Detection of N-glycolylneuraminic acid biomarkers in sera from patients with ovarian cancer using an engineered N-glycolylneuraminic acid-specific lectin SubB2M.

N-glycolylneuraminic acid (Neu5Gc)-containing glycans are a prominent form of aberrant glycosylation found in human tumor cells and have been proposed as cancer biomarkers. The B subunit of the subtilase cytotoxin (SubB) produced by Shiga toxigenic Escherichia coli recognises Neu5Gc containing glycans. We have previously engineered this lectin, SubB2M, for greater specificity and enhanced recognition of Neu5Gc-containing glycans. Here we further explore the utility of SubB2M to detect Neu5Gc tumor biomarkers in sera from patients with ovarian cancer. Using surface plasmon resonance (SPR) we show that SubB2M can detect the established ovarian cancer biomarker, CA125, in a highly sensitive and specific fashion in the context of human serum. These studies established conditions for screening serum samples from patients with ovarian cancer for Neu5Gc glycans. We found that serum from patients with all stages of ovarian cancer had significantly elevated mean levels of Neu5Gc glycans compared to normal controls. Serum from patients with late stage disease (stages IIIC, IV) had uniformly elevated levels of Neu5Gc glycans. Detection of Neu5Gc-glycans using SubB2M has the potential to be used as a diagnostic ovarian cancer biomarker, as well as a tool for monitoring treatment and disease progression in late stage disease.

A synthetic review of notoedres species mites and mange.

Notoedric mange, caused by obligately parasitic sarcoptiform Notoedres mites, is associated with potentially fatal dermatitis with secondary systemic disease in small mammals, felids and procyonids among others, as well as an occasional zoonosis. We describe clinical spectra in non-chiropteran hosts, review risk factors and summarize ecological and epidemiological studies. The genus is disproportionately represented on rodents. Disease in felids and procyonids ranges from very mild to death. Knowledge of the geographical distribution of the mites is highly inadequate, with focal hot spots known for Notoedres cati in domestic cats and bobcats. Predisposing genetic and immunological factors are not known, except that co-infection with other parasites and anticoagulant rodenticide toxicoses may contribute to severe disease. Treatment of individual animals is typically successful with macrocytic lactones such as selamectin, but herd or wildlife population treatment has not been undertaken. Transmission requires close contact and typically is within a host species. Notoedric mange can kill half all individuals in a population and regulate host population below non-diseased density for decades, consistent with frequency-dependent transmission or spillover from other hosts. Epidemics are increasingly identified in various hosts, suggesting global change in suitable environmental conditions or increased reporting bias.

Nanoscale conductive pattern of the homoepitaxial AlGaN/GaN transistor.

The gallium nitride (GaN)-based buffer/barrier mode of growth and morphology, the transistor electrical response (25-310 °C) and the nanoscale pattern of a homoepitaxial AlGaN/GaN high electron mobility transistor (HEMT) have been investigated at the micro and nanoscale. The low channel sheet resistance and the enhanced heat dissipation allow a highly conductive HEMT transistor (Ids > 1 A mm(-1)) to be defined (0.5 A mm(-1) at 300 °C). The vertical breakdown voltage has been determined to be ∼850 V with the vertical drain-bulk (or gate-bulk) current following the hopping mechanism, with an activation energy of 350 meV. The conductive atomic force microscopy nanoscale current pattern does not unequivocally follow the molecular beam epitaxy AlGaN/GaN morphology but it suggests that the FS-GaN substrate presents a series of preferential conductive spots (conductive patches). Both the estimated patches density and the apparent random distribution appear to correlate with the edge-pit dislocations observed via cathodoluminescence. The sub-surface edge-pit dislocations originating in the FS-GaN substrate result in barrier height inhomogeneity within the HEMT Schottky gate producing a subthreshold current.

How big is my neighborhood? Individual and contextual effects on perceptions of neighborhood scale.

Neighborhood is a social and geographic concept that plays an increasingly important role in research and practice that address disparities in health and well-being of populations. However, most studies of neighborhoods, as well as community initiatives geared toward neighborhood improvement, make simplifying assumptions about boundaries, often relying on census geography to operationalize the neighborhood units. This study used geographic information system (GIS) tools to gather and analyze neighborhood maps drawn by residents of low-income communities in 10 cities. The median resident map size was approximately 30 percent smaller than the median census tract, but 25 percent of residents viewed their neighborhood as quite small (less than one-fifth of the typical census tract). Multi-level modeling showed significant within context variation in perceived neighborhood scale. Longer term residents with higher education and income and who were more engaged in the neighborhood held more expansive views. But there were also contextual influences with higher density and mixed use areas associated with smaller perceived neighborhoods, and higher collective efficacy associated with larger neighborhood sizes. Artificially imposed neighborhood units may misrepresent resident experience, but GIS tools can be used to craft more authentic neighborhood definitions for research and practice.

Rational engineering of an improved adenosine deaminase 2 enzyme for weaponizing T-cell therapies.

Adenosine is a potent immunosuppressive metabolite that accumulates in the extracellular space within solid tumors and inhibits the antitumor function of native immune cell responses as well as chimeric antigen receptor (CAR) T-cell therapies. Here, we show that engineered human cells can degrade extracellular adenosine through secretion of adenosine deaminase (ADA) enzymes-a possible therapeutic enhancement for CAR T cells. We first determine that the high-activity ADA1 isoform is naturally intracellularly restricted and show that the addition of canonical or computationally predicted secretory peptides did not allow for improved secretion. We did, however, determine that the lower-activity ADA2 isoform is naturally secreted. Thus, we utilized phylogenetic-based structural comparisons to guide a mutational survey of ADA2 active site residues, which when coupled with a high-throughput screen for enhanced ADA2-mediated extracellular adenosine rate allowed isolation of the most catalytically efficient ADA2 variant reported to date. When expressed by human cells, this variant exhibits 30× higher extracellular adenosine degradation activity than the wild-type enzyme. Finally, we demonstrate that Jurkat and CAR T cells engineered to express this secreted, high-activity ADA2 variant can degrade significant amounts of extracellular adenosine in vitro.

Corrigendum to "Rational engineering of an improved adenosine deaminase 2 enzyme for weaponizing T-cell therapies": [Immuno-Oncology and Technology 10 (2023) 100394].

[This corrects the article DOI: 10.1016/j.iotech.2023.100394.].

Nanoscale investigation of AlGaN/GaN-on-Si high electron mobility transistors.

AlGaN/GaN HEMTs are devices which are strongly influenced by surface properties such as donor states, roughness or any kind of inhomogeneity. The electron gas is only a few nanometers away from the surface and the transistor forward and reverse currents are considerably affected by any variation of surface property within the atomic scale. Consequently, we have used the technique known as conductive AFM (CAFM) to perform electrical characterization at the nanoscale. The AlGaN/GaN HEMT ohmic (drain and source) and Schottky (gate) contacts were investigated by the CAFM technique. The estimated area of these highly conductive pillars (each of them of approximately 20-50 nm radius) represents around 5% of the total contact area. Analogously, the reverse leakage of the gate Schottky contact at the nanoscale seems to correlate somehow with the topography of the narrow AlGaN barrier regions producing larger currents.

United States (US) multi-center study to assess the validity and reliability of the Spinal Cord Independence Measure (SCIM III).

STUDY DESIGN: Multi-center, prospective, cohort study. OBJECTIVES: To assess the validity and reliability of the Spinal Cord Independence Measure (SCIM III) in measuring functional ability in persons with spinal cord injury (SCI). SETTING: Inpatient rehabilitation hospitals in the United States (US). METHODS: Functional ability was measured with the SCIM III during the first week of admittance into inpatient acute rehabilitation and within one week of discharge from the same rehabilitation program. Motor and sensory neurologic impairment was measured with the American Spinal Injury Association Impairment Scale. The Functional Independence Measure (FIM), the default functional measure currently used in most US hospitals, was used as a comparison standard for the SCIM III. Statistical analyses were used to test the validity and reliability of the SCIM III. RESULTS: Total agreement between raters was above 70% on most SCIM III tasks and all κ-coefficients were statistically significant (P<0.001). The coefficients of Pearson correlation between the paired raters were above 0.81 and intraclass correlation coefficients were above 0.81. Cronbach's-α was above 0.7, with the exception of the respiration task. The coefficient of Pearson correlation between the FIM and SCIM III was 0.8 (P<0.001). For the respiration and sphincter management subscale, the SCIM III was more responsive to change, than the FIM (P<0.0001). CONCLUSION: Overall, the SCIM III is a reliable and valid measure of functional change in SCI. However, improved scoring instructions and a few modifications to the scoring categories may reduce variability between raters and enhance clinical utility.

The integrated stress response contributes to tRNA synthetase-associated peripheral neuropathy.

Dominant mutations in ubiquitously expressed transfer RNA (tRNA) synthetase genes cause axonal peripheral neuropathy, accounting for at least six forms of Charcot-Marie-Tooth (CMT) disease. Genetic evidence in mouse and Drosophila models suggests a gain-of-function mechanism. In this study, we used in vivo, cell type­specific transcriptional and translational profiling to show that mutant tRNA synthetases activate the integrated stress response (ISR) through the sensor kinase GCN2 (general control nonderepressible 2). The chronic activation of the ISR contributed to the pathophysiology, and genetic deletion or pharmacological inhibition of Gcn2 alleviated the peripheral neuropathy. The activation of GCN2 suggests that the aberrant activity of the mutant tRNA synthetases is still related to translation and that inhibiting GCN2 or the ISR may represent a therapeutic strategy in CMT.

An immunoaffinity liquid chromatography-tandem mass spectrometry assay for detection of endogenous aggrecan fragments in biological fluids: Use as a biomarker for aggrecanase activity and cartilage degradation.

The degradation of articular cartilage by aggrecanases (ADAMTS-4 and ADAMTS-5) plays a significant role in the pathology of osteoarthritis (OA). To monitor aggrecanase activity in OA, we have developed a sensitive, accurate, and versatile assay for detection of two specific cleavage sites on aggrecan. The assay uses an immunoaffinity-based liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to detect cleavage at the (374)ARGS site and the (1820)AGEG site. The dynamic range of the assay is more than three orders of magnitude, with interassay precision less than 15%. It has been successfully applied to various biological fluids and species, including rat, bovine, dog, and human. The assay has been analytically qualified for use in human urine and synovial fluid (SF). The limits of detection (LODs) for ARGS in urine and SF are 2.5 and 10 pg/ml, respectively, whereas the LOD for AGEG is 20 pg/ml in SF. Analysis of these biomarkers from OA subjects and normal healthy volunteers revealed a significant elevation of both markers in OA. Similarly, in a rat model of cartilage degradation, both ARGS and AGEG were elevated, demonstrating the utility of these biomarkers for translational research. These data suggest that the ARGS and AGEG biomarkers developed have potential as measures of aggrecanase activity in OA and may contribute to our understanding of OA pathology.

Alterations in serum MMP-8, MMP-9, IL-12p40 and IL-23 in multiple sclerosis patients treated with interferon-beta1b.

BACKGROUND: Interferon-beta1b (IFN-beta1b), an effective treatment for multiple sclerosis (MS), lessens disease severity in MS patients. However, the mechanisms of its immunoregulatory and anti-inflammatory effects in MS remain only partially understood. Matrix metalloproteinases (MMP) and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) are involved in blood brain barrier disruption and formation of MS lesions. Th1/Th17 cytokines e.g. interleukins IL-12p40, IL-17, and IL-23, are associated with MS disease activity and are significant players in pathogenesis of MS. OBJECTIVE: During a 1-year prospective study, we serially measured serum MMP-8, MMP-9, TIMP-1, IL-12p40, IL-17, and IL-23 in 24 patients with relapsing-remitting MS. We compared the results to clinical course and to brain magnetic resonance imaging. IFN-beta1b decreased serum MMP-8 and MMP-9 (not TIMP-1). RESULTS: The sustained treatment with IFN-beta1b attenuated the pro-inflammatory environment by significantly reducing the serum IL-12p40, IL-23, and showed a trend for decreasing IL-17. Decreased serum MMP-8, MMP-9, IL-12 and IL-23 levels were correlated with a decrease in the number of contrast-enhanced T2-weighted lesions. CONCLUSION: Early treatment of MS with IFN-beta1b may stabilize clinical disease by attenuating levels of inflammatory cytokines and MMPs. Serial measurement of inflammatory mediators may serve as sensitive markers to gauge therapeutic responses to IFN-beta1b during the first year of treatment.

A dimensional investigation of error-related negativity (ERN) and self-reported psychiatric symptoms.

Alterations in error processing are implicated in a range of DSM-defined psychiatric disorders. For instance, obsessive-compulsive disorder (OCD) and generalised anxiety disorder show enhanced electrophysiological responses to errors-i.e. error-related negativity (ERN)-while others like schizophrenia have an attenuated ERN. However, as diagnostic categories in psychiatry are heterogeneous and also highly intercorrelated, the precise mapping of ERN enhancements/impairments is unclear. To address this, we recorded electroencephalograms (EEG) from 196 participants who performed the Flanker task and collected scores on 9 questionnaires assessing psychiatric symptoms to test if a dimensional framework could reveal specific transdiagnostic clinical manifestations of error processing dysfunctions. Contrary to our hypothesis, we found non-significant associations between ERN amplitude and symptom severity of OCD, trait anxiety, depression, social anxiety, impulsivity, eating disorders, alcohol addiction, schizotypy and apathy. A transdiagnostic approach did nothing to improve signal; there were non-significant associations between all three transdiagnostic dimensions (anxious-depression, compulsive behaviour and intrusive thought, and social withdrawal) and ERN magnitude. In these same individuals, we replicated a previously published transdiagnostic association between goal-directed learning and compulsive behaviour and intrusive thought. Possible explanations discussed are (i) that associations between the ERN and psychopathology might be smaller than previously assumed, (ii) that these associations might depend on a greater level of symptom severity than other transdiagnostic cognitive biomarkers, or (iii) that task parameters, such as the ratio of compatible to incompatible trials, might be crucial for ensuring the sensitivity of the ERN to clinical phenomena.

Exploiting species specificity to understand the tropism of a human-specific toxin.

Many pathogens produce virulence factors that are specific toward their natural host. Clinically relevant methicillin-resistant Staphylococcus aureus (MRSA) isolates are highly adapted to humans and produce an array of human-specific virulence factors. One such factor is LukAB, a recently identified pore-forming toxin that targets human phagocytes by binding to the integrin component CD11b. LukAB exhibits strong tropism toward human, but not murine, CD11b. Here, phylogenetics and biochemical studies lead to the identification of an 11-residue domain required for the specificity of LukAB toward human CD11b, which is sufficient to render murine CD11b compatible with toxin binding. CRISPR-mediated gene editing was used to replace this domain, resulting in a "humanized" mouse. In vivo studies revealed that the humanized mice exhibit enhanced susceptibility to MRSA bloodstream infection, a phenotype mediated by LukAB. Thus, these studies establish LukAB as an important toxin for MRSA bacteremia and describe a new mouse model to study MRSA pathobiology.

The pilin O-glycosylation pathway of pathogenic Neisseria is a general system that glycosylates AniA, an outer membrane nitrite reductase.

O-Glycosylation is emerging as a common posttranslational modification of surface exposed proteins in bacterial mucosal pathogens. In pathogenic Neisseria an O-glycosylation pathway modifies a single abundant protein, pilin, the subunit protein that forms pili. Here, we identify an additional outer membrane glycoprotein in pathogenic Neisseria, the nitrite reductase AniA, that is glycosylated in its C-terminal repeat region by the pilin glycosylation pathway. To our knowledge, this is the first report of a general O-glycosylation pathway in a prokaryote. We also show that AniA displays polymorphisms in residues that map to the surface of the protein. A frame-shift mutation abolishes AniA expression in 34% of Neisseria meningitidis strains surveyed, however, all Neisseria gonorrhoeae strains examined are predicted to express AniA, implying a crucial role for AniA in gonococcal biology.

Refinements in husbandry, care and common procedures for non-human primates: Ninth report of the BVAAWF/FRAME/RSPCA/UFAW Joint Working Group on Refinement.

Preface Whenever animals are used in research, minimizing pain and distress and promoting good welfare should be as important an objective as achieving the experimental results. This is important for humanitarian reasons, for good science, for economic reasons and in order to satisfy the broad legal principles in international legislation. It is possible to refine both husbandry and procedures to minimize suffering and improve welfare in a number of ways, and this can be greatly facilitated by ensuring that up-to-date information is readily available. The need to provide such information led the British Veterinary Association Animal Welfare Foundation (BVAAWF), the Fund for the Replacement of Animals in Medical Experiments (FRAME), the Royal Society for the Prevention of Cruelty to Animals (RSPCA) and the Universities Federation for Animal Welfare (UFAW) to establish a Joint Working Group on Refinement (JWGR) in the UK. The chair is Professor David Morton and the secretariat is provided by the RSPCA. This report is the ninth in the JWGR series. The RSPCA is opposed to the use of animals in experiments that cause pain, suffering, distress or lasting harm and together with FRAME has particular concerns about the continued use of non-human primates. The replacement of primate experiments is a primary goal for the RSPCA and FRAME. However, both organizations share with others in the Working Group, the common aim of replacing primate experiments wherever possible, reducing suffering and improving welfare while primate use continues. The reports of the refinement workshops are intended to help achieve these aims. This report produced by the British Veterinary Association Animal Welfare Foundation (BVAAWF)/Fund for the Replacement of Animals in Medical Experiments (FRAME)/Royal Society for the Prevention of Cruelty to Animals (RSPCA)/Universities Federation for Animal Welfare (UFAW) Joint Working Group on Refinement (JWGR) sets out practical guidance on refining the husbandry and care of non-human primates (hereinafter primates) and on minimizing the adverse effects of some common procedures. It provides a valuable resource to help understand the physical, social and behavioural characteristics and needs of individual primates, and is intended to develop and complement the existing literature and legislative guidelines. Topics covered include refinements in housing, husbandry and common procedures such as restraint, identification and sampling, with comprehensive advice on issues such as primate communication, assessing and facilitating primate wellbeing, establishing and maintaining social groups, environmental and nutritional enrichment and animal passports. The most commonly used species are the key focus of this resource, but its information and recommendations are generally applicable to other species, provided that relevant individual species characteristics are taken into account.

Clinico-pathologic disparities of breast cancer in Hispanic/Latina women.

BACKGROUND: Breast cancer is the leading cause of cancer death in Hispanic/Latina women nationwide. Limited cancer research has been conducted in this population. El Paso, Texas is a large border city with a population of around 900,000, of which 85% are Latinos and would provide a suitable setting for this study. The aim of this study is to evaluate ethnic differences and cancer characteristics in Hispanic/latina women with breast cancer. METHODS: After IRB approval, we retrospectively analyzed the variables of patients with breast cancer treated consecutively at a large tertiary medical center in El Paso, TX between 2005-2015. Descriptive statistics, bivariate, and multivariable analyses were conducted. RESULTS: 1,252 patients were identified. Mean age at diagnosis was 57 years. 1074 were Hispanics/Latinas (86%). When comparing Hispanics versus non-Hispanics, 31% of Hispanics compared to 24% Non-Hispanics were diagnosed at age <50 (P = 0.043). More Hispanics are uninsured (34%) compared to Non-Hispanics (25%) (p = 0.008). Hispanics presenting with advanced stages were more likely to be uninsured (P = 0.02). CONCLUSIONS: This analysis confirms that Hispanics/Latinas are diagnosed with breast cancer at a younger age and are more commonly uninsured than Non-Hispanics. We did not observe significant differences in the prevalence of ER+, triple negative or Her2 -neu positive disease or stages at presentation between the 2 groups in this cohort, however the non-Hispanic group was constituted only 14% of the studied population. A larger multi-institutional comparative study is being conducted to confirm these findings.

Contextual characteristics associated with the perceived neighbourhood scale in a cross-sectional study in a large urban centre in Brazil.

INTRODUCTION: Health outcomes have been associated with physical and social characteristics of neighbourhoods, but little is known about the relationship between contextual factors and perceived neighbourhood scale. OBJECTIVE: To identify the contextual factors associated with self-perceived neighbourhood scale. METHODS: We analysed data from a cross-sectional population-based study in Belo Horizonte, Brazil, that took place in 2008-2009. The dependent variable was perceived neighbourhood, encoded as an ordinal scale based on a brief description of the concept of the neighbourhood, and two independent scales relating distance, expressed in terms of geography and time. Street connectivity, demographic density and residents' perceptions of the neighbourhoods' physical and social environment were used as contextual predictors. Individual characteristics were used as covariates. Multilevel ordinal logistic regression models estimated the association between perceived neighbourhood scale and contextual characteristics. RESULTS: Residents that perceive better walkability (OR 2.96; 95% CI 1.29 to 3.82) and high amounts of violence (OR 1.35; 95% CI 1.12 to 1.62) perceived their neighbourhoods to be larger, even after adjusting for individual characteristics. CONCLUSION: There are contextual factors that are associated with self-perceived neighbourhood scale. Careful definition of neighbourhood scale is a key factor in improving the results of eco-epidemiological studies. Although these findings must be further explored in other studies, these results can contribute to a better understanding of an appropriate choice of neighbourhood scale, especially for cities in Latin America.

An ovine hepatorenal fibrocystic model of a Meckel-like syndrome associated with dysmorphic primary cilia and TMEM67 mutations.

Meckel syndrome (MKS) is an inherited autosomal recessive hepatorenal fibrocystic syndrome, caused by mutations in TMEM67, characterized by occipital encephalocoele, renal cysts, hepatic fibrosis, and polydactyly. Here we describe an ovine model of MKS, with kidney and liver abnormalities, without polydactyly or occipital encephalocoele. Homozygous missense p.(Ile681Asn; Ile687Ser) mutations identified in ovine TMEM67 were pathogenic in zebrafish phenotype rescue assays. Meckelin protein was expressed in affected and unaffected kidney epithelial cells by immunoblotting, and in primary cilia of lamb kidney cyst epithelial cells by immunofluorescence. In contrast to primary cilia of relatively consistent length and morphology in unaffected kidney cells, those of affected cyst-lining cells displayed a range of short and extremely long cilia, as well as abnormal morphologies, such as bulbous regions along the axoneme. Putative cilia fragments were also consistently located within the cyst luminal contents. The abnormal ciliary phenotype was further confirmed in cultured interstitial fibroblasts from affected kidneys. These primary cilia dysmorphologies and length control defects were significantly greater in affected cells compared to unaffected controls. In conclusion, we describe abnormalities involving primary cilia length and morphology in the first reported example of a large animal model of MKS, in which we have identified TMEM67 mutations.