Structural Elucidation of Ubiquitin via Gas-Phase Ion/Ion Cross-Linking Reactions Using Sodium-Cationized Reagents Coupled with Infrared Multiphoton Dissociation.

Accurate structural determination of proteins is critical to understanding their biological functions and the impact of structural disruption on disease progression. Gas-phase cross-linking mass spectrometry (XL-MS) via ion/ion reactions between multiply charged protein cations and singly charged cross-linker anions has previously been developed to obtain low-resolution structural information on proteins. This method significantly shortens experimental time relative to conventional solution-phase XL-MS but has several technical limitations: (1) the singly deprotonated N-hydroxysulfosuccinimide (sulfo-NHS)-based cross-linker anions are restricted to attachment at neutral amine groups of basic amino acid residues and (2) analyzing terminal cross-linked fragment ions is insufficient to unambiguously localize sites of linker attachment. Herein, we demonstrate enhanced structural information for alcohol-denatured A-state ubiquitin obtained from an alternative gas-phase XL-MS approach. Briefly, singly sodiated ethylene glycol bis(sulfosuccinimidyl succinate) (sulfo-EGS) cross-linker anions enable covalent cross-linking at both ammonium and amine groups. Additionally, covalently modified internal fragment ions, along with terminal b-/y-type counterparts, improve the determination of linker attachment sites. Molecular dynamics simulations validate experimentally obtained gas-phase conformations of denatured ubiquitin. This method has identified four cross-linking sites across 8+ ubiquitin, including two new sites in the N-terminal region of the protein that were originally inaccessible in prior gas-phase XL approaches. The two N-terminal cross-linking sites suggest that the N-terminal half of ubiquitin is more compact in gas-phase conformations. By comparison, the two C-terminal linker sites indicate the signature transformation of this region of the protein from a native to a denatured conformation. Overall, the results suggest that the solution-phase secondary structures of the A-state ubiquitin are conserved in the gas phase. This method also provides sufficient sensitivity to differentiate between two gas-phase conformers of the same charge state with subtle structural variations.

T2 Black Synovitis in Musculoskeletal MRI: Disease Spectrum and Imaging Characteristics of Joint Diseases

The synovium may be affected by a wide spectrum of disorders, including inflammatory, infectious, degenerative, traumatic, hemorrhagic, and tumorous conditions. Magnetic resonance imaging (MRI) is a valuable imaging modality to characterize synovial disorders. Most abnormal lesions appear as areas of nonspecific high signal intensity on T2-weighted images (T2-WI) due to high water content or increased perfusion. However, T2 hypointensity can be attributed to blood components of varying ages, calcification, inorganic crystals, fibrosis, caseous necrosis and/or amyloid deposition. Hypointense lesions on T2-WI are infrequent and additional clinical and imaging characteristics can help to limit the list of differential diagnoses, which may include tenosynovial giant cell tumor, synovial chondromatosis, rheumatoid arthritis, tuberculous arthritis, chronic tophaceous gout, amyloid arthropathy, synovial hemangioma, lipoma arborescens and hemosiderotic synovitis. Recently, susceptibility weighted imaging has been developed and may contribute to more accurate diagnosis for deoxygenated blood and calcium. We review the MRI features of hypointense synovial lesions on MRI and emphasize the characteristic findings that suggest a correct diagnosis.

Relative Quantification of Lipid Isomers in Imaging Mass Spectrometry Using Gas-Phase Charge Inversion Ion/Ion Reactions and Infrared Multiphoton Dissociation.

Accurate structural identification of lipids in imaging mass spectrometry is critical to properly contextualizing spatial distributions with tissue biochemistry. Gas-phase charge inversion ion/ion reactions alter the ion type prior to dissociation to allow for more structurally informative fragmentation and improve lipid identification at the isomeric level. In this work, infrared multiphoton dissociation (IRMPD) was interfaced with a commercial hybrid Qh-FT-ICR mass spectrometer to enable the rapid fragmentation of gas-phase charge inversion ion/ion reaction products at every pixel in imaging mass spectrometry experiments. An ion/ion reaction between phosphatidylcholine (PC) monocations generated from rat brain tissue via matrix-assisted laser desorption/ionization (MALDI) and 1,4-phenylenediproprionic acid reagent dianions generated via electrospray ionization (ESI) followed by IRMPD of the resulting product ion complex produces selective fatty acyl chain cleavages indicative of fatty acyl carbon compositions in the lipid. Ion/ion reaction images using this workflow allow for mapping of the relative spatial distribution of multiple PC isomers under a single sum composition lipid identification. Lipid isomers display significantly different relative spatial distributions within rat brain tissue, highlighting the importance of resolving isomers in imaging mass spectrometry experiments.

Extraskeletal Osteochondroma in the Posterior Neck of a Middle-Aged Female: A Case Report.

Extraskeletal osteochondroma, a variant of chondroma, typically arises in the para-articular location of hands and feet. It is a rare disease and is particularly uncommon when joint components are not involved or localized away from joints. Herein, we report a case of extraskeletal osteochondroma in the posterior neck of a 66-year-old female. The characteristic radiologic finding of our case is presented, along with the typical findings of the disease and review of related literature reports.

Diagnostic performance of electron-density dual-energy CT in detection of cervical disc herniation in comparison with standard gray-scale CT and virtual non-calcium images.

OBJECTIVES: To assess the diagnostic performance of dual-energy CT (DECT) with electron-density (ED) image reconstruction compared with standard CT (SC) and virtual non-calcium (VNCa) image CT reconstruction for detecting cervical disc herniation. METHODS: This cross-sectional study was approved by the IRB. We enrolled 64 patients (336 intervertebral discs from C2/3 to C7/T1; mean age, 55 years; 17 women and 47 men) who underwent DECT with spectral reconstruction and 3-T MRI within 2 weeks between January 2018 and June 2020. Four radiologists independently evaluated the first image set of randomized SC, VNCa, and ED images to detect cervical disc herniation. After 8 weeks, the readers re-evaluated the second and the last image sets with an 8-week interval. MRI evaluations performed by two other experienced served as the reference standard. Comparing diagnostic performance between each images set was evaluated by a generalized estimating equation. RESULTS: A total of 233 cervical disc herniations were noted on MRI. For detecting cervical disc herniation, electron-density images showed higher sensitivity (94% [219/233; 95% CI, 90-97] vs. 76% [177/233; 70-81] vs. 69% [160/233; 62-76]) (p < 0.001) and similar specificity (90% [93/103; 83-95] vs. 89% [92/103; 82-96] vs. 90% [93/103; 83-95]) (p > 0.05) as SC and VNCa images, respectively. Inter-reader agreement for cervical disc herniation calculated among the four readers was moderate for all image sets (κ = 0.558 for ED, κ = 0.422 for SC, and κ = 0.449 for VNCa). CONCLUSION: DECT with ED reconstruction can improve cervical disc herniation detection and diagnostic confidence compared with SC and VNCa images. KEY POINTS: • Intervertebral discs with high material density are well visualized on electron-density images obtained from dual-energy CT. • Electron-density images showed much higher sensitivity and diagnostic accuracy than standard CT and virtual non-calcium images for the detection of cervical disc herniation. • Electron-density images can have false-negative results, especially for disc herniation with high signal intensity on T2W images and can show pseudo-disc extrusion at the lower cervical spine.

Improved segmented-scan spectral stitching for stable isotope resolved metabolomics (SIRM) by ultra-high-resolution Fourier transform mass spectrometry.

We have implemented a linear ion trap (LIT)-based SIM-stitching method for ultra-high-resolution Fourier transform mass spectrometry (FTMS) that increases the S/N over a wide m/z range compared to non-segmented wide full-scan (WFS) spectra. Here we described an improved segmented spectral scan stitching method that was based on quadrupole mass filter (QMF)-SIM, which overcame previous limitations of ion signal loss in LIT. This allowed for accurate representation of isotopologue distributions, both at natural abundance and in stable isotope-resolved metabolomics (SIRM)-based experiments. We also introduced a new spectral binning method that provided more precise and resolution-independent bins for irreversibly noise-suppressed FTMS spectra. We demonstrated a substantial improvement in S/N and sensitivity (typically > 10-fold) for 13C labeled lipid extracts of human macrophages grown as three-dimensional (3D) cell culture, with detection of an increased number of 13C isotopologue ions. The method also enabled analysis of extracts from very limited biological samples.

Systemic effects of fluoroscopically guided epidural steroid injection with dexamethasone.

BACKGROUND: Epidural steroid injections (ESIs) have been widely used in managing spinal pain. Dexamethasone has recently emerged as a useful drug in this setting, relative to particulate steroids, although the associated systemic effects have not been fully elucidated. This study aimed to investigate the incidences and types of systemic effects after fluoroscopically guided ESI with dexamethasone. METHODS: This retrospective study included 888 ESIs with dexamethasone (fluoroscopically guided at the cervical and lumbosacral levels) performed on 825 patients during January to June 2017. Data regarding systemic effects were collected via telephone interviews using a standardized questionnaire at 2 weeks after the procedure. Data on patient demographic, clinical, and procedural characteristics were collected and analyzed to identify factors that were associated with systemic effects. All statistical analyses were performed using the chi-squared test. RESULTS: Among the 825 patients, 40 patients (4.8%) experienced systemic effects during the 2-week follow-up period. The most common systemic effect was facial flushing (12 patients, 1.5%), which was followed by urticaria (7 patients, 0.8%) and insomnia (7 patients, 0.8%). A history of spine surgery was significantly associated with the occurrence of systemic effects (P = 0.036). Systemic effects were significantly more common for injections at the cervical level than at the lumbar level (P = 0.019). CONCLUSIONS: Approximately 4.8% of the patients who underwent ESI with dexamethasone experienced minor and transient systemic effects. These effects were more common in patients who had undergone a previous spine surgery or received a cervical ESI.

Exosomal lipids for classifying early and late stage non-small cell lung cancer.

Lung cancer is the leading cause of cancer deaths in the United States. Patients with early stage lung cancer have the best prognosis with surgical removal of the tumor, but the disease is often asymptomatic until advanced disease develops, and there are no effective blood-based screening methods for early detection of lung cancer in at-risk populations. We have explored the lipid profiles of blood plasma exosomes using ultra high-resolution Fourier transform mass spectrometry (UHR-FTMS) for early detection of the prevalent non-small cell lung cancers (NSCLC). Exosomes are nanovehicles released by various cells and tumor tissues to elicit important biofunctions such as immune modulation and tumor development. Plasma exosomal lipid profiles were acquired from 39 normal and 91 NSCLC subjects (44 early stage and 47 late stage). We have applied two multivariate statistical methods, Random Forest (RF) and Least Absolute Shrinkage and Selection Operator (LASSO) to classify the data. For the RF method, the Gini importance of the assigned lipids was calculated to select 16 lipids with top importance. Using the LASSO method, 7 features were selected based on a grouped LASSO penalty. The Area Under the Receiver Operating Characteristic curve for early and late stage cancer versus normal subjects using the selected lipid features was 0.85 and 0.88 for RF and 0.79 and 0.77 for LASSO, respectively. These results show the value of RF and LASSO for metabolomics data-based biomarker development, which provide robust an independent classifiers with sparse data sets. Application of LASSO and Random Forests identifies lipid features that successfully distinguish early stage lung cancer patient from healthy individuals.

Efficacy and outcome predictors of fluoroscopy-guided facet joint injection for spondylolysis.

OBJECTIVE: To assess the efficacy of fluoroscopy-guided facet joint injection for symptomatic spondylolysis and to analyze the outcome predictors. MATERIALS AND METHODS: This study included 108 patients who underwent fluoroscopy-guided facet joint steroid injection for symptomatic spondylolysis with follow-up medical records from January 2013 to December 2016. Among them, 47 patients underwent concomitant epidural steroid injection. Response to injection was assessed at the initial follow-up. The symptom-free interval was analyzed using the Kaplan-Meier method. Outcome predictors were statistically analyzed using independent t test and Chi-square test. RESULTS: The facet joint injection was effective in 52 of 108 (48.1%) patients at initial follow-up. Recurrence was reported in 21 of these 52 (40.4%) patients. For the 52 patients in whom facet joint injection was effective, the median symptom-free interval was 298 days (95% confidence interval, 29-567 days). No significant difference was found in the response between the group with facet joint injection only (n = 61) and the group with facet joint injection and additional ESI (n = 47). Additionally, no significant outcome predictors were detected. CONCLUSIONS: Fluoroscopy-guided facet joint injection may potentially be an effective therapy for providing prolonged pain relief and avoiding unnecessary surgery for symptomatic spondylolysis without significant outcome predictors.

Prevalence of Frailty and Aging-Related Health Conditions in Older Koreans in Rural Communities: a Cross-Sectional Analysis of the Aging Study of Pyeongchang Rural Area.

Frailty has been previously studied in Western countries and the urban Korean population; however, the burden of frailty and geriatric conditions in the aging populations of rural Korean communities had not yet been determined. Thus, we established a population-based prospective study of adults aged ≥ 65 years residing in rural communities of Korea between October 2014 and December 2014. All participants underwent comprehensive geriatric assessment that encompassed the assessment of cognitive and physical function, depression, nutrition, and body composition using bioimpedance analysis. We determined the prevalence of frailty based on the Cardiovascular Health Study (CHS) and Korean version of FRAIL (K-FRAIL) criteria, as well as geriatric conditions. We recruited 382 adults (98% of eligible adults; mean age: 74 years; 56% women). Generally, sociodemographic characteristics were similar to those of the general rural Korean population. Common geriatric conditions included instrumental activity of daily living disability (39%), malnutrition risk (38%), cognitive dysfunction (33%), multimorbidity (32%), and sarcopenia (28%), while dismobility (8%), incontinence (8%), and polypharmacy (3%) were less common conditions. While more individuals were classified as frail according to the K-FRAIL criteria (27%) than the CHS criteria (17%), the CHS criteria were more strongly associated with prevalent geriatric conditions. Older Koreans living in rural communities have a significant burden of frailty and geriatric conditions that increase the risk of functional decline, poor quality of life, and mortality. The current study provides a basis to guide public health professionals and policy-makers in prioritizing certain areas of care and designing effective public health interventions to promote healthy aging of this vulnerable population.

Acute benign vertebral compression fractures: "see-through sign" on contrast-enhanced MR images.

PURPOSE: To retrospectively evaluate the diagnostic role of the contrast-enhanced MRI (CE-MRI) for differentiation between benign VCFs and malignant VCFs focusing on the internal transparent trabecular bone on CE-MRI (the "see-through sign"). MATERIALS AND METHODS: The institutional review board approved this study and informed consent was waived due to the retrospective nature of the study. From January 2012 to December 2013, all 149 consecutive benign or malignant VCF patients were enrolled for consideration in this study from a CE-MRI database. In the first analysis, four radiologists independently evaluated the presence or absence of the see-through sign. The see-through sign was defined as internal transparent trabecular bone morphology on CE-MRI. The intraclass correlation coefficient (ICC), percentage agreement, and Fleiss's kappa statistics were obtained. RESULTS: Fifty-seven patients (M:F = 27:30; mean age, 63 years; age range, 20-88 years) who diagnosed as acute benign (n = 24) and malignant (n = 33) VCFs were finally included for the analysis. The results of all readers showed that the see-through sign was associated with acute benign VCFs (p < 0.05). The sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) of the see-through sign ranged from 75-96, 70-88, 66-85, 81-97 %, respectively. The inter-observer reliability of the see-through sign was sufficient with ICC = 0.847, percentage agreement = 78.9, and κ = 0.578. CONCLUSION: The see-through sign on CE-MRI is featured in acute benign VCFs, and it can be a useful finding to differentiate between benign and malignant VCFs.

Migratory defect of mesencephalic dopaminergic neurons in developing reeler mice.

Reelin, an extracellular glycoprotein has an important role in the proper migration and positioning of neurons during brain development. Lack of reelin causes not only disorganized lamination of the cerebral and cerebellar cortex but also malpositioning of mesencephalic dopaminergic (mDA) neurons. However, the accurate role of reelin in the migration and positioning of mDA neurons is not fully elucidated. In this study, reelin-deficient reeler mice exhibited a significant loss of mDA neurons in the substantia nigra pars compacta (SNc) and a severe alteration of cell distribution in the retrorubal field (RRF). This abnormality was also found in Dab1-deficinet, yotari mice. Stereological analysis revealed that total number of mDA neurons was not changed compared to wild type, suggesting that the loss of mDA neurons in reeler may not be due to the neurogenesis of mDA neurons. We also found that formation of PSA-NCAM-positive tangential nerve fibers rather than radial glial fibers was greatly reduced in the early developmental stage (E14.5) of reeler. These findings provide direct evidence that the alteration in distribution pattern of mDA neurons in the reeler mesencephalon mainly results from the defect of the lateral migration using tangential fibers as a scaffold.