Innate type 2 immunity controls hair follicle commensalism by Demodex mites.

Demodex mites are commensal parasites of hair follicles (HFs). Normally asymptomatic, inflammatory outgrowth of mites can accompany malnutrition, immune dysfunction, and aging, but mechanisms restricting Demodex outgrowth are not defined. Here, we show that control of mite HF colonization in mice required group 2 innate lymphoid cells (ILC2s), interleukin-13 (IL-13), and its receptor, IL-4Ra-IL-13Ra1. HF-associated ILC2s elaborated IL-13 that attenuated HFs and epithelial proliferation at anagen onset; in their absence, Demodex colonization led to increased epithelial proliferation and replacement of gene programs for repair by aberrant inflammation, leading to the loss of barrier function and HF exhaustion. Humans with rhinophymatous acne rosacea, an inflammatory condition associated with Demodex, had increased HF inflammation with decreased type 2 cytokines, consistent with the inverse relationship seen in mice. Our studies uncover a key role for skin ILC2s and IL-13, which comprise an immune checkpoint that sustains cutaneous integrity and restricts pathologic infestation by colonizing HF mites.

A high-resolution transcriptomic and spatial atlas of cell types in the whole mouse brain.

The mammalian brain consists of millions to billions of cells that are organized into many cell types with specific spatial distribution patterns and structural and functional properties1-3. Here we report a comprehensive and high-resolution transcriptomic and spatial cell-type atlas for the whole adult mouse brain. The cell-type atlas was created by combining a single-cell RNA-sequencing (scRNA-seq) dataset of around 7 million cells profiled (approximately 4.0 million cells passing quality control), and a spatial transcriptomic dataset of approximately 4.3 million cells using multiplexed error-robust fluorescence in situ hybridization (MERFISH). The atlas is hierarchically organized into 4 nested levels of classification: 34 classes, 338 subclasses, 1,201 supertypes and 5,322 clusters. We present an online platform, Allen Brain Cell Atlas, to visualize the mouse whole-brain cell-type atlas along with the single-cell RNA-sequencing and MERFISH datasets. We systematically analysed the neuronal and non-neuronal cell types across the brain and identified a high degree of correspondence between transcriptomic identity and spatial specificity for each cell type. The results reveal unique features of cell-type organization in different brain regions-in particular, a dichotomy between the dorsal and ventral parts of the brain. The dorsal part contains relatively fewer yet highly divergent neuronal types, whereas the ventral part contains more numerous neuronal types that are more closely related to each other. Our study also uncovered extraordinary diversity and heterogeneity in neurotransmitter and neuropeptide expression and co-expression patterns in different cell types. Finally, we found that transcription factors are major determinants of cell-type classification and identified a combinatorial transcription factor code that defines cell types across all parts of the brain. The whole mouse brain transcriptomic and spatial cell-type atlas establishes a benchmark reference atlas and a foundational resource for integrative investigations of cellular and circuit function, development and evolution of the mammalian brain.

The microbiome stabilizes circadian rhythms in the gut.

The gut microbiome is well known to impact host physiology and health. Given widespread control of physiology by circadian clocks, we asked how the microbiome interacts with circadian rhythms in the Drosophila gut. The microbiome did not cycle in flies fed ad libitum, and timed feeding (TF) drove limited cycling only in clockless per01 flies. However, TF and loss of the microbiome influenced the composition of the gut cycling transcriptome, independently and together. Moreover, both interventions increased the amplitude of rhythmic gene expression, with effects of TF at least partly due to changes in histone acetylation. Contrary to expectations, timed feeding rendered animals more sensitive to stress. Analysis of microbiome function in circadian physiology revealed that germ-free flies reset more rapidly with shifts in the light:dark cycle. We propose that the microbiome stabilizes cycling in the host gut to prevent rapid fluctuations with changing environmental conditions.

Adequate bone healing after supplementary fixation of periprosthetic total knee arthroplasty fractures using Luque cerclage wiring: a retrospective case series.

PURPOSE: Cerclage wiring is a well-known supplemental fixation technique that can be used in many types of fractures. With the tendency toward minimally invasive approaches in the management of periprosthetic total knee arthroplasty (TKA) fractures, and with absence of a dedicated study that reports the results of cerclage wiring in TKA fractures in particular, the aim of this retrospective study is to report the outcomes of supplementary cerclage wiring using simple Luque wires in fractures of distal femur associated with TKA. METHOD: Eighteen cases, with a mean age of 77.2 years had complete follow-up data and had their radiographs and clinical data assessed for this study. Patients received cerclage wiring along with plates, retrograde nailing or around cracked femoral shaft overlying revision TKA femoral stem during the surgical management of periprosthetic TKA distal femur fractures. RESULTS: Fracture healing with adequate callus formation occurred in all 18 cases at a mean of 11.4 weeks postoperatively. None of the cases had any vascular injury, and after a mean clinical follow-up of 51 weeks, none of the cases had nonunion or hardware complications. One case had postoperative periprosthetic infection that developed 8 months after full fracture healing and had a two-stage revision using revision stemmed TKA and protective cerclage wiring with successful eradication of infection. CONCLUSION: Supplementary cerclage wiring in distal femur TKA fractures can aid in enhanced bone healing with minimal complications, provided that adequate reduction and rigid fixation were achieved. This study reflects the level of evidence IV.

A Randomized Trial Comparing the Specific Carbohydrate Diet to a Mediterranean Diet in Adults With Crohn's Disease.

BACKGROUND & AIMS: This study compared the effectiveness of the Specific Carbohydrate Diet (SCD) to the Mediterranean diet (MD) as treatment for Crohn's disease (CD) with mild to moderate symptoms. METHODS: Adult patients with CD and with mild-to-moderate symptoms were randomly assigned 1:1 to consume the MD or SCD for 12 weeks. For the first 6 weeks, participants received prepared meals and snacks according to their assigned diet. After 6 weeks, participants were instructed to follow the diet independently. The primary outcome was symptomatic remission at week 6. Key secondary outcomes at week 6 included fecal calprotectin (FC) response (FC <250 µg/g and reduction by >50% among those with baseline FC >250 µg/g) and C-reactive protein (CRP) response (high-sensitivity CRP <5 mg/L and >50% reduction from baseline among those with high-sensitivity CRP >5 mg/L). RESULTS: The study randomized 194 patients, and 191 were included in the efficacy analyses. The percentage of participants who achieved symptomatic remission at week 6 was not superior with the SCD (SCD, 46.5%; MD, 43.5%; P = .77). FC response was achieved in 8 of 23 participants (34.8%) with the SCD and in 4 of 13 participants (30.8%) with the MD (P = .83). CRP response was achieved in 2 of 37 participants (5.4%) with the SCD and in 1 of 28 participants (3.6%) with the MD (P = .68). CONCLUSIONS: The SCD was not superior to the MD to achieve symptomatic remission, FC response, and CRP response. CRP response was uncommon. Given these results, the greater ease of following the MD and other health benefits associated with the MD, the MD may be preferred to the SCD for most patients with CD with mild to moderate symptoms. ClinicalTrials.gov Identifier: NCT03058679.

Outcomes and Predictors of Sustained Remission After Drug Withdrawal in Pediatric Crohn Disease.

OBJECTIVES: The primary aim of this study was to determine the proportion of pediatric Crohn disease (CD) subjects in sustained drug-free remission 52 weeks after stopping pharmacological therapy. We also aimed to explore the effects of the Crohn Disease Exclusion Diet (CDED) and microbiome composition on remission. METHODS: We performed a prospective study following 18 CD patients ages 13-21 years in deep clinical remission withdrawing from immunomodulator (n = 7) or anti-TNFα (n = 11) monotherapy at two tertiary care centers. Stool for calprotectin and microbiome analyses was collected over 52 weeks. Participants followed either the CDED or free diet after drug withdrawal. The primary endpoint was sustained relapse-free drug-free remission (calprotectin <250 µg/g) at 52 weeks. RESULTS: Seventeen participants were followed through 52 weeks with 11 (64.7%) in sustained remission. There was no improvement in remission among participants following the CDED (5/9; 55.6%), P = 0.63. By 104 weeks, only 8 (47.1 %) participants remained off immunosuppressive therapies. Analysis of shotgun metagenomic sequence data revealed that taxonomic and gene function abundance in the gut microbiome was relatively stable for participants in remission and relapse. However, a predictive model incorporating gut microbial gene pathway abundance for amino sugar/nucleotide sugar metabolism and galactose metabolism from baseline samples predicted relapse at 52 weeks with 80% accuracy. CONCLUSIONS: After withdrawal of immunomodulator or anti-TNFα monotherapy among a small cohort of pediatric CD subjects in deep remission, nearly 65% sustained remission at 52 weeks. Baseline microbiome alterations predicted relapse. Large prospective studies are needed to better understand outcomes after treatment de-escalation.

Persistence of the Probiotic Lacticaseibacillus rhamnosus Strain GG (LGG) in an In Vitro Model of the Gut Microbiome.

The consumption of probiotics is widely encouraged due to reports of their positive effects on human health. In particular, Lacticaseibacillus rhamnosus strain GG (LGG) is an approved probiotic that has been reported to improve health outcomes, especially for gastrointestinal disorders. However, how LGG cooperates with the gut microbiome has not been fully explored. To understand the interaction between LGG and its ability to survive and grow within the gut microbiome, this study introduced LGG into established microbial communities using an in vitro model of the colon. LGG was inoculated into the simulated ascending colon and its persistence in, and transit through the subsequent transverse and descending colon regions was monitored over two weeks. The impact of LGG on the existing bacterial communities was investigated using 16S rRNA sequencing and short-chain fatty acid analysis. LGG was able to engraft and proliferate in the ascending region for at least 10 days but was diminished in the transverse and descending colon regions with little effect on short-chain fatty acid abundance. These data suggest that the health benefits of the probiotic LGG rely on its ability to transiently engraft and modulate the host microbial community.

Diversification of Unprotected Alicyclic Amines by C-H Bond Functionalization: Decarboxylative Alkylation of Transient Imines.

Despite extensive efforts by many practitioners in the field, methods for the direct α-C-H bond functionalization of unprotected alicyclic amines remain rare. A new advance in this area utilizes N-lithiated alicyclic amines. These readily accessible intermediates are converted to transient imines through the action of a simple ketone oxidant, followed by alkylation with a ß-ketoacid under mild conditions to provide valuable ß-amino ketones with unprecedented ease. Regioselective α'-alkylation is achieved for substrates with existing α-substituents. The method is further applicable to the convenient one-pot synthesis of polycyclic dihydroquinolones through the incorporation of a SN Ar step.

Maternal reflective functioning, interpersonal violence-related posttraumatic stress disorder, and risk for psychopathology in early childhood.

The aim of this study was to examine associations between maternal mentalization, interactive behavior, and child symptoms in families in which mothers suffer from interpersonal violence-related posttraumatic stress disorder (IPV-PTSD). Fifty-six mothers and children (aged 12-42 months) including mothers with a diagnosis of IPV-PTSD were studied. Mentalization was measured by the Parental Reflective Functioning (PRF) Scale. Interactive behavior during free-play was measured via the CARE-Index. Child symptoms were measured by the Infant-Toddler Social and Emotional Assessment (ITSEA). Data analyses included non-parametric correlations and multiple linear regression. Results showed that lower IPV-PTSD and higher Maternal Reflective Functioning (MRF) were related to greater maternal sensitivity. Lower MRF and greater controlling behavior were related to child dysregulation. MRF was found to be lower in the subgroup of IPV-PTSD when the child's father was the perpetrator of IPV. Both MRF and interactive behavior are thus likely to be important targets for intervention during sensitive periods of early social-emotional development.

The Role of the US Surgeon in Global Care: Melding the Reality at Home with Global Need.

There is a great mismatch between surgeon workforce capacity in the US and other high income countries (HICs) and that in low and lower middle income countries (LMICs). Many surgeons in HICs are willing to try to be of assistance in LMICs. It is not intuitive, though, exactly how such assistance is best delivered. Similarly, the body of literature describing what is known about the needs in LMICs may not be in the usual cadre of journals and sources accessed by many practicing surgeons. Consequently, many surgeons who are capable and willing to help in LMICs are often not sure how their abilities might be best used.This essay presents a very brief overview of what is known about those needs, then presents some commentary on how the practicing surgeon in the US and other HICs may be best utilized, with particular attention to the short term trip model. Deployment in the short term trip model is often the most practical and available means of making this effort for HIC surgeons. This model has come under significant criticism in recent years, often for good reason, but it is argued that details of the implementation of that model can determine its applicability to developmental needs. Given the practicality of short term deployments for HIC surgeons, it behooves Ministries of Health and NGOs to examine how trips of this nature can be incorporated into the overall bigger picture of surgical development.This essay aims to help the perspective of the HIC surgeon as s/he seeks to contribute to the development of surgical access and quality for the approximately five billion people in the world who do not have adequate access to surgical care.

Factors Affecting Burn Contracture Outcome in Developing Countries: A Review of 2506 Patients.

INTRODUCTION: Burn contractures hinder joint mobility, resulting in functional impairment and reduced quality of life. This is of greater significance in developing countries where there are fewer resources for assistance with such impairments. Contracture release reduces deformity, but multiple factors affect the extent of postsurgical improvements and outcomes. Elucidating these factors may enable surgeons to better care for burn patients. This study assesses factors that impact burn contracture resolution in developing nations. METHODS: A retrospective review of 2506 burn contractures was performed using information extracted from a large nongovernment organization (ReSurge International) database from Nepal, India, and Zambia. Data points included age, type of burn, time elapsed between injury and release, and extent of final release achieved based on preoperative and postoperative images of hand (n = 1960), elbow (n = 371), and knee (n = 176) contractures. Hand improvement was scored based on digit/wrist involvement (severity of dysfunction) and joint extension capability (functionality); elbow and knee improvement were calculated using preoperative and postoperative joint angles. Multivariate analysis was performed. RESULTS: Hands burned by hot liquid had greater functionality after surgery than open-fire burns (P < 0.01). Improvement in severity of dysfunction and functionality were inversely correlated to age (P < 0.01) and time until surgery (P < 0.01). Elbow improvement decreased as age increased (P < 0.01). Postoperative increase of knee extension decreased for each year elapsed between injury and surgery (P < 0.01). CONCLUSIONS: Burn type, age when burned, and timing of surgery were significant factors affecting hand outcomes, whereas age affected elbow outcomes, and time elapsed until surgery affected knee results. An algorithm was formulated to enable physicians in developing countries with limited resources to triage patients and optimize patient outcomes.

Futsch/MAP1B mRNA is a translational target of TDP-43 and is neuroprotective in a Drosophila model of amyotrophic lateral sclerosis.

TDP-43 is an RNA-binding protein linked to amyotrophic lateral sclerosis (ALS) that is known to regulate the splicing, transport, and storage of specific mRNAs into stress granules. Although TDP-43 has been shown to interact with translation factors, its role in protein synthesis remains unclear, and no in vivo translation targets have been reported to date. Here we provide evidence that TDP-43 associates with futsch mRNA in a complex and regulates its expression at the neuromuscular junction (NMJ) in Drosophila. In the context of TDP-43-induced proteinopathy, there is a significant reduction of futsch mRNA at the NMJ compared with motor neuron cell bodies where we find higher levels of transcript compared with controls. TDP-43 also leads to a significant reduction in Futsch protein expression at the NMJ. Polysome fractionations coupled with quantitative PCR experiments indicate that TDP-43 leads to a futsch mRNA shift from actively translating polysomes to nontranslating ribonuclear protein particles, suggesting that in addition to its effect on localization, TDP-43 also regulates the translation of futsch mRNA. We also show that futsch overexpression is neuroprotective by extending life span, reducing TDP-43 aggregation, and suppressing ALS-like locomotor dysfunction as well as NMJ abnormalities linked to microtubule and synaptic stabilization. Furthermore, the localization of MAP1B, the mammalian homolog of Futsch, is altered in ALS spinal cords in a manner similar to our observations in Drosophila motor neurons. Together, our results suggest a microtubule-dependent mechanism in motor neuron disease caused by TDP-43-dependent alterations in futsch mRNA localization and translation in vivo.

Unassigning bacterial species for microbiome studies.

The method of 16S rRNA marker gene sequencing has fueled microbiome research and continues to be relevant. A perceived weakness of the method is that taxonomic assignments are not possible to make at the rank of species. We show that by working to rule out bacterial or archaeal species membership, we can provide an answer that is more accurate and useful. The Unassigner software operates on 16S rRNA marker gene data and computes a rule-out probability for species membership using a beta-binomial distribution. We demonstrate that our approach is accurate based on full-genome comparisons. Our method is consistent with existing approaches and dramatically improves on them based on the percentage of reads it can associate with a species in a sample. The software is available at https://github.com/PennChopMicrobiomeProgram/unassigner.IMPORTANCEWhile existing methods do not provide reliable species-level assignments for 16S rRNA marker gene data, the Unassigner software solves this problem by ruling out species membership, allowing researchers to reason at the species level.

A quantitative approach to measure and predict microbiome response to antibiotics.

Although antibiotics induce sizable perturbations in the human microbiome, we lack a systematic and quantitative method to measure and predict the microbiome's response to specific antibiotics. Here, we introduce such a method, which takes the form of a microbiome response index (MiRIx) for each antibiotic. Antibiotic-specific MiRIx values quantify the overall susceptibility of the microbiota to an antibiotic, based on databases of bacterial phenotypes and published data on intrinsic antibiotic susceptibility. We applied our approach to five published microbiome studies that carried out antibiotic interventions with vancomycin, metronidazole, ciprofloxacin, amoxicillin, and doxycycline. We show how MiRIx can be used in conjunction with existing microbiome analytical approaches to gain a deeper understanding of the microbiome response to antibiotics. Finally, we generate antibiotic response predictions for the oral, skin, and gut microbiome in healthy humans. Our approach is implemented as open-source software and is readily applied to microbiome data sets generated by 16S rRNA marker gene sequencing or shotgun metagenomics. IMPORTANCE: Antibiotics are potent influencers of the human microbiome and can be a source for enduring dysbiosis and antibiotic resistance in healthcare. Existing microbiome data analysis methods can quantify perturbations of bacterial communities but cannot evaluate whether the differences are aligned with the expected activity of a specific antibiotic. Here, we present a novel method to quantify and predict antibiotic-specific microbiome changes, implemented in a ready-to-use software package. This has the potential to be a critical tool to broaden our understanding of the relationship between the microbiome and antibiotics.

Surgery for Crohn's Disease Is Associated With a Dysbiotic Microbiome and Metabolome: Results From Two Prospective Cohorts.

BACKGROUND & AIMS: Crohn's disease is associated with alterations in the gut microbiome and metabolome described as dysbiosis. We characterized the microbial and metabolic consequences of ileal resection, the most common Crohn's disease surgery. METHODS: Patients with and without intestinal resection were identified from the Diet to Induce Remission in Crohn's Disease and Study of a Prospective Adult Research Cohort with Inflammatory Bowel Disease studies. Stool samples were analyzed with shotgun metagenomics sequencing. Fecal butyrate was measured with 1H nuclear magnetic resonance spectroscopy. Fecal bile acids and plasma 7α-hydroxy-4-cholesten-3-one (C4) was measured with mass spectrometry. RESULTS: Intestinal resection was associated with reduced alpha diversity and altered beta diversity with increased Proteobacteria and reduced Bacteroidetes and Firmicutes. Surgery was associated with higher representation of genes in the KEGG pathway for ABC transporters and reduction in genes related to bacterial metabolism. Surgery was associated with reduced concentration of the But gene but this did not translate to reduced fecal butyrate concentration. Surgery was associated with decreased abundance of bai operon genes, with increased plasma C4 concentration, increased primary bile acids and reduced secondary bile acids, including isoLCA. Additionally, Egerthella lenta, Adlercreutzia equalofaciens, and Gordonibacter pamelaeae were lower in abundance among patients with prior surgery in both cohorts. CONCLUSIONS: In 2 different populations, prior surgery in Crohn's disease is associated with altered fecal microbiome. Patients who had undergone ileal resection had reduction in the potentially beneficial bacteria E lenta and related actinobacteria and secondary bile acids, including isoLCA, suggesting that these could be biomarkers of patients at higher risk for disease progression.

Disruption of intestinal oxygen balance in acute colitis alters the gut microbiome.

The juxtaposition of well-oxygenated intestinal colonic tissue with an anerobic luminal environment supports a fundamentally important relationship that is altered in the setting of intestinal injury, a process likely to be relevant to diseases such as inflammatory bowel disease. Herein, using two-color phosphorometry to non-invasively quantify both intestinal tissue and luminal oxygenation in real time, we show that intestinal injury induced by DSS colitis reduces intestinal tissue oxygenation in a spatially defined manner and increases the flux of oxygen from the tissue into the gut lumen. By characterizing the composition of the microbiome in both DSS colitis-affected gut and in a bioreactor containing a stable human fecal community exposed to microaerobic conditions, we provide evidence that the increased flux of oxygen into the gut lumen augments glycan degrading bacterial taxa rich in glycoside hydrolases which are known to inhabit gut mucosal surface. Continued disruption of the intestinal mucus barrier through such a mechanism may play a role in the perpetuation of the intestinal inflammatory process.

Effects of dietary zinc on the gut microbiome and resistome of the gestating cow and neonatal calf.

Zinc is an essential trace element required in the diet of all species. While the effects of zinc have been studied in growing calves, little is known about the effect of zinc on the microbiota of the gestating cow or her neonatal calf. Understanding factors that shape the gut health of neonatal animals and evaluating the effect of dietary supplements in adult gestating animals is important in promoting animal health and informing feeding practices. The aims of this study were to determine the effect of dietary zinc on the microbiota and resistome of the gestating cow and calf. Gestating cows received standard (40 ppm) or high (205 ppm) dietary zinc levels from dry off to calving. Fecal samples were collected from cows upon enrollment and at calving and from neonatal calves. Fecal samples underwent 16S rRNA sequencing and a subset also underwent shotgun metagenomic sequencing. The effect of zinc supplementation on the diversity and composition of the cow and calf microbiome and resistome was assessed. Alpha and beta diversity and composition of the microbiota were significantly altered over time but not by treatment in the cows, with alpha diversity decreasing and 14 genera found at significantly higher relative abundances at calving compared to enrollment. Levels of 27 antimicrobial resistance genes significantly increased over time. Only a small number of taxa were differentially expressed at calving in treatment and control groups, including Faecalibacterium, Bacteroides, Turicibacter, and Bifidobacterium pseudolongum. No effect of the dam's treatment group was observed on the diversity or composition of the neonatal calf microbiota. The calf resistome, which was relatively rich and diverse compared to the cow, was also unaffected by the dam's treatment group. The impact of high levels of dietary zinc thus appeared to be minimal, with no observed changes in alpha or beta diversity, and few changes in the relative abundance of a small number of taxa and antimicrobial resistance genes.

The evolutionarily ancient FOXA transcription factors shape the murine gut microbiome via control of epithelial glycosylation.

Evolutionary adaptation of multicellular organisms to a closed gut created an internal microbiome differing from that of the environment. Although the composition of the gut microbiome is impacted by diet and disease state, we hypothesized that vertebrates promote colonization by commensal bacteria through shaping of the apical surface of the intestinal epithelium. Here, we determine that the evolutionarily ancient FOXA transcription factors control the composition of the gut microbiome by establishing favorable glycosylation on the colonic epithelial surface. FOXA proteins bind to regulatory elements of a network of glycosylation enzymes, which become deregulated when Foxa1 and Foxa2 are deleted from the intestinal epithelium. As a direct consequence, microbial composition shifts dramatically, and spontaneous inflammatory bowel disease ensues. Microbiome dysbiosis was quickly reversed upon fecal transplant into wild-type mice, establishing a dominant role for the host epithelium, in part mediated by FOXA factors, in controlling symbiosis in the vertebrate holobiont.