, liver, spleen, kidneys, bowel, urinary bladder, gluteus muscle and malignant lesions. The results from MSG and placebo scans were compared by paired analysis of the ROI data. Results: A total of 142 pathological lesions along with normal tissues were analyzed. As hypothesized a priori, there was a significant decrease in maximal standardized uptake values corrected for lean body mass (SULmax) on images obtained following MSG administration in the parotids (24 ± 14%, P = 0.001), submandibular glands (35 ± 11%, P<0.001) and kidneys (23 ± 26%, P = 0.014). Significant decreases were also observed in lacrimal glands (49 ± 13%, P<0.001), liver (15 ± 6%, P<0.001), spleen (28 ± 13%, P = 0.001) and bowel (44 ± 13%, P<0.001). Mildly lower blood pool SULmean was observed after MSG administration (decrease of 11 ± 13%, P = 0.021). However, significantly lower radiotracer uptake in terms of SULmean, SULpeak, and SULmax was observed in malignant lesions on scans performed after MSG administration compared to the placebo studies (SULmax median decrease 33%, range -1 to 75%, P<0.001). No significant adverse events occurred and vital signs were stable following placebo or MSG administration. Conclusion: Orally administered MSG significantly decreased salivary gland, kidney and other normal organ PSMA radiotracer uptake in human subjects, using [¹⁸F]DCFPyL as an exemplar. However, MSG caused a corresponding reduction in tumor uptake, which may limit the benefits of this approach for diagnostic and therapeutic applications.

31 October 2019

Trade between Central and Eastern Europe and sub-Saharan Africa has increased significantly in the last decade and a half. There is a strong case to be made for greater economic re-engagement, especially in terms of investment, that has the potential to support inclusive growth in both regions.

Invitation Only Research Event

Event participants

Professor Nick Binedell, Founding Director and Sasol Chair of Strategic Management, Gordon Institute of Business Science (GIBS), University of Pretoria

Research Event

Event participants

Hon. Manuel José Nunes Júnior, Minister of State for Economic Coordination, Republic of Angola
Chair: Dr Alex Vines OBE, Managing Director, Ethics, Risk & Resilience; Director, Africa Programme, Chatham House

Research Event

Event participants

Tutu Alicante, Executive Director, EG Justice
Chair: Dr Alex Vines OBE, Managing Director, Ethics, Risk & Resilience; Director, Africa Programme, Chatham House

Research Event

Event participants

Paul Mashatile, Treasurer General, African National Congress (ANC)

Recurrent urinary tract infections (UTIs) pose a significant burden on the health care system. Underlying mechanisms predisposing children to UTIs and associated changes in the urinary proteome are not well understood. We aimed to investigate the urinary proteome of a subset of children who have vesicoureteral reflux (VUR) and recurrent UTIs because of their risk of developing infection-related renal damage. Improving diagnostic modalities to identify UTI risk factors would significantly alter the clinical management of children with VUR. We profiled the urinary proteomes of 22 VUR patients with low grade VUR (1–3 out of 5), a history of recurrent UTIs, and renal scarring, comparing them to those obtained from 22 age-matched controls. Urinary proteins were analyzed by mass spectrometry followed by protein quantitation based on spectral counting. Of the 2,551 proteins identified across both cohorts, 964 were robustly quantified, as defined by meeting criteria with spectral count (SC) ≥2 in at least 7 patients in either VUR or control cohort. Eighty proteins had differential expression between the two cohorts, with 44 proteins significantly up-regulated and 36 downregulated (q <0.075, FC ≥1.2). Urinary proteins involved in inflammation, acute phase response (APR), modulation of extracellular matrix (ECM), and carbohydrate metabolism were altered among the study cohort.

Male infertility is widespread and estimated to affect 1 in 20 men. Although in some cases the etiology of the condition is well understood, for at least 50% of men, the underlying cause is yet to be classified. Male infertility, or subfertility, is often diagnosed by looking at total sperm produced, motility of the cells and overall morphology. Although counting spermatozoa and their associated motility is routine, morphology assessment is highly subjective, mainly because of the procedure being based on microscopic examination. A failure to diagnose male-infertility or sub-fertility has led to a situation where assisted conception is often used unnecessarily. As such, biomarkers of male infertility are needed to help establish a more consistent diagnosis. In the present study, we compared nuclear extracts from both high- and low-quality spermatozoa by LC-MS/MS based proteomic analysis. Our data shows that nuclear retention of specific proteins is a common facet among low-quality sperm cells. We demonstrate that the presence of Topoisomerase 2A in the sperm head is highly correlated to poor head morphology. Topoisomerase 2A is therefore a potential new biomarker for confirming male infertility in clinical practice.

The peroxisome is a subcellular organelle that functions in essential metabolic pathways, including biosynthesis of plasmalogens, fatty acid β-oxidation of very-long-chain fatty acids, and degradation of hydrogen peroxide. Peroxisome biogenesis disorders (PBDs) manifest as severe dysfunction in multiple organs, including the central nervous system (CNS), but the pathogenic mechanisms in PBDs are largely unknown. Because CNS integrity is coordinately established and maintained by neural cell interactions, we here investigated whether cell-cell communication is impaired and responsible for the neurological defects associated with PBDs. Results from a noncontact co-culture system consisting of primary hippocampal neurons with glial cells revealed that a peroxisome-deficient astrocytic cell line secretes increased levels of brain-derived neurotrophic factor (BDNF), resulting in axonal branching of the neurons. Of note, the BDNF expression in astrocytes was not affected by defects in plasmalogen biosynthesis and peroxisomal fatty acid β-oxidation in the astrocytes. Instead, we found that cytosolic reductive states caused by a mislocalized catalase in the peroxisome-deficient cells induce the elevation in BDNF secretion. Our results suggest that peroxisome deficiency dysregulates neuronal axogenesis by causing a cytosolic reductive state in astrocytes. We conclude that astrocytic peroxisomes regulate BDNF expression and thereby support neuronal integrity and function.

Coenzyme Q (Qn) is a vital lipid component of the electron transport chain that functions in cellular energy metabolism and as a membrane antioxidant. In the yeast Saccharomyces cerevisiae, coq1–coq9 deletion mutants are respiratory-incompetent, sensitive to lipid peroxidation stress, and unable to synthesize Q6. The yeast coq10 deletion mutant is also respiratory-deficient and sensitive to lipid peroxidation, yet it continues to produce Q6 at an impaired rate. Thus, Coq10 is required for the function of Q6 in respiration and as an antioxidant and is believed to chaperone Q6 from its site of synthesis to the respiratory complexes. In several fungi, Coq10 is encoded as a fusion polypeptide with Coq11, a recently identified protein of unknown function required for efficient Q6 biosynthesis. Because “fused” proteins are often involved in similar biochemical pathways, here we examined the putative functional relationship between Coq10 and Coq11 in yeast. We used plate growth and Seahorse assays and LC-MS/MS analysis to show that COQ11 deletion rescues respiratory deficiency, sensitivity to lipid peroxidation, and decreased Q6 biosynthesis of the coq10Δ mutant. Additionally, immunoblotting indicated that yeast coq11Δ mutants accumulate increased amounts of certain Coq polypeptides and display a stabilized CoQ synthome. These effects suggest that Coq11 modulates Q6 biosynthesis and that its absence increases mitochondrial Q6 content in the coq10Δcoq11Δ double mutant. This augmented mitochondrial Q6 content counteracts the respiratory deficiency and lipid peroxidation sensitivity phenotypes of the coq10Δ mutant. This study further clarifies the intricate connection between Q6 biosynthesis, trafficking, and function in mitochondrial metabolism.

The millions of customers, the short deadline: how the public service's digital revolution will start.

Kotaro Hama
Apr 1, 2020; 61:523-536
Patient-Oriented and Epidemiological Research

Dong-Jae Jun
May 1, 2020; 61:746-757
Research Articles

Yipeng Sui
May 1, 2020; 61:696-706
Research Articles

M John Chapman
Apr 15, 2020; 0:jlr.P119000543v1-jlr.P119000543
Patient-Oriented and Epidemiological Research

Sookyoung Jeon
Apr 1, 2020; 61:470-479
Reviews

Once more for 2017 as Henry Burrell, Karen Khan and Scott Carey bid farewell to this wonderful year (ahem) with musings on Apple's sexy new iMac Pro. Who is it for, how much is it and does this mean there is no Mac Pro in 2018?

We then tackle Netflix's Twitter shaming of its users and why Spotify got away with it earlier in the year. How comfortable are we all when we realise how much data companies really have on us?

In light of this, Apple bought Shazam - most likely for the data sets as much as the tech and the talent. What form will it take in Apple as another UK tech company is acquired?

See acast.com/privacy for privacy and opt-out information.

Atherogenic LDL particles are physicochemically and metabolically heterogeneous. Can bioactive lipid cargo differentiate LDL subclasses, and thus potential atherogenicity?  What is the effect of statin treatment? Obese, hypertriglyceridemic, hypercholesterolemic males (n=12; Lp(a) <10 mg/dL) received pitavastatin calcium (4mg/day) for 180 days in a single-phase, unblinded study. The lipidomic profiles (23 lipid classes) of five LDL subclasses fractionated from baseline and post-statin plasmas were determined by LC-MS. At baseline and on statin treatment, very small dense LDL (LDL5) was preferentially enriched (up to 3-fold) in specific lysophospholipids (lysophosphatidylcholine (LPC); lysophosphatidylinositol (LPI); lyso-platelet activating factor (LPC(O)); 9,0.2 and 0.14 mol/mol apoB respectively; all p<0.001 versus LDL1-4), suggesting  elevated inflammatory potential per particle. In contrast, lysophosphatidylethanolamine was uniformly distributed among LDL subclasses. Statin treatment markedly reduced absolute plasma concentrations of all LDL subclasses (up to 33.5%), including LPC, LPI and LPC(O) contents (up to -52%), consistent with reduction in cardiovascular risk. Despite such reductions, lipotoxic ceramide load per particle in LDL1-5 (1.5 - 3 mol/mol apoB; 3 - 7 mmol/mol phosphatidylcholine) was either conserved or elevated. Bioactive lipids may constitute biomarkers for the cardiometabolic risk associated with specific LDL subclasses in atherogenic dyslipidemia at baseline, and with residual risk on statin therapy.

31 March 2020

6 April 2020

16 April 2020

Genetic and genomic research has greatly advanced our understanding of heart disease. Yet, comprehensive, in-depth, quantitative maps of protein expression in hearts of living humans are still lacking. Using samples obtained during valve replacement surgery in patients with mitral valve prolapse (MVP), we set out to define inter-chamber differences, the intersect of proteomic data with genetic or genomic datasets, and the impact of left atrial dilation on the proteome of patients with no history of atrial fibrillation (AF).  We collected biopsies from right atria (RA), left atria (LA) and left ventricle (LV) of seven male patients with mitral valve regurgitation with dilated LA but no history of AF. Biopsy samples were analyzed by high-resolution mass spectrometry (MS), where peptides were pre-fractionated by reverse phase high-pressure liquid chromatography prior to MS measurement on a Q-Exactive-HF Orbitrap instrument. We identified 7,314 proteins based on 130,728 peptides. Results were confirmed in an independent set of biopsies collected from three additional individuals. Comparative analysis against data from post-mortem samples showed enhanced quantitative power and confidence level in samples collected from living hearts. Our analysis, combined with data from genome wide association studies suggested candidate gene associations to MVP, identified higher abundance in ventricle for proteins associated with cardiomyopathies and revealed the dilated LA proteome, demonstrating differential representation of molecules previously associated with AF, in non-AF hearts. This is the largest dataset of cardiac protein expression from human samples collected in vivo. It provides a comprehensive resource that allows insight into molecular fingerprints of MVP and facilitates novel inferences between genomic data and disease mechanisms. We propose that over-representation of proteins in ventricle is consequent not to redundancy but to functional need, and conclude that changes in abundance of proteins known to associate with AF are not sufficient for arrhythmogenesis.

Protein cross-linking and the analysis of cross-linked peptides by mass spectrometry is currently receiving much attention. Not only is this approach applied to isolated complexes to provide information about spatial arrangements of proteins but it is also increasingly applied to entire cells and their organelles. As in quantitative proteomics, the application of isotopic labelling further makes it possible to monitor quantitative changes in the protein-protein interactions between different states of a system. Here, we cross-linked mitochondria from Saccharomyces cerevisiae grown on either glycerol- or glucose-containing medium to monitor protein-protein interactions under non-fermentative and fermentative conditions. We investigated qualitatively the protein-protein interactions of the 400 most abundant proteins applying stringent data-filtering criteria, i.e. a minimum of two cross-linked peptide spectrum matches and a cut-off in the spectrum scoring of the used search engine. The cross-linker BS3 proved to be equally suited for connecting proteins in all compartments of mitochondria when compared with its water-insoluble but membrane-permeable derivative DSS. We also applied quantitative cross-linking to mitochondria of both the growth conditions using stable-isotope labelled BS3. Significant differences of cross-linked proteins under glycerol and glucose conditions were detected, however, mainly due to the different copy numbers of these proteins in mitochondria under both the conditions. Results obtained from the glycerol condition indicate that the internal NADH:ubiquinone oxidoreductase Ndi1 is part of an electron transport chain supercomplex. We have also detected several hitherto uncharacterized proteins and identified their interaction partners. Among those, Min8 was found to be associated with cytochrome c oxidase. BN-PAGE analyses of min8 mitochondria suggest that Min8 promotes the incorporation of Cox12 into cytochrome c oxidase.

27 November 2019

16 April 2020

Invitation Only Research Event

Ceramides (Cers) with ultralong (~32-carbon) chains and -esterified linoleic acid, composing a subclass called omega-O-acylceramides (acylCers), are indispensable components of the skin barrier. Normal barriers typically contain acylCer concentrations of ~10 mol%; diminished concentrations, along with altered or missing long periodicity lamellar phase (LPP), and increased permeability accompany an array of skin disorders, including atopic dermatitis, psoriasis, and ichthyoses. We developed model membranes to investigate the effects of the acylCer structure and concentration on skin lipid organization and permeability. The model membrane systems contained six to nine Cer subclasses as well as fatty acids, cholesterol, and cholesterol sulfate; acylCer content—namely, acylCers containing sphingosine (Cer EOS), dihydrosphingosine (Cer EOdS), and phytosphingosine (Cer EOP) ranged from zero to 30 mol%. Systems with normal physiologic concentrations of acylCer mixture mimicked the permeability and nanostructure of human skin lipids (with regard to LPP, chain order, and lateral packing). The models also showed that the sphingoid base in acylCer significantly affects the membrane architecture and permeability and that Cer EOP, notably, is a weaker barrier component than Cer EOS and Cer EOdS. Membranes with diminished or missing acylCers displayed some of the hallmarks of diseased skin lipid barriers (i.e., lack of LPP, less ordered lipids, less orthorhombic chain packing, and increased permeability). These results could inform the rational design of new and improved strategies for the barrier-targeted treatment of skin diseases.

GPR120 is implicated as a lipid receptor in the oro-sensory detection of dietary fatty acids. However, the effects of GPR120 activation on dietary fat intake or obesity are not clearly understood. We investigated to determine whether the binding of TUG891, a novel GPR120 agonist, to lingual GPR120 modulates fat preference in mice. We explored the effects of TUG891 on obesity-related hormones and conducted behavioral choice tests on mice to better understand the physiologic relevance of the action of TUG891. In cultured mouse and human taste bud cells (TBCs), TUG891 induced a rapid increase in Ca²⁺ by acting on GPR120. A long-chain dietary fatty acid, linoleic acid (LA), also recruited Ca²⁺ via GPR120 in human and mouse TBCs. Both TUG891 and LA induced ERK1/2 phosphorylation and enhanced in vitro release of glucagon-like peptide-1 from cultured human and mouse TBCs. In situ application of TUG891 onto the tongue of anesthetized mice triggered the secretion of pancreatobiliary juice, probably via the tongue-brain-gut axis. Furthermore, lingual application of TUG891 altered circulating concentrations of cholecystokinin and adipokines, associated with decreased circulating LDL, in conscious mice. In behavioral tests, mice exhibited a spontaneous preference for solutions containing either TUG891 or LA instead of a control. However, addition of TUG891 to a solution containing LA significantly curtailed fatty acid preference. Our study demonstrates that TUG891 binds to lingual GPR120 receptors, activates the tongue-brain-gut axis, and modulates fat preference. These findings may support the development of new fat taste analogs that can change the approach to obesity prevention and treatment.

Whether HDL is associated with dementia risk is unclear. In addition to apoA1, other apolipoproteins are found in HDL, creating subspecies of HDL that may have distinct metabolic properties. We measured apoA1, apoC3, and apoJ levels in plasma and apoA1 levels in HDL that contains or lacks apoE, apoJ, or apoC3 using a modified sandwich ELISA in a case-cohort study nested within the Ginkgo Evaluation of Memory Study. We included 995 randomly selected participants and 521 participants who developed dementia during a mean of 5.1 years of follow-up. The level of total apoA1 was not significantly related to dementia risk, regardless of the coexistence of apoC3, apoJ, or apoE. Higher levels of total plasma apoC3 were associated with better cognitive function at baseline (difference in Modified Mini-Mental State Examination scores tertile 3 vs. tertile 1: 0.60; 95% CI: 0.23, 0.98) and a lower dementia risk (adjusted hazard ratio tertile 3 vs. tertile 1: 0.73; 95% CI: 0.55, 0.96). Plasma concentrations of apoA1 in HDL and its apolipoprotein-defined subspecies were not associated with cognitive function at baseline or with the risk of dementia during follow-up. Similar studies in other populations are required to better understand the association between apoC3 and Alzheimer’s disease pathology.

Plasma lipoprotein (a) [Lp(a)] levels are largely determined by variation in the LPA gene, which codes for apo(a). Genome-wide association studies (GWASs) have identified nonsynonymous variants in LPA that associate with low Lp(a) levels, although their effect on apo(a) function is unknown. We investigated two such variants, R990Q and R1771C, which were present in four null Lp(a) individuals, for structural and functional effects. Sequence alignments showed the R990 and R1771 residues to be highly conserved and homologous to each other and to residues associated with plasminogen deficiency. Structural modeling showed both residues to make several polar contacts with neighboring residues that would be ablated on substitution. Recombinant expression of the WT and R1771C apo(a) in liver and kidney cells showed an abundance of an immature form for both apo(a) proteins. A mature form of apo(a) was only seen with the WT protein. Imaging of the recombinant apo(a) proteins in conjunction with markers of the secretory pathway indicated a poor transit of R1771C into the Golgi. Furthermore, the R1771C mutant displayed a glycosylation pattern consistent with ER, but not Golgi, glycosylation. We conclude that R1771 and the equivalent R990 residue facilitate correct folding of the apo(a) kringle structure and mutations at these positions prevent the proper folding required for full maturation and secretion. To our knowledge, this is the first example of nonsynonymous variants in LPA being causative of a null Lp(a) phenotype.

Zinc metallopeptidase STE24 (ZMPSTE24) is essential for the conversion of farnesyl–prelamin A to mature lamin A, a key component of the nuclear lamina. In the absence of ZMPSTE24, farnesyl–prelamin A accumulates in the nucleus and exerts toxicity, causing a variety of disease phenotypes. By ~4 months of age, both male and female Zmpste24^–/– mice manifest a near-complete loss of adipose tissue, but it has never been clear whether this phenotype is a direct consequence of farnesyl–prelamin A toxicity in adipocytes. To address this question, we generated a conditional knockout Zmpste24 allele and used it to create adipocyte-specific Zmpste24–knockout mice. To boost farnesyl–prelamin A levels, we bred in the "prelamin A–only" Lmna allele. Gene expression, immunoblotting, and immunohistochemistry experiments revealed that adipose tissue in these mice had decreased Zmpste24 expression along with strikingly increased accumulation of prelamin A. In male mice, Zmpste24 deficiency in adipocytes was accompanied by modest changes in adipose stores (an 11% decrease in body weight, a 23% decrease in body fat mass, and significantly smaller gonadal and inguinal white adipose depots). No changes in adipose stores were detected in female mice, likely because prelamin A expression in adipose tissue is lower in female mice. Zmpste24 deficiency in adipocytes did not alter the number of macrophages in adipose tissue, nor did it alter plasma levels of glucose, triglycerides, or fatty acids. We conclude that ZMPSTE24 deficiency in adipocytes, and the accompanying accumulation of farnesyl–prelamin A, reduces adipose tissue stores, but only modestly and only in male mice.

Niemann-Pick disease type C (NPC) disease is a lipid-storage disorder that is caused by mutations in the genes encoding NPC proteins and results in lysosomal cholesterol accumulation. 2-Hydroxypropyl-β-cyclodextrin (CD) has been shown to reduce lysosomal cholesterol levels and enhance sterol homeostatic responses, but CD’s mechanism of action remains unknown. Recent work provides evidence that CD stimulates lysosomal exocytosis, raising the possibility that lysosomal cholesterol is released in exosomes. However, therapeutic concentrations of CD do not alter total cellular cholesterol, and cholesterol homeostatic responses at the ER are most consistent with increased ER membrane cholesterol. To address these disparate findings, here we used stable isotope labeling to track the movement of lipoprotein cholesterol cargo in response to CD in NPC1-deficient U2OS cells. Although released cholesterol was detectable, it was not associated with extracellular vesicles. Rather, we demonstrate that lysosomal cholesterol trafficks to the plasma membrane (PM), where it exchanges with lipoprotein-bound cholesterol in a CD-dependent manner. We found that in the absence of suitable extracellular cholesterol acceptors, cholesterol exchange is abrogated, cholesterol accumulates in the PM, and reesterification at the ER is increased. These results support a model in which CD promotes intracellular redistribution of lysosomal cholesterol, but not cholesterol exocytosis or efflux, during the restoration of cholesterol homeostatic responses.

Bile acids (BAs) facilitate intestinal absorption of lipid-soluble nutrients and modulate various metabolic pathways through the farnesoid X receptor (FXR) and Takeda G-protein-coupled receptor 5. These receptors are targets for therapy in cholestatic and metabolic diseases. However, dissimilarities in BA metabolism between humans and mice complicate translation of preclinical data. Cytochrome P450 family 2 subfamily c polypeptide 70 (CYP2C70) was recently proposed to catalyze the formation of rodent-specific muricholic acids (MCAs). With CRISPR/Cas9-mediated somatic genome editing, we generated an acute hepatic Cyp2c70 knockout mouse model (Cyp2c70^ako) to clarify the role of CYP2C70 in BA metabolism in vivo and evaluate whether its activity modulates effects of pharmacologic FXR activation on cholesterol homeostasis. In Cyp2c70^ako mice, chenodeoxycholic acid (CDCA) increased at the expense of βMCA, resulting in a more hydrophobic human-like BA pool. Tracer studies demonstrated that, in vivo, CYP2C70 catalyzes the formation of βMCA primarily by sequential 6β-hydroxylation and C7-epimerization of CDCA, generating αMCA as an intermediate metabolite. Physiologically, the humanized BA composition in Cyp2c70^ako mice blunted the stimulation of fecal cholesterol disposal in response to FXR activation compared with WT mice, predominantly due to reduced stimulation of transintestinal cholesterol excretion. Thus, deletion of hepatic Cyp2c70 in adult mice translates into a human-like BA pool composition and impacts the response to pharmacologic FXR activation. This Cyp2c70^ako mouse model may be a useful tool for future studies of BA signaling and metabolism that informs human disease development and treatment.

X-linked adrenoleukodystrophy (X-ALD) is an inherited disorder caused by deleterious mutations in the ABCD1 gene. The ABCD1 protein transports very long-chain FAs (VLCFAs) from the cytosol into the peroxisome where the VLCFAs are degraded through β-oxidation. ABCD1 dysfunction leads to VLCFA accumulation in individuals with X-ALD. FAs are activated by esterification to CoA before metabolic utilization. However, the intracellular pools and metabolic profiles of individual acyl-CoA esters have not been fully analyzed. In this study, we profiled the acyl-CoA species in fibroblasts from X-ALD patients and in ABCD1-deficient HeLa cells. We found that hexacosenoyl (26:1)-CoA, but not hexacosanoyl (26:0)-CoA, was the most abundantly concentrated among the VLCFA-CoA species in these cells. We also show that 26:1-CoA is mainly synthesized from oleoyl-CoA, and the metabolic turnover rate of 26:1-CoA was almost identical to that of oleoyl-CoA in both WT and ABCD1-deficient HeLa cells. The findings of our study provide precise quantitative and metabolic information of each acyl-CoA species in living cells. Our results suggest that VLCFA is endogenously synthesized as VLCFA-CoA through a FA elongation pathway and is then efficiently converted to other metabolites, such as phospholipids, in the absence of ABCD1.

Alcoholic liver disease (ALD) is the most prevalent type of chronic liver disease with significant morbidity and mortality worldwide. ALD begins with simple hepatic steatosis and progresses to alcoholic steatohepatitis, fibrosis, and cirrhosis. The severity of hepatic steatosis is highly associated with the development of later stages of ALD. This review explores the disturbances of alcohol-induced hepatic lipid metabolism through altered hepatic lipid uptake, de novo lipid synthesis, fatty acid oxidation, hepatic lipid export, and lipid droplet formation and catabolism. In addition, we review emerging data on the contributions of genetics and bioactive lipid metabolism in alcohol-induced hepatic lipid accumulation.

The autosomal dominant disorder Schnyder corneal dystrophy (SCD) is caused by mutations in UbiA prenyltransferase domain-containing protein-1 (UBIAD1), which uses geranylgeranyl pyrophosphate (GGpp) to synthesize the vitamin K₂ subtype menaquinone-4 (MK-4). SCD is characterized by opacification of the cornea, owing to aberrant build-up of cholesterol in the tissue. We previously discovered that sterols stimulate association of UBIAD1 with ER-localized HMG-CoA reductase, which catalyzes a rate-limiting step in the synthesis of cholesterol and nonsterol isoprenoids, including GGpp. Binding to UBIAD1 inhibits sterol-accelerated ER-associated degradation (ERAD) of reductase and permits continued synthesis of GGpp in cholesterol-replete cells. GGpp disrupts UBIAD1-reductase binding and thereby allows for maximal ERAD of reductase as well as ER-to-Golgi translocation of UBIAD1. SCD-associated UBIAD1 is refractory to GGpp-mediated dissociation from reductase and remains sequestered in the ER to inhibit ERAD. Here, we report development of a biochemical assay for UBIAD1-mediated synthesis of MK-4 in isolated membranes and intact cells. Using this assay, we compared enzymatic activity of WT UBIAD1 with that of SCD-associated variants. Our studies revealed that SCD-associated UBIAD1 exhibited reduced MK-4 synthetic activity, which may result from its reduced affinity for GGpp. Sequestration in the ER protects SCD-associated UBIAD1 from autophagy and allows intracellular accumulation of the mutant protein, which amplifies the inhibitory effect on reductase ERAD. These findings have important implications not only for the understanding of SCD etiology but also for the efficacy of cholesterol-lowering statin therapy, which becomes limited, in part, because of UBIAD1-mediated inhibition of reductase ERAD.

The pregnane X receptor (PXR) is a nuclear receptor that can be activated by numerous drugs and xenobiotic chemicals. PXR thereby functions as a xenobiotic sensor to coordinately regulate host responses to xenobiotics by transcriptionally regulating many genes involved in xenobiotic metabolism. We have previously reported that PXR has pro-atherogenic effects in animal models, but how PXR contributes to atherosclerosis development in different tissues or cell types remains elusive. In this study, we generated an LDL receptor-deficient mouse model with myeloid-specific PXR deficiency (PXR^MyeLDLR^–/–) to elucidate the role of macrophage PXR signaling in atherogenesis. The myeloid PXR deficiency did not affect metabolic phenotypes and plasma lipid profiles, but PXR^MyeLDLR^–/– mice had significantly decreased atherosclerosis at both aortic root and brachiocephalic arteries compared with control littermates. Interestingly, the PXR deletion did not affect macrophage adhesion and migration properties, but reduced lipid accumulation and foam cell formation in the macrophages. PXR deficiency also led to decreased expression of the scavenger receptor CD36 and impaired lipid uptake in macrophages of the PXR^MyeLDLR^–/– mice. Further, RNA-Seq analysis indicated that treatment with a prototypical PXR ligand affects the expression of many atherosclerosis-related genes in macrophages in vitro. These findings reveal a pivotal role of myeloid PXR signaling in atherosclerosis development and suggest that PXR may be a potential therapeutic target in atherosclerosis management.

Lipid rafts are highly ordered regions of the plasma membrane that are enriched in cholesterol and sphingolipids and play important roles in many cells. In hematopoietic stem and progenitor cells (HSPCs), lipid rafts house receptors critical for normal hematopoiesis. Lipid rafts also can bind and sequester kinases that induce negative feedback pathways to limit proliferative cytokine receptor cycling back to the cell membrane. Modulation of lipid rafts occurs through an array of mechanisms, with optimal cholesterol efflux one of the major regulators. As such, cholesterol homeostasis also regulates hematopoiesis. Increased lipid raft content, which occurs in response to changes in cholesterol efflux in the membrane, can result in prolonged receptor occupancy in the cell membrane and enhanced signaling. In addition, certain diseases, like diabetes, may contribute to lipid raft formation and affect cholesterol retention in rafts. In this review, we explore the role of lipid raft-related mechanisms in hematopoiesis and CVD (specifically, atherosclerosis) and discuss how defective cholesterol efflux pathways in HSPCs contribute to expansion of lipid rafts, thereby promoting myelopoiesis and thrombopoiesis. We also discuss the utility of cholesterol acceptors in contributing to lipid raft regulation and disruption, and highlight the potential to manipulate these pathways for therapeutic gain in CVD as well as other disorders with aberrant hematopoiesis.

Large numeric values consisting of 16 digits in SAS might be incorrect when written to Hadoop for columns defined with the BIGINT data type.  This problem was introduced in SAS 9