Distinct metabolic programs established in the thymus control effector functions of γδ T cell subsets in tumor microenvironments.

Metabolic programming controls immune cell lineages and functions, but little is known about γδ T cell metabolism. Here, we found that γδ T cell subsets making either interferon-γ (IFN-γ) or interleukin (IL)-17 have intrinsically distinct metabolic requirements. Whereas IFN-γ+ γδ T cells were almost exclusively dependent on glycolysis, IL-17+ γδ T cells strongly engaged oxidative metabolism, with increased mitochondrial mass and activity. These distinct metabolic signatures were surprisingly imprinted early during thymic development and were stably maintained in the periphery and within tumors. Moreover, pro-tumoral IL-17+ γδ T cells selectively showed high lipid uptake and intracellular lipid storage and were expanded in obesity and in tumors of obese mice. Conversely, glucose supplementation enhanced the antitumor functions of IFN-γ+ γδ T cells and reduced tumor growth upon adoptive transfer. These findings have important implications for the differentiation of effector γδ T cells and their manipulation in cancer immunotherapy.

Ascaris lumbricoides and ticks associated with sensitization to galactose α1,3-galactose and elicitation of the alpha-gal syndrome.

BACKGROUND: IgE to galactose alpha-1,3 galactose (alpha-gal) causes alpha-gal syndrome (delayed anaphylaxis after ingestion of mammalian meat). Development of sensitization has been attributed to tick bites; however, the possible role of other parasites has not been well studied. OBJECTIVE: Our aims were to assess the presence, relative abundances, and site of localization of alpha-gal-containing proteins in common ectoparasites and endoparasites endemic in an area of high prevalence of alpha-gal syndrome, as well as to investigate the ability of ascaris antigens to elicit a reaction in a humanized rat basophil in vitro sensitization model. METHODS: Levels of total IgE, Ascaris-specific IgE, and alpha-gal IgE were measured in sera from patients with challenge-proven alpha-gal syndrome and from controls without allergy. The presence, concentration, and localization of alpha-gal in parasites were assessed by ELISA, Western blotting, and immunohistochemistry. The ability of Ascaris lumbricoides antigen to elicit IgE-dependent reactivity was demonstrated by using the RS-ATL8 basophil reporter system. RESULTS: Alpha-gal IgE level correlated with A lumbricoides-specific IgE level. Alpha-gal protein at 70 to 130 kDa was detected in A lumbricoides at concentrations higher than those found in Rhipicephalus evertsi and Amblyomma hebraeum ticks. Immunohistochemistry was used to localize alpha-gal in tick salivary acini and the helminth gut. Non-alpha-gal-containing A lumbricoides antigens activated RS-ATL8 basophils primed with serum from subjects with alpha-gal syndrome. CONCLUSION: We demonstrated the presence, relative abundances, and site of localization of alpha-gal-containing proteins in parasites. The activation of RS-ATL8 IgE reporter cells primed with serum from subjects with alpha-gal syndrome on exposure to non-alpha-gal-containing A lumbricoides proteins indicates a possible role of exposure to A lumbricoides in alpha-gal sensitization and clinical reactivity.

Transcriptomic Analysis of Glycosylation and Neuroregulatory Pathways in Rodent Models in Response to Psychedelic Molecules.

The potential for psychedelic molecules in impacting cognitive flexibility has long been supported and acknowledged across scientific reports. In the current study, an approach leveraging knowledge-based gene-set information analysis has been adopted to explore the potential impact of psychedelic molecules on both glycosylation, (a post-translational modifications (PTM)) and on neuro-regulatory pathways. Though limitations and restrictions rise from the scarcity of publicly available 'omics' data, targeted analysis enabled us to identify a number of key glycogenes (Hexb, Hs6st2, Col9a2, B3gat2, Mgat5, Bgn) involved the structural organization of extracellular matrix and neuroprotective factors (Kl, Pomc, Oxt, Gal, Avp, Cartpt) which play vital roles in neuron protection, development as well as synaptic stability. In response to psychedelic molecules, we found that these genes and associated pathways are transcriptional altered in rodent models. The approach used indicates the potential to exploit existing datasets for hypothesis generation and testing for the molecular processes which play a role in the physiological response to psychedelic molecule effects. These reported findings, which focused on alterations in glycogenes and neuro-regulatory factors may provide a novel range of biomarkers to track the beneficial, as well as potential toxicological effects of psychedelic molecules.

Plasmonic nanodiscs on vanadium dioxide thin films for tunable luminescence enhancement.

We propose an alternative method to dynamically tune luminescence enhancement in the near infrared spectral range using noble metal nanostructures on top of phase change material vanadium dioxide (VO2) thin films. The VO2 phase change is used to tune the nanodisc plasmon resonance providing a luminescence modification mechanism. We employ a model to calculate the emission of quantum emitters, such as dye molecules, in hybrid systems comprising single silver (Ag) nanodiscs on top of a thin layer of VO2. The model considers different dipole orientations and positions with respect to the nanostructure-VO2 film and determines the degree of observable luminescence modification. In the NIR spectral region, the observable photoluminescence of Alexa Dyes in the hybrid systems at room temperature is enhanced by more than a factor of 2.5 as compared to the same system without plasmonic particles. An additional photoluminescence enhancement by more than a factor of 2 can be achieved with the Ag nanodisc-VO2 film systems after the phase transition of the VO2. These systems can be used for tunable luminescence modification and for compensation of thermally induced luminescence quenching. Through optimization of the Ag nanodisc-VO2 film system, luminescence enhancements of up to a factor of 4 can be seen in the metallic VO2 compared to the semiconducting phase and would therefore compensate for a thermal quenching of up to 70% between room temperature and 70° C, rendering the hybrid systems as promising candidates for improved photon management in optoelectronic devices where elevated temperatures minimize the efficiencies of such devices.

Integrating Fiber Optic Data in Numerical Reservoir Simulation Using Intelligent Optimization Workflow.

A novel workflow is presented for integrating fiber optic Distributed Temperature Sensor (DTS) data in numerical simulation model for the Cyclic Steam Stimulation (CSS) process, using an intelligent optimization routine that automatically learns and improves from experience. As the steam-oil relationship is the main driver for forecasting and decision-making in thermal recovery operations, knowledge of downhole steam distribution across the well over time can optimize injection and production. This study uses actual field data from a CSS operation in a heavy oil field in California, and the value of integrating DTS in the history matching process is illustrated as it allows the steam distribution to be accurately estimated along the entire length of the well. The workflow enables the simultaneous history match of water, oil, and temperature profiles, while capturing the reservoir heterogeneity and the actual physics of the injection process, and ultimately reducing the uncertainty in the predictive models. A novel stepwise grid-refinement approach coupled with an evolutionary optimization algorithm was implemented to improve computational efficiency and predictive accuracy. DTS surveillance also made it possible to detect a thermal communication event due to steam channeling in real-time, and even assess the effectiveness of the remedial workover to resolve it, demonstrating the value of continuous fiber optic monitoring.

Induced stem cell neoplasia in a cnidarian by ectopic expression of a POU domain transcription factor.

The evolutionary origin of stem cell pluripotency is an unresolved question. In mammals, pluripotency is limited to early embryos and is induced and maintained by a small number of key transcription factors, of which the POU domain protein Oct4 is considered central. Clonal invertebrates, by contrast, possess pluripotent stem cells throughout their life, but the molecular mechanisms that control their pluripotency are poorly defined. To address this problem, we analyzed the expression pattern and function of Polynem (Pln), a POU domain gene from the marine cnidarian Hydractinia echinata. We show that Pln is expressed in the embryo and adult stem cells of the animal and that ectopic expression in epithelial cells induces stem cell neoplasms and loss of epithelial tissue. Neoplasm cells downregulated the transgene but expressed the endogenous Pln gene and also Nanos, Vasa, Piwi and Myc, which are all known cnidarian stem cell markers. Retinoic acid treatment caused downregulation of Pln and the differentiation of neoplasm cells to neurosensory and epithelial cells. Pln downregulation by RNAi led to differentiation. Collectively, our results suggest an ancient role of POU proteins as key regulators of animal stem cells.

Development of a convenient competitive ELISA for the detection of the free and protein-bound nonhuman galactosyl-α-(1,3)-galactose epitope based on highly specific chicken single-chain antibody variable-region fragments.

The presence of the nonhuman galactosyl-α-(1,3)-galactose (Gal-α-(1,3)-Gal) carbohydrate epitope on a number of recombinant therapeutic proteins has recently been reported, renewing interest in this immunogenic carbohydrate epitope. It is well-known that this motif is the primary contributing factor in hyperacute rejection of porcine organ xenograft, due to the existence of natural antibodies against this epitope in human serum. Though the number of epitopes on recombinant glycoproteins may be low when compared directly to whole tissue, circulating anti-Gal-α-R immunoglobulins can still induce anaphylaxis. Therefore, there is a need for rapid and convenient methods for detection and monitoring of this epitope in biopharmaceuticals produced in recombinant mammalian systems. To this end, we have generated immune-challenged chicken single-chain antibody variable-region fragment (scFv) libraries targeting the Gal-α-(1,3)-Gal motif and have selected a panel of scFv's that bind the target. We have used one of these antibodies to develop a competitive ELISA for both free and protein-bound Gal-α-(1,3)-Gal and have demonstrated that the ELISA is specific for the target and can be used to determine the loading of the target on glycoproteins. This competitive ELISA will provide a convenient method of detecting and quantifying Gal-α-(1,3)-Gal on therapeutic glycoproteins.

Effect of nanoparticle stabilization and physicochemical properties on exposure outcome: acute toxicity of silver nanoparticle preparations in zebrafish (Danio rerio).

Nanotechnology has vast potential for expanded development and novel application in numerous sectors of society. With growing use and applications, substantial production volumes and associated environmental release can be anticipated. Exposure effect of nanoparticles (NP) on biological systems may be intrinsic to their physicochemical properties introducing unknown associated risk. Herein, we expand the knowledge of health and environmental impact of silver nanoparticles (AgNPs), testing the acute toxicity of 14 AgNP preparations on developing zebrafish embryos (Danio rerio). Toxicological end points, including mortality, hatching rate, and heart rate were recorded. Concentration, stabilization agent and physicochemical properties were monitored as contributing outcome factors. Our findings indicate wide ranging LC50 24 h postfertilization values (0.487 ppm (0.315, 0.744 95% CI) to 47.89 ppm (18.45, 203.49 95% CI)), and indicate surface charge and ionic dissolution as key contributory factors in AgNP exposure outcome.

Bacterial Staining and Profiling for Glycan Interactions on Glycan Microarrays for t-Test Calculation.

The use of glycan microarrays to study carbohydrate interactions of bacterial cells is of great interest owing to the key roles these interactions play in bacterial colonization and infection of a host. In this chapter, the methods to fluorescently stain Gram-positive or Gram-negative bacteria and profiling them for glycan interactions using glycan microarrays are described in detail. The application of the Student's t-test to glycan microarray data using an example data set comparing glycan microarray binding of an Acinetobacter baumannii wild type and mutant strain is also described in step-by-step detail.

Gastrointestinal Symptoms in Autism Spectrum Disorder: A Systematic Review.

This systematic review aims to offer an updated understanding of the relationship between gastrointestinal symptoms (GIS) and autism spectrum disorder (ASD) in children and adolescents. The databases PsycINFO, Medline, Cinahl, and ERIC were searched using keywords, and relevant literature was hand-searched. Papers (n = 3319) were systematically screened and deemed eligible if they were empirical studies published in English since 2014 and measured the GIS of individuals with ASD who were under 18 years old. Thirty studies were included in the final review. The study findings were synthesized under eight themes, including the prevalence and nature of GIS and their relationship with developmental regression, language and communication, ASD severity, challenging behavior, comorbid psychopathology, sleep problems, and sensory issues. The review found that GIS were common and that there was contradictory evidence concerning their relationship with co-occurring conditions. It also identified evidence of some causal relationships that support the existence of the gut-immune-brain pathways. Future research needs to use large prospective designs and objective and standardized GIS measurements to provide a nuanced understanding of GIS in the context of ASD.

Transcriptomic Analysis of Respiratory Tissue and Cell Line Models to Examine Glycosylation Machinery during SARS-CoV-2 Infection.

Glycosylation, being the most abundant post-translational modification, plays a profound role affecting expression, localization and function of proteins and macromolecules in immune response to infection. Presented are the findings of a transcriptomic analysis performed using high-throughput functional genomics data from public repository to examine the altered transcription of the human glycosylation machinery in response to SARS-CoV-2 stimulus and infection. In addition to the conventional in silico functional enrichment analysis methods we also present results from the manual analysis of biomedical literature databases to bring about the biological significance of glycans and glycan-binding proteins in modulating the host immune response during SARS-CoV-2 infection. Our analysis revealed key immunomodulatory lectins, proteoglycans and glycan epitopes implicated in exerting both negative and positive downstream inflammatory signaling pathways, in addition to its vital role as adhesion receptors for SARS-CoV-2 pathogen. A hypothetical correlation of the differentially expressed human glycogenes with the altered host inflammatory response and the cytokine storm-generated in response to SARS-CoV-2 pathogen is proposed. These markers can provide novel insights into the diverse roles and functioning of glycosylation pathways modulated by SARS-CoV-2, provide avenues of stratification, treatment, and targeted approaches for COVID-19 immunity and other viral infectious agents.

An integrative network analysis framework for identifying molecular functions in complex disorders examining major depressive disorder as a test case.

In addition to the psychological depressive phenotype, major depressive disorder (MDD) patients are also associated with underlying immune dysregulation that correlates with metabolic syndrome prevalent in depressive patients. A robust integrative analysis of biological pathways underlying the dysregulated neural connectivity and systemic inflammatory response will provide implications in the development of effective strategies for the diagnosis, management and the alleviation of associated comorbidities. In the current study, focusing on MDD, we explored an integrative network analysis methodology to analyze transcriptomic data combined with the meta-analysis of biomarker data available throughout public databases and published scientific peer-reviewed articles. Detailed gene set enrichment analysis and complex protein-protein, gene regulatory and biochemical pathway analysis has been undertaken to identify the functional significance and potential biomarker utility of differentially regulated genes, proteins and metabolite markers. This integrative analysis method provides insights into the molecular mechanisms along with key glycosylation dysregulation underlying altered neutrophil-platelet activation and dysregulated neuronal survival maintenance and synaptic functioning. Highlighting the significant gap that exists in the current literature, the network analysis framework proposed reduces the impact of data gaps and permits the identification of key molecular signatures underlying complex disorders with multiple etiologies such as within MDD and presents multiple treatment options to address their molecular dysfunction.

Glucagon receptor signaling regulates weight loss via central KLB receptor complexes.

Glucagon regulates glucose and lipid metabolism and promotes weight loss. Thus, therapeutics stimulating glucagon receptor (GCGR) signaling are promising for obesity treatment; however, the underlying mechanism(s) have yet to be fully elucidated. We previously identified that hepatic GCGR signaling increases circulating fibroblast growth factor 21 (FGF21), a potent regulator of energy balance. We reported that mice deficient for liver Fgf21 are partially resistant to GCGR-mediated weight loss, implicating FGF21 as a regulator of glucagon's weight loss effects. FGF21 signaling requires an obligate coreceptor (ß-Klotho, KLB), with expression limited to adipose tissue, liver, pancreas, and brain. We hypothesized that the GCGR-FGF21 system mediates weight loss through a central mechanism. Mice deficient for neuronal Klb exhibited a partial reduction in body weight with chronic GCGR agonism (via IUB288) compared with controls, supporting a role for central FGF21 signaling in GCGR-mediated weight loss. Substantiating these results, mice with central KLB inhibition via a pharmacological KLB antagonist, 1153, also displayed partial weight loss. Central KLB, however, is dispensable for GCGR-mediated improvements in plasma cholesterol and liver triglycerides. Together, these data suggest GCGR agonism mediates part of its weight loss properties through central KLB and has implications for future treatments of obesity and metabolic syndrome.

Glycobiomimics and glycobiosensors.

Following steady advances in analytical technologies, our knowledge in glycomics is now increasing rapidly. Over the last decade, specific glycans have been described that are associated with a range of diseases, such as cancer and inflammation, with host-pathogen interactions and with various stages during stem cell development and differentiation. Simultaneously, deeper structural insight has been gained on glycosylated biopharmaceutical protein therapeutics manufactured in CHO (Chinese-hamster ovary) and other cell systems. This glycomic information is highly relevant for clinicians and biomanufacturing industries as a new class of glycobiomarkers emerges. However, current methods of glycoanalysis are primarily research tools and are not suitable for point-of-care on-site detection and analysis, or sensor devices. Lectin-based glycan detection provides the most promising approach to fill these gaps. However, the limited availability of lectins with high specificity and sensitivity for specific glycan motifs presents one of the main challenges in building reliable glycobiosensors. Recent reports have demonstrated the use of recombinant protein engineering, phage display and aptamer technologies in the production of lectin mimics, as well as the construction of biosensors that are capable of rapidly detecting glycan motifs at low levels in both a labelled and label-free manner. These are primarily proof-of-principle reports at this stage, but some of the approaches, either alone or in combination, will lead to functional glycobiosensors in the coming years which will be valuable tools for the clinical, biopharmaceutical and life science research communities.

Glyco-biosensors: recent advances and applications for the detection of free and bound carbohydrates.

The field of biosensor development now encompasses several areas specifically geared toward the rapid and sensitive detection, identification, and quantification of target analytes. In contrast to the more mature research and development of nucleic acid and protein biosensors, the development of 'glyco-biosensors' for detecting carbohydrates and conjugates of carbohydrates (glycoconjugates) is at a relatively nascent stage. The application of glyco-biosensors aims to open novel analytical and diagnostic avenues, encompassing industrial bioprocesses, biomedical and clinical applications. This area of research has been greatly aided by advancement brought by interdisciplinary mergers of engineering, biology, chemistry and physical sciences and enabling the miniaturization of detection platforms. In this review, we briefly introduce the need for glyco-biosensors, discuss current analytical technologies, and examine advances in glyco-biosensor approaches aimed at the detection and/or quantification of glycoconjugates or carbohydrates derived from glycoconjugates since 2005.

Genome, Environment, Microbiome and Metabolome in Autism (GEMMA) Study Design: Biomarkers Identification for Precision Treatment and Primary Prevention of Autism Spectrum Disorders by an Integrated Multi-Omics Systems Biology Approach.

Autism Spectrum Disorder (ASD) affects approximately 1 child in 54, with a 35-fold increase since 1960. Selected studies suggest that part of the recent increase in prevalence is likely attributable to an improved awareness and recognition, and changes in clinical practice or service availability. However, this is not sufficient to explain this epidemiological phenomenon. Research points to a possible link between ASD and intestinal microbiota because many children with ASD display gastro-intestinal problems. Current large-scale datasets of ASD are limited in their ability to provide mechanistic insight into ASD because they are predominantly cross-sectional studies that do not allow evaluation of perspective associations between early life microbiota composition/function and later ASD diagnoses. Here we describe GEMMA (Genome, Environment, Microbiome and Metabolome in Autism), a prospective study supported by the European Commission, that follows at-risk infants from birth to identify potential biomarker predictors of ASD development followed by validation on large multi-omics datasets. The project includes clinical (observational and interventional trials) and pre-clinical studies in humanized murine models (fecal transfer from ASD probands) and in vitro colon models. This will support the progress of a microbiome-wide association study (of human participants) to identify prognostic microbiome signatures and metabolic pathways underlying mechanisms for ASD progression and severity and potential treatment response.

γδ T cells in cancer: a small population of lymphocytes with big implications.

γδ T cells are a small population of mostly tissue-resident lymphocytes, with both innate and adaptive properties. These unique features make them particularly attractive candidates for the development of new cellular therapy targeted against tumor development. Nevertheless, γδ T cells may play dual roles in cancer, promoting cancer development on the one hand, while participating in antitumor immunity on the other hand. In mice, γδ T-cell subsets preferentially produce IL-17 or IFN-γ. While antitumor functions of murine γδ T cells can be attributed to IFN-γ+ γδ T cells, recent studies have implicated IL-17+ γδ T cells in tumor growth and metastasis. However, in humans, IL-17-producing γδ T cells are rare and most studies have attributed a protective role to γδ T cells against cancer. In this review, we will present the current knowledge and most recent findings on γδ T-cell functions in mouse models of tumor development and human cancers. We will also discuss their potential as cellular immunotherapy against cancer.

Vulnerability to stress: Personality facet of vulnerability is associated with cardiovascular adaptation to recurring stress.

It is increasingly suggested that personality traits are critical to understanding patterns of cardiovascular stress adaptation. However, studies have focused on higher-order traits with no research having examined underlying facet effects to repeated stress. The examination of facets provides a more granular examination, which has the potential to identify specific personality components that are relevant within the context of psychophysiological stress adaptation. This study objective was to determine if the underlying facets which encapsulate the dimension of emotional stability, are associated with cardiovascular adaptation to recurring stress. Continuous cardiovascular monitoring and psychometric measures were collated from 79 healthy young male and female adults, across a protocol of recurring active stress tasks. Multiple regression analysis revealed that the facet of vulnerability was associated with systolic and diastolic blood pressure adaptation across the protocol. More specifically, vulnerability was negatively associated with adaptation to recurring stress, such that those highest in vulnerability displayed a sensitization to the recurring stressor. No significant effects emerged for any other facet. Importantly, this research adds to the existing literature examining stress adaptation and has implications for future research on the relevance of examining facet effects. This study is the first to implicate the personality facet of vulnerability which encapsulates an individual's tendency to feel unable to cope with stress and becoming hopeless when faced with emergency situations, in the context of cardiovascular stress adaptation. Taken together, this study suggests that the facet of vulnerability is a critical component to consider in the context of cardiovascular stress adaptation.

Activity-based selection of a proteolytic species using ribosome display.

We have examined the potential of displaying a protease species in vitro using ribosome display and demonstrate specific capture on the basis of its catalytic activity. Using a model bacterial cysteine protease, sortase A (SrtA), we show that this enzyme can be functionally expressed in vitro. By overlap PCR we constructed ribosome display templates with the SrtA open reading frame fused to a C terminal glycine-serine rich flexible linker and a tether derived from eGFP. Using the broad range cysteine protease irreversible inhibitor E-64 linked to acrylic beads, we show that we can isolate SrtA ribosome display ternary complexes, and recover their encoding mRNA by RT-PCR. This recovery was lost when applied to a SrtA catalytically inactive mutant, or could be alleviated by competition with free inhibitor. This sensitive technique could be further developed to allow the screening of proteases against putative inhibitors and/or the identification of novel proteolytic species.

The neuropeptide genes TAC1, TAC3, TAC4, VIP and PACAP(ADCYAP1), and susceptibility to multiple sclerosis.

The related immunomodulatory neuropeptides vasoactive intestinal peptide (VIP) and pituitary adenylyl cyclase activating peptide (PACAP; gene symbol ADCYAP1) have recently been proposed as novel therapeutics for the treatment of multiple sclerosis (MS). These neuropeptides, as well as those belonging to the tachykinin family exert pleiotropic effects, many of which are of relevance to central nervous system inflammation. In the present study, we have analysed 14 single nucleotide polymorphisms (SNPs) and 4 microsatellite markers in the VIP, ADCYAP1, TAC3 and TAC4 genes for susceptibility to MS in a case-control collection from Northern Ireland. Following correction for multiple comparisons, we did not find any significant associations between single polymorphic markers or multiple-marker haplotypes and susceptibility to MS. Furthermore, we analysed 2 SNPs in the TAC1 gene in a set of Sardinian trio MS families, based on our previous observation of association of these SNPs with MS in the Northern Irish (Genes Immun. 2005, 6, 265-270). Analysis of these SNPs in the Sardinians was not significant though a similar trend to that originally observed in the Northern Irish was present. Meta-analysis of the Sardinian and Northern Irish TAC1 SNP genotype data revealed a Mantel-Haenszel Common OR Estimate for the TAC1 intron 1 SNP rs2072100 of 0.76 (95% CI 0.63-0.92; P=0.005; A allele) and for the TAC1 promoter SNP rs7793277 of 0.76 (95% CI 0.615-0.95; P=0.014; C allele). Our data advocate a need for further exploration of the TAC1 gene region in MS.