Homeostatic IL-13 in healthy skin directs dendritic cell differentiation to promote TH2 and inhibit TH17 cell polarization.

The signals driving the adaptation of type 2 dendritic cells (DC2s) to diverse peripheral environments remain mostly undefined. We show that differentiation of CD11blo migratory DC2s-a DC2 population unique to the dermis-required IL-13 signaling dependent on the transcription factors STAT6 and KLF4, whereas DC2s in lung and small intestine were STAT6-independent. Similarly, human DC2s in skin expressed an IL-4 and IL-13 gene signature that was not found in blood, spleen and lung DCs. In mice, IL-13 was secreted homeostatically by dermal innate lymphoid cells and was independent of microbiota, TSLP or IL-33. In the absence of IL-13 signaling, dermal DC2s were stable in number but remained CD11bhi and showed defective activation in response to allergens, with diminished ability to support the development of IL-4+GATA3+ helper T cells (TH), whereas antifungal IL-17+RORγt+ TH cells were increased. Therefore, homeostatic IL-13 fosters a noninflammatory skin environment that supports allergic sensitization.

Differentiation of type 1 ILCs from a common progenitor to all helper-like innate lymphoid cell lineages.

Innate lymphoid cells (ILCs) are a recently recognized group of lymphocytes that have important functions in protecting epithelial barriers against infections and in maintaining organ homeostasis. ILCs have been categorized into three distinct groups, transcriptional circuitry and effector functions of which strikingly resemble the various T helper cell subsets. Here, we identify a common, Id2-expressing progenitor to all interleukin 7 receptor-expressing, "helper-like" ILC lineages, the CHILP. Interestingly, the CHILP differentiated into ILC2 and ILC3 lineages, but not into conventional natural killer (cNK) cells that have been considered an ILC1 subset. Instead, the CHILP gave rise to a peculiar NKp46(+) IL-7Rα(+) ILC lineage that required T-bet for specification and was distinct of cNK cells or other ILC lineages. Such ILC1s coproduced high levels of IFN-γ and TNF and protected against infections with the intracellular parasite Toxoplasma gondii. Our data significantly advance our understanding of ILC differentiation and presents evidence for a new ILC lineage that protects barrier surfaces against intracellular infections.

Neuro-mesenchymal units control ILC2 and obesity via a brain-adipose circuit.

Signals from sympathetic neurons and immune cells regulate adipocytes and thereby contribute to fat tissue biology. Interactions between the nervous and immune systems have recently emerged as important regulators of host defence and inflammation1-4. Nevertheless, it is unclear whether neuronal and immune cells co-operate in brain-body axes to orchestrate metabolism and obesity. Here we describe a neuro-mesenchymal unit that controls group 2 innate lymphoid cells (ILC2s), adipose tissue physiology, metabolism and obesity via a brain-adipose circuit. We found that sympathetic nerve terminals act on neighbouring adipose mesenchymal cells via the ß2-adrenergic receptor to control the expression of glial-derived neurotrophic factor (GDNF) and the activity of ILC2s in gonadal fat. Accordingly, ILC2-autonomous manipulation of the GDNF receptor machinery led to alterations in ILC2 function, energy expenditure, insulin resistance and propensity to obesity. Retrograde tracing and chemical, surgical and chemogenetic manipulations identified a sympathetic aorticorenal circuit that modulates ILC2s in gonadal fat and connects to higher-order brain areas, including the paraventricular nucleus of the hypothalamus. Our results identify a neuro-mesenchymal unit that translates cues from long-range neuronal circuitry into adipose-resident ILC2 function, thereby shaping host metabolism and obesity.

Interleukin-15-Dependent T-Cell-like Innate Intraepithelial Lymphocytes Develop in the Intestine and Transform into Lymphomas in Celiac Disease.

The nature of gut intraepithelial lymphocytes (IELs) lacking antigen receptors remains controversial. Herein we showed that, in humans and in mice, innate intestinal IELs expressing intracellular CD3 (iCD3(+)) differentiate along an Id2 transcription factor (TF)-independent pathway in response to TF NOTCH1, interleukin-15 (IL-15), and Granzyme B signals. In NOTCH1-activated human hematopoietic precursors, IL-15 induced Granzyme B, which cleaved NOTCH1 into a peptide lacking transcriptional activity. As a result, NOTCH1 target genes indispensable for T cell differentiation were silenced and precursors were reprogrammed into innate cells with T cell marks including intracellular CD3 and T cell rearrangements. In the intraepithelial lymphoma complicating celiac disease, iCD3(+) innate IELs acquired gain-of-function mutations in Janus kinase 1 or Signal transducer and activator of transcription 3, which enhanced their response to IL-15. Overall we characterized gut T cell-like innate IELs, deciphered their pathway of differentiation and showed their malignant transformation in celiac disease.

Author Correction: Light-entrained and brain-tuned circadian circuits regulate ILC3s and gut homeostasis.

An Amendment to this paper has been published and can be accessed via a link at the top of the paper.

Light-entrained and brain-tuned circadian circuits regulate ILC3s and gut homeostasis.

Group 3 innate lymphoid cells (ILC3s) are major regulators of inflammation, infection, microbiota composition and metabolism1. ILC3s and neuronal cells have been shown to interact at discrete mucosal locations to steer mucosal defence2,3. Nevertheless, it is unclear whether neuroimmune circuits operate at an organismal level, integrating extrinsic environmental signals to orchestrate ILC3 responses. Here we show that light-entrained and brain-tuned circadian circuits regulate enteric ILC3s, intestinal homeostasis, gut defence and host lipid metabolism in mice. We found that enteric ILC3s display circadian expression of clock genes and ILC3-related transcription factors. ILC3-autonomous ablation of the circadian regulator Arntl led to disrupted gut ILC3 homeostasis, impaired epithelial reactivity, a deregulated microbiome, increased susceptibility to bowel infection and disrupted lipid metabolism. Loss of ILC3-intrinsic Arntl shaped the gut 'postcode receptors' of ILC3s. Strikingly, light-dark cycles, feeding rhythms and microbial cues differentially regulated ILC3 clocks, with light signals being the major entraining cues of ILC3s. Accordingly, surgically or genetically induced deregulation of brain rhythmicity led to disrupted circadian ILC3 oscillations, a deregulated microbiome and altered lipid metabolism. Our work reveals a circadian circuitry that translates environmental light cues into enteric ILC3s, shaping intestinal health, metabolism and organismal homeostasis.

CkP1 bacteriophage, a S16-like myovirus that recognizes Citrobacter koseri lipopolysaccharide through its long tail fibers.

Citrobacter koseri is an emerging Gram-negative bacterial pathogen, which causes urinary tract infections. We isolated and characterized a novel S16-like myovirus CKP1 (vB_CkoM_CkP1), infecting C. koseri. CkP1 has a host range covering the whole C. koseri species, i.e., all strains that were tested, but does not infect other species. Its linear 168,463-bp genome contains 291 coding sequences, sharing sequence similarity with the Salmonella phage S16. Based on surface plasmon resonance and recombinant green florescence protein fusions, the tail fiber (gp267) was shown to decorate C. koseri cells, binding with a nanomolar affinity, without the need of accessory proteins. Both phage and the tail fiber specifically bind to bacterial cells by the lipopolysaccharide polymer. We further demonstrate that CkP1 is highly stable towards different environmental conditions of pH and temperatures and is able to control C. koseri cells in urine samples. Altogether, CkP1 features optimal in vitro characteristics to be used both as a control and detection agent towards drug-resistant C. koseri infections. KEY POINTS: • CkP1 infects all C. koseri strains tested • CkP1 recognizes C. koseri lipopolysaccharide through its long tail fiber • Both phage CkP1 and its tail fiber can be used to treat or detect C. koseri pathogens.

Orthorexia nervosa in yoga practitioners: relationship with personality, attitudes about appearance, and yoga engagement.

PURPOSE: Disordered eating symptoms and a high prevalence of orthorexia nervosa can be found in yoga practitioners. Given that yoga is increasingly used as a complementary treatment for eating disorders (ED), understanding the relationship between yoga practice and the development of disordered eating is crucial to guide treatment recommendations. The goal of this work is, therefore, to study the relationships between orthorexia nervosa (ON) and potential risk factors for ON, in an international sample of experienced yoga practitioners. METHOD: An online questionnaire that included several psychometric instruments was responded by 469 yoga practitioners. Instruments used were the Teruel orthorexia scale, Yoga immersion scale, Passion scale, Frost multidimensional perfectionism scale, Self-discipline scale of NEO-PI-R, Drive for thinness scale of EDI, and Beliefs about appearance scale. Descriptive statistics, correlational analysis and multiple regression were used to evaluate relationships between ON and the other variables. RESULTS: The main predictors of orthorexia nervosa were the drive for thinness and a healthy orthorexia, suggesting that, like in anorexia and bulimia, orthorexic individuals are also concerned with food quantity and physical appearance, rather than just food quality. CONCLUSIONS: The potential effects of yoga on eating behaviours and attitudes of long-term practitioners, particularly the high prevalence of orthorexia nervosa and the concern for physical appearance, should be taken into consideration when using yoga as prevention or treatment for eating disorders. LEVEL OF EVIDENCE: Level V, descriptive cross-sectional study.

The neurotrophic factor receptor RET drives haematopoietic stem cell survival and function.

Haematopoiesis is a developmental cascade that generates all blood cell lineages in health and disease. This process relies on quiescent haematopoietic stem cells capable of differentiating, self renewing and expanding upon physiological demand. However, the mechanisms that regulate haematopoietic stem cell homeostasis and function remain largely unknown. Here we show that the neurotrophic factor receptor RET (rearranged during transfection) drives haematopoietic stem cell survival, expansion and function. We find that haematopoietic stem cells express RET and that its neurotrophic factor partners are produced in the haematopoietic stem cell environment. Ablation of Ret leads to impaired survival and reduced numbers of haematopoietic stem cells with normal differentiation potential, but loss of cell-autonomous stress response and reconstitution potential. Strikingly, RET signals provide haematopoietic stem cells with critical Bcl2 and Bcl2l1 surviving cues, downstream of p38 mitogen-activated protein (MAP) kinase and cyclic-AMP-response element binding protein (CREB) activation. Accordingly, enforced expression of RET downstream targets, Bcl2 or Bcl2l1, is sufficient to restore the activity of Ret null progenitors in vivo. Activation of RET results in improved haematopoietic stem cell survival, expansion and in vivo transplantation efficiency. Remarkably, human cord-blood progenitor expansion and transplantation is also improved by neurotrophic factors, opening the way for exploration of RET agonists in human haematopoietic stem cell transplantation. Our work shows that neurotrophic factors are novel components of the haematopoietic stem cell microenvironment, revealing that haematopoietic stem cells and neurons are regulated by similar signals.

Maternal retinoids control type 3 innate lymphoid cells and set the offspring immunity.

The impact of nutritional status during fetal life on the overall health of adults has been recognized; however, dietary effects on the developing immune system are largely unknown. Development of secondary lymphoid organs occurs during embryogenesis and is considered to be developmentally programmed. Secondary lymphoid organ formation depends on a subset of type 3 innate lymphoid cells (ILC3) named lymphoid tissue inducer (LTi) cells. Here we show that mouse fetal ILC3s are controlled by cell-autonomous retinoic acid (RA) signalling in utero, which pre-sets the immune fitness in adulthood. We found that embryonic lymphoid organs contain ILC progenitors that differentiate locally into mature LTi cells. Local LTi cell differentiation was controlled by maternal retinoid intake and fetal RA signalling acting in a haematopoietic cell-autonomous manner. RA controlled LTi cell maturation upstream of the transcription factor RORγt. Accordingly, enforced expression of Rorgt restored maturation of LTi cells with impaired RA signalling, whereas RA receptors directly regulated the Rorgt locus. Finally, we established that maternal levels of dietary retinoids control the size of secondary lymphoid organs and the efficiency of immune responses in the adult offspring. Our results reveal a molecular link between maternal nutrients and the formation of immune structures required for resistance to infection in the offspring.

Disordered eating behaviours and correlates in yoga practitioners: a systematic review.

PURPOSE: Yoga has been increasingly used as a complementary therapy for eating disorders. However, it is still not clear whether yoga is effective in the prevention and treatment of eating disorders, as some studies suggest that yoga practitioners show elevated levels of disordered eating behaviours. The goal of this systematic review is, thus, to analyse the occurrence of disordered eating behaviours and correlates in yoga practitioners. METHOD: PRISMA guidelines for systematic reviews were used. Search was conducted in several databases and specific journals. RESULTS: Twelve articles, all cross-sectional, were identified, following PRISMA guidelines. Results across studies were inconsistent. Yoga practice was usually associated with healthier eating behaviours, lower disordered eating symptoms, and higher positive body image and body satisfaction, suggesting that yoga practitioners may be at a lower risk of developing eating disorders. However, other studies suggested that a high dosage of yoga practice may be associated with a higher prevalence of disordered eating behaviours. CONCLUSIONS: As yoga is increasingly used as therapy for eating disorders, understanding the relationship between yoga dosage and disordered eating behaviours is critical to guide treatment recommendations and establish yoga as a valuable complementary therapy. LEVEL OF EVIDENCE: Level I, systematic review.

CD5 expression is regulated during human T-cell activation by alternative polyadenylation, PTBP1, and miR-204.

T lymphocytes stimulated through their antigen receptor (TCR) preferentially express mRNA isoforms with shorter 3´ untranslated regions (3´-UTRs) derived from alternative pre-mRNA cleavage and polyadenylation (APA). However, the physiological relevance of APA programs remains poorly understood. CD5 is a T-cell surface glycoprotein that negatively regulates TCR signaling from the onset of T-cell activation. CD5 plays a pivotal role in mediating outcomes of cell survival or apoptosis, and may prevent both autoimmunity and cancer. In human primary T lymphocytes and Jurkat cells we found three distinct mRNA isoforms encoding CD5, each derived from distinct poly(A) signals (PASs). Upon T-cell activation, there is an overall increase in CD5 mRNAs with a specific increase in the relative expression of the shorter isoforms. 3´-UTRs derived from these shorter isoforms confer higher reporter expression in activated T cells relative to the longer isoform. We further show that polypyrimidine tract binding protein (PTB/PTBP1) directly binds to the proximal PAS and PTB siRNA depletion causes a decrease in mRNA derived from this PAS, suggesting an effect on stability or poly(A) site selection to circumvent targeting of the longer CD5 mRNA isoform by miR-204. These mechanisms fine-tune CD5 expression levels and thus ultimately T-cell responses.

RNA polymerase II kinetics in polo polyadenylation signal selection.

Regulated alternative polyadenylation is an important feature of gene expression, but how gene transcription rate affects this process remains to be investigated. polo is a cell-cycle gene that uses two poly(A) signals in the 3' untranslated region (UTR) to produce alternative messenger RNAs that differ in their 3'UTR length. Using a mutant Drosophila strain that has a lower transcriptional elongation rate, we show that transcription kinetics can determine alternative poly(A) site selection. The physiological consequences of incorrect polo poly(A) site choice are of vital importance; transgenic flies lacking the distal poly(A) signal cannot produce the longer transcript and die at the pupa stage due to a failure in the proliferation of the precursor cells of the abdomen, the histoblasts. This is due to the low translation efficiency of the shorter transcript produced by proximal poly(A) site usage. Our results show that correct polo poly(A) site selection functions to provide the correct levels of protein expression necessary for histoblast proliferation, and that the kinetics of RNA polymerase II have an important role in the mechanism of alternative polyadenylation.

SDHB/SDHA immunohistochemistry in pheochromocytomas and paragangliomas: a multicenter interobserver variation analysis using virtual microscopy: a Multinational Study of the European Network for the Study of Adrenal Tumors (ENS@T).

Despite the established role of SDHB/SDHA immunohistochemistry as a valuable tool to identify patients at risk for familial succinate dehydrogenase-related pheochromocytoma/paraganglioma syndromes, the reproducibility of the assessment methods has not as yet been determined. The aim of this study was to investigate interobserver variability among seven expert endocrine pathologists using a web-based virtual microscopy approach in a large multicenter pheochromocytoma/paraganglioma cohort (n=351): (1) 73 SDH mutated, (2) 105 non-SDH mutated, (3) 128 samples without identified SDH-x mutations, and (4) 45 with incomplete SDH molecular genetic analysis. Substantial agreement among all the reviewers was observed either with a two-tiered classification (SDHB κ=0.7338; SDHA κ=0.6707) or a three-tiered classification approach (SDHB κ=0.6543; SDHA κ=0.7516). Consensus was achieved in 315 cases (89.74%) for SDHB immunohistochemistry and in 348 cases (99.15%) for SDHA immunohistochemistry. Among the concordant cases, 62 of 69 (~90%) SDHB-/C-/D-/AF2-mutated cases displayed SDHB immunonegativity and SDHA immunopositivity, 3 of 4 (75%) with SDHA mutations showed loss of SDHA/SDHB protein expression, whereas 98 of 105 (93%) non-SDH-x-mutated counterparts demonstrated retention of SDHA/SDHB protein expression. Two SDHD-mutated extra-adrenal paragangliomas were scored as SDHB immunopositive, whereas 9 of 128 (7%) tumors without identified SDH-x mutations, 6 of 37 (~16%) VHL-mutated, as well as 1 of 21 (~5%) NF1-mutated tumors were evaluated as SDHB immunonegative. Although 14 out of those 16 SDHB-immunonegative cases were nonmetastatic, an overall significant correlation between SDHB immunonegativity and malignancy was observed (P=0.00019). We conclude that SDHB/SDHA immunohistochemistry is a reliable tool to identify patients with SDH-x mutations with an additional value in the assessment of genetic variants of unknown significance. If SDH molecular genetic analysis fails to detect a mutation in SDHB-immunonegative tumor, SDHC promoter methylation and/or VHL/NF1 testing with the use of targeted next-generation sequencing is advisable.

Evaluation of the responsiveness pattern to caffeine through a smart data-driven ECG non-linear multi-band analysis.

This study aimed to explore more efficient ways of administering caffeine to the body by investigating the impact of caffeine on the modulation of the nervous system's activity through the analysis of electrocardiographic signals (ECG). An ECG non-linear multi-band analysis using Discrete Wavelet Transform (DWT) was employed to extract various features from healthy individuals exposed to different caffeine consumption methods: expresso coffee (EC), decaffeinated coffee (ED), Caffeine Oral Films (OF_caffeine), and placebo OF (OF_placebo). Non-linear feature distributions representing every ECG minute time series have been selected by PCA with different variance percentages to serve as inputs for 23 machine learning models in a leave-one-out cross-validation process for analyzing the behavior differences between ED/EC and OF_placebo/OF_caffeine groups, respectively, over time. The study generated 50-point accuracy curves per model, representing the discrimination power between groups throughout the 50 min. The best model accuracies for ED/EC varied between 30 and 70 %, (using the decision tree classifier) and OF_placebo/OF_caffeine ranged from 62 to 84 % (using Fine Gaussian). Notably, caffeine delivery through OFs demonstrated effective capacity compared to its placebo counterpart, as evidenced by significant differences in accuracy curves between OF_placebo/OF_caffeine. Caffeine delivery via OFs also exhibited rapid dissolution efficiency and controlled release rate over time, distinguishing it from EC. The study supports the potential of caffeine delivery through Caffeine OFs as a superior technology compared to traditional methods by means of ECG analysis. It highlights the efficiency of OFs in controlling the release of caffeine and underscores their promise for future caffeine delivery systems.

Genomic Analysis of Two Novel Bacteriophages Infecting Acinetobacter beijerinckii and halotolerans Species.

Bacteriophages are the most diverse genetic entities on Earth. In this study, two novel bacteriophages, nACB1 (Podoviridae morphotype) and nACB2 (Myoviridae morphotype), which infect Acinetobacter beijerinckii and Acinetobacter halotolerans, respectively, were isolated from sewage samples. The genome sequences of nACB1 and nACB2 revealed that their genome sizes were 80,310 bp and 136,560 bp, respectively. Comparative analysis showed that both genomes are novel members of the Schitoviridae and the Ackermannviridae families, sharing ≤ 40% overall nucleotide identities with any other phages. Interestingly, among other genetic features, nACB1 encoded a very large RNA polymerase, while nACB2 displayed three putative depolymerases (two capsular depolymerases and one capsular esterase) encoded in tandem. This is the first report of phages infecting A. halotolerans and beijerinckii human pathogenic species. The findings regarding these two phages will allow us to further explore phage-Acinetobacter interactions and the genetic evolution for this group of phages.

Short-Term Effects of Climate Change on Planktonic Heterotrophic Prokaryotes in a Temperate Coastal Lagoon: Temperature Is Good, Ultraviolet Radiation Is Bad, and CO2 Is Neutral.

Planktonic heterotrophic prokaryotes (HProks) are a pivotal functional group in marine ecosystems and are highly sensitive to environmental variability and climate change. This study aimed to investigate the short-term effects of increasing carbon dioxide (CO2), ultraviolet radiation (UVR), and temperature on natural assemblages of HProks in the Ria Formosa coastal lagoon during winter. Two multi-stressor microcosm experiments were used to evaluate the isolated and combined effects of these environmental changes on HProk abundance, production, growth, and mortality rates. The isolated and combined effects of increased CO2 on HProks were not significant. However, HProk production, cellular activity, instantaneous growth rate, and mortality rate were negatively influenced by elevated UVR and positively influenced by warming. Stronger effects were detected on HProk mortality in relation to specific growth rate, leading to higher HProk net growth rates and abundance under elevated UVR and lower values under warming conditions.

Maternal γδ T cells shape offspring pulmonary type 2 immunity in a microbiota-dependent manner.

Immune development is profoundly influenced by vertically transferred cues. However, little is known about how maternal innate-like lymphocytes regulate offspring immunity. Here, we show that mice born from γδ T cell-deficient (TCRδ-/-) dams display an increase in first-breath-induced inflammation, with a pulmonary milieu selectively enriched in type 2 cytokines and type 2-polarized immune cells, when compared with the progeny of γδ T cell-sufficient dams. Upon helminth infection, mice born from TCRδ-/- dams sustain an increased type 2 inflammatory response. This is independent of the genotype of the pups. Instead, the offspring of TCRδ-/- dams harbors a distinct intestinal microbiota, acquired during birth and fostering, and decreased levels of intestinal short-chain fatty acids (SCFAs), such as pentanoate and hexanoate. Importantly, exogenous SCFA supplementation inhibits type 2 innate lymphoid cell function and suppresses first-breath- and infection-induced inflammation. Taken together, our findings unravel a maternal γδ T cell-microbiota-SCFA axis regulating neonatal lung immunity.

Identification of the first germline HRPT2 whole-gene deletion in a patient with primary hyperparathyroidism.

OBJECTIVE: Germline mutations in the HRPT2 gene are associated with the hereditary hyperparathyroidism-jaw tumour syndrome (HPT-JT) and a subset of familial isolated hyperparathyroidism (FIHP). Somatic HRPT2 mutations are detected in sporadic parathyroid carcinomas and less frequently in cystic adenomas. The purpose of this study was to investigate the underlying HRPT2 defect in a young patient with symptomatic hyperparathyroidism due to an apparently sporadic parathyroid adenoma with cystic features. DESIGN AND METHODS: HRPT2 mutations in the patient's genomic and parathyroid tumour DNA were screened by PCR-based sequencing. Tumour loss of heterozygosity (LOH) at the HRPT2 locus was assessed with microsatellite markers. A large germline HRPT2 deletion was investigated by real-time quantitative PCR analysis (qPCR). Genomic DNA losses were also appraised by chromosomal comparative genomic hybridization (cCGH). RESULTS: No germline HRPT2 point mutation was detected by direct sequencing. A novel hemizygous HRPT2 somatic mutation (c.32delA) was identified in the tumour. Apparent constitutional homozygosity for HRPT2 flanking microsatellite markers, and absence of LOH at a distal marker, suggested a large germline deletion. Gene dose mapping by qPCR unveiled a de novo deletion of the whole HRPT2 gene and adjacent loci (<9·3 Mb in size). cCGH confirmed germline DNA loss involving the HRPT2 locus. CONCLUSIONS: We report the first large germline deletion of the HRPT2 gene, which was not detectable by conventional PCR-based sequencing methods. This finding emphasizes that qPCR should be implemented in HRPT2 molecular analysis, which may improve genetic assessment and clinical management of patients with FIHP and HPT-JT.