Negative Fragment Research Articles

Generalized Correspondence Analysis for Three-Valued Logics

Correspondence analysis is Kooi and Tamminga’s universal approach which generates in one go sound and complete natural deduction systems with independent inference rules for tabular extensions of many-valued functionally incomplete logics. Originally, this method was applied to Asenjo–Priest’s paraconsistent logic of paradox LP. As a result, one has natural deduction systems for all the logics obtainable from the basic three-valued connectives of LP (which is built in the $$ \{\vee ,\wedge ,\lnot \} $$ -language) by the addition of unary and binary connectives. Tamminga has also applied this technique to the paracomplete analogue of LP, strong Kleene logic $$ \mathbf K_3 $$ . In this paper, we generalize these results for the negative fragments of LP and $$ \mathbf K_3 $$ , respectively. Thus, the method of correspondence analysis works for the logics which have the same negations as LP or $$ \mathbf K_3 $$ , but either have different conjunctions or disjunctions or even don’t have them as well at all. Besides, we show that correspondence analyses for the negative fragments of $$ \mathbf K_3 $$ and LP, respectively, are also suitable without any changes for the negative fragments of Heyting’s logic $$ \mathbf G_3 $$ and its dual $$ \mathbf DG_3 $$ (which have different interpretations of negation than $$ \mathbf K_3 $$ and LP).

Brr6 plays a role in gene recruitment and transcriptional regulation at the nuclear envelope.

Correlation between transcriptional regulation and positioning of genes at the nuclear envelope is well established in eukaryotes, but the mechanisms involved are not well understood. We show that brr6-1, a mutant of the essential yeast envelope transmembrane protein Brr6p, impairs normal positioning and expression of the PAB1 and FUR4-GAL1,10,7 loci. Similarly, expression of a dominant negative nucleoplasmic Brr6 fragment in wild-type cells reproduced many of the brr6-1 effects. Histone chromatin immunoprecipitation (ChIP) experiments showed decreased acetylation at the key histone H4K16 residue in the FUR4-GAL1,10,7 region in brr6-1. Importantly, blocking deacetylation significantly suppressed selected brr6-1 phenotypes. ChIPseq with FLAG-tagged Brr6 fragments showed enrichment at FUR4 and several other genes that showed striking changes in brr6-1 RNAseq data. These associations depended on a Brr6 putative zinc finger domain. Importantly, artificially tethering the GAL1 locus to the envelope suppressed the brr6-1 effects on GAL1 and FUR4 expression and increased H4K16 acetylation between GAL1 and FUR4 in the mutant. Together these results argue that Brr6 interacts with chromatin, helping to maintain normal chromatin architecture and transcriptional regulation of certain loci at the nuclear envelope.

Is habitat fragmentation good for biodiversity?

Habitat loss is a primary threat to biodiversity across the planet, yet contentious debate has ensued on the importance of habitat fragmentation ‘per se’ (i.e., altered spatial configuration of habitat for a given amount of habitat loss). Based on a review of landscape-scale investigations, Fahrig (2017; Ecological responses to habitat fragmentation per se. Annual Review of Ecology, Evolution, and Systematics 48:1-23) reports that biodiversity responses to habitat fragmentation ‘per se’ are more often positive rather than negative and concludes that the widespread belief in negative fragmentation effects is a ‘zombie idea’. We show that Fahrig's conclusions are drawn from a narrow and potentially biased subset of available evidence, which ignore much of the observational, experimental and theoretical evidence for negative effects of altered habitat configuration. We therefore argue that Fahrig's conclusions should be interpreted cautiously as they could be misconstrued by policy makers and managers, and we provide six arguments why they should not be applied in conservation decision-making. Reconciling the scientific disagreement, and informing conservation more effectively, will require research that goes beyond statistical and correlative approaches. This includes a more prudent use of data and conceptual models that appropriately partition direct vs indirect influences of habitat loss and altered spatial configuration, and more clearly discriminate the mechanisms underpinning any changes. Incorporating these issues will deliver greater mechanistic understanding and more predictive power to address the conservation issues arising from habitat loss and fragmentation.

Analysis of human C24 bile acids metabolome in serum and urine based on enzyme digestion of conjugated bile acids and LC-MS determination of unconjugated bile acids

Host-gut microbiota metabolic interactions are closely associated with health and disease. A manifestation of such co-metabolism is the vast structural diversity of bile acids (BAs) involving both oxidative stereochemistry and conjugation. Herein, we describe the development and validation of a LC-MS-based method for the analysis of human C24 BA metabolome in serum and urine. The method has high throughput covering the discrimination of oxidative stereochemistry of unconjugated species in a 15-min analytical cycle. The validated quantitative performance provided an indirect way to ascertain the conjugation patterns of BAs via enzyme-digestion protocols that incorporated the enzymes, sulfatase, β-glucuronidase, and choloylglycine hydrolase. Application of the method has led to the detection of at least 70 unconjugated BAs including 27 known species and 43 newly found species in the post-prandial serum and urine samples from 7 nonalcoholic steatohepatitis patients and 13 healthy volunteers. Newly identified unconjugated BAs included 3α, 12β-dihydroxy-5β-cholan-24-oic acid, 12α-hydroxy-3-oxo-5β-cholan-24-oic acid, and 3α, 7α, 12β-trihydroxy-5β-cholan-24-oic acid. High-definition negative fragment spectra of the other major unknown species were acquired to facilitate future identification endeavors. An extensive conjugation pattern is the major reason for the "invisibility" of the newly found BAs to other common analytical methods. Metabolomic analysis of the total unconjugated BA profile in combination with analysis of their conjugation patterns and urinary excretion tendencies have provided substantial insights into the interconnected roles of host and gut microbiota in maintaining BA homeostasis. It was proposed that the urinary total BA profile may serve as an ideal footprint for the functional status of the host-gut microbial BA co-metabolism. In summary, this work provided a powerful tool for human C24 BA metabolome analysis that bridges the gap between GC-MS techniques in the past age and LC-MS techniques currently prevailing in biomedical researches. Further applications of the present method in clinical, translational research, and other biomedical explorations will continue to boost the construction of a host-gut microbial co-metabolism network of BAs and thus facilitate the decryption of BA-mediated host-gut microbiota crosstalk in health and diseases. Graphical abstract ᅟ.

Combining Rules and Ontologies into Clopen Knowledge Bases

We propose Clopen Knowledge Bases (CKBs) as a new formalism combining Answer Set Programming (ASP) with ontology languages based on first-order logic. CKBs generalize the prominent r-hybrid and DL+LOG languages of Rosati, and are more flexible for specification of problems that combine open-world and closed-world reasoning. We argue that the guarded negation fragment of first-order logic(GNFO)—a very expressive fragment that subsumes many prominent ontology languages like Description Logics (DLs) and the guarded fragment—is an ontology language that can be used in CKBs while enjoying decidability for basic reasoning problems. We further show how CKBs can be used with expressive DLs of the ALC family, and obtain worst-case optimal complexity results in this setting. For DL-based CKBs, we define a fragment called separable CKBs (which still strictly subsumes r-hybrid and DL+LOG knowledge bases), and show that they can be rather efficiently translated into standard ASP programs. This approach allows us to perform basic inference from separable CKBs by reusing existing efficient ASP solvers. We have implemented the approach for separable CKBs containing ontologies in the DL ALCH, and present in this paper some promising empirical results for real-life data. They show that our approach provides a dramatic improvement over a naive implementation based on a translation of such CKBs into dl-programs.

MP70-16 SEMA3C DRIVES CANCER GROWTH AND TREATMENT RESISTANCE VIA COGNATE LIGAND-INDEPENDENT ACTIVATION

You have accessJournal of UrologyProstate Cancer: Basic Research & Pathophysiology III1 Apr 2018MP70-16 SEMA3C DRIVES CANCER GROWTH AND TREATMENT RESISTANCE VIA COGNATE LIGAND-INDEPENDENT ACTIVATION Ario Takeuchi, Masaki Shiota, Junichi Inokuchi, Katsunori Tatsugami, and Masatoshi Eto Ario TakeuchiArio Takeuchi More articles by this author , Masaki ShiotaMasaki Shiota More articles by this author , Junichi InokuchiJunichi Inokuchi More articles by this author , Katsunori TatsugamiKatsunori Tatsugami More articles by this author , and Masatoshi EtoMasatoshi Eto More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2018.02.2260AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Our gene expression profiling data identified a member of the secreted class 3 semaphorins, SEMA3C, to be highly expressed in CRPC and AR pathway inhibitor-recurrent tumors. Increased expression of SEMA3C is associated with poor prognosis and tumor progression in a number of cancers. In prostate cancer, SEMA3C promotes cell migration and invasion in vitro and SEMA3C expression is a predictive marker for biochemical recurrence (BCR). Therefore, we investigated whether SEMA3C could be a key growth factor that drives CRPC progression and treatment resistance. METHODS The objective of this study was to evaluate SEMA3C as a therapeutic target for advanced prostate cancer and the physiological consequences of loss- and gain-of-function of SEMA3C. The expression of SEMA3C in a tissue microarray representing a total of 280 prostate cancer specimens was evaluated to determine association of SEMA3C expression with castration and treatment resistance. For in vivo orthotopic model, when mice bearing LNCaP tumors reached a tumor volume over 200 mm3 or serum PSA levels reached greater than 75 ng/ml, castration was performed via the scrotum under isoflurane anesthesia. Treatment commenced when PSA recovered to pre-castration levels. Mice were randomized into 4 groups for treatment with SEMA3C ASO or Scr with or without ENZ. For ASO treatment, Scr ASO or SEMA3C ASO was intra-peritoneally injected once daily for 7 days followed by three weekly treatments thereafter. For ENZ treatment, ENZ or vehicle was intragastrically administered once daily 5 days per week. RESULTS Here we show SEMA3C drives activation of multiple RTKs including EGFR, ErbB2, and c-MET in a cognate ligand-independent manner. High SEMA3C expression is associated with metastatic castration-resistant prostate cancer (CRPC). SEMA3C promotes cancer growth and castrate resistant progression while SEMA3C inhibition delays CRPC and Enzalutamide-resistant progression. Proteolytic processing of SEMA3C generates a dominant negative sema domain-containing decoy fragment that suppresses cell growth and RTK signaling. Intratumoral lentiviral delivery of this decoy protein inhibits CRPC progression. CONCLUSIONS SEMA3C inhibition represents a novel therapeutic strategy for treatment of advanced prostate cancer. © 2018FiguresReferencesRelatedDetails Volume 199Issue 4SApril 2018Page: e940 Advertisement Copyright & Permissions© 2018MetricsAuthor Information Ario Takeuchi More articles by this author Masaki Shiota More articles by this author Junichi Inokuchi More articles by this author Katsunori Tatsugami More articles by this author Masatoshi Eto More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Isomer Information from Ion Mobility Separation of High-Mannose Glycan Fragments

Extracted arrival time distributions of negative ion CID-derived fragments produced prior to traveling-wave ion mobility separation were evaluated for their ability to provide structural information on N-linked glycans. Fragmentation of high-mannose glycans released from several glycoproteins, including those from viral sources, provided over 50 fragments, many of which gave unique collisional cross-sections and provided additional information used to assign structural isomers. For example, cross-ring fragments arising from cleavage of the reducing terminal GlcNAc residue on Man8GlcNAc2 isomers have unique collision cross-sections enabling isomers to be differentiated in mixtures. Specific fragment collision cross-sections enabled identification of glycans, the antennae of which terminated in the antigenic α-galactose residue, and ions defining the composition of the 6-antenna of several of the glycans were also found to have different cross-sections from isomeric ions produced in the same spectra. Potential mechanisms for the formation of the various ions are discussed and the estimated collisional cross-sections are tabulated.Graphical ᅟ

Frightened by the perpetrator’s voice: Startle responsivity and cognitive processing predict earwitness speaker identification

This study was inspired by the case of a robbery victim who was startled and reminded of the crime upon hearing a stranger’s voice, while not clearly recognizing the speaker. To investigate whether specific voices can modulate startle reactions and thereby predict speaker identification, we presented an audio hijack scenario to 84 participants and afterwards asked them to identify the perpetrator among neutral and negative speech fragments, while measuring flash-evoked eye-blink startle responses. Furthermore, we addressed data-driven cognitive processing during the audio scenario as a potential moderator in voice discrimination. Negative speech and the perpetrator’s voice led to potentiated startle. Enhanced startle was positively associated with voice discrimination, but only in neutral speech fragments. In negative fragments, this association was weakened as a function of self-reported levels of data-driven processing during encoding. Thus, startle responses can generally predict accurate voice recognition, but speech emotionality and cognitive processing moderate this relationship.

Interpretation and Simulation of Negative Ion Mass Spectra of Some Phosphorus Organoelement Compounds

Negative ion mass spectra for a series of organophosphorus compounds were obtained and negative ion fragmentation processes were treated theoretically. Using O-isopropyl and O-pinacolyl methylphosphonofluoridates as examples, electron affinities of molecules and their fragments were estimated using the UB3LYP/6-311+G(d,p) quantum-chemical approach and energetically more favorable and characteristic routes of dissociative electron attachment, including simple bond cleavage and rearrangements, were determined. Based on the obtained experimental and theoretical data, hypothetic fragmentation patterns were proposed and a special algorithm was compiled to predict negative ion mass spectra for some groups of organophosphorus compounds, such as О-alkyl methylphosphonofluoridates, О,О-alkyl phosphonodichloridates, and О,О′-dialkyl phosphonochloridates. The simulated mass spectra showed a good agreement with the experimental ones, confirming reasonable reliability of the proposed algorithm.

Decoupling habitat fragmentation from habitat loss: butterfly species mobility obscures fragmentation effects in a naturally fragmented landscape of lake islands.

Since the publication of the theory ofisland biogeography, ecologists have postulated that fragmentation of continuous habitat presents a prominent threat to species diversity. However, negative fragmentation effects may be artifacts; the result of species diversity declining with habitat loss, and habitat loss correlating positively with degree of fragmentation. In this study, we used butterfly assemblages on islands of Lake of the Woods, Ontario, Canada to decouple habitat fragmentation from habitat loss and test two competing hypotheses: (1) the island effect hypothesis, which predicts that decreasing fragment size and increasing fragmentisolation reduces species diversity beyond the effects of habitat loss, and (2) the habitat amount hypothesis, which negates fragmentation effects and predicts that only total habitat area determines the diversity of species persisting on fragmented landscapes. Using eight independent size classes of islands (ranging from 0.1 to 8.0ha) that varied in number of islands while holding total area constant, species diversity comparisons, species accumulation curves, and species-area relationship extrapolations demonstrated that smaller insular habitats contained at least as many butterfly species as continuous habitat. However, when highly mobile species occurring on islands without their larval food plants were excluded from analyses, island effects on potentially reproducing species became apparent. Similarily, generalized linear models suggested that effects of island isolation and vascular plant richness on insularbutterfly richness were confounded by species of high mobility. We conclude that inter-fragment movements of highly mobile species may obscure important fragmentation effects on potentially reproducing populations, questioning support for the habitat amount hypothesis.

Dissociation of 2-oxopropanoic acid by low energy electrons

SynopsisElectron attachment to molecular target can lead to a variety of negative fragment ions. We experimentally probe the main dissociative electron attachment (DEA) channels in 2-oxopropanoic acid (CH3 ― CO ― COOH) - an important trace component in the Earth’s atmosphere. In our studies we utilise nearly mono-energetic electrons at electron energies from 0 eV up to 15 eV.

P.4.003 - Testing familial transmission of smoking with two different research designs

This study was inspired by the case of a robbery victim who was startled and reminded of the crime upon hearing a stranger’s voice, while not clearly recognizing the speaker. To investigate whether specific voices can modulate startle reactions and thereby predict speaker identification, we presented an audio hijack scenario to 84 participants and afterwards asked them to identify the perpetrator among neutral and negative speech fragments, while measuring flash-evoked eye-blink startle responses. Furthermore, we addressed data-driven cognitive processing during the audio scenario as a potential moderator in voice discrimination. Negative speech and the perpetrator’s voice led to potentiated startle. Enhanced startle was positively associated with voice discrimination, but only in neutral speech fragments. In negative fragments, this association was weakened as a function of self-reported levels of data-driven processing during encoding. Thus, startle responses can generally predict accurate voice recognition, but speech emotionality and cognitive processing moderate this relationship.

Rejection of the Renal Allograft in the Absence of Demonstrable Antibody and Complement.

Recent literature has stressed the prominent role of antibodies in graft loss. This study was designed to assess a growing perception that T cell-mediated rejection (TCMR) is no longer clinically relevant. Five hundred forty-five renal allograft recipients over a 3-year period were screened for biopsies with: (a) TCMR including borderline change (BL), (b) negative complement protein C4 degradation fragment, and (c) absence of donor-specific antibody at time of transplant, within 30 days of the biopsy, and up to 4 measurements at later time points. These stringent requirements identified 28 "pure" cases of late TCMR/BL. Low-grade glomerulitis, peritubular capillaritis, or chronic transplant glomerulopathy were found in 9/28 (32%) biopsies. Serum creatinine showed complete short-term remission in 7/10 (70%) BL and 9/18 (50%) TCMR patients 1 month postbiopsy. Yet, both treated and untreated patients demonstrated further decline in graft function as assessed by serum creatinine and estimated glomerular filtration rate. Late TCMR seen in 7.9% of biopsies can contribute to significant deterioration of graft function in patients in whom the dominant contribution of antibody-mediated injury has been reasonably excluded. Our data also reinforce existing literature showing that microvascular lesions do not have absolute specificity for a diagnosis of antibody-mediated rejection.

Theoretical study of low-energy electron scattering with GeH2

We present a comprehensive study of electron collisions with germylene (GeH2 using the UK molecular R-matrix codes for electron energies ranging from 0.01 to 10 eV. The calculations are performed within the static-exchange, static-exchange-polarization, and 17-state close-coupling approximations. The elastic integral, differential, momentum transfer cross sections and the excitation cross sections from the ground state to the six low-lying electron excited states are presented. We found three Feshbach resonances and one Core-excited resonance. These resonances reveal the probability of anion formation by an electron attachment process and further decay to neutral and negative ion fragments. The electronic and rotational excitation cross sections for e-GeH2 scattering are reported for the first time. The cross-section dataset obtained from the present calculations are expected to be sufficiently accurate and comprehensive for most current modeling applications involving neutral GeH2.

Cytocompatibility of a self-adhesive gutta-percha root-filling material

Context:A novel root-filling material based on the incorporation of ultrafine alkaline bioactive glass particles (bioactive gutta-percha, [BGP]) was developed to work without sealer.Aim:In the present study, the objective was to verify the in vitro biological response to this material by assessing its cytocompatibility.Materials and Methods:Prototypes of BGP were compared to conventional gutta-percha (GP), dense polystyrene beads as a negative control and fragments of latex as a positive control. Extracts of each material were prepared according to ISO 10993-5:2009, and human osteoblast-like cells in primary culture were exposed to all extracts for 24 h. Cell viability was assayed sequentially for three different parameters: mitochondrial activity, membrane integrity, and cell density.Statistical Analysis Used:Nonparametric analysis (using Kruskal–Wallis test combined with post hoc Dunn's test) was performed for comparison among groups, with significance established at 5%.Results:BGP reduced mitochondrial activity to 62% of control, but presented no toxicity on membrane integrity and proliferation assays. BGP effect on metabolism was dose-dependent and reduced to acceptable levels with dilution.Conclusion:The novel GP material presented slight dose-dependent effects on cell metabolism but did not affect cell survival.

Resonant electron capture by aspartame and aspartic acid molecules

The processes for dissociative electron capture are the key mechanisms for decomposition of biomolecules, proteins in particular, under interaction with low-energy electrons. Molecules of aspartic acid and aspartame, i.e. modified dipeptides, were studied herein to define the impact of the side functional groups on peptide chain decomposition in resonant electron-molecular reactions. The processes of formation and decomposition of negative ions of both aspartame and aspartic acid were studied by mass spectrometry of negative ions under resonant electron capture. The obtained mass spectra were interpreted under thermochemical analysis by quantum chemical calculations. Main channels of negative molecular ions fragmentation were found and characteristic fragment ions were identified. The СООН fragment of the side chain in aspartic acid is shown to play a key role like the carboxyl group in amino acids and aliphatic oligopeptides. Copyright © 2016 John Wiley & Sons, Ltd.

Travelling-wave ion mobility mass spectrometry and negative ion fragmentation of hybrid and complex N-glycans.

Nitrogen collisional cross sections (CCSs) of hybrid and complex glycans released from the glycoproteins IgG, gp120 (from human immunodeficiency virus), ovalbumin, α1-acid glycoprotein and thyroglobulin were measured with a travelling-wave ion mobility mass spectrometer using dextran as the calibrant. The utility of this instrument for isomer separation was also investigated. Some isomers, such as Man3 GlcNAc3 from chicken ovalbumin and Man3 GlcNAc3 Fuc1 from thyroglobulin could be partially resolved and identified by their negative ion fragmentation spectra obtained by collision-induced decomposition (CID). Several other larger glycans, however, although existing as isomers, produced only asymmetric rather than separated arrival time distributions (ATDs). Nevertheless, in these cases, isomers could often be detected by plotting extracted fragment ATDs of diagnostic fragment ions from the negative ion CID spectra obtained in the transfer cell of the Waters Synapt mass spectrometer. Coincidence in the drift times of all fragment ions with an asymmetric ATD profile in this work, and in the related earlier paper on high-mannose glycans, usually suggested that separations were because of conformers or anomers, whereas symmetrical ATDs of fragments showing differences in drift times indicated isomer separation. Although some significant differences in CCSs were found for the smaller isomeric glycans, the differences found for the larger compounds were usually too small to be analytically useful. Possible correlations between CCSs and structural types were also investigated, and it was found that complex glycans tended to have slightly smaller CCSs than high-mannose glycans of comparable molecular weight. In addition, biantennary glycans containing a core fucose and/or a bisecting GlcNAc residue fell on different mobility-m/z trend lines to those glycans not so substituted with both of these substituents contributing to larger CCSs. Copyright © 2016 John Wiley & Sons, Ltd.

Fragmentation effects on dead wood-dependent species associated with disturbed forest habitats: implications for stump harvesting

ABSTRACTStump harvesting after clear-felling may be detrimental for species’ landscape-scale persistence. Sensitivity to it is most likely due to reductions in habitat density, resulting in a non-linear relationship between population size and total habitat amount (i.e. fragmentation effects). Here we summarize theoretical expectations and empirical findings on fragmentation effects in stumps and other types of dead wood in disturbed forest habitats and draw conclusions about the consequences of stump harvesting. Within disturbed patches, some negative fragmentation effects have been observed for beetles and lichens, but most studies have found a linear relationship between habitat amount and population size. At landscape scale, evidence of fragmentation effects in disturbed forest habitats has been detected in some published investigations, but none of them focused on stumps. Thus, although organisms associated with disturbed forest habitats are assumed to be strong dispersers, they may still be sensitive to habitat fragmentation. We conclude that stump harvesting at a moderate level is not likely to increase risks for landscape-scale species extinctions markedly, despite clearly negative local effects. However, due to large uncertainties, adaptive management with monitoring of dead wood-dependent species should be applied wherever stump harvesting is more extensive.

N-glycan MALDI Imaging Mass Spectrometry on Formalin-Fixed Paraffin-Embedded Tissue Enables the Delineation of Ovarian Cancer Tissues

Ovarian cancer is a fatal gynaecological malignancy in adult women with a five-year overall survival rate of only 30%. Glycomic and glycoproteomic profiling studies have reported extensive protein glycosylation pattern alterations in ovarian cancer. Therefore, spatio-temporal investigation of these glycosylation changes may unearth tissue-specific changes that occur in the development and progression of ovarian cancer. A novel method for investigating tissue-specific N-linked glycans is using matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) on formalin-fixed paraffin-embedded (FFPE) tissue sections that can spatially profile N-glycan compositions released from proteins in tissue-specific regions. In this study, tissue regions of interest (e.g. tumor, stroma, adipose tissue and necrotic areas) were isolated from FFPE tissue sections of advanced serous ovarian cancers (n = 3). PGC-LC-ESI-MS/MS and MALDI-MSI were used as complementary techniques to firstly generate structural information on the tissue-specific glycans in order to then obtain high resolution images of the glycan structure distribution in ovarian cancer tissue. The N-linked glycan repertoires carried by the proteins in these tissue regions were structurally characterized for the first time in FFPE ovarian cancer tissue regions, using enzymatic peptide-N-glycosidase F (PNGase F) release of N-glycans. The released glycans were analyzed by porous graphitized carbon liquid chromatography (PGC-LC) and collision induced electrospray negative mode MS fragmentation analysis. The N-glycan profiles identified by this analysis were then used to determine the location and distribution of each N-glycan on FFPE ovarian cancer sections that were treated with PNGase F using high resolution MALDI-MSI. A tissue-specific distribution of N-glycan structures identified particular regions of the ovarian cancer sections. For example, high mannose glycans were predominantly expressed in the tumor tissue region whereas complex/hybrid N-glycans were significantly abundant in the intervening stroma. Therefore, tumor and non-tumor tissue regions were clearly demarcated solely on their N-glycan structure distributions.

Key Role for the 12-Hydroxy Group in the Negative Ion Fragmentation of Unconjugated C24 Bile Acids.

Host-gut microbial interactions contribute to human health and disease states and an important manifestation resulting from this cometabolism is a vast diversity of bile acids (BAs). There is increasing interest in using BAs as biomarkers to assess the health status of individuals and, therefore, an increased need for their accurate separation and identification. In this study, the negative ion fragmentation behaviors of C24 BAs were investigated by UPLC-ESI-QTOF-MS. The step-by-step fragmentation analysis revealed a distinct fragmentation mechanism for the unconjugated BAs containing a 12-hydroxyl group. The unconjugated BAs lacking 12-hydroxylation fragmented via dehydration and dehydrogenation. In contrast, the 12-hydroxylated ones, such as deoxycholic acid (DCA) and cholic acid (CA), employed dissociation routes including dehydration, loss of carbon monoxide or carbon dioxide, and dehydrogenation. All fragmentations of the 12-hydroxylated unconjugated BAs, characterized by means of stable isotope labeled standards, were associated with the rotation of the carboxylate side chain and the subsequent rearrangements accompanied by proton transfer between 12-hydroxyl and 24-carboxyl groups. Compared to DCA, CA underwent further cleavages of the steroid skeleton. Accordingly, the effects of stereochemistry on the fragmentation pattern of CA were investigated using its stereoisomers. Based on the knowledge gained from the fragmentation analysis, a novel BA, 3β,7β,12α-trihydroxy-5β-cholanic acid, was identified in the postprandial urine samples of patients with nonalcoholic steatohepatitis. The analyses used in this study may contribute to a better understanding of the chemical diversity of BAs and the molecular basis of human liver diseases that involve BA synthesis, transport, and metabolism.