Ability of γδ T cells to modulate the Foxp3 T cell response is dependent on adenosine.

Whether γδ T cells inhibit or enhance the Foxp3 T cell response depends upon their activation status. The critical enhancing effector in the supernatant is adenosine. Activated γδ T cells express adenosine receptors at high levels, which enables them to deprive Foxp3+ T cells of adenosine, and to inhibit their expansion. Meanwhile, cell-free supernatants of γδ T cell cultures enhance Foxp3 T cell expansion. Thus, inhibition and enhancement by γδ T cells of Foxp3 T cell response are a reflection of the balance between adenosine production and absorption by γδ T cells. Non-activated γδ T cells produce adenosine but bind little, and thus enhance the Foxp3 T cell response. Activated γδ T cells express high density of adenosine receptors and have a greatly increased ability to bind adenosine. Extracellular adenosine metabolism and expression of adenosine receptor A2ARs by γδ T cells played a major role in the outcome of γδ and Foxp3 T cell interactions. A better understanding of the functional conversion of γδ T cells could lead to γδ T cell-targeted immunotherapies for related diseases

Bifidobacterium breve UCC2003 employs multiple transcriptional regulators to control metabolism of particular human milk oligosaccharides

Bifidobacterial carbohydrate metabolism has been studied in considerable detail for a variety of both plant and human-derived glycans, particularly involving the bifidobacterial prototype Bifidobacterium breve UCC2003. We recently elucidated the metabolic pathways by which the human milk oligosaccharide (HMO) constituents lacto-N-tetraose (LNT), lacto-N-neotetraose (LNnT) and lacto-N-biose (LNB) are utilized by B. breve UCC2003. However, to date no work has been carried out on the regulatory mechanisms that control expression of the genetic loci involved in these HMO metabolic pathways. In the current study, we describe the characterization of three transcriptional regulators and corresponding operator and associated (inducible) promoter sequences, the latter governing transcription of the genetic elements involved in LN(n)T/LNB metabolism. The activity of these regulators is dependent on the release of specific monosaccharides, which are believed to act as allosteric effectors, and which are derived from the corresponding HMOs targeted by the particular locus.Importance Human milk oligosaccharides (HMOs) are a key factor in the development of the breastfed infant microbiota. They function as prebiotics, selecting for a specific range of microbes, including a number of infant-associated species of bifidobacteria, which are thought to provide a range of health benefits to the infant host. While much research has been carried out on elucidating the mechanisms of HMO metabolism in infant-associated bifidobacteria, there is to date very little understanding of the transcriptional regulation of these pathways. The current study reveals a multi-component transcriptional regulation system that controls the recently-identified pathways of HMO metabolism in the infant-associated Bifidobacterium breve prototype strain UCC2003. This not only provides insight into the regulatory mechanisms present in other infant-associated bifidobacteria, but also provides an example of a network of sequential steps regulating microbial carbohydrate metabolism

Role of gammadelta T cells in protecting normal airway function

Since their discovery 15 years ago, the role of γδ T cells has remained somewhat elusive. Responses of γδ T cells have been found in numerous infectious and non-infectious diseases. New evidence points to γδ T cells' functioning in the airways to maintain normal airway responsiveness or tone. In the lung, distinct subsets of γδ T cell subsets seem to have specific roles, one subset promoting allergic inflammation, the other serving a protective role

PI 3-kinase- and ERK-MAPK-dependent mechanisms underlie Glucagon-Like Peptide-1-mediated activation of Sprague Dawley colonic myenteric neurons

Background: Glucagon-like peptide (GLP-1) can modify colonic function, with beneficial effects reported in the functional bowel disorder, irritable bowel syndrome (IBS). IBS pathophysiology is characterized by hyper-activation of the hypothalamic-pituitary-adrenal stress axis and altered microbial profiles. This study aims to characterize the neuronal and functional effects of GLP-1 in healthy rat colons to aid understanding of its beneficial effects in moderating bowel dysfunction. Methods: Immunofluorescent and calcium imaging of myenteric neurons prepared from Sprague Dawley rat colons was carried out to elucidate the neuromodulatory actions of the GLP-1 receptor agonist, exendin-4 (Ex-4). Colonic contractile activity was assessed using organ bath physiological recordings. Key results: Ex-4 induced an elevation of intracellular calcium arising from store release and influx via voltage-gated calcium channels. Ex-4 activated both ERK-MAPK and PI 3-kinase signaling cascades. Neuronal activation was found to underlie suppression of contractile activity in colonic circular muscle. Although the stress hormone, corticotropin-releasing factor (CRF) potentiated the neuronal response to Ex-4, and the functional effects of Ex-4 on colonic circular muscle activity were not altered. Conclusions and inferences: Ex-4 evoked neurally regulated suppression of rat colonic circular muscle activity. In myenteric neurons, the neurostimulatory effects of Ex-4 were dependent upon activation of PI 3-kinase and ERK-MAPK signaling cascades. No further change in circular muscle function was noted in the presence of CRF suggesting that stress does not impact on colonic function in health. Further studies in a model of IBS are needed to determine whether mechanisms are modified in the context of bowel dysfunction

MicroRNA profiling reveals marker of motor neuron disease in ALS models

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder marked by the loss of motor neurons (MNs) in the brain and spinal cord, leading to fatally debilitating weakness. Because this disease predominantly affects MNs, we aimed to characterize the distinct expression profile of that cell type to elucidate underlying disease mechanisms and to identify novel targets that inform on MN health during ALS disease time course. microRNAs (miRNAs) are short, noncoding RNAs that can shape the expression profile of a cell and thus often exhibit cell-type-enriched expression. To determine MN-enriched miRNA expression, we used Cre recombinase-dependent miRNA tagging and affinity purification in mice. By defining thein vivomiRNA expression of MNs, all neurons, astrocytes, and microglia, we then focused on MN-enriched miRNAs via a comparative analysis and found that they may functionally distinguish MNs postnatally from other spinal neurons. Characterizing the levels of the MN-enriched miRNAs in CSF harvested from ALS models of MN disease demonstrated that one miRNA (miR-218) tracked with MN loss and was responsive to an ALS therapy in rodent models. Therefore, we have used cellular expression profiling tools to define the distinct miRNA expression of MNs, which is likely to enrich future studies of MN disease. This approach enabled the development of a novel, drug-responsive marker of MN disease in ALS rodents.SIGNIFICANCE STATEMENTAmyotrophic lateral sclerosis (ALS) is a neurodegenerative disease in which motor neurons (MNs) in the brain and spinal cord are selectively lost. To develop tools to aid in our understanding of the distinct expression profiles of MNs and, ultimately, to monitor MN disease progression, we identified small regulatory microRNAs (miRNAs) that were highly enriched or exclusive in MNs. The signal for one of these MN-enriched miRNAs is detectable in spinal tap biofluid from an ALS rat model, where its levels change as disease progresses, suggesting that it may be a clinically useful marker of disease status. Furthermore, rats treated with ALS therapy have restored expression of this MN RNA marker, making it an MN-specific and drug-responsive marker for ALS rodents.</jats:p

Leptin modifies the prosecretory and prokinetic effects of the inflammatory cytokine interleukin-6 on colonic function in Sprague–Dawley rats

Leptin ameliorates the prosecretory and prokinetic effects of the pro-inflammatory cytokine interleukin-6 on rat colon. Leptin also suppresses the neurostimulatory effects of irritable bowel syndrome plasma, which has elevated concentrations of interleukin-6, on enteric neurons. This may indicate a regulatory role for leptin in immune-mediated bowel dysfunction. In addition to its role in regulating energy homeostasis, the adipokine leptin modifies gastrointestinal (GI) function. Indeed, leptin-resistant obese humans and leptin-deficient obese mice exhibit altered GI motility. In the functional GI disorder irritable bowel syndrome (IBS), circulating leptin concentrations are reported to differ from those of healthy control subjects. Additionally, IBS patients display altered cytokine profiles, including elevated circulating concentrations of the pro-inflammatory cytokine interleukin-6 (IL-6), which bears structural homology and similarities in intracellular signalling to leptin. This study aimed to investigate interactions between leptin and IL-6 in colonic neurons and their possible contribution to IBS pathophysiology. The functional effects of leptin and IL-6 on colonic contractility and absorptosecretory function were assessed in organ baths and Ussing chambers in Sprague–Dawley rat colon. Calcium imaging and immunohistochemical techniques were used to investigate the neural regulation of GI function by these signalling molecules. Our findings provide a neuromodulatory role for leptin in submucosal neurons, where it inhibited the stimulatory effects of IL-6. Functionally, this translated to suppression of IL-6-evoked potentiation of veratridine-induced secretory currents. Leptin also attenuated IL-6-induced colonic contractions, although it had little direct effect on myenteric neurons. Calcium responses evoked by IBS plasma in both myenteric and submucosal neurons were also suppressed by leptin, possibly through interactions with IL-6, which is elevated in IBS plasma. As leptin has the capacity to ameliorate the neurostimulatory effects of soluble mediators in IBS plasma and modulated IL-6-evoked changes in bowel function, leptin may have a role in immune-mediated bowel dysfunction in IBS patients

Nuclear membrane-localised NOX4D generates pro-survival ROS in FLT3-ITD-expressing AML

Internal tandem duplication of the juxtamembrane domain of FMS-like tyrosine kinase 3 (FLT3-ITD) is the most prevalent genetic aberration present in 20-30% of acute myeloid leukaemia (AML) cases and is associated with a poor prognosis. FLT3-ITD expressing cells express elevated levels of NADPH oxidase 4 (NOX4)generated pro-survival hydrogen peroxide (H2O2) contributing to increased levels of DNA oxidation and double strand breaks. NOX4 is constitutively active and has been found to have various isoforms expressed at multiple locations within a cell. The purpose of this study was to investigate the expression, localisation and regulation of NOX4 28 kDa splice variant, NOX4D. NOX4D has previously been shown to localise to the nucleus and nucleolus in various cell types and is implicated in the generation of reactive oxygen species (ROS) and DNA damage. Here, we demonstrate that FLT3ITD expressing-AML patient samples as well as -cell lines express the NOX4D isoform resulting in elevated H2O2 levels compared to FLT3-WT expressing cells, as quantified by flow cytometry. Cell fractionation indicated that NOX4D is nuclear membrane-localised in FLT3-ITD expressing cells. Treatment of MV4-11 cells with receptor trafficking inhibitors, tunicamycin and brefeldin A, resulted in deglycosylation of NOX4 and NOX4D. Inhibition of the FLT3 receptor revealed that the FLT3-ITD oncogene is responsible for the production of NOX4D-generated H2O2 in AML. We found that inhibition of the PI3K/AKT and STAT5 pathways resulted in down-regulation of NOX4D-generated pro-survival ROS. Taken together these findings indicate that nuclear membrane-localised NOX4D-generated pro-survival H2O2 may be contributing to genetic instability in FLT3-ITD expressing AML