Erratum: Gut microbial metabolites limit the frequency of autoimmune T cells and protect against type 1 diabetes.

This corrects the article DOI: 10.1038/ni.3713.

Gut microbial metabolites limit the frequency of autoimmune T cells and protect against type 1 diabetes.

Gut dysbiosis might underlie the pathogenesis of type 1 diabetes. In mice of the non-obese diabetic (NOD) strain, we found that key features of disease correlated inversely with blood and fecal concentrations of the microbial metabolites acetate and butyrate. We therefore fed NOD mice specialized diets designed to release large amounts of acetate or butyrate after bacterial fermentation in the colon. Each diet provided a high degree of protection from diabetes, even when administered after breakdown of immunotolerance. Feeding mice a combined acetate- and butyrate-yielding diet provided complete protection, which suggested that acetate and butyrate might operate through distinct mechanisms. Acetate markedly decreased the frequency of autoreactive T cells in lymphoid tissues, through effects on B cells and their ability to expand populations of autoreactive T cells. A diet containing butyrate boosted the number and function of regulatory T cells, whereas acetate- and butyrate-yielding diets enhanced gut integrity and decreased serum concentration of diabetogenic cytokines such as IL-21. Medicinal foods or metabolites might represent an effective and natural approach for countering the numerous immunological defects that contribute to T cell-dependent autoimmune diseases.

Shot noise-mitigated secondary electron imaging with ion count-aided microscopy.

Modern science is dependent on imaging on the nanoscale, often achieved through processes that detect secondary electrons created by a highly focused incident charged particle beam. Multiple types of measurement noise limit the ultimate trade-off between the image quality and the incident particle dose, which can preclude useful imaging of dose-sensitive samples. Existing methods to improve image quality do not fundamentally mitigate the noise sources. Furthermore, barriers to assigning a physically meaningful scale make the images qualitative. Here, we introduce ion count-aided microscopy (ICAM), which is a quantitative imaging technique that uses statistically principled estimation of the secondary electron yield. With a readily implemented change in data collection, ICAM substantially reduces source shot noise. In helium ion microscopy, we demonstrate 3[Formula: see text] dose reduction and a good match between these empirical results and theoretical performance predictions. ICAM facilitates imaging of fragile samples and may make imaging with heavier particles more attractive.

Baricitinib and ß-Cell Function in Patients with New-Onset Type 1 Diabetes.

BACKGROUND: Janus kinase (JAK) inhibitors, including baricitinib, block cytokine signaling and are effective disease-modifying treatments for several autoimmune diseases. Whether baricitinib preserves ß-cell function in type 1 diabetes is unclear. METHODS: In this phase 2, double-blind, randomized, placebo-controlled trial, we assigned patients with type 1 diabetes diagnosed during the previous 100 days to receive baricitinib (4 mg once per day) or matched placebo orally for 48 weeks. The primary outcome was the mean C-peptide level, determined from the area under the concentration-time curve, during a 2-hour mixed-meal tolerance test at week 48. Secondary outcomes included the change from baseline in the glycated hemoglobin level, the daily insulin dose, and measures of glycemic control assessed with the use of continuous glucose monitoring. RESULTS: A total of 91 patients received baricitinib (60 patients) or placebo (31 patients). The median of the mixed-meal-stimulated mean C-peptide level at week 48 was 0.65 nmol per liter per minute (interquartile range, 0.31 to 0.82) in the baricitinib group and 0.43 nmol per liter per minute (interquartile range, 0.13 to 0.63) in the placebo group (P = 0.001). The mean daily insulin dose at 48 weeks was 0.41 U per kilogram of body weight per day (95% confidence interval [CI], 0.35 to 0.48) in the baricitinib group and 0.52 U per kilogram per day (95% CI, 0.44 to 0.60) in the placebo group. The levels of glycated hemoglobin were similar in the two trial groups. However, the mean coefficient of variation of the glucose level at 48 weeks, as measured by continuous glucose monitoring, was 29.6% (95% CI, 27.8 to 31.3) in the baricitinib group and 33.8% (95% CI, 31.5 to 36.2) in the placebo group. The frequency and severity of adverse events were similar in the two trial groups, and no serious adverse events were attributed to baricitinib or placebo. CONCLUSIONS: In patients with type 1 diabetes of recent onset, daily treatment with baricitinib over 48 weeks appeared to preserve ß-cell function as estimated by the mixed-meal-stimulated mean C-peptide level. (Funded by JDRF International and others; BANDIT Australian New Zealand Clinical Trials Registry number, ACTRN12620000239965.).

T cell regulation mediated by interaction of soluble CD52 with the inhibitory receptor Siglec-10.

Functionally diverse T cell populations interact to maintain homeostasis of the immune system. We found that human and mouse antigen-activated T cells with high expression of the lymphocyte surface marker CD52 suppressed other T cells. CD52(hi)CD4(+) T cells were distinct from CD4(+)CD25(+)Foxp3(+) regulatory T cells. Their suppression was mediated by soluble CD52 released by phospholipase C. Soluble CD52 bound to the inhibitory receptor Siglec-10 and impaired phosphorylation of the T cell receptor-associated kinases Lck and Zap70 and T cell activation. Humans with type 1 diabetes had a lower frequency and diminished function of CD52(hi)CD4(+) T cells responsive to the autoantigen GAD65. In diabetes-prone mice of the nonobese diabetic (NOD) strain, transfer of lymphocyte populations depleted of CD52(hi) cells resulted in a substantially accelerated onset of diabetes. Our studies identify a ligand-receptor mechanism of T cell regulation that may protect humans and mice from autoimmune disease.

A tethered ligand assay to probe SARS-CoV-2:ACE2 interactions.

SignificanceIn the dynamic environment of the airways, where SARS-CoV-2 infections are initiated by binding to human host receptor ACE2, mechanical stability of the viral attachment is a crucial fitness advantage. Using single-molecule force spectroscopy techniques, we mimic the effect of coughing and sneezing, thereby testing the force stability of SARS-CoV-2 RBD:ACE2 interaction under physiological conditions. Our results reveal a higher force stability of SARS-CoV-2 binding to ACE2 compared to SARS-CoV-1, causing a possible fitness advantage. Our assay is sensitive to blocking agents preventing RBD:ACE2 bond formation. It will thus provide a powerful approach to investigate the modes of action of neutralizing antibodies and other agents designed to block RBD binding to ACE2 that are currently developed as potential COVID-19 therapeutics.

Quantifying the proportion of different cell types in the human cortex using DNA methylation profiles.

BACKGROUND: Due to interindividual variation in the cellular composition of the human cortex, it is essential that covariates that capture these differences are included in epigenome-wide association studies using bulk tissue. As experimentally derived cell counts are often unavailable, computational solutions have been adopted to estimate the proportion of different cell types using DNA methylation data. Here, we validate and profile the use of an expanded reference DNA methylation dataset incorporating two neuronal and three glial cell subtypes for quantifying the cellular composition of the human cortex. RESULTS: We tested eight reference panels containing different combinations of neuronal- and glial cell types and characterised their performance in deconvoluting cell proportions from computationally reconstructed or empirically derived human cortex DNA methylation data. Our analyses demonstrate that while these novel brain deconvolution models produce accurate estimates of cellular proportions from profiles generated on postnatal human cortex samples, they are not appropriate for the use in prenatal cortex or cerebellum tissue samples. Applying our models to an extensive collection of empirical datasets, we show that glial cells are twice as abundant as neuronal cells in the human cortex and identify significant associations between increased Alzheimer's disease neuropathology and the proportion of specific cell types including a decrease in NeuNNeg/SOX10Neg nuclei and an increase of NeuNNeg/SOX10Pos nuclei. CONCLUSIONS: Our novel deconvolution models produce accurate estimates for cell proportions in the human cortex. These models are available as a resource to the community enabling the control of cellular heterogeneity in epigenetic studies of brain disorders performed on bulk cortex tissue.

Dietary patterns during pregnancy and maternal and birth outcomes in women with type 1 diabetes: the Environmental Determinants of Islet Autoimmunity (ENDIA) study.

AIMS/HYPOTHESIS: Dietary patterns characterised by high intakes of vegetables may lower the risk of pre-eclampsia and premature birth in the general population. The effect of dietary patterns in women with type 1 diabetes, who have an increased risk of complications in pregnancy, is not known. The aim of this study was to investigate the relationship between dietary patterns and physical activity during pregnancy and maternal complications and birth outcomes in women with type 1 diabetes. We also compared dietary patterns in women with and without type 1 diabetes. METHODS: Diet was assessed in the third trimester using a validated food frequency questionnaire in participants followed prospectively in the multi-centre Environmental Determinants of Islet Autoimmunity (ENDIA) study. Dietary patterns were characterised by principal component analysis. The Pregnancy Physical Activity Questionnaire was completed in each trimester. Data for maternal and birth outcomes were collected prospectively. RESULTS: Questionnaires were completed by 973 participants during 1124 pregnancies. Women with type 1 diabetes (n=615 pregnancies with dietary data) were more likely to have a 'fresh food' dietary pattern than women without type 1 diabetes (OR 1.19, 95% CI 1.07, 1.31; p=0.001). In women with type 1 diabetes, an increase equivalent to a change from quartile 1 to 3 in 'fresh food' dietary pattern score was associated with a lower risk of pre-eclampsia (OR 0.37, 95% CI 0.17, 0.78; p=0.01) and premature birth (OR 0.35, 95% CI 0.20, 0.62, p<0.001). These associations were mediated in part by BMI and HbA1c. The 'processed food' dietary pattern was associated with an increased birthweight (ß coefficient 56.8 g, 95% CI 2.8, 110.8; p=0.04). Physical activity did not relate to outcomes. CONCLUSIONS/INTERPRETATION: A dietary pattern higher in fresh foods during pregnancy was associated with sizeable reductions in risk of pre-eclampsia and premature birth in women with type 1 diabetes.

Evaluation of nanopore sequencing for epigenetic epidemiology: a comparison with DNA methylation microarrays.

Most epigenetic epidemiology to date has utilized microarrays to identify positions in the genome where variation in DNA methylation is associated with environmental exposures or disease. However, these profile less than 3% of DNA methylation sites in the human genome, potentially missing affected loci and preventing the discovery of disrupted biological pathways. Third generation sequencing technologies, including Nanopore sequencing, have the potential to revolutionize the generation of epigenetic data, not only by providing genuine genome-wide coverage but profiling epigenetic modifications direct from native DNA. Here we assess the viability of using Nanopore sequencing for epidemiology by performing a comparison with DNA methylation quantified using the most comprehensive microarray available, the Illumina EPIC array. We implemented a CRISPR-Cas9 targeted sequencing approach in concert with Nanopore sequencing to profile DNA methylation in three genomic regions to attempt to rediscover genomic positions that existing technologies have shown are differentially methylated in tobacco smokers. Using Nanopore sequencing reads, DNA methylation was quantified at 1779 CpGs across three regions, providing a finer resolution of DNA methylation patterns compared to the EPIC array. The correlation of estimated levels of DNA methylation between platforms was high. Furthermore, we identified 12 CpGs where hypomethylation was significantly associated with smoking status, including 10 within the AHRR gene. In summary, Nanopore sequencing is a valid option for identifying genomic loci where large differences in DNAm are associated with a phenotype and has the potential to advance our understanding of the role differential methylation plays in the etiology of complex disease.

The quality of care delivered to residents in long-term care in Australia: an indicator-based review of resident records (CareTrack Aged study).

BACKGROUND: This study estimated the prevalence of evidence-based care received by a population-based sample of Australian residents in long-term care (LTC) aged ≥ 65 years in 2021, measured by adherence to clinical practice guideline (CPG) recommendations. METHODS: Sixteen conditions/processes of care amendable to estimating evidence-based care at a population level were identified from prevalence data and CPGs. Candidate recommendations (n = 5609) were extracted from 139 CPGs which were converted to indicators. National experts in each condition rated the indicators via the RAND-UCLA Delphi process. For the 16 conditions, 236 evidence-based care indicators were ratified. A multi-stage sampling of LTC facilities and residents was undertaken. Trained aged-care nurses then undertook manual structured record reviews of care delivered between 1 March and 31 May 2021 (our record review period) to assess adherence with the indicators. RESULTS: Care received by 294 residents with 27,585 care encounters in 25 LTC facilities was evaluated. Residents received care for one to thirteen separate clinical conditions/processes of care (median = 10, mean = 9.7). Adherence to evidence-based care indicators was estimated at 53.2% (95% CI: 48.6, 57.7) ranging from a high of 81.3% (95% CI: 75.6, 86.3) for Bladder and Bowel to a low of 12.2% (95% CI: 1.6, 36.8) for Depression. Six conditions (skin integrity, end-of-life care, infection, sleep, medication, and depression) had less than 50% adherence with indicators. CONCLUSIONS: This is the first study of adherence to evidence-based care for people in LTC using multiple conditions and a standardised method. Vulnerable older people are not receiving evidence-based care for many physical problems, nor care to support their mental health nor for end-of-life care. The six conditions in which adherence with indicators was less than 50% could be the focus of improvement efforts.

Investigating the missing-wedge problem in small-angle X-ray scattering tensor tomography across real and reciprocal space.

Small-angle-scattering tensor tomography is a technique for studying anisotropic nanostructures of millimetre-sized samples in a volume-resolved manner. It requires the acquisition of data through repeated tomographic rotations about an axis which is subjected to a series of tilts. The tilt that can be achieved with a typical setup is geometrically constrained, which leads to limits in the set of directions from which the different parts of the reciprocal space map can be probed. Here, we characterize the impact of this limitation on reconstructions in terms of the missing wedge problem of tomography, by treating the problem of tensor tomography as the reconstruction of a three-dimensional field of functions on the unit sphere, represented by a grid of Gaussian radial basis functions. We then devise an acquisition scheme to obtain complete data by remounting the sample, which we apply to a sample of human trabecular bone. Performing tensor tomographic reconstructions of limited data sets as well as the complete data set, we further investigate and validate the missing wedge problem by investigating reconstruction errors due to data incompleteness across both real and reciprocal space. Finally, we carry out an analysis of orientations and derived scalar quantities, to quantify the impact of this missing wedge problem on a typical tensor tomographic analysis. We conclude that the effects of data incompleteness are consistent with the predicted impact of the missing wedge problem, and that the impact on tensor tomographic analysis is appreciable but limited, especially if precautions are taken. In particular, there is only limited impact on the means and relative anisotropies of the reconstructed reciprocal space maps.

Characteristics of large granular lymphocyte leukemia associated with variable common immunodeficiency disorders: A study of 12 cases.

OBJECTIVES: Common Variable Immunodeficiency Disorders (CVID) and Large Granular Lymphocytes leukemia (LGLL) exhibit diverse clinical manifestations including infections, dysimmunity, and lymphoproliferation. Recent decades have seen the discovery of new genes in the lymphopoiesis pathway, such as JAK-STAT. This case series supplemented by a literature review aims to describe clinical and biological characteristics of patients with both CIVD and LGLL. METHODOLOGY: Patients were included through a call for comments to French and Belgian centers and through a literature review via PubMed. Clinical characteristics were compared to two large French cohort involving CVID and LGLL patients. RESULTS: Twelve patients were included. In all cases, CVID precedes LLGL (median diagnosis delay for LLGL was 7 years). Most cases presented with splenomegaly and autoimmune cytopenia. Ten out of 12 patients underwent splenectomy during follow up. CONCLUSIONS: Patients with LGLL and CVID differ from patients without immune deficiency in term of clinical presentation and prognosis. We suggest CVID may act as a trigger of LGL lymphocytosis, due to endogenous and exogenous antigenic pressure leading to the selection of a dominant LGL clone and stimulation of the JAK-STAT pathway. The role of splenomegaly and splenectomy in LGLL onset warrant further investigation in future studies.

The virtual dermatopathology mentorship.

The Covid pandemic necessitated changes in traditional mentorship and educational roles. We offer our experience in structuring a virtual dermatopathology mentorship in the midst of the pandemic. Virtual mentorships have many benefits including ease of access, lack of geographic restrictions, and financial savings, though there still remains an important and irreplaceable role for in-person relationships.

Laser welding of fiber array units.

We report the results of fabricating fiber array unit (FAU) connectors using a near IR laser welding process, locking fibers in proper position on planar glass substrates and forming strong glass-to-glass bonds, followed by final assembly using lower coefficient of thermal expansion (CTE) epoxies. A thin metal film deposited on the glass substrate provides the absorption required to attain interfacial temperatures suitable for glass-to-glass bonding. This method allows the elimination of dedicated expensive V-groove plates while still maintaining very good fiber placement accuracy. The use of epoxy is minimized to simply securing macro packaging components and protecting fibers from environmental pressure, temperature, and humidity variation. The thermal expansion properties of the epoxy used were essential for the long-term FAU reliability.

Patient outcome quality indicators for older persons in acute care: original development data using interRAI AC-CGA.

BACKGROUND: A range of strategies are available that can improve the outcomes of older persons particularly in relation to basic activities of daily living during and after an acute care (AC) episode. This paper outlines the original development of outcome-oriented quality indicators (QIs) in relation to common geriatric syndromes and function for the care of the frail aged hospitalized in acute general medical wards. METHODS: Design QIs were developed using evidence from literature, expert opinion, field study data and a formal voting process. A systematic literature review of literature identified existing QIs (there were no outcome QIs) and evidence of interventions that improve older persons' outcomes in AC. Preliminary indicators were developed by two expert panels following consideration of the evidence. After analysis of the data from field testing (indicator prevalence, variability across sites), panel meetings refined the QIs prior to a formal voting process. SETTING: Data was collected in nine Australian general medical wards. PARTICIPANTS: Patients aged 70 years and over, consented within 24 h of admission to the AC ward. MEASUREMENTS: The interRAI Acute Care - Comprehensive Geriatric Assessment (interRAI AC-CGA) was administered at admission and discharge; a daily risk assessment in hospital; 28-day phone follow-up and chart audit. RESULTS: Ten outcome QIs were established which focused on common geriatric syndromes and function for the care of the frail aged hospitalized in acute general medical wards. CONCLUSION: Ten outcome QIs were developed. These QIs can be used to identify areas where specific action will lead to improvements in the quality of care delivered to older persons in hospital.

Sialoglycan recognition is a common connection linking acidosis, zinc, and HMGB1 in sepsis.

Blood pH is tightly maintained between 7.35 and 7.45, and acidosis (pH <7.3) indicates poor prognosis in sepsis, wherein lactic acid from anoxic tissues overwhelms the buffering capacity of blood. Poor sepsis prognosis is also associated with low zinc levels and the release of High mobility group box 1 (HMGB1) from activated and/or necrotic cells. HMGB1 added to whole blood at physiological pH did not bind leukocyte receptors, but lowering pH with lactic acid to mimic sepsis conditions allowed binding, implying the presence of natural inhibitor(s) preventing binding at normal pH. Testing micromolar concentrations of divalent cations showed that zinc supported the robust binding of sialylated glycoproteins with HMGB1. Further characterizing HMGB1 as a sialic acid-binding lectin, we found that optimal binding takes place at normal blood pH and is markedly reduced when pH is adjusted with lactic acid to levels found in sepsis. Glycan array studies confirmed the binding of HMGB1 to sialylated glycan sequences typically found on plasma glycoproteins, with binding again being dependent on zinc and normal blood pH. Thus, HMGB1-mediated hyperactivation of innate immunity in sepsis requires acidosis, and micromolar zinc concentrations are protective. We suggest that the potent inflammatory effects of HMGB1 are kept in check via sequestration by plasma sialoglycoproteins at physiological pH and triggered when pH and zinc levels fall in late stages of sepsis. Current clinical trials independently studying zinc supplementation, HMGB1 inhibition, or pH normalization may be more successful if these approaches are combined and perhaps supplemented by infusions of heavily sialylated molecules.

Impact of Positive Cultures During the Second Stage of a Two-Stage Exchange: Systematic Review and Meta-Analysis.

BACKGROUND: Two-stage exchange is a frequently performed procedure in patients who have a periprosthetic joint infection. Positive cultures when performing the second stage are perceived as a risk factor for reinfection. This study aimed to determine the impact of positive cultures during the second stage on the outcome of patients undergoing a 2-stage septic exchange and the impact of stopping the antibiotic treatment before reimplantation. METHODS: We systematically searched four databases. We performed a meta-analysis on the risk of complications after positive cultures during second stage and a subgroup analysis by antibiotic holiday period. We included 24 studies. RESULTS: Failure in the positive group was 37.0% and in the negative group was 13.7% with an odds ratio (OR) of 4.05. In the subgroup analysis by antibiotic holidays, failure rate without holidays was 15% and with holidays was 17.3% (P = .21). Failure in each group was higher when cultures were positive (without holidays, 25 versus 12.2%, P = .0003, and with holidays 41.1 versus 12.7%, P < .0001), but the risk of failure when cultures were positive was higher in the holiday group (OR 4.798) than in the nonholiday group (OR 2.225) in comparison to those patients who were culture negative at the second stage. CONCLUSIONS: Microbiological eradication at second stage was not obtained in 18% of cases and it was associated with a higher failure rate. In patients with positive cultures, withholding antibiotic treatment was associated with lower failure rate. Further studies to define the antibiotic strategy in 2-stage exchange procedure are necessary.

Gut microbiota maturity mediates the protective effect of siblings on food allergy.

BACKGROUND: The mechanisms underlying the protective effect of older siblings on allergic disease remain unclear but may relate to the infant gut microbiota. OBJECTIVE: We sought to investigate whether having older siblings decreases the risk of IgE-mediated food allergy by accelerating the maturation of the infant gut microbiota. METHODS: In a birth cohort assembled using an unselected antenatal sampling frame (n = 1074), fecal samples were collected at 1 month, 6 months, and 1 year, and food allergy status at 1 year was determined by skin prick test and in-hospital food challenge. We used 16S rRNA gene amplicon sequencing to derive amplicon sequence variants. Among a random subcohort (n = 323), microbiota-by-age z scores at each time point were calculated using fecal amplicon sequence variants to represent the gut microbiota maturation over the first year of life. RESULTS: A greater number of siblings was associated with a higher microbiota-by-age z score at age 1 year (ß  = 0.15 per an additional sibling; 95% CI, 0.05-0.24; P = .003), which was in turn associated with decreased odds of food allergy (odds ratio, 0.45; 95% CI, 0.33-0.61; P < .001). Microbiota-by-age z scores mediated 63% of the protective effect of siblings. Analogous associations were not observed at younger ages. CONCLUSIONS: The protective effect of older siblings on the risk of developing IgE-mediated food allergy during infancy is substantially mediated by advanced maturation of the gut microbiota at age 1 year.

Key aspects of soft tissue management in fracture-related infection: recommendations from an international expert group.

A judicious, well-planned bone and soft tissue debridement remains one of the cornerstones of state-of-the-art treatment of fracture-related infection (FRI). Meticulous surgical excision of all non-viable tissue can, however, lead to the creation of large soft tissue defects. The management of these defects is complex and numerous factors need to be considered when selecting the most appropriate approach. This narrative review summarizes the current evidence with respect to soft tissue management in patients diagnosed with FRI. Specifically we discuss the optimal timing for tissue closure following debridement in cases of FRI, the need for negative microbiological culture results from the surgical site as a prerequisite for definitive wound closure, the optimal type of flap in case of large soft tissue defects caused by FRI and the role of negative pressure wound therapy (NPWT) in FRI. Finally, recommendations are made with regard to soft tissue management in FRI that should be useful for clinicians in daily clinical practice.Level of evidence Level V.