Glucose Challenge Uncovers Temporal Fungibility of Metabolic Homeostasis Throughout the Day.

Rhythmicity is a central feature of behavioral and biological processes including metabolism, however, the mechanisms of metabolite cycling are poorly understood. A robust oscillation in a network of key metabolite pathways downstream of glucose is described in humans, then these pathways mechanistically probed through purpose-built 13C6-glucose isotope tracing in Drosophila every 4h. A temporal peak in biosynthesis was noted by broad labelling of pathways downstream of glucose in wild-type flies shortly following lights on. Krebs cycle labelling was generally increased in a hyperactive mutant (fumin) along with glycolysis labelling primarily observed at dawn. Surprisingly, neither underlying feeding rhythms nor the presence of food explains the rhythmicity of glucose processing across genotypes. These results are consistent with clinical data demonstrating detrimental effects of mis-timed energy intake. This approach provides a window into the dynamic range of metabolic processing ability through the day and mechanistic basis for exploring circadian metabolic homeostasis in disease states.

Altered Metabolism During the Dark Period in Drosophila Short Sleep Mutants.

Sleep is an almost universally required state in biology. Disrupted sleep has been associated with adverse health risks including metabolic perturbations. Sleep is in part regulated via circadian mechanisms, however, metabolic dysfunction at different times of day arising from sleep disruption is unclear. We used targeted liquid chromatography-mass spectrometry to probe metabolic alterations using high-resolution temporal sampling of two Drosophila short sleep mutants, fumin and sleepless, across a circadian day. Discriminant analyses revealed overall distinct metabolic profiles for mutants when compared to a wild type dataset. Altered levels of metabolites involved in nicotinate/nicotinamide, alanine, aspartate, and glutamate, glyoxylate and dicarboxylate metabolism, and the TCA cycle were observed in mutants suggesting increased energetic demands. Furthermore, rhythmicity analyses revealed fewer 24 hr rhythmic metabolites in both mutants. Interestingly, mutants displayed two major peaks in phases while wild type displayed phases that were less concerted. In contrast to 24 hr rhythmic metabolites, an increase in the number of 12 hr rhythmic metabolites was observed in fumin while sleepless displayed a decrease. These results support that decreased sleep alters the overall metabolic profile with short sleep mutants displaying altered metabolite levels associated with a number of pathways in addition to altered neurotransmitter levels.

Chronic sleep loss sensitizes Drosophila melanogaster to nitrogen stress.

Chronic sleep loss profoundly impacts metabolic health and shortens lifespan, but studies of the mechanisms involved have focused largely on acute sleep deprivation.1,2 To identify metabolic consequences of chronically reduced sleep, we conducted unbiased metabolomics on heads of three adult Drosophila short-sleeping mutants with very different mechanisms of sleep loss: fumin (fmn), redeye (rye), and sleepless (sss).3,4,5,6,7 Common features included elevated ornithine and polyamines, with lipid, acyl-carnitine, and TCA cycle changes suggesting mitochondrial dysfunction. Studies of excretion demonstrate inefficient nitrogen elimination in adult sleep mutants, likely contributing to their polyamine accumulation. Increasing levels of polyamines, particularly putrescine, promote sleep in control flies but poison sleep mutants. This parallels the broadly enhanced toxicity of high dietary nitrogen load from protein in chronically sleep-restricted Drosophila, including both sleep mutants and flies with hyper-activated wake-promoting neurons. Together, our results implicate nitrogen stress as a novel mechanism linking chronic sleep loss to adverse health outcomes-and perhaps for linking food and sleep homeostasis at the cellular level in healthy organisms.

Spectrum of imaging findings in hyperplastic cholecystosis and potential mimics.

Adenomyomatosis and cholesterolosis of the gallbladder, collectively termed hyperplastic cholecystosis, are commonly encountered incidental findings on imaging studies performed for a variety of indications including biliary colic or nonspecific abdominal pain. These pathologies are rarely the source of symptoms, generally considered benign and do not require further work-up. However, their imaging characteristics can overlap with more sinister conditions that should not be missed. In this review, the imaging findings of adenomyomatosis and cholesterolosis will be reviewed followed by other gallbladder pathologies that might mimic these conditions radiologically. Important differentiating factors will be discussed that can aid the radiologist in making a more confident imaging diagnosis.

Interventional Radiology Management of Adult Liver Transplant Complications.

Liver transplant remains the definitive therapy for patients with end-stage liver disease. Outcomes have continued to improve, in part owing to interventions used to treat posttransplant complications involving the hepatic arteries, portal vein, hepatic veins or inferior vena cava (IVC), and biliary system. Significant hepatic artery stenosis can be treated with angioplasty or stent placement to prevent thrombosis and biliary ischemic complications. Hepatic arterioportal fistula and hepatic artery pseudoaneurysm are rare complications that can often be treated with endovascular means. Treatment of hepatic artery thrombosis can have mixed results. Portal vein stenosis can be treated with venoplasty or more commonly stent placement. The rarer portal vein thrombosis can also be treated with endovascular techniques. Hepatic venous outflow stenosis of the hepatic veins or IVC is amenable to venoplasty or stent placement. Complications of the bile ducts are the most encountered complication after liver transplant. When not amenable to endoscopic intervention, biliary stricture, bile leak, and ischemic cholangiopathy can be treated with percutaneous transhepatic cholangiography with biliary drainage and other interventions. New techniques have further improved care for these patients. Transsplenic portal vein recanalization has improved transplant candidacy for patients with chronic portal vein thrombosis. Spontaneous splenorenal shunt and splenic artery steal syndrome (nonocclusive hepatic artery hypoperfusion syndrome) remain complicated topics, and the role of endovascular embolization is developing. When patients have recurrence of cirrhosis after transplant, most commonly due to viral hepatitis, transjugular intrahepatic portosystemic shunt (TIPS) may be required to treat symptoms of portal hypertension. Online supplemental material is available for this article. ©RSNA, 2022.

Non-Vascular Considerations When Interpreting Extremity Arterial and Venous Examinations.

Peripheral arterial and venous examinations are performed regularly in vascular labs and interpreted by physicians of different specialities. Many vascular examinations have nonvascular pathology that is either inadvertently imaged by the sonographer or imaged with intent as it relates to patient's symptoms. It is prudent for every reader of vascular studies to be acquainted with the sonographic appearance of these non-vascular lesions to enable appropriate and optimal interpretation that has a direct bearing on patient's clinical care. Our review includes a discussion of the nonvascular pathologies like lymph nodes, soft tissue edema, soft tissue fluid collections, musculotendinous injuries, soft tissue masses, and joint and bursal pathologies that may be encountered during interpretation of vascular exams. The pictorial essay includes a discussion of their sonographic appearances and pitfalls in interpretation. Multiple illustrative examples and sonographic images of the non-vascular pathologies found during interpretation of vascular studies have been utilized to highlight their appearances.

Advanced MRI Protocols to Discriminate Glioma From Treatment Effects: State of the Art and Future Directions.

In the follow-up treatment of high-grade gliomas (HGGs), differentiating true tumor progression from treatment-related effects, such as pseudoprogression and radiation necrosis, presents an ongoing clinical challenge. Conventional MRI with and without intravenous contrast serves as the clinical benchmark for the posttreatment surveillance imaging of HGG. However, many advanced imaging techniques have shown promise in helping better delineate the findings in indeterminate scenarios, as posttreatment effects can often mimic true tumor progression on conventional imaging. These challenges are further confounded by the histologic admixture that can commonly occur between tumor growth and treatment-related effects within the posttreatment bed. This review discusses the current practices in the surveillance imaging of HGG and the role of advanced imaging techniques, including perfusion MRI and metabolic MRI.

Rhythmic glucose metabolism regulates the redox circadian clockwork in human red blood cells.

Circadian clocks coordinate mammalian behavior and physiology enabling organisms to anticipate 24-hour cycles. Transcription-translation feedback loops are thought to drive these clocks in most of mammalian cells. However, red blood cells (RBCs), which do not contain a nucleus, and cannot perform transcription or translation, nonetheless exhibit circadian redox rhythms. Here we show human RBCs display circadian regulation of glucose metabolism, which is required to sustain daily redox oscillations. We found daily rhythms of metabolite levels and flux through glycolysis and the pentose phosphate pathway (PPP). We show that inhibition of critical enzymes in either pathway abolished 24-hour rhythms in metabolic flux and redox oscillations, and determined that metabolic oscillations are necessary for redox rhythmicity. Furthermore, metabolic flux rhythms also occur in nucleated cells, and persist when the core transcriptional circadian clockwork is absent in Bmal1 knockouts. Thus, we propose that rhythmic glucose metabolism is an integral process in circadian rhythms.

CRY1-CBS binding regulates circadian clock function and metabolism.

Circadian disruption influences metabolic health. Metabolism modulates circadian function. However, the mechanisms coupling circadian rhythms and metabolism remain poorly understood. Here, we report that cystathionine ß-synthase (CBS), a central enzyme in one-carbon metabolism, functionally interacts with the core circadian protein cryptochrome 1 (CRY1). In cells, CBS augments CRY1-mediated repression of the CLOCK/BMAL1 complex and shortens circadian period. Notably, we find that mutant CBS-I278T protein, the most common cause of homocystinuria, does not bind CRY1 or regulate its repressor activity. Transgenic CbsZn/Zn  mice, while maintaining circadian locomotor activity period, exhibit reduced circadian power and increased expression of E-BOX outputs. CBS function is reciprocally influenced by CRY1 binding. CRY1 modulates enzymatic activity of the CBS. Liver extracts from Cry1-/- mice show reduced CBS activity that normalizes after the addition of exogenous wild-type (WT) CRY1. Metabolomic analysis of WT, CbsZn/Zn , Cry1-/- , and Cry2-/- samples highlights the metabolic importance of endogenous CRY1. We observed temporal variation in one-carbon and transsulfuration pathways attributable to CRY1-induced CBS activation. CBS-CRY1 binding provides a post-translational switch to modulate cellular circadian physiology and metabolic control.

Identification of Bladder Diverticular Stone on 99mTc-Methyl Diphosphonate Bone Scintigraphy and SPECT/CT.

We report a bladder diverticular stone with increased 99mTc-methyl diphosphonate uptake on bone scintigraphy and SPECT/CT. Diverticular stone is a known risk factor for bladder malignancy. The deposition of 99mTc-methyl diphosphonate on the crystal surface of the diverticular stone is a rare phenomenon but of clinical significance. Cystolitholapaxy is indicated to remove the diverticular stone and to reduce the risk of bladder cancer.

Circadian and Sleep Metabolomics Across Species.

Under normal circadian function, metabolic control is temporally coordinated across tissues and behaviors with a 24-h period. However, circadian disruption results in negative consequences for metabolic homeostasis including energy or redox imbalances. Yet, circadian disruption has become increasingly prevalent within today's society due to many factors including sleep loss. Metabolic consequences of both have been revealed by metabolomics analyses of circadian biology and sleep. Specifically, two primary analytical platforms, mass spectrometry and nuclear magnetic resonance spectroscopy, have been used to study molecular clock and sleep influences on overall metabolic rhythmicity. For example, human studies have demonstrated the prevalence of metabolic rhythms in human biology, as well as pan-metabolome consequences of sleep disruption. However, human studies are limited to peripheral metabolic readouts primarily through minimally invasive procedures. For further tissue- and organism-specific investigations, a number of model systems have been studied, based upon the conserved nature of both the molecular clock and sleep across species. Here we summarize human studies as well as key findings from metabolomics studies using mice, Drosophila, and zebrafish. While informative, a limitation in existing literature is a lack of interpretation regarding dynamic synthesis or catabolism within metabolite pools. To this extent, future work incorporating isotope tracers, specific metabolite reporters, and single-cell metabolomics may provide a means of exploring dynamic activity in pathways of interest.

Adrenal Tumors Found During Staging and Surveillance for Colorectal Cancer: Benign Incidentalomas or Metastatic Disease?

OBJECTIVE: To evaluate the incidence of adrenal metastases in patient with colorectal cancer (CRC) and determine the clinical and radiographic features associated metastatic CRC to the adrenal glands. MATERIALS AND METHODS: The review of consecutive adults with newly diagnosed CRC found to have adrenal tumors > 1 cm in size on staging or surveillance CT scans with at least two scans to evaluate progression or stability of disease. RESULTS: Fifty-eight of 856 (6.8%) CRC patients had an adrenal tumor. Forty-three patients (74%) with 46 adrenal tumors had benign adrenal tumors, and 15 (26%) patients with 17 adrenal tumors had metastatic disease. On univariate analysis, patients with metastatic CRC had larger adrenal tumors (26.7 mm vs 12.4 mm, p < 0.01), a higher mean CEA (239 ng/mL vs 14.2 ng/mL, p = 0.03), and were more likely to have other sites of metastatic disease seen on imaging 8/43 (19%) vs 14/15 (93%), p < 0.01. On multivariable analysis, adrenal tumor size > 1.8 cm (OR 49.6 CI 8-306), CEA > 2.5 ng/mL (OR 15.8 CI 1.7-144) and other metastatic disease seen on imaging (OR 68.1 CI 7-661) were independently associated with adrenal metastases. CONCLUSION: CRC patients with small adrenal tumors, normal CEA levels and no evidence of other metastatic disease are unlikely to have spread to the adrenal glands. Adrenal tumors found during staging and surveillance of CRC patients should be evaluated with appropriate imaging and biochemical analysis.

Targeting glutamine metabolism slows soft tissue sarcoma growth.

Tumour cells frequently utilize glutamine to meet bioenergetic and biosynthetic demands of rapid cell growth. However, glutamine dependence can be highly variable between in vitro and in vivo settings, based on surrounding microenvironments and complex adaptive responses to glutamine deprivation. Soft tissue sarcomas (STSs) are mesenchymal tumours where cytotoxic chemotherapy remains the primary approach for metastatic or unresectable disease. Therefore, it is critical to identify alternate therapies to improve patient outcomes. Using autochthonous STS murine models and unbiased metabolomics, we demonstrate that glutamine metabolism supports sarcomagenesis. STS subtypes expressing elevated glutaminase (GLS) levels are highly sensitive to glutamine starvation. In contrast to previous studies, treatment of autochthonous tumour-bearing animals with Telaglenastat (CB-839), an orally bioavailable GLS inhibitor, successfully inhibits undifferentiated pleomorphic sarcoma (UPS) tumour growth. We reveal glutamine metabolism as critical for sarcomagenesis, with CB-839 exhibiting potent therapeutic potential.

Differences in Growth Rate on CT of Adrenal Adenomas and Malignant Adrenal Nodules.

OBJECTIVE. The purpose of this study is to determine the differences in growth rate of adrenal adenomas and malignant adrenal nodules. MATERIALS AND METHODS. This was a retrospective review of adults with an adrenal nodule seen at two different abdominal or chest CT examinations or PET/CT examinations. Patients in the adenoma group were included if they had a CT, MRI, or pathologic diagnosis of an adrenal adenoma. Patients in the malignant group were included if they had a pathologically proven malignant adrenal nodule. Nodule growth was defined as a change in the largest axial diameter greater than or equal to 3 mm. Growth rate was calculated by dividing the change in the longest axial diameter by the time between the first and last imaging examination. RESULTS. There were 105 adenomas and 26 malignant nodules. Of the 105 adenomas, 34 (32.4%; 95% CI, 23.6-42.2%) grew, three (2.9%; 95% CI, 0.6-8.1%) became smaller, and 68 (64.8%; 95% CI, 54.8-73.8%), were unchanged in size. All 26 (100%; 95% CI, 89.1-100%) malignant nodules grew. The mean (± SD) growth rate of adenomas was 1.0 ± 0.67 mm/year (range, 0.3-2.8 mm/year), compared with 58.4 ± 78.5 mm/year (range, 5.8-395.4 mm/year) for malignant nodules (p < 0.001). A growth rate of 3 mm/year distinguished adenomas from malignant nodules with a sensitivity of 100% (95% CI, 86.8-100%) and a specificity of 100% (95% CI, 96.6-100%). CONCLUSION. Approximately one-third of radiologically proven adrenal adenomas grew, all of which grew at a rate less than 3 mm/year. All malignant adrenal nodules grew, and all at a rate greater than 5 mm/year.

Astrocyte HIF-2α supports learning in a passive avoidance paradigm under hypoxic stress.

BACKGROUND: The brain is extensively vascularized, uses20% of the body's oxygen, and is highly sensitive to changes in oxygen. While synaptic plasticity and memory are impaired in healthy individuals by exposure to mild hypoxia, aged individuals appear to be even more sensitive. Aging is associated with progressive failure in pulmonary and cardiovascular systems, exposing the aged to both chronic and superimposed acute hypoxia. The HIF proteins, the "master regulators" of the cellular response to hypoxia, are robustly expressed in neurons and astrocytes. Astrocytes support neurons and synaptic plasticity via complex metabolic and trophic mechanisms. The activity of HIF proteins in the brain is diminished with aging, and the increased exposure to chronic and acute hypoxia with aging combined with diminished HIF activity may impair synaptic plasticity. PURPOSE: Herein, we test the hypothesis that astrocyte HIF supports synaptic plasticity and learning upon hypoxia. MATERIALS AND METHODS: An Astrocyte-specific HIF loss-of-function model was employed, where knock-out of HIF-1α or HIF-2α in GFAP expressing cells was accomplished by cre-mediated recombination. Animals were tested for behavioral (open field and rotarod), learning (passive avoidance paradigm), and electrophysiological (long term potentiation) responses to mild hypoxic challenge. RESULTS: In an astrocyte-specific HIF loss-of-function model followed by mild hypoxia, we identified that the depletion of HIF-2α resulted in an impaired passive avoidance learning performance. This was accompanied by an attenuated response to induction in long-term potentiation (LTP), suggesting that the hippocampal circuitry was perturbed upon hypoxic exposure following HIF-2α loss in astrocytes, and not due to hippocampal cell death. We investigated HIF-regulated trophic and metabolic target genes and found that they were not regulated by HIF-2α, suggesting that these specific targets may not be involved in mediating the phenotypes observed. CONCLUSION: Together, these results point to a role for HIF-2α in the astrocyte's regulatory role in synaptic plasticity and learning under hypoxia and suggest that even mild, acute hypoxic challenges can impair cognitive performance in the aged population who harbor impaired HIF function.

Cyclooxygenase-2, Asymmetric Dimethylarginine, and the Cardiovascular Hazard From Nonsteroidal Anti-Inflammatory Drugs.

BACKGROUND: Large-scale, placebo-controlled trials established that nonsteroidal anti-inflammatory drugs confer a cardiovascular hazard: this has been attributed to depression of cardioprotective products of cyclooxygenase (COX)-2, especially prostacyclin. An alternative mechanism by which nonsteroidal anti-inflammatory drugs might constrain cardioprotection is by enhancing the formation of methylarginines in the kidney that would limit the action of nitric oxide throughout the vasculature. METHODS: Targeted and untargeted metabolomics were used to investigate the effect of COX-2 deletion or inhibition in mice and in osteoarthritis patients exposed to nonsteroidal anti-inflammatory drugs on the l-arginine/nitric oxide pathway. RESULTS: Analysis of the plasma and renal metabolome was performed in postnatal tamoxifen-inducible Cox-2 knockout mice, which exhibit normal renal function and blood pressure. This revealed no changes in arginine and methylarginines compared with their wild-type controls. Moreover, the expression of genes in the l-arginine/nitric oxide pathway was not altered in the renal medulla or cortex of tamoxifen inducible Cox-2 knockout mice. Therapeutic concentrations of the selective COX-2 inhibitors, rofecoxib, celecoxib, and parecoxib, none of which altered basal blood pressure or renal function as reflected by plasma creatinine, failed to elevate plasma arginine and methylarginines in mice. Finally, plasma arginine or methylarginines were not altered in osteoarthritis patients with confirmed exposure to nonsteroidal anti-inflammatory drugs that inhibit COX-1 and COX-2. By contrast, plasma asymmetrical dimethylarginine was increased in mice infused with angiotensin II sufficient to elevate blood pressure and impair renal function. Four weeks later, blood pressure, plasma creatinine, and asymmetrical dimethylarginine were restored to normal levels. The increase in asymmetrical dimethylarginine in response to infusion with angiotensin II in celecoxib-treated mice was also related to transient impairment of renal function. CONCLUSIONS: Plasma methylarginines are not altered by COX-2 deletion or inhibition but rather are elevated coincident with renal compromise.

Extraction and Analysis of Pan-metabolome Polar Metabolites by Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry (UPLC-MS/MS).

Modern triple quadrupole mass spectrometers provide the ability to detect and quantify a large number of metabolites using tandem mass spectrometry (MS/MS). Liquid chromatography (LC) is advantageous, as it does not require derivatization procedures and a large diversity in physiochemical characteristics of analytes can be accommodated through a variety of column chemistries. Recently, the comprehensive optimization of LC-MS metabolomics using design of experiments (COLMeD) approach has been described and used by our group to develop robust LC-MS workflows (Rhoades and Weljie, 2016). The optimized LC-MS/MS method described here has been utilized extensively for metabolomics analysis of polar metabolites. Typically, tissue or biofluid samples are extracted using a modified Bligh-Dyer protocol (Bligh and Dyer, 1959; Tambellini et al., 2013). The protocol described herein describes this workflow using targeted polar metabolite multiple reaction monitoring (MRM) from tissues and biofluids via ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). This workflow has been utilized extensively for chronometabolic analysis (Krishnaiah et al., 2017), with applications generalized to other types of analyses as well (Sengupta et al., 2017; Sivanand et al., 2017).