SIRT5 variants from patients with mitochondrial disease are associated with reduced SIRT5 stability and activity, but not with neuropathology.

SIRT5 is a sirtuin deacylase that represents the major activity responsible for removal of negatively-charged lysine modifications, in the mitochondrial matrix and elsewhere in the cell. In benign cells and mouse models, under basal non-stressed conditions, the phenotypes of SIRT5 deficiency are generally quite subtle. Here, we identify two homozygous SIRT5 variants in human patients suffering from severe mitochondrial disease. Both variants, P114T and L128V, are associated with reduced SIRT5 protein stability and impaired biochemical activity, with no evidence of neomorphic or dominant negative properties. The crystal structure of the P114T enzyme was solved and shows only subtle deviations from wild-type. Via CRISPR-Cas9, we generate a mouse model that recapitulates the human P114T mutation; homozygotes show reduced SIRT5 levels and activity, but no obvious metabolic abnormalities, neuropathology or other gross evidence of severe disease. We conclude that these human SIRT5 variants most likely represent severe hypomorphs, and are likely not the primary pathogenic cause of the neuropathology observed in the patients.

Functional and molecular profiling of fasted piglets reveals decreased energy metabolic function and cell proliferation in the small intestine.

The small intestine requires energy to exert its important role in nutrient uptake and barrier function. Pigs are an important source of food and a model for humans. Young piglets and infants can suffer from periods of insufficient food intake. Whether this functionally affects the small intestinal epithelial cell (IEC) metabolic capacity and how this may be associated with an increased vulnerability to intestinal disease is unknown. We therefore performed a 48-h fasting intervention in young piglets. After feeding a standard weaning diet for 2 wk, 6-wk-old piglets (n = 16/group) were fasted for 48 h, and midjejunal IECs were collected upon euthanasia. Functional metabolism of isolated IECs was analyzed with the Seahorse XF analyzer and gene expression was assessed using RNA-sequencing. Fasting decreased the mitochondrial and glycolytic function of the IECs by 50% and 45%, respectively (P < 0.0001), signifying that overall metabolic function was decreased. The RNA-sequencing results corroborated our functional metabolic measurements, showing that particularly pathways related to mitochondrial energy production were decreased. Besides oxidative metabolic pathways, decreased cell-cycle progression pathways were most regulated in the fasted piglets, which were confirmed by 43% reduction of Ki67-stained cells (P < 0.05). Finally, the expression of barrier function genes was reduced upon fasting. In conclusion, we found that the decreased IEC energy metabolic function in response to fasting is supported by a decreased gene expression of mitochondrial pathways and is likely linked to the observed decreased intestinal cell proliferation and barrier function, providing insight into the vulnerability of piglets, and infants, to decreased food intake.NEW & NOTEWORTHY Fasting is identified as one of the underlying causes potentiating diarrhea development, both in piglets and humans. With this study, we demonstrate that fasting decreases the metabolism of intestinal epithelial cells, on a functional and transcriptional level. Transcriptional and histological data also show decreased intestinal cell proliferation. As such, fasting-induced intestinal energy shortage could contribute to intestinal dysfunction upon fasting.

X-linked creatine transporter (SLC6A8) deficiency in females: Difficult to recognize, but a potentially treatable disease.

Creatine transporter deficiency (CTD), caused by pathogenic variants in SLC6A8, is the second most common cause of X-linked intellectual disability. Symptoms include intellectual disability, epilepsy, and behavioral disorders and are caused by reduced cerebral creatine levels. Targeted treatment with oral supplementation is available, however the treatment efficacy is still being investigated. There are clinical and theoretical indications that heterozygous females with CTD respond better to supplementation treatment than hemizygous males. Unfortunately, heterozygous females with CTD often have more subtle and uncharacteristic clinical and biochemical phenotypes, rendering diagnosis more difficult. We report a new female case who presented with learning disabilities and seizures. After determining the diagnosis with molecular genetic testing confirmed by proton magnetic resonance spectroscopy (1H-MRS), the patient was treated with supplementation treatment including creatine, arginine, and glycine. After 28 months of treatment, the patient showed prominent clinical improvement and increased creatine levels in the brain. Furthermore, we provide a review of the 32 female cases reported in the current literature including a description of phenotypes, genotypes, diagnostic approaches, and effects of supplementation treatment. Based on this, we find that supplementation treatment should be tested in heterozygous female patients with CTD, and a prospective treatment underlines the importance of diagnosing these patients. The diagnosis should be suspected in a broad clinical spectrum of female patients and can only be made by molecular genetic testing. 1H-MRS of cerebral creatine levels is essential for establishing the diagnosis in females, and especially valuable when assessing variants of unknown significance.

Hepatic Fibrosis Evaluated in Patients with Type 2 Diabetes with and without Chronic Kidney Disease.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD), and particularly liver fibrosis, has been suggested as a risk factor of chronic kidney disease (CKD). Given that NAFLD affects every fourth person globally, better insight is needed. Our aim was to investigate the association between hepatic fibrosis and CKD in patients with type 2 diabetes and to compare different methods for diagnosing liver fibrosis in this study population. METHODS: Cross-sectional study including patients with type 2 diabetes with CKD stages 3-5 (N = 50) or without CKD (N = 50). CKD was defined as estimated glomerular filtration rate <60 mL/min/1.73 m2 with or without proteinuria. Three methods were used to detect significant liver fibrosis defined as either ≥8 kilopascal measured by transient elastography (FibroScan®), fibrosis-4 (FIB-4) score ≥2.67, or NAFLD fibrosis score (NFS) >0.675. RESULTS: Significant liver fibrosis was found in 38% and 28% of the patients with and without CKD, respectively, using at least one of the three methods. Both FIB-4 score and NFS were significantly higher in patients with CKD (p < 0.0009 and p < 0.0001, respectively), although insignificant after adjustments for age, sex, body mass index, and duration of diabetes. In patients without CKD, a significant association between steatosis and fibrosis was observed (p = 0.0007). CONCLUSION: Our data do not support any strong independent association between liver fibrosis and established CKD as assessed by FibroScan, FIB-4 score, and NFS, respectively.

Single and Joined Behaviour of Circulating Biomarkers and Metabolic Parameters in High-Fit and Low-Fit Healthy Females.

Biomarkers are important in the assessment of health and disease, but are poorly studied in still healthy individuals with a (potential) different risk for metabolic disease. This study investigated, first, how single biomarkers and metabolic parameters, functional biomarker and metabolic parameter categories, and total biomarker and metabolic parameter profiles behave in young healthy female adults of different aerobic fitness and, second, how these biomarkers and metabolic parameters are affected by recent exercise in these healthy individuals. A total of 102 biomarkers and metabolic parameters were analysed in serum or plasma samples from 30 young, healthy, female adults divided into a high-fit (V̇O2peak ≥ 47 mL/kg/min, N = 15) and a low-fit (V̇O2peak ≤ 37 mL/kg/min, N = 15) group, at baseline and overnight after a single bout of exercise (60 min, 70% V̇O2peak). Our results show that total biomarker and metabolic parameter profiles were similar between high-fit and low-fit females. Recent exercise significantly affected several single biomarkers and metabolic parameters, mostly related to inflammation and lipid metabolism. Furthermore, functional biomarker and metabolic parameter categories corresponded to biomarker and metabolic parameter clusters generated via hierarchical clustering models. In conclusion, this study provides insight into the single and joined behavior of circulating biomarkers and metabolic parameters in healthy females, and identified functional biomarker and metabolic parameter categories that may be used for the characterisation of human health physiology.

FOXO3A-short is a novel regulator of non-oxidative glucose metabolism associated with human longevity.

Intronic single-nucleotide polymorphisms (SNPs) in FOXO3A are associated with human longevity. Currently, it is unclear how these SNPs alter FOXO3A functionality and human physiology, thereby influencing lifespan. Here, we identify a primate-specific FOXO3A transcriptional isoform, FOXO3A-Short (FOXO3A-S), encoding a major longevity-associated SNP, rs9400239 (C or T), within its 5' untranslated region. The FOXO3A-S mRNA is highly expressed in the skeletal muscle and has very limited expression in other tissues. We find that the rs9400239 variant influences the stability and functionality of the primarily nuclear protein(s) encoded by the FOXO3A-S mRNA. Assessment of the relationship between the FOXO3A-S polymorphism and peripheral glucose clearance during insulin infusion (Rd clamp) in a cohort of Danish twins revealed that longevity T-allele carriers have markedly faster peripheral glucose clearance rates than normal lifespan C-allele carriers. In vitro experiments in human myotube cultures utilizing overexpression of each allele showed that the C-allele represses glycolysis independently of PI3K signaling, while overexpression of the T-allele represses glycolysis only in a PI3K-inactive background. Supporting this finding inducible knockdown of the FOXO3A-S C-allele in cultured myotubes increases the glycolytic rate. We conclude that the rs9400239 polymorphism acts as a molecular switch which changes the identity of the FOXO3A-S-derived protein(s), which in turn alters the relationship between FOXO3A-S and insulin/PI3K signaling and glycolytic flux in the skeletal muscle. This critical difference endows carriers of the FOXO3A-S T-allele with consistently higher insulin-stimulated peripheral glucose clearance rates, which may contribute to their longer and healthier lifespans.

High-Resolution Motion-corrected 7.0-T MRI to Derive Morphologic Measures from the Human Cerebellum in Vivo.

Background The human cerebellum has a large, highly folded cortical sheet. Its visualization is important for various disorders, including multiple sclerosis and spinocerebellar ataxias. The derivation of the cerebellar cortical surface in vivo is impeded by its high foliation. Purpose To image the cerebellar cortex, including its foliations and lamination, in less than 20 minutes, reconstruct the cerebellocortical surface, and extract cortical measures with use of motion-corrected, high-spatial-resolution 7.0-T MRI. Materials and Methods In this prospective study, conducted between February 2021 and July 2022, healthy participants underwent an examination with either a 0.19 × 0.19 × 0.5-mm3, motion-corrected fast low-angle shot (FLASH) sequence (14.5 minutes) or a whole-cerebellum 0.4 × 0.4 × 0.4-mm3, motion-corrected magnetization-prepared 2 rapid gradient-echo (MP2RAGE) sequence (18.5 minutes) at 7.0 T. Four participants underwent an additional FLASH sequence without motion correction. FLASH and MP2RAGE sequences were used to visualize the cerebellar cortical layers, derive cerebellar gray and white matter segmentations, and examine their fidelity. Quantitative measures were compared using repeated-measures analyses of variance or paired t tests. Results Nine participants (median age, 36 years [IQR, 25-42 years; range, 21-62 years]; five women) underwent examination with the FLASH sequence. Nine participants (median age, 37 years [IQR, 34-42 years; range, 25-62 years]; five men) underwent examination with the MP2RAGE sequence. A susceptibility difference between the expected location of the granular and molecular cerebellar layers was visually detected in the FLASH data in all participants. The segmentations derived from the whole-cerebellum MP2RAGE sequence showed the characteristic anatomic features of the cerebellum, like the transverse fissures and splitting folds. The cortical surface area (median, 949 cm2 [IQR, 825-1021 cm2]) was 1.8 times larger, and the cortical thickness (median, 0.88 mm [IQR, 0.81-0.93 mm]) was five times thinner than previous in vivo estimates and closer to ex vivo reference data. Conclusion In vivo imaging of the cerebellar cortical layers and surface and derivation of quantitative measures was feasible in a clinically acceptable acquisition time with use of motion-corrected 7.0-T MRI. Published under a CC BY 4.0 license. Supplemental material is available for this article. See also the editorial by Dietrich in this issue.

Nonalcoholic Fatty Liver Disease in Patients with Type 2 Diabetes and Chronic Kidney Disease.

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) is suggested as a risk factor for chronic kidney disease (CKD). The incidence of NAFLD is rising globally in parallel to the increasing incidences of obesity and type 2 diabetes. Diabetes remains the leading cause of CKD, but the co-existence of NAFLD, CKD, and type 2 diabetes is not well elucidated. Here, we evaluated the prevalence of NAFLD in patients with type 2 diabetes with and without CKD. METHODS: This was a cross-sectional study including 50 patients with type 2 diabetes and CKD stages 3-5 (no dialysis), and 50 patients with type 2 diabetes without CKD. Liver fat content was estimated by proton magnetic resonance spectroscopy and magnetic resonance imaging proton density fat fraction. NAFLD was defined as liver fat fraction ≥5.6% according to guidelines. RESULTS: Mean age was 72 ± 4.9 years in patients with CKD and 65.9 ± 7.8 years in patients without CKD (p < 0.0001). Three out of four participants were men. BMI was 28.6 ± 3.5 kg/m2 and 27 ± 4.0 kg/m2 in patients with and without CKD, respectively (p = 0.0087). NAFLD was identified in 22 (44%) patients with CKD and 19 (38%) patients without CKD (p = 0.6845). Median (IQR) liver fat fraction was 4.7% (3.0-8.5) and 4.1% (2.9-7.7) in patients with and without CKD, respectively (difference in geometric means 5.3%, 95% CI -23; 45, p = 0.7463). CONCLUSION: These findings do not support any association between NAFLD and CKD (stages 3-5) in patients with type 2 diabetes.

Mastering the game of Stratego with model-free multiagent reinforcement learning.

We introduce DeepNash, an autonomous agent that plays the imperfect information game Stratego at a human expert level. Stratego is one of the few iconic board games that artificial intelligence (AI) has not yet mastered. It is a game characterized by a twin challenge: It requires long-term strategic thinking as in chess, but it also requires dealing with imperfect information as in poker. The technique underpinning DeepNash uses a game-theoretic, model-free deep reinforcement learning method, without search, that learns to master Stratego through self-play from scratch. DeepNash beat existing state-of-the-art AI methods in Stratego and achieved a year-to-date (2022) and all-time top-three ranking on the Gravon games platform, competing with human expert players.

Improved detection limits of J-coupled neurometabolites in the human brain at 7 T with a J-refocused sLASER sequence.

In a standard spin echo, the time evolution due to homonuclear couplings is not reversed, leading to echo time (TE)-dependent modulation of the signal amplitude and signal loss in the case of overlapping multiplet resonances. This has an adverse effect on quantification of several important metabolites such as glutamate and glutamine. Here, we propose a J-refocused variant of the sLASER sequence (J-sLASER) to improve quantification of J-coupled metabolites at ultrahigh field (UHF). The use of the sLASER sequence is particularly advantageous at UHF as it minimizes chemical shift displacement error and results in relatively homogenous refocusing. We simulated the MRS signal from brain metabolites over a broad range of TE values with sLASER and J-sLASER, and showed that the signal of J-coupled metabolites was increased with J-sLASER with TE values up to ~80 ms. We further simulated "brain-like" spectra with both sequences at the shortest TE available on our scanner. We showed that, despite the slightly longer TE, the J-sLASER sequence results in significantly lower Cramer-Rao lower bounds (CRLBs) for J-coupled metabolites compared with those obtained with sLASER. Following phantom validation, we acquired spectra from two brain regions in 10 healthy volunteers (age 38 ± 15 years) using both sequences. We showed that using J-sLASER results in a decrease of CRLBs for J-coupled metabolites. In particular, we measured a robust ~38% decrease in the mean CRLB (glutamine) in parietal white matter and posterior cingulate cortex (PCC). We further showed, in 10 additional healthy volunteers (age 34 ± 15 years), that metabolite quantification following two separate acquisitions with J-sLASER in the PCC was repeatable. The improvement in quantification of glutamine may in turn improve the independent quantification of glutamate, the main excitatory neurotransmitter in the brain, and will simultaneously help to track possible modulations of glutamine, which is a key player in the glutamatergic cycle in astrocytes.

Add-On MEmaNtine to Dopamine Antagonism to Improve Negative Symptoms at First Psychosis- the AMEND Trial Protocol.

Background: Antipsychotic drugs are primarily efficacious in treating positive symptoms by blocking the dopamine D2 receptor, but they fail to substantially improve negative symptoms and cognitive deficits. The limited efficacy may be attributed to the fact that the pathophysiology of psychosis involves multiple neurotransmitter systems. In patients with chronic schizophrenia, memantine, a non-competitive glutamatergic NMDA receptor antagonist, shows promise for ameliorating negative symptoms and improving cognition. Yet, it is unknown how memantine modulates glutamate levels, and memantine has not been investigated in patients with first-episode psychosis. Aims: This investigator-initiated double-blinded randomized controlled trial is designed to (1) test the clinical effects on negative symptoms of add-on memantine to antipsychotic medication, and (2) neurobiologically characterize the responders to add-on memantine. Materials and Equipment: Antipsychotic-naïve patients with first-episode psychosis will be randomized to 12 weeks treatment with [amisulpride + memantine] or [amisulpride + placebo]. We aim for a minimum of 18 patients in each treatment arm to complete the trial. Brain mapping will be performed before and after 12 weeks focusing on glutamate and neuromelanin in predefined regions. Regional glutamate levels will be probed with proton magnetic resonance spectroscopy (MRS), while neuromelanin signal will be mapped with neuromelanin-sensitive magnetic resonance imaging (MRI). We will also perform structural and diffusion weighted, whole-brain MRI. MRS and MRI will be performed at an ultra-high field strength (7 Tesla). Alongside, participants undergo clinical and neuropsychological assessments. Twenty matched healthy controls will undergo similar baseline- and 12-week examinations, but without receiving treatment. Outcome Measures: The primary endpoint is negative symptom severity. Secondary outcomes comprise: (i) clinical endpoints related to cognition, psychotic symptoms, side effects, and (ii) neurobiological endpoints related to regional glutamate- and neuromelanin levels, and structural brain changes. Anticipated Results: We hypothesize that add-on memantine to amisulpride will be superior to amisulpride monotherapy in reducing negative symptoms, and that this effect will correlate with thalamic glutamate levels. Moreover, we anticipate that add-on memantine will restore regional white matter integrity and improve cognitive functioning. Perspectives: By combining two licensed, off-patent drugs, AMEND aims to optimize treatment of psychosis while investigating the memantine response. Alongside, AMEND will provide neurobiological insights to effects of dual receptor modulation, which may enable future stratification of patients with first-episode psychosis before initial antipsychotic treatment. Clinical Trial Registration: [ClinicalTrials.gov], identifier [NCT04789915].

Improving brain B0 shimming using an easy and accessible multi-coil shim array at ultra-high field.

OBJECT: Improve shimming capabilities of ultra-high field systems, with addition of an accessible low-complexity B0 shim array for head MRI at 7 T. MATERIALS AND METHODS: An eight channel B0 shim coil array was designed as a tradeoff between shimming improvement and construction complexity, to provide an easy to use shim array that can be employed with the standard 7 T head coil. The array was interfaced using an open-source eight-channel shim amplifier rack. Improvements in field homogeneity for whole-brain and slice-based shimming were compared to standard second-order shimming, and to more complex higher order dynamic shimming and shim arrays with 32 and 48 channels. RESULTS: The eight-channel shim array provided 12% improvement in whole brain static shimming and provided 33% improvement when using slice-based shimming. With this, the eight-channel array performed similar to third-order dynamic shimming (without the need for higher order eddy current compensation). More complex shim arrays with 32 and 48 channels performed better, but require a dedicated RF coil. DISCUSSION: The designed eight-channel shim array provides a low-complexity and low-cost approach for improving B0 field shimming on an ultra-high field system. In both static and dynamic shimming, it provides improved B0 homogeneity over standard shimming.

Accuracy investigations for volumetric head-motion navigators with and without EPI at 7 T.

PURPOSE: Accuracy investigation of volumetric navigators for motion correction, with emphasis on geometric EPI distortions at ultrahigh field. METHODS: High-resolution Dixon images were collected in different head positions and reconstructed to water, fat, T2 *, and B0 maps. Resolution reduction was performed, and the T2 * and B0 maps were used to apply effects of TE and EPI distortions to simulate various volumetric water and fat navigators. Registrations of the simulated navigators were compared with registrations of the original high-resolution images. RESULTS: Increased accuracy was observed with increased spatial resolution for non-EPI navigators. When using EPI, the distortions had a negative effect on registration accuracy, which was most noticeable for high-resolution navigators. Parallel imaging helped to alleviate those caveats to a certain extent, and 5-fold acceleration gave close to similar accuracy to non-EPI in most cases. Shortening the TE by partial Fourier sampling was shown to be mostly beneficial, except for water navigators with long readout durations. The EPI blip direction had an influence on navigator accuracy, and positive blip gradient polarities (yielding mostly image stretching frontally) typically gave the best accuracy for water navigators, whereas no clear recommendation could be made for fat navigators. Generally, fat EPI navigators had lower accuracy than water EPI navigators with otherwise similar parameters. CONCLUSIONS: Echo planar imaging has been widely used for MRI navigators, but the induced distortions reduce navigator accuracy at ultrahigh field. This study can help protocol optimization and guide the complex tradeoff between resolution and EPI acceleration in navigator parameter setup.

Proton metabolic mapping of the brain at 7 T using a two-dimensional free induction decay-echo-planar spectroscopic imaging readout with lipid suppression.

The increased signal-to-noise ratio (SNR) and chemical shift dispersion at high magnetic fields (≥7 T) have enabled neuro-metabolic imaging at high spatial resolutions. To avoid very long acquisition times with conventional magnetic resonance spectroscopic imaging (MRSI) phase-encoding schemes, solutions such as pulse-acquire or free induction decay (FID) sequences with short repetition time and inner volume selection methods with acceleration (echo-planar spectroscopic imaging [EPSI]), have been proposed. With the inner volume selection methods, limited spatial coverage of the brain and long echo times may still impede clinical implementation. FID-MRSI sequences benefit from a short echo time and have a high SNR per time unit; however, contamination from strong extra-cranial lipid signals remains a problem that can hinder correct metabolite quantification. L2-regularization can be applied to remove lipid signals in cases with high spatial resolution and accurate prior knowledge. In this work, we developed an accelerated two-dimensional (2D) FID-MRSI sequence using an echo-planar readout and investigated the performance of lipid suppression by L2-regularization, an external crusher coil, and the combination of these two methods to compare the resulting spectral quality in three subjects. The reduction factor of lipid suppression using the crusher coil alone varies from 2 to 7 in the lipid region of the brain boundary. For the combination of the two methods, the average lipid area inside the brain was reduced by 2% to 38% compared with that of unsuppressed lipids, depending on the subject's region of interest. 2D FID-EPSI with external lipid crushing and L2-regularization provides high in-plane coverage and is suitable for investigating brain metabolite distributions at high fields.

Identifying the source of spurious signals caused by B0 inhomogeneities in single-voxel 1 H MRS.

PURPOSE: Single-voxel MRS (SV MRS) requires robust volume localization as well as optimized crusher and phase-cycling schemes to reduce artifacts arising from signal outside the volume of interest. However, due to local magnetic field gradients (B0 inhomogeneities), signal that was dephased by the crusher gradients during acquisition might rephase, leading to artifacts in the spectrum. Here, we analyzed this mechanism, aiming to identify the source of signals arising from unwanted coherence pathways (spurious signals) in SV MRS from a B0 map. METHODS: We investigated all possible coherence pathways associated with imperfect localization in a semi-localized by adiabatic selective refocusing (semi-LASER) sequence for potential rephasing of signals arising from unwanted coherence pathways by a local magnetic field gradient. We searched for locations in the B0 map where the signal dephasing due to external (crusher) and internal (B0 ) field gradients canceled out. To confirm the mechanism, SV-MR spectra (TE = 31 ms) and 3D-CSI data with the same volume localization as the SV experiments were acquired from a phantom and 2 healthy volunteers. RESULTS: Our analysis revealed that potential sources of spurious signals were scattered over multiple locations throughout the brain. This was confirmed by 3D-CSI data. Moreover, we showed that the number of potential locations where spurious signals could originate from monotonically decreases with crusher strength. CONCLUSION: We proposed a method to identify the source of spurious signals in SV 1 H MRS using a B0 map. This can facilitate MRS sequence design to be less sensitive to experimental artifacts.

Locus Coeruleus Shows a Spatial Pattern of Structural Disintegration in Parkinson's Disease.

BACKGROUND: Parkinson's disease (PD) causes a loss of neuromelanin-positive, noradrenergic neurons in the locus coeruleus (LC), which has been implicated in nonmotor dysfunction. OBJECTIVES: We used "neuromelanin sensitive" magnetic resonance imaging (MRI) to localize structural disintegration in the LC and its association with nonmotor dysfunction in PD. METHODS: A total of 42 patients with PD and 24 age-matched healthy volunteers underwent magnetization transfer weighted (MTw) MRI of the LC. The contrast-to-noise ratio of the MTw signal (CNRMTw ) was used as an index of structural LC integrity. We performed slicewise and voxelwise analyses to map spatial patterns of structural disintegration, complemented by principal component analysis (PCA). We also tested for correlations between regional CNRMTw and severity of nonmotor symptoms. RESULTS: Mean CNRMTw of the right LC was reduced in patients relative to controls. Voxelwise and slicewise analyses showed that the attenuation of CNRMTw was confined to the right mid-caudal LC and linked regional CNRMTw to nonmotor symptoms. CNRMTw attenuation in the left mid-caudal LC was associated with the orthostatic drop in systolic blood pressure, whereas CNRMTw attenuation in the caudal most portion of right LC correlated with apathy ratings. PCA identified a bilateral component that was more weakly expressed in patients. This component was characterized by a gradient in CNRMTw along the rostro-caudal and dorso-ventral axes of the nucleus. The individual expression score of this component reflected the overall severity of nonmotor symptoms. CONCLUSION: A spatially heterogeneous disintegration of LC in PD may determine the individual expression of specific nonmotor symptoms such as orthostatic dysregulation or apathy. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson Movement Disorder Society.

Extracellular flux analyses reveal differences in mitochondrial PBMC metabolism between high-fit and low-fit females.

Analyzing metabolism of peripheral blood mononuclear cells (PBMCs) can possibly serve as a cellular metabolic read-out for lifestyle factors and lifestyle interventions. However, the impact of PBMC composition on PBMC metabolism is not yet clear, neither is the differential impact of a longer-term lifestyle factor versus a short-term lifestyle intervention. We investigated the effect of aerobic fitness level and a recent exercise bout on PBMC metabolism in females. PBMCs from 31 young female adults divided into a high-fit (V̇o2peak ≥ 47 mL/kg/min, n = 15) and low-fit (V̇o2peak ≤ 37 mL/kg/min, n = 16) groups were isolated at baseline and overnight after a single bout of exercise (60 min, 70% V̇o2peak). Oxygen consumption rate (OCR) and glycolytic rate (GR) were measured using extracellular flux (XF) assays and PBMC subsets were characterized using fluorescence-activated cell sorting (FACS). Basal OCR, FCCP-induced OCR, spare respiratory capacity, ATP-linked OCR, and proton leak were significantly higher in high-fit than in low-fit females (all P < 0.01), whereas no significant differences in glycolytic rate (GR) were found (all P > 0.05). A recent exercise bout did not significantly affect GR or OCR parameters (all P > 0.05). The overall PBMC composition was similar between high-fit and low-fit females. Mitochondrial PBMC function was significantly higher in PBMCs from high-fit than from low-fit females, which was unrelated to PBMC composition and not impacted by a recent bout of exercise. Our study reveals a link between PBMC metabolism and levels of aerobic fitness, increasing the relevance of PBMC metabolism as a marker to study the impact of lifestyle factors on human health.NEW & NOTEWORTHY Mitochondrial metabolism was significantly higher in PBMCs from high-fit than from low-fit females. This was unrelated to PBMC composition and not impacted by a recent bout of exercise. Our study reveals a link between PBMC metabolism and levels of aerobic fitness, increasing the relevance of PBMC metabolism as a marker to study the impact of lifestyle factors on human health.

The female mouse is resistant to mild vitamin B3 deficiency.

PURPOSE: Vitamin B3 provides nicotinamide adenine dinucleotide (NAD+), an essential coenzyme in oxidoreductase reactions. Severe vitamin B3 deficiency leads to the disease Pellagra, while mild vitamin B3 deficiency has been linked to age-related and metabolic diseases. Mild vitamin B3 deficiency is understudied, especially in females. Therefore, we examined how female mice responded to a diet that induced mild vitamin B3 deficiency in male mice. METHODS: Female C57BL/6RccHsd mice were subjected for 18 weeks to a diet without vitamin B3 and low but sufficient tryptophan (0.115%) (0NR) and were compared to control female mice on the same diet with the reference dose of vitamin B3 (30NR, 30 mg nicotinamide riboside/ kg diet). RESULTS: In the female mice, no differences between the two dietary groups were found in liver nicotinamide mononucleotide (NMN) levels, body composition, whole body energy and substrate metabolism measured by indirect calorimetry, or liver triacylglycerol metabolism. Expression of seven genes that previously were shown to respond to mild vitamin B3 deficiency in male white adipose tissue were not differentially expressed between the female dietary groups, neither was insulin sensitivity. CONCLUSION: We concluded that the female 0NR mice were not vitamin B3 deficient; the role of age, sex and health status is discussed. Demonstrated by clear differences between females and males, the latter showing mild deficiency under the same conditions, this study highlights the importance of studying both sexes.

The Effect of a Single Bout of Exercise on Vitamin B2 Status Is Not Different between High- and Low-Fit Females.

High-fitness individuals have been suggested to be at risk of a poor vitamin B2 (riboflavin) status due to a potentially higher vitamin B2 demand, as measured by the erythrocyte glutathione reductase (EGR) activation coefficient (EGRAC). Longer-term exercise interventions have been shown to result in a lower vitamin B2 status, but studies are contradictory. Short-term exercise effects potentially contribute to discrepancies between studies but have only been tested in limited study populations. This study investigated if vitamin B2 status, measured by EGRAC, is affected by a single exercise bout in females who differ in fitness levels, and that represents long-term physical activity. At baseline and overnight after a 60-min cycling bout at 70% V·O2peak, EGR activity and EGRAC were measured in 31 young female adults, divided into a high-fit (V·O2peak ≥ 47 mL/kg/min, N = 15) and low-fit (V·O2peak ≤ 37 mL/kg/min, N = 16) group. A single exercise bout significantly increased EGR activity in high-fit and low-fit females (Ptime = 0.006). This response was not affected by fitness level (Ptime*group = 0.256). The effect of exercise on EGRAC was not significant (Ptime = 0.079) and not influenced by EGR activity. The exercise response of EGRAC was not significantly different between high-fit and low-fit females (Ptime*group = 0.141). Thus, a single exercise bout increased EGR activity, but did not affect EGRAC, indicating that vitamin B2 status was not affected. The exercise response on EGRAC and EGR did not differ between high-fit and low-fit females.

Butyrate Alters Pyruvate Flux and Induces Lipid Accumulation in Cultured Colonocytes.

Butyrate is considered the primary energy source of colonocytes and has received wide attention due to its unique health benefits. Insight into the mechanistic effects of butyrate on cellular and metabolic function relies mainly on research in in-vitro-cultured cells. However, cells in culture differ from those in vivo in terms of metabolic phenotype and nutrient availability. For translation, it is therefore important to understand the impact of different nutrients on the effects of butyrate. We investigated the metabolic consequences of butyrate exposure under various culturing conditions, with a focus on the interaction between butyrate and glucose. To investigate whether the effects of butyrate were different between cells with high and low mitochondrial capacity, we cultured HT29 cells under either low- (0.5 mM) or high- (25 mM) glucose conditions. Low-glucose culturing increased the mitochondrial capacity of HT29 cells compared to high-glucose (25 mM) cultured HT29 cells. Long-term exposure to butyrate did not alter mitochondrial bioenergetics, but it decreased glycolytic function, regardless of glucose availability. In addition, both high- and low-glucose-grown HT29 cells showed increased lipid droplet accumulation following long-term butyrate exposure. Acute exposure of cultured cells (HT29 and Caco-2) to butyrate increased their oxygen consumption rate (OCR). A simultaneous decrease in extracellular acidification rate (ECAR) was observed. Furthermore, in the absence of glucose, OCR did not increase in response to butyrate. These results lead us to believe that butyrate itself was not responsible for the observed increase in OCR, but, instead, butyrate stimulated pyruvate flux into mitochondria. Indeed, blocking of the mitochondrial pyruvate carrier prevented a butyrate-induced increase in oxygen consumption. Taken together, our results indicate that butyrate itself is not oxidized in cultured cells but instead alters pyruvate flux and induces lipid accumulation.