Coupling Microdroplet-Based Sample Preparation, Multiplexed Isobaric Labeling, and Nanoflow Peptide Fractionation for Deep Proteome Profiling of the Tissue Microenvironment.

There is increasing interest in developing in-depth proteomic approaches for mapping tissue heterogeneity in a cell-type-specific manner to better understand and predict the function of complex biological systems such as human organs. Existing spatially resolved proteomics technologies cannot provide deep proteome coverage due to limited sensitivity and poor sample recovery. Herein, we seamlessly combined laser capture microdissection with a low-volume sample processing technology that includes a microfluidic device named microPOTS (microdroplet processing in one pot for trace samples), multiplexed isobaric labeling, and a nanoflow peptide fractionation approach. The integrated workflow allowed us to maximize proteome coverage of laser-isolated tissue samples containing nanogram levels of proteins. We demonstrated that the deep spatial proteomics platform can quantify more than 5000 unique proteins from a small-sized human pancreatic tissue pixel (∼60,000 µm2) and differentiate unique protein abundance patterns in pancreas. Furthermore, the use of the microPOTS chip eliminated the requirement for advanced microfabrication capabilities and specialized nanoliter liquid handling equipment, making it more accessible to proteomic laboratories.

The impact of heat stress on the human plasma lipidome.

The year of 2023 displayed the highest average global temperatures since it has been recorded-the duration and severity of extreme heat are projected to increase. Rising global temperatures represent a major public health threat, especially to occupations exposed to hot environments, such as construction and agricultural workers, and first responders. Despite efforts of the scientific community, there is still a need to characterize the pathophysiological processes leading to heat related illness and develop biomarkers that can predict its onset. Here, we performed a plasma lipidomic analysis on male and female subjects who underwent heat tolerance testing (HTT), consisting of a 2-h treadmill walk at 5 km/h with 2% inclination at a controlled temperature of 40°C. We identified 995 lipids from 27 classes, with nearly half of all detected lipids being responsive to HTT. Lipid classes related to substrate utilization were predominantly affected by HTT, with a downregulation of triacylglycerols and upregulation of free fatty acids and acyl-carnitines (CARs). We additionally examined correlations between changes in plasma lipids by using the physiological strain index (PSI). Here, even chain CAR 4:0, 14:0 and 16:1, suggested by-products of incomplete beta oxidation, and diacylglycerols displayed the highest correlation to PSI. PSI did not correlate with plasma lactate levels, suggesting that correlations between even chain CARs and PSI is related to metabolic efficiency versus physical exertion. Overall, our results show that HTT has a strong impact on the plasma lipidome and that metabolic inefficiencies may underlie heat intolerance.

PeakQC: A Software Tool for Omics-Agnostic Automated Quality Control of Mass Spectrometry Data.

Mass spectrometry is broadly employed to study complex molecular mechanisms in various biological and environmental fields, enabling 'omics' research such as proteomics, metabolomics, and lipidomics. As study cohorts grow larger and more complex with dozens to hundreds of samples, the need for robust quality control (QC) measures through automated software tools becomes paramount to ensure the integrity, high quality, and validity of scientific conclusions from downstream analyses and minimize the waste of resources. Since existing QC tools are mostly dedicated to proteomics, automated solutions supporting metabolomics are needed. To address this need, we developed the software PeakQC, a tool for automated QC of MS data that is independent of omics molecular types (i.e., omics-agnostic). It allows automated extraction and inspection of peak metrics of precursor ions (e.g., errors in mass, retention time, arrival time) and supports various instrumentations and acquisition types, from infusion experiments or using liquid chromatography and/or ion mobility spectrometry front-end separations and with/without fragmentation spectra from data-dependent or independent acquisition analyses. Diagnostic plots for fragmentation spectra are also generated. Here, we describe and illustrate PeakQC's functionalities using different representative data sets, demonstrating its utility as a valuable tool for enhancing the quality and reliability of omics mass spectrometry analyses.

Assessing Digital Breast Tomosynthesis Impact on Early Cancer Detection: Insights from Consecutive Screening.

Background Analysis of how digital breast tomosynthesis (DBT) screening affects consecutive screening performance is important to estimate its future value in screening. Purpose To evaluate whether DBT contributes to early detection of breast cancer by assessing cancer detection rates (CDRs), including the fraction of invasive cancers and cancer subtypes in consecutive routine digital mammography (DM). Materials and Methods The paired prospective Malmö Breast Tomosynthesis Screening Trial (MBTST) was performed between January 2010 and February 2015. Participating women underwent one-view DBT and two-view DM at one screening occasion. In this secondary analysis, women were followed up through their first (DM1) and second (DM2) consecutive two-view DM screening rounds after MBTST participation. Cancer diagnoses were identified by referencing records. A logistic regression model, adjusted for age, was used to calculate the odds of luminal A-like cancers with use of the MBTST as reference. Results There were 14 848 final participants in the MBTST (median age, 57 years [IQR, 49-65 years]). Of those, 12 876 women were screened in DM1 (median age, 58 years [IQR, 50-66 years]) and 10 883 were screened in DM2 (median age, 59 years [IQR, 51-67 years]). Compared with CDRs in the trial of 6.5 of 1000 women (95% CI: 5.2, 7.9) for DM and 8.7 of 1000 women (95% CI: 7.3, 10.3) for DBT, the CDR was lower in DM1 (4.6 of 1000 women [95% CI: 3.6, 5.9]) and DM2 (5.3 of 1000 women [95% CI: 4.1, 6.9]). The proportion of invasive cancers was 84.9% (118 of 139 cancers) in the MBTST; the corresponding numbers were 66% (39 of 59 cancers) for DM1 and 83% (50 of 60 cancers) for DM2. The odds of luminal A-like cancers were lower in DM1 at 0.28 (95% CI: 0.12, 0.66 [P = .004]) but not in DM2 at 0.80 (95% CI: 0.40, 1.58 [P = .52]) versus screening in the MBTST. Conclusion CDR and the fraction of invasive cancers were lower in DM1 and then increased in DM2 following the MBTST, indicating earlier cancer detection mainly due to increased detection of luminal A-like cancers in DBT screening. Clinical trials registration no. NCT01091545 © RSNA, 2024 See also the editorial by Hooley and Philpotts in this issue.

Influence of the Menstrual Cycle and Training on the Performance of a Perturbed Single-Leg Squatting Task in Female Collegiate Athletes.

Background: Anterior cruciate ligament (ACL) injuries often occur when an athlete experiences an unexpected disruption, or perturbation, during sports. ACL injury rates may also be influenced by the menstrual cycle. Purpose: To determine whether training adaptations to knee control and muscle activity during a perturbed single-leg squatting (SLS) task depend on menstrual cycle phase in female athletes. Study Design: Controlled laboratory study. Methods: A total of 21 healthy female collegiate athletes (current or former [<3 years]) who competed in 9 different sports performed an SLS task in which they attempted to match their knee position (user signal) to a target signal. The protocol consisted of a 9-condition pretest, 5 sets of 3 training trials, and a 9-condition posttest. One perturbation was delivered in each condition by altering the resistance of the device. Sagittal knee control (absolute error between the target signal and user signal) was assessed using a potentiometer. Muscle activity during perturbed squat cycles was normalized to maximal activation and to corresponding muscle activity during unperturbed squat cycles (%unperturbed) within the same test condition. Athletes performed the protocol during a distinct menstrual cycle phase (early follicular [EF], late follicular [LF], midluteal [ML]). Two-way mixed analysis of variance was used to determine the effects of the menstrual cycle and training on knee control and muscle activity during task performance. Venous blood was collected for hormonal analysis, and a series of health questionnaires and anthropometric measures were also assessed to determine differences among the menstrual cycle groups. Results: After training, athletes demonstrated better knee control during the perturbed squat cycles (lower absolute error, P < .001) and greater soleus feedback responses to the perturbation (%unperturbed, P = .035). Better knee control was demonstrated in the ML phase versus the EF phase during unperturbed and perturbed squat cycles (P < .039 for both). Quadriceps activation was greater in the ML phase compared with the EF and LF phases, both immediately before and after the perturbation (P < .001 for all). Conclusion: Athletes learned to improve knee control during the perturbed performance regardless of menstrual cycle phase. The best knee control and greatest quadriceps activation during the perturbed squatting task was found in the ML phase. Clinical Relevance: These findings may correspond to a lower incidence of ACL injury in the luteal phase and alterations in exercise performance across the menstrual cycle.

A fast and sensitive size-exclusion chromatography method for plasma extracellular vesicle proteomic analysis.

Extracellular vesicles (EVs) carry diverse biomolecules derived from their parental cells, making their components excellent biomarker candidates. However, purifying EVs is a major hurdle in biomarker discovery since current methods require large amounts of samples, are time-consuming and typically have poor reproducibility. Here we describe a simple, fast, and sensitive EV fractionation method using size exclusion chromatography (SEC) on a fast protein liquid chromatography (FPLC) system. Our method uses a Superose 6 Increase 5/150, which has a bed volume of 2.9 mL. The FPLC system and small column size enable reproducible separation of only 50 µL of human plasma in 15 min. To demonstrate the utility of our method, we used longitudinal samples from a group of individuals who underwent intense exercise. A total of 838 proteins were identified, of which, 261 were previously characterized as EV proteins, including classical markers, such as cluster of differentiation (CD)9 and CD81. Quantitative analysis showed low technical variability with correlation coefficients greater than 0.9 between replicates. The analysis captured differences in relevant EV proteins involved in response to physical activity. Our method enables fast and sensitive fractionation of plasma EVs with low variability, which will facilitate biomarker studies in large clinical cohorts.

A compendium of multi-omics data illuminating host responses to lethal human virus infections.

Human infections caused by viral pathogens trigger a complex gamut of host responses that limit disease, resolve infection, generate immunity, and contribute to severe disease or death. Here, we present experimental methods and multi-omics data capture approaches representing the global host response to infection generated from 45 individual experiments involving human viruses from the Orthomyxoviridae, Filoviridae, Flaviviridae, and Coronaviridae families. Analogous experimental designs were implemented across human or mouse host model systems, longitudinal samples were collected over defined time courses, and global multi-omics data (transcriptomics, proteomics, metabolomics, and lipidomics) were acquired by microarray, RNA sequencing, or mass spectrometry analyses. For comparison, we have included transcriptomics datasets from cells treated with type I and type II human interferon. Raw multi-omics data and metadata were deposited in public repositories, and we provide a central location linking the raw data with experimental metadata and ready-to-use, quality-controlled, statistically processed multi-omics datasets not previously available in any public repository. This compendium of infection-induced host response data for reuse will be useful for those endeavouring to understand viral disease pathophysiology and network biology.

Low-frequency electrically induced exercise after spinal cord injury: Physiologic challenge to skeletal muscle and feasibility for long-term use.

CONTEXT: Skeletal muscle has traditionally been considered a "force generator": necessary for purposes of locomotion, but expendable for non-ambulators who use wheelchairs, such as people with a spinal cord injury (SCI). Active skeletal muscle plays an indispensable role in regulating systemic metabolic functions, even in people with paralysis, but because of severe osteoporosis, high tetanic muscle forces induced with high frequency electrical stimulation may be risky for some individuals. The purpose of this study was to compare the physiologic muscle properties incurred by two low force/low frequency repetitive stimulation protocols (1 and 3 Hz); and, to assess the acceptability of each protocol among people with SCI. METHODS: Ten individuals with chronic SCI (12.9 years) and 11 individuals without SCI (NonSCI) participated in the study. Participants received either 1 or 3 Hz stimulation to the quadriceps muscle on Day 1, then the converse on Day 2. Each session consisted of 1000 stimulus pulses. RESULTS: The initial and maximum forces were similar for the 1 and 3 Hz frequencies. The fatigue index (FI) for SCI and NonSCI groups were lower (P < 0.007) for 3 Hz than for 1 Hz (0.34 ± 0.17 versus 0.65 ± 0.16 and 0.72 ± 0.14 versus 0.87 ± 0.07, respectively). CONCLUSION: The 3 Hz stimulation offered the greatest physiological challenge and was perceived as more acceptable for long term use among people with SCI.

Computational tools and algorithms for ion mobility spectrometry-mass spectrometry.

Ion mobility spectrometry-mass spectrometry (IMS-MS or IM-MS) is a powerful analytical technique that combines the gas-phase separation capabilities of IM with the identification and quantification capabilities of MS. IM-MS can differentiate molecules with indistinguishable masses but different structures (e.g., isomers, isobars, molecular classes, and contaminant ions). The importance of this analytical technique is reflected by a staged increase in the number of applications for molecular characterization across a variety of fields, from different MS-based omics (proteomics, metabolomics, lipidomics, etc.) to the structural characterization of glycans, organic matter, proteins, and macromolecular complexes. With the increasing application of IM-MS there is a pressing need for effective and accessible computational tools. This article presents an overview of the most recent free and open-source software tools specifically tailored for the analysis and interpretation of data derived from IM-MS instrumentation. This review enumerates these tools and outlines their main algorithmic approaches, while highlighting representative applications across different fields. Finally, a discussion of current limitations and expectable improvements is presented.

Regulation of ß-cell death by ADP-ribosylhydrolase ARH3 via lipid signaling in insulitis.

BACKGROUND: Lipids are regulators of insulitis and ß-cell death in type 1 diabetes development, but the underlying mechanisms are poorly understood. Here, we investigated how the islet lipid composition and downstream signaling regulate ß-cell death. METHODS: We performed lipidomics using three models of insulitis: human islets and EndoC-ßH1 ß cells treated with the pro-inflammatory cytokines interlukine-1ß and interferon-γ, and islets from pre-diabetic non-obese mice. We also performed mass spectrometry and fluorescence imaging to determine the localization of lipids and enzyme in islets. RNAi, apoptotic assay, and qPCR were performed to determine the role of a specific factor in lipid-mediated cytokine signaling. RESULTS: Across all three models, lipidomic analyses showed a consistent increase of lysophosphatidylcholine species and phosphatidylcholines with polyunsaturated fatty acids and a reduction of triacylglycerol species. Imaging assays showed that phosphatidylcholines with polyunsaturated fatty acids and their hydrolyzing enzyme phospholipase PLA2G6 are enriched in islets. In downstream signaling, omega-3 fatty acids reduce cytokine-induced ß-cell death by improving the expression of ADP-ribosylhydrolase ARH3. The mechanism involves omega-3 fatty acid-mediated reduction of the histone methylation polycomb complex PRC2 component Suz12, upregulating the expression of Arh3, which in turn decreases cell apoptosis. CONCLUSIONS: Our data provide insights into the change of lipidomics landscape in ß cells during insulitis and identify a protective mechanism by omega-3 fatty acids. Video Abstract.

Mapping microhabitats of lignocellulose decomposition by a microbial consortium.

The leaf-cutter ant fungal garden ecosystem is a naturally evolved model system for efficient plant biomass degradation. Degradation processes mediated by the symbiotic fungus Leucoagaricus gongylophorus are difficult to characterize due to dynamic metabolisms and spatial complexity of the system. Herein, we performed microscale imaging across 12-µm-thick adjacent sections of Atta cephalotes fungal gardens and applied a metabolome-informed proteome imaging approach to map lignin degradation. This approach combines two spatial multiomics mass spectrometry modalities that enabled us to visualize colocalized metabolites and proteins across and through the fungal garden. Spatially profiled metabolites revealed an accumulation of lignin-related products, outlining morphologically unique lignin microhabitats. Metaproteomic analyses of these microhabitats revealed carbohydrate-degrading enzymes, indicating a prominent fungal role in lignocellulose decomposition. Integration of metabolome-informed proteome imaging data provides a comprehensive view of underlying biological pathways to inform our understanding of metabolic fungal pathways in plant matter degradation within the micrometer-scale environment.

A fast and sensitive size-exclusion chromatography method for plasma extracellular vesicle proteomic analysis.

Extracellular vesicles (EVs) carry diverse biomolecules derived from their parental cells, making their components excellent biomarker candidates. However, purifying EVs is a major hurdle in biomarker discovery since current methods require large amounts of samples, are time-consuming and typically have poor reproducibility. Here we describe a simple, fast, and sensitive EV fractionation method using size exclusion chromatography (SEC) on a fast protein liquid chromatography (FPLC) system. Our method uses a Superose 6 Increase 5/150, which has a bed volume of 2.9 mL. The FPLC system and small column size enable reproducible separation of only 50 µL of human plasma in 15 minutes. To demonstrate the utility of our method, we used longitudinal samples from a group of individuals that underwent intense exercise. A total of 838 proteins were identified, of which, 261 were previously characterized as EV proteins, including classical markers, such as cluster of differentiation (CD)9 and CD81. Quantitative analysis showed low technical variability with correlation coefficients greater than 0.9 between replicates. The analysis captured differences in relevant EV-proteins involved in response to physical activity. Our method enables fast and sensitive fractionation of plasma EVs with low variability, which will facilitate biomarker studies in large clinical cohorts.

Deciphering ApoE Genotype-Driven Proteomic and Lipidomic Alterations in Alzheimer's Disease Across Distinct Brain Regions.

Alzheimer's disease (AD) is a neurodegenerative disease with a complex etiology influenced by confounding factors such as genetic polymorphisms, age, sex, and race. Traditionally, AD research has not prioritized these influences, resulting in dramatically skewed cohorts such as three times the number of Apolipoprotein E (APOE) ε4-allele carriers in AD relative to healthy cohorts. Thus, the resulting molecular changes in AD have previously been complicated by the influence of apolipoprotein E disparities. To explore how apolipoprotein E polymorphism influences AD progression, 62 post-mortem patients consisting of 33 AD and 29 controls (Ctrl) were studied to balance the number of ε4-allele carriers and facilitate a molecular comparison of the apolipoprotein E genotype. Lipid and protein perturbations were assessed across AD diagnosed brains compared to Ctrl brains, ε4 allele carriers (APOE4+ for those carrying 1 or 2 ε4s and APOE4- for non-ε4 carriers), and differences in ε3ε3 and ε3ε4 Ctrl brains across two brain regions (frontal cortex (FCX) and cerebellum (CBM)). The region-specific influences of apolipoprotein E on AD mechanisms showcased mitochondrial dysfunction and cell proteostasis at the core of AD pathophysiology in the post-mortem brains, indicating these two processes may be influenced by genotypic differences and brain morphology.

Assessment of Beverage Trends and Replacing Nondairy Caloric Beverages with Milk at Meals across Childhood Improves Intake of Key Nutrients at Risk of Inadequate Consumption: An NHANES Modeling Study.

Background: Milk is a key source of important nutrients including the nutrients of public health concern. However, most Americans do not meet current (dairy) United States Department of Agriculture (USDA) dietary guideline recommendations, and the intake has been declining. Objective: The aim of this study was to investigate milk and beverage intake trends and nutrient intakes from these products in United States children aged 6-18 y and to model the effect of isocaloric substitution of nondairy beverages with milk. Methods: Data from National Health and Nutrition Examination Survey (NHANES) 2001-2018 for children age 6-8 (N = 4696), 9-13 (N = 8117) and 14-18 y (N = 8514) were used with milk and other beverage intakes determined from the first 24-h in-person dietary recall. Nutrient intake was determined using the NHANES cycle-specific total nutrient intake files. Nutrient modeling was performed by isocaloric substitution with milk of all nondairy beverages consumed during lunch and dinner meals combined. Sample-weighted analyses were performed using SAS 9.4. Results: Between ages 6-8 and 14-18 y, daily intake of milk and flavored milk decreased by 10% and 62%, respectively, while daily intake of caloric beverages excluding milk increased by 96%. Daily intake from caloric beverages and milk combined decreased for fiber, protein, fat, saturated fat, calcium, magnesium, potassium, vitamin A, and vitamin D and increased for energy, carbohydrates, added sugars, and folate between ages 6-8 y and 14-18 y. Isocaloric substitution of all caloric nondairy beverages at meals with milk (using nutrient contribution of USDA milk, not further specified (NFS)) resulted in increases in protein, fat, saturated fat, calcium, magnesium, potassium, sodium, vitamin A, folate, vitamin B12, and vitamin D and decreases in carbohydrate, fiber, and added sugar. Conclusion: These findings provide additional evidence to support dietary recommendations for milk, and efforts should be made on behalf of leading health professionals and childhood meal programs to highlight milk as a beverage of choice in children and adolescents.

Identification of a specific exporter that enables high production of aconitic acid in Aspergillus pseudoterreus.

Aconitic acid is an unsaturated tricarboxylic acid that is attractive for its potential use in manufacturing biodegradable and biocompatible polymers, plasticizers, and surfactants. Previously Aspergillus pseudoterreus was engineered as a platform to produce aconitic acid by deleting the cadA (cis-aconitic acid decarboxylase) gene in the itaconic acid biosynthetic pathway. In this study, the aconitic acid transporter gene (aexA) was identified using comparative global discovery proteomics analysis between the wild-type and cadA deletion strains. The protein AexA belongs to the Major Facilitator Superfamily (MFS). Deletion of aexA almost abolished aconitic acid secretion, while its overexpression led to a significant increase in aconitic acid production. Transportation of aconitic acid across the plasma membrane is a key limiting step in its production. In vitro, proteoliposome transport assay further validated AexA's function and substrate specificity. This research provides new approaches to efficiently pinpoint and characterize exporters of fungal organic acids and accelerate metabolic engineering to improve secretion capability and lower the cost of bioproduction.

The Accuracy of Ankle Eccentric Torque Control Explains Dynamic Postural Control During the Y-Balance Test.

Background: The Y-Balance Test (YBT), especially the posteromedial (PM) reach direction (PM-YBT), is able to identify dynamic postural control deficits in those who have ankle instability. However, there still exists a need to understand how sensorimotor function at the ankle explains the performance during the PM-YBT. Hypothesis/Purpose: The purpose of this study was to determine whether the ability to accurately control eccentric ankle torque explained PM-YBT performance. It was hypothesized that eccentric dorsiflexion/plantarflexion torque control would be positively related to the maximum reach distance (MRD) of PM-YBT. Study Design: Cross-sectional study. Methods: Twelve healthy subjects performed the PM-YBT, maximum voluntary isometric contractions (MVIC) for both dorsiflexion and plantarflexion muscle strength, and then the torque control testing of the ankle. The torque control testing provided a target torque level on a screen in front of the subject and passive rotations of the ankle joint in the sagittal plane at 10 deg/sec between plantarflexion to dorsiflexion. Subjects were then instructed to eccentrically contract the dorsiflexors and plantar flexors to generate torque while the ankle joint rotated. The accuracy of torque control during eccentric dorsiflexion and plantarflexion by calculating absolute errors, the area between the target torque and the produced torque were evaluated. Tibialis anterior and soleus muscle activities were simultaneously recorded during testing. A step-wise linear regression model was used to determine the best model predicted the MRD of the PM-YBT (PM-MRD). Results: A step-wise linear regression developed a model explaining only eccentric dorsiflexion torque control predicted higher PM-MRD score (R2 = 44%, F1,10 = 7.94, ß = -0.67, p = 0.02). Conclusion: The accuracy of torque control during eccentric dorsiflexion predicts better performance in the PM-YBT. Level of Evidence: 3b.

Elucidating regulatory processes of intense physical activity by multi-omics analysis.

BACKGROUND: Physiological and biochemical processes across tissues of the body are regulated in response to the high demands of intense physical activity in several occupations, such as firefighting, law enforcement, military, and sports. A better understanding of such processes can ultimately help improve human performance and prevent illnesses in the work environment. METHODS: To study regulatory processes in intense physical activity simulating real-life conditions, we performed a multi-omics analysis of three biofluids (blood plasma, urine, and saliva) collected from 11 wildland firefighters before and after a 45 min, intense exercise regimen. Omics profiles post- versus pre-exercise were compared by Student's t-test followed by pathway analysis and comparison between the different omics modalities. RESULTS: Our multi-omics analysis identified and quantified 3835 proteins, 730 lipids and 182 metabolites combining the 3 different types of samples. The blood plasma analysis revealed signatures of tissue damage and acute repair response accompanied by enhanced carbon metabolism to meet energy demands. The urine analysis showed a strong, concomitant regulation of 6 out of 8 identified proteins from the renin-angiotensin system supporting increased excretion of catabolites, reabsorption of nutrients and maintenance of fluid balance. In saliva, we observed a decrease in 3 pro-inflammatory cytokines and an increase in 8 antimicrobial peptides. A systematic literature review identified 6 papers that support an altered susceptibility to respiratory infection. CONCLUSION: This study shows simultaneous regulatory signatures in biofluids indicative of homeostatic maintenance during intense physical activity with possible effects on increased infection susceptibility, suggesting that caution against respiratory diseases could benefit workers on highly physical demanding jobs.

Direct Transverse Sinus Puncture for Transvenous Coil Embolization of Vein of Galen Malformations: Innovating Existing Techniques.

BACKGROUND AND IMPORTANCE: Vein of Galen malformations (VOGMs) are complex arteriovenous malformations in neonates and young children. Recent advances in endovascular interventions have drastically improved treatment and clinical outcomes in what was previously high-morbidity, high-mortality disease. The high-flow shunt pathophysiology in VOGMs can lead to dynamic changes in the malformation angioarchitecture, and over time patients can develop jugular bulb stenosis. In the setting of inaccessible transvenous access to the malformation for endovascular embolization in cases where transarterial embolization is inadequate, a combined surgical and endovascular technique must be used. We present the first successful modern-day application of direct puncture through transverse sinus for transvenous embolization of a VOGM. CLINICAL PRESENTATION: We present 2 unique cases of complex VOGM malformations in patients who had previously undergone staged endovascular embolization for reduction of flow within the malformation. On follow-up, in both cases, there was development of severe sigmoid sinus and jugular bulb stenosis, increasing intracranial venous congestion and causing marked clinical deterioration. The stenosis prevented traditional transvenous access and treatment. We describe a direct puncture transverse sinus access using a burr hole approach for endovascular transvenous embolization in both cases with successful clinical outcomes. CONCLUSION: Direct access using burr hole craniotomy to the transverse sinus for transvenous endovascular embolization is a safe approach in the setting of severe jugular bulb stenosis for treatment of VOGMs. This technique can be done efficiently to achieve complete flow elimination in the malformation, in cases where that is called for, without significant risks or complications related to the approach.

Impact of breast density on diagnostic accuracy in digital breast tomosynthesis versus digital mammography: results from a European screening trial.

BACKGROUND: The diagnostic accuracy of digital breast tomosynthesis (DBT) and digital mammography (DM) in breast cancer screening may vary per breast density subgroup. The purpose of this study was to evaluate which women, based on automatically assessed breast density subgroups, have the greatest benefit of DBT compared with DM in the prospective Malmö Breast Tomosynthesis Screening Trial. MATERIALS AND METHODS: The prospective European, Malmö Breast Tomosynthesis Screening Trial (n = 14,848, Jan. 27, 2010-Feb. 13, 2015) compared one-view DBT and two-view DM, with consensus meeting before recall. Breast density was assessed in this secondary analysis with the automatic software Laboratory for Individualized Breast Radiodensity Assessment. DBT and DM's diagnostic accuracies were compared by breast density quintiles of breast percent density (PD) and absolute dense area (DA) with confidence intervals (CI) and McNemar's test. The association between breast density and cancer detection was analyzed with logistic regression, adjusted for ages < 55 and ≥ 55 years and previous screening participation. RESULTS: In total, 14,730 women (median age: 58 years; inter-quartile range = 16) were included in the analysis. Sensitivity was higher and specificity lower for DBT compared with DM for all density subgroups. The highest breast PD quintile showed the largest difference in sensitivity and specificity at 81.1% (95% CI 65.8-90.5) versus 43.2% (95% CI 28.7-59.1), p < .001 and 95.5% (95% CI 94.7-96.2) versus 97.2% (95% CI 96.6-97.8), p < 0.001, respectively. Breast PD quintile was also positively associated with cancer detected via DBT at odds ratio 1.24 (95% CI 1.09-1.42, p = 0.001). CONCLUSION: Women with the highest breast density had the greatest benefit from digital breast tomosynthesis compared with digital mammography with increased sensitivity at the cost of slightly lower specificity. These results may influence digital breast tomosynthesis's use in an individualized screening program stratified by, for instance, breast density. TRIAL REGISTRATION: Trial registration at https://www. CLINICALTRIALS: gov : NCT01091545, registered March 24, 2010.

Exercise Prescription Principles among Physicians and Physical Therapists for Patients with Impaired Glucose Control: A Cross-Sectional Study.

Exercise confers a multitude of benefits with limited adverse side effects, making it a powerful "medication" for a plethora of diseases. In people living with uncontrolled glucose levels, exercise can be an effective "medication" to assist in the management of hyperglycemia. We sought to survey healthcare providers (physicians and physical therapists) to determine the current state of exercise recommendation for people with glucose control issues. Healthcare providers were surveyed from six academic medical centers in the Midwest to determine the recommended exercise parameters (type, frequency, duration, intensity, and timing) for patients with glucose control issues. Data from 209 practitioners who completed the survey were used for analysis. Chi-square tests were used to determine differences in exercise recommendations between physical therapists (PTs) and physicians (MD/DOs). PTs and MD/DOs recommended similar exercise parameters. Of all respondents, 78.9% recommended exercise to patients with glucose control issues. Respondents who considered themselves to be active exercisers were more likely to recommend exercise than those who were not exercisers. Only 6.1% of all respondents recommended post-meal exercise. Healthcare providers overwhelmingly recommended exercise for people with glucose control issues, but the "timing" is not congruent with best practice recommendations.