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Software that easily helps higher education instructors to remove poor quality items and set appropriate grade boundaries is generally lacking. To address these challenges, the SmartStandardSet system provides a graphical-user interface for removing defective items, weighting student scores using a two-parameter model IRT score analysis, and a mechanism for standard-setting. We evaluated the system through a series of six interviews with teachers and six focus groups involving 19 students to understand how key stakeholders would view the use of the tool in practice. Generally, both groups of participants reported high levels of feasibility, accuracy, and utility in SmartStandardSet’s statistical scoring of items and score calculation for test-takers. Teachers indicated the data displays would help them improve future test items; students indicated the system would be fairer and would motivate greater effort on more difficult test items. However, both groups had concerns about implementing the system without institutional policy endorsement. Students specifically were concerned that academics may set grade boundaries on arbitrary and invalid grounds. Our results provide useful insights into the perceived benefits of using the tool for standard setting, and suggest concrete next steps for gaining wider acceptance that will be the focus of future work.

Systemic autoinflammatory diseases (SAIDs) represent a spectrum of genetically heterogeneous inflammatory disorders. Some SAID-associated genes are located in chromosome 16, including familial Mediterranean fever gene (MEFV) and nucleotide-binding oligomerization domain 2 [NOD2] gene that are linked to Crohn’s disease, Blau syndrome, and Yao syndrome. These disorders share overlapping clinical phenotypes, and genotyping is diagnostically helpful and distinctive. Using next generation sequencing in SAIDs, digenic variants or combinations of more genetic variants in different genes can be detected, and they may be related to the MEFV and NOD2 genes. These variants may contribute to heterogeneous phenotypes in an individual, complicating the diagnosis and therapy. An awareness of the clinical significance of the digenic or combined gene variants is important in the era of genomic medicine.

Abstract Oncogenic KRAS plays a vital role in controlling tumor metabolism by enhancing aerobic glycolysis. Obesity driven by chronic consumption of high-fat diet (HFD) is a major risk factor for oncogenic KRAS-mediated pancreatic ductal adenocarcinoma (PDAC). However, the role of HFD in KRAS-mediated metabolic reprogramming has been obscure. Here, by using genetically engineered mouse models expressing an endogenous level of KRASG12D in pancreatic acinar cells, we demonstrate that hyperactivation of KRASG12D by obesogenic HFD, as compared to carbohydrate-rich diet, is responsible for enhanced aerobic glycolysis that associates with critical pathogenic responses in the path towards PDAC. Ablation of Cox-2 attenuates KRAS hyperactivation leading to the reversal of both aggravated aerobic glycolysis and high-grade dysplasia under HFD challenge. Our data highlight a pivotal role of the cooperative interaction between obesity-ensuing HFD and oncogenic KRAS in driving the heightened aerobic glycolysis during pancreatic tumorigenesis and suggest that in addition to directly targeting KRAS and aerobic glycolysis pathway, strategies to target the upstream of KRAS hyperactivation may bear important therapeutic value.

Abstract The combination therapy of lumacaftor and ivacaftor (Orkambi®) is approved for patients bearing the major cystic fibrosis (CF) mutation: ΔF508. It has been predicted that Orkambi® could treat patients with rarer mutations of similar “theratype”; however, a standardized approach confirming efficacy in these cohorts has not been reported. Here, we demonstrate that patients bearing the rare mutation: c.3700 A>G, causing protein misprocessing and altered channel function—similar to ΔF508‐CFTR, are unlikely to yield a robust Orkambi® response. While in silico and biochemical studies confirmed that this mutation could be corrected and potentiated by lumacaftor and ivacaftor, respectively, this combination led to a minor in vitro response in patient‐derived tissue. A CRISPR/Cas9‐edited bronchial epithelial cell line bearing this mutation enabled studies showing that an “amplifier” compound, effective in increasing the levels of immature CFTR protein, augmented the Orkambi® response. Importantly, this “amplifier” effect was recapitulated in patient‐derived nasal cultures—providing the first evidence for its efficacy in augmenting Orkambi® in tissues harboring a rare CF‐causing mutation. We propose that this multi‐disciplinary approach, including creation of CRISPR/Cas9‐edited cells to profile modulators together with validation using primary tissue, will facilitate therapy development for patients with rare CF mutations.

We have previously identified NOD2 genotype and inflammatory bowel diseases (IBD) phenotype, as associated with shifts in the ileal microbiome ("dysbiosis") in a patient cohort. Here we report an integrative analysis of an expanded number of Crohn's disease (CD) related genetic defects in innate immune function (NOD2, ATG16L1, IRGM, CARD9, XBP1, ORMDL3) and composition of the ileal microbiome by combining the initial patient cohort (Batch 1, 2005-2010, n = 165) with a second consecutive patient cohort (Batch 2, 2010-2012, n = 118). These combined patient cohorts were composed of three non-overlapping phenotypes: 1.) 106 ileal CD subjects undergoing initial ileocolic resection for diseased ileum, 2.) 88 IBD colitis subjects without ileal disease (predominantly ulcerative colitis but also Crohn's colitis and indeterminate colitis, and 3.) 89 non-IBD subjects. Significant differences (FDR < 0.05) in microbiota were observed between macroscopically disease unaffected and affected regions of resected ileum in ileal CD patients. Accordingly, analysis of the effects of genetic and clinical factors were restricted to disease unaffected regions of the ileum. Beta-diversity differed across the three disease categories by PERMANOVA (p < 0.001), whereas no significant differences in alpha diversity were noted. Using negative binomial models, we confirmed significant effects of IBD phenotype, C. difficile infection, and NOD2 genotype on ileal dysbiosis in the expanded analysis. The relative abundance of the Proteobacteria phylum was positively associated with ileal CD and colitis phenotypes, but negatively associated with NOD2R genotype. Additional associations with ORMDL3 and XBP1 were detected at the phylum/subphylum level. IBD medications, such as immunomodulators and anti-TNFα agents, may have a beneficial effect on reversing dysbiosis associated with the IBD phenotype. Exploratory analysis comparing microbial composition of the disease unaffected region of the resected ileum between 27 ileal CD patients who subsequently developed endoscopic recurrence within 6-12 months versus 34 patients who did not, suggested that microbial biomarkers in the resected specimen helped stratify patients with respect to risk of post-surgical recurrence.

Studies of colonoscopic fecal microbiota transplant (FMT) in patients with recurrent CDI, indicate that this is a very effective treatment for preventing further relapses. In order to provide this service at Stony Brook University Hospital, we initiated an open-label prospective study of single colonoscopic FMT among patients with ≥ 2 recurrences of CDI, with the intention of monitoring microbial composition in the recipient before and after FMT, as compared with their respective donor. We also initiated a concurrent open label prospective trial of single colonoscopic FMT of patients with ulcerative colitis (UC) not responsive to therapy, after obtaining an IND permit (IND 15642). To characterize how FMT alters the fecal microbiota in patients with recurrent Clostridia difficile infections (CDI) and/or UC, we report the results of a pilot microbiome analysis of 11 recipients with a history of 2 or more recurrences of C. difficile infections without inflammatory bowel disease (CDI-only), 3 UC recipients with recurrent C. difficile infections (CDI + UC), and 5 UC recipients without a history of C. difficile infections (UC-only).V3V4 Illumina 16S ribosomal RNA (rRNA) gene sequencing was performed on the pre-FMT, 1-week post-FMT, and 3-months post-FMT recipient fecal samples along with those collected from the healthy donors. Fitted linear mixed models were used to examine the effects of Group (CDI-only, CDI + UC, UC-only), timing of FMT (Donor, pre-FMT, 1-week post-FMT, 3-months post-FMT) and first order Group*FMT interactions on the diversity and composition of fecal microbiota. Pairwise comparisons were then carried out on the recipient vs. donor and between the pre-FMT, 1-week post-FMT, and 3-months post-FMT recipient samples within each group.Significant effects of FMT on overall microbiota composition (e.g., beta diversity) were observed for the CDI-only and CDI + UC groups. Marked decreases in the relative abundances of the strictly anaerobic Bacteroidetes phylum, and two Firmicutes sub-phyla associated with butyrate production (Ruminococcaceae and Lachnospiraceae) were observed between the CDI-only and CDI + UC recipient groups. There were corresponding increases in the microaerophilic Proteobacteria phylum and the Firmicutes/Bacilli group in the CDI-only and CDI + UC recipient groups. At a more granular level, significant effects of FMT were observed for 81 genus-level operational taxonomic units (OTUs) in at least one of the three recipient groups (p

[This corrects the article DOI: 10.1371/journal.pone.0153125.].

Incidence and mortality rates of colorectal carcinoma (CRC) are higher in African Americans (AAs) than in Caucasian Americans (CAs). Deficient micronutrient intake due to dietary restrictions in racial/ethnic populations can alter genetic and molecular profiles leading to dysregulated methylation patterns and the inheritance of somatic to germline mutations.Total DNA and RNA samples of paired tumor and adjacent normal colon tissues were prepared from AA and CA CRC specimens. Reduced Representation Bisulfite Sequencing (RRBS) and RNA sequencing were employed to evaluate total genome methylation of 5'-regulatory regions and dysregulation of gene expression, respectively. Robust analysis was conducted using a trimming-and-retrieving scheme for RRBS library mapping in conjunction with the BStool toolkit.DNA from the tumor of AA CRC patients, compared to adjacent normal tissues, contained 1,588 hypermethylated and 100 hypomethylated differentially methylated regions (DMRs). Whereas, 109 hypermethylated and 4 hypomethylated DMRs were observed in DNA from the tumor of CA CRC patients; representing a 14.6-fold and 25-fold change, respectively. Specifically; CHL1, 4 anti-inflammatory genes (i.e., NELL1, GDF1, ARHGEF4, and ITGA4), and 7 miRNAs (of which miR-9-3p and miR-124-3p have been implicated in CRC) were hypermethylated in DNA samples from AA patients with CRC. From the same sample set, RNAseq analysis revealed 108 downregulated genes (including 14 ribosomal proteins) and 34 upregulated genes (including POLR2B and CYP1B1 [targets of miR-124-3p]) in AA patients with CRC versus CA patients.DNA methylation profile and/or products of its downstream targets could serve as biomarker(s) addressing racial health disparity.

Mutation of the adenomatous polyposis coli (APC gene), an early event in the adenoma-carcinoma sequence, is present in 70-80% of sporadic human colorectal adenomas and carcinomas. To test the hypothesis that mutation of the APC gene alters microbial interactions with host intestinal mucosa prior to the development of polyposis, culture-independent methods (targeted qPCR assays and Illumina sequencing of the 16S rRNA gene V1V2 hypervariable region) were used to compare the intestinal microbial composition of 30 six-week old C57BL/6 APCMin/+ and 30 congenic wild type (WT) mice. The results demonstrate that similar to 12-14 week old APCMin/+ mice with intestinal neoplasia, 6 week old APCMin/+ mice with no detectable neoplasia, exhibit an increased relative abundance of Bacteroidetes spp in the colon. Parallel mouse RNA sequence analysis, conducted on a subset of proximal colonic RNA samples (6 APCMin/+, 6 WT) revealed 130 differentially expressed genes (DEGs, fold change ≥ 2, FDR

In order to assess potential associations between autism spectrum disorder (ASD) phenotype, functional GI disorders and fecal microbiota, we recruited simplex families, which had only a single ASD proband and neurotypical (NT) siblings, through the Simons Simplex Community at the Interactive Autism Network (SSC@IAN). Fecal samples and metadata related to functional GI disorders and diet were collected from ASD probands and NT siblings of ASD probands (age 7-14). Functional gastrointestinal disorders (FGID) were assessed using the parent-completed ROME III questionnaire for pediatric FGIDs, and problem behaviors were assessed using the Child Behavior Check List (CBCL). Targeted quantitative polymerase chain reaction (qPCR) assays were conducted on selected taxa implicated in ASD, including Sutterella spp., Bacteroidetes spp. and Prevotella spp. Illumina sequencing of the V1V2 and the V1V3 regions of the bacterial 16S rRNA genes from fecal DNA was performed to an average depth of 208,000 and 107,000 high-quality reads respectively. Twenty-five of 59 ASD children and 13 of 44 NT siblings met ROME III criteria for at least one FGID. Functional constipation was more prevalent in ASD (17 of 59) compared to NT siblings (6 of 44, P = 0.035). The mean CBCL scores in NT siblings with FGID, ASD children with FGID and ASD without FGID were comparably higher (58-62 vs. 44, P < 0.0001) when compared to NT children without FGID. There was no significant difference in macronutrient intake between ASD and NT siblings. There was no significant difference in ASD severity scores between ASD children with and without FGID. No significant difference in diversity or overall microbial composition was detected between ASD children with NT siblings. Exploratory analysis of the 16S rRNA sequencing data, however, identified several low abundance taxa binned at the genus level that were associated with ASD and/or first order ASD*FGID interactions (FDR

Adherent-invasive Escherichia coli (AIEC) strains are detected more frequently within mucosal lesions of patients with Crohn's disease (CD). The AIEC phenotype consists of adherence and invasion of intestinal epithelial cells and survival within macrophages of these bacteria in vitro. Our aim was to identify candidate transcripts that distinguish AIEC from non-invasive E. coli (NIEC) strains and might be useful for rapid and accurate identification of AIEC by culture-independent technology. We performed comparative RNA-Sequence (RNASeq) analysis using AIEC strain LF82 and NIEC strain HS during exponential and stationary growth. Differential expression analysis of coding sequences (CDS) homologous to both strains demonstrated 224 and 241 genes with increased and decreased expression, respectively, in LF82 relative to HS. Transition metal transport and siderophore metabolism related pathway genes were up-regulated, while glycogen metabolic and oxidation-reduction related pathway genes were down-regulated, in LF82. Chemotaxis related transcripts were up-regulated in LF82 during the exponential phase, but flagellum-dependent motility pathway genes were down-regulated in LF82 during the stationary phase. CDS that mapped only to the LF82 genome accounted for 747 genes. We applied an in silico subtractive genomics approach to identify CDS specific to AIEC by incorporating the genomes of 10 other previously phenotyped NIEC. From this analysis, 166 CDS mapped to the LF82 genome and lacked homology to any of the 11 human NIEC strains. We compared these CDS across 13 AIEC, but none were homologous in each. Four LF82 gene loci belonging to clustered regularly interspaced short palindromic repeats region (CRISPR)--CRISPR-associated (Cas) genes were identified in 4 to 6 AIEC and absent from all non-pathogenic bacteria. As previously reported, AIEC strains were enriched for pdu operon genes. One CDS, encoding an excisionase, was shared by 9 AIEC strains. Reverse transcription quantitative polymerase chain reaction assays for 6 genes were conducted on fecal and ileal RNA samples from 22 inflammatory bowel disease (IBD), and 32 patients without IBD (non-IBD). The expression of Cas loci was detected in a higher proportion of CD than non-IBD fecal and ileal RNA samples (p

BACKGROUND: Colorectal cancer (CRC) has the third highest mortality rates among the US population. According to the most recent concept of carcinogenesis, human tumors are organized hierarchically, and the top of it is occupied by malignant stem cells (cancer stem cells, CSCs, or cancer-initiating cells, CICs), which possess unlimited self-renewal and tumor-initiating capacities and high resistance to conventional therapies. To reflect the complexity and diversity of human tumors and to provide clinically and physiologically relevant cancer models, large banks of characterized patient-derived low-passage cell lines, and especially CIC-enriched cell lines, are urgently needed. PRINCIPAL FINDINGS: Here we report the establishment of a novel CIC-enriched, highly tumorigenic and clonogenic colon cancer cell line, CR4, derived from liver metastasis. This stable cell line was established by combining 3D culturing and 2D culturing in stem cell media, subcloning of cells with particular morphology, co-culture with carcinoma associated fibroblasts (CAFs) and serial transplantation to NOD/SCID mice. Using RNA-Seq complete transcriptome profiling of the tumorigenic fraction of the CR4 cells in comparison to the bulk tumor cells, we have identified about 360 differentially expressed transcripts, many of which represent stemness, pluripotency and resistance to treatment. Majority of the established CR4 cells express common markers of stemness, including CD133, CD44, CD166, EpCAM, CD24 and Lgr5. Using immunocytochemical, FACS and western blot analyses, we have shown that a significant ratio of the CR4 cells express key markers of pluripotency markers, including Sox-2, Oct3/4 and c-Myc. Constitutive overactivation of ABC transporters and NF-kB and absence of tumor suppressors p53 and p21 may partially explain exceptional drug resistance of the CR4 cells. CONCLUSIONS: The highly tumorigenic and clonogenic CIC-enriched CR4 cell line may provide an important new tool to support the discovery of novel diagnostic and/or prognostic biomarkers as well as the development of more effective therapeutic strategies.

Crohn's disease (CD) is a chronic inflammatory disease of the gastrointestinal tract. Genetic polymorphisms can confer CD risk and influence disease phenotype. Indoleamine 2,3 dioxygenase-1 (IDO1) is one of the most over-expressed genes in CD and mediates potent anti-inflammatory effects via tryptophan metabolism along the kynurenine pathway. We aimed to determine whether non-synonymous polymorphisms in IDO1 or IDO2 (a gene paralog) are important either as CD risk alleles or as modifiers of CD phenotype.Utilizing a prospectively collected database, clinically phenotyped CD patients (n = 734) and non-IBD controls (n = 354) were genotyped for established IDO1 and IDO2 non-synonymous single nucleotide polymorphisms (SNPs) and novel genetic variants elucidated in the literature. Allelic frequencies between CD and non-IBD controls were compared. Genotype-phenotype analysis was conducted. IDO1 enzyme activity was assessed by calculating the serum kynurenine to tryptophan ratio (K/T).IDO1 SNPs were rare (1.7% non-IBD vs 1.1% CD; p = NS) and not linked to Crohn's disease diagnosis in this population. IDO1 SNPs did however associate with a severe clinical course, presence of perianal disease, extraintestinal manifestations and a reduced serum K/T ratio during active disease suggesting lower IDO1 function. IDO2 minor allele variants were common and one of them, rs45003083, associated with reduced risk of Crohn's disease (p = 0.025). No IDO2 SNPs associated with a particular Crohn's disease clinical phenotype.This work highlights the functional importance of IDO enzymes in human Crohn's disease and establishes relative rates of IDO genetic variants in a US population.

Abnormalities of the intestinal microbiota are implicated in the pathogenesis of Crohn's disease (CD) and ulcerative colitis (UC), two spectra of inflammatory bowel disease (IBD). However, the high complexity and low inter-individual overlap of intestinal microbial composition are formidable barriers to identifying microbial taxa representing this dysbiosis. These difficulties might be overcome by an ecologic analytic strategy to identify modules of interacting bacteria (rather than individual bacteria) as quantitative reproducible features of microbial composition in normal and IBD mucosa. We sequenced 16S ribosomal RNA genes from 179 endoscopic lavage samples from different intestinal regions in 64 subjects (32 controls, 16 CD and 16 UC patients in clinical remission). CD and UC patients showed a reduction in phylogenetic diversity and shifts in microbial composition, comparable to previous studies using conventional mucosal biopsies. Analysis of weighted co-occurrence network revealed 5 microbial modules. These modules were unprecedented, as they were detectable in all individuals, and their composition and abundance was recapitulated in an independent, biopsy-based mucosal dataset 2 modules were associated with healthy, CD, or UC disease states. Imputed metagenome analysis indicated that these modules displayed distinct metabolic functionality, specifically the enrichment of oxidative response and glycan metabolism pathways relevant to host-pathogen interaction in the disease-associated modules. The highly preserved microbial modules accurately classified IBD status of individual patients during disease quiescence, suggesting that microbial dysbiosis in IBD may be an underlying disorder independent of disease activity. Microbial modules thus provide an integrative view of microbial ecology relevant to IBD.

ABSTRACT Autism is a heterogeneous group of complex developmental disabilities that result from a number of possible etiologies. There are a well-known number of comorbidities associated with autism spectrum disorders (ASD), including, commonly, gastrointestinal (GI) pathology, which can include variable combinations of constipation, diarrhea, abdominal pain, gastroesophageal reflux, and vomiting. An American Academy of Pediatrics consensus panel has recommended that prospective studies be carried out to determine the prevalence of GI disorders in ASD and their pathophysiologic basis. In a recent article, Williams et al. [B. L. Williams, M. Hornig, T. Parekh, and W. I. Lipkin, mBio 3(1):e00261-11, 2012] have provided one such study of autism with GI comorbidities by presenting evidence of Sutterella species in ileal mucosal biopsy specimens from patients diagnosed with ASD but not in control children with GI symptoms, suggesting a specific role for Sutterella in ASD. Sutterella sequences represented ~1 to 7% of the total bacterial sequences, and this is a very large effect size on the ileal mucosal composition of the autism phenotype, rivaling or perhaps exceeding the effect size of the ileal Crohn’s disease phenotype. This study opens a new field of investigation to study the etiology or consequences of GI comorbidities in ASD.

Previous genome-wide expression studies have highlighted distinct gene expression patterns in inflammatory bowel disease (IBD) compared to control samples, but the interpretation of these studies has been limited by sample heterogeneity with respect to disease phenotype, disease activity, and anatomic sites. To further improve molecular classification of inflammatory bowel disease phenotypes we focused on a single anatomic site, the disease unaffected proximal ileal margin of resected ileum, and three phenotypes that were unlikely to overlap: ileal Crohn's disease (ileal CD), ulcerative colitis (UC), and control patients without IBD. Whole human genome (Agilent) expression profiling was conducted on two independent sets of disease-unaffected ileal samples collected from the proximal margin of resected ileum. Set 1 (47 ileal CD, 27 UC, and 25 Control non-IBD patients) was used as the training set and Set 2 was subsequently collected as an independent test set (10 ileal CD, 10 UC, and 10 control non-IBD patients). We compared the 17 gene signatures selected by four different feature-selection methods to distinguish ileal CD phenotype with non-CD phenotype. The four methods yielded different but overlapping solutions that were highly discriminating. All four of these methods selected FOLH1 as a common feature. This gene is an established biomarker for prostate cancer, but has not previously been associated with Crohn's disease. Immunohistochemical staining confirmed increased expression of FOLH1 in the ileal epithelium. These results provide evidence for convergent molecular abnormalities in the macroscopically disease unaffected proximal margin of resected ileum from ileal CD subjects.

The aim of this study was to integrate human clinical, genotype, mRNA microarray and 16 S rRNA sequence data collected on 84 subjects with ileal Crohn's disease, ulcerative colitis or control patients without inflammatory bowel diseases in order to interrogate how host-microbial interactions are perturbed in inflammatory bowel diseases (IBD). Ex-vivo ileal mucosal biopsies were collected from the disease unaffected proximal margin of the ileum resected from patients who were undergoing initial intestinal surgery. Both RNA and DNA were extracted from the mucosal biopsy samples. Patients were genotyped for the three major NOD2 variants (Leufs1007, R702W, and G908R) and the ATG16L1T300A variant. Whole human genome mRNA expression profiles were generated using Agilent microarrays. Microbial composition profiles were determined by 454 pyrosequencing of the V3-V5 hypervariable region of the bacterial 16 S rRNA gene. The results of permutation based multivariate analysis of variance and covariance (MANCOVA) support the hypothesis that host mucosal Paneth cell and xenobiotic metabolism genes play an important role in host microbial interactions.

We tested the hypothesis that Crohn's disease (CD)-related genetic polymorphisms involved in host innate immunity are associated with shifts in human ileum-associated microbial composition in a cross-sectional analysis of human ileal samples. Sanger sequencing of the bacterial 16S ribosomal RNA (rRNA) gene and 454 sequencing of 16S rRNA gene hypervariable regions (V1-V3 and V3-V5), were conducted on macroscopically disease-unaffected ileal biopsies collected from 52 ileal CD, 58 ulcerative colitis and 60 control patients without inflammatory bowel diseases (IBD) undergoing initial surgical resection. These subjects also were genotyped for the three major NOD2 risk alleles (Leu1007fs, R708W, G908R) and the ATG16L1 risk allele (T300A). The samples were linked to clinical metadata, including body mass index, smoking status and Clostridia difficile infection. The sequences were classified into seven phyla/subphyla categories using the Naïve Bayesian Classifier of the Ribosome Database Project. Centered log ratio transformation of six predominant categories was included as the dependent variable in the permutation based MANCOVA for the overall composition with stepwise variable selection. Polymerase chain reaction (PCR) assays were conducted to measure the relative frequencies of the Clostridium coccoides - Eubacterium rectales group and the Faecalibacterium prausnitzii spp. Empiric logit transformations of the relative frequencies of these two microbial groups were included in permutation-based ANCOVA. Regardless of sequencing method, IBD phenotype, Clostridia difficile and NOD2 genotype were selected as associated (FDR ≤ 0.05) with shifts in overall microbial composition. IBD phenotype and NOD2 genotype were also selected as associated with shifts in the relative frequency of the C. coccoides--E. rectales group. IBD phenotype, smoking and IBD medications were selected as associated with shifts in the relative frequency of F. prausnitzii spp. These results indicate that the effects of genetic and environmental factors on IBD are mediated at least in part by the enteric microbiota.

OBJECTIVES/GOALS: Serious video games are designed for skill-building and are increasingly being used for healthcare interventions with adolescents and young adults (AYAs). The study goal was to identify AYAs’ preferred game features, by demographic groups, to inform the development of a game to improve AYA’s engagement in their congenital heart disease (CHD) care. METHODS/STUDY POPULATION: Pediatric patients, 12-18 years old, completed surveys at a routine CHD care visit. Participants rated their likelihood of using games to learn CHD management skills (5-point Likert) and preferences for ten game features commonly used, such as: personalization (make your own avatar) and levels (unlock new, advanced stages as you do better). Participants selected one of three response options: 1=would make me less interested in the game, 2=doesn’t matter, 3=would make me more interested in the game. Descriptives and frequencies assessed interest in different game features. Chi-square tests were used to identify potential differences in game feature preferences by gender identity, age group (early/mid-adolescence vs. late adolescence), and race and ethnicity. RESULTS/ANTICIPATED RESULTS: Of 83 participants who completed surveys, the mean age was 15 years old (12-18; SD=1.73), 55% were male, 79% were Non-Hispanic White, and 70% were interested in video games for gaining CHD management skills. The top-rated game features were: levels (78%; unlock advanced stages), conflict (74%; face challenges), personalization (70%; create avatar), and story (70%; journey-based). The three lowest-ranked features were: time (29%; restricted time to complete challenge), competition (47%; score/play against others), strategy (53%; plan to reach goal). No significant differences in game feature preferences were found by demographic characteristics. DISCUSSION/SIGNIFICANCE: Most AYAs with CHD were interested in games, offering a promising avenue for future healthcare interventions. Given no significantly different preferences by demographics, the game may not require tailoring game features for certain groups. However, additional research with diverse participants is needed to fully inform game development.

Introduction Atrial fibrillation is highly prevalent in patients on chronic dialysis. It is unclear whether anticoagulant therapy for stroke prevention is beneficial in these patients. Vitamin K-antagonists (VKA) remain the predominant anticoagulant choice. Importantly, anticoagulation remains inconsistently used and a possible benefit remains untested in randomised clinical trials comparing oral anticoagulation with no treatment in patients on chronic dialysis. The Danish Warfarin-Dialysis (DANWARD) trial aims to investigate the safety and efficacy of VKAs in patients with atrial fibrillation on chronic dialysis. The hypothesis is that VKA treatment compared with no treatment is associated with stroke risk reduction and overall benefit.Methods and analysis The DANWARD trial is an investigator-initiated trial at 13 Danish dialysis centres. In an open-label randomised clinical trial study design, a total of 718 patients with atrial fibrillation on chronic dialysis will be randomised in a 1:1 ratio to receive either standard dose VKA targeting an international normalised ratio of 2.0–3.0 or no oral anticoagulation. Principal analyses will compare the risk of a primary efficacy endpoint, stroke or transient ischaemic attack and a primary safety endpoint, major bleeding, in patients allocated to VKA treatment and no treatment, respectively. The first patient was randomised in October 2019. Patients will be followed until 1 year after the inclusion of the last patient.Ethics and dissemination The study protocol was approved by the Regional Research Ethics Committee (journal number H-18050839) and the Danish Medicines Agency (case number 2018101877). The trial is conducted in accordance with the Helsinki declaration and standards of Good Clinical Practice. Study results will be disseminated to participating sites, at research conferences and in peer-reviewed journals.Trial registration numbers NCT03862859, EUDRA-CT 2018-000484-86 and CTIS ID 2022-502500-75-00.

Abstract Objective This study aimed to compare older adults reporting dizziness to those not reporting dizziness regarding health‐related quality of life (HRQL), distress due to dizziness, and balance confidence. A secondary aim was to investigate potential association between HRQL, number of falls, balance confidence, and distress due to dizziness. Methods Patients coming for bone density measurements answered questions regarding occurrence of dizziness. Patients reporting dizziness on a daily or weekly basis were considered eligible and invited for investigation at the Ear, Nose and Throat clinic at Södra Älvsborg Hospital, Sweden. Patients not reporting dizziness were considered eligible as controls. All patients answered the Dizziness Handicap Inventory (DHI), Activity Balance Confidence Scale (ABC‐scale), and Euro‐QoL‐5D‐3L questionnaires. Results A total of 55 dizzy patients came for physical investigation and answered the questionnaires and 47 non‐dizzy participants only answered the questionnaires. The dizzy participants reported lower levels of balance confidence, lower HRQL, more prior falls, and higher levels of distress due to dizziness than the non‐dizzy controls. Lower levels of balance confidence and higher level of distress due to dizziness were each associated with lower HRQL. Conclusion Dizziness, unsteadiness, and low balance confidence are associated with HRQL in a negative way. This is important to consider when measuring HRQL in a senior population, since a sensation of unsteadiness may indirectly contribute to low HRQL together with other symptoms. Level of evidence 2b.

Abstract Objective The present study aimed to relate the strength, assistance with walking, rising from a chair, climbing stairs, and falls (SARC-F) score with the presence or absence of fragility fracture in the population over 60 years of age. Methods The risk of sarcopenia was determined through the application of the SARC-F questionnaire, and the patients were divided into 2 groups, according to the occurrence or not of fragility fracture (n = 100). Results Thirty-two cases of distal radius fractures and eighteen cases of proximal femur fractures were identified. A higher score on the SARC-F is determinant between having or not a fragility fracture, estimating that for each point in the score there is a 70% increase in the chance of a patient having a fracture, regardless of age, gender, and body mass index (BMI). Conclusion There was a direct correlation between a higher score on the SARC-F and an increase in the chance of fragility fracture.

Abstract Human induced pluripotent stem cell-derived kidney organoids have potential for disease modeling and to be developed into clinically transplantable auxiliary tissue. However, they lack a functional vasculature, and the sparse endogenous endothelial cells (ECs) are lost upon prolonged culture in vitro, limiting maturation and applicability. Here, we use intracoelomic transplantation in chicken embryos followed by single-cell RNA sequencing and advanced imaging platforms to induce and study vasculogenesis in kidney organoids. We show expansion of human organoid-derived ECs that reorganize into perfused capillaries and form a chimeric vascular network with host-derived blood vessels. Ligand-receptor analysis infers extensive potential interactions of human ECs with perivascular cells upon transplantation, enabling vessel wall stabilization. Perfused glomeruli display maturation and morphogenesis to capillary loop stage. Our findings demonstrate the beneficial effect of vascularization on not only epithelial cell types, but also the mesenchymal compartment, inducing the expansion of ´on target´ perivascular stromal cells, which in turn are required for further maturation and stabilization of the neo-vasculature. The here described vasculogenic capacity of kidney organoids will have to be deployed to achieve meaningful glomerular maturation and kidney morphogenesis in vitro.

Abstract Background Dizziness and vertigo affect around 15% of adults annually and represent common reasons for contacting health services, accounting for around 3% of all emergency department visits worldwide. Vertigo is also associated with excessive use of diagnostic imaging and emergency care and decreased productivity, primarily because of work absenteeism. Vestibular rehabilitation is an evidence-based treatment for chronic dizziness and supervised group exercise therapy has recently been shown to be effective after vestibular neuritis, a common cause of acute onset vertigo. However, such interventions are not readily available and there is a need for more easily accessible tools. The purpose of this study is to investigate the effects on vestibular symptoms of a 6-week online vestibular rehabilitation tool after acute onset vertigo, with the aim of aiding vestibular rehabilitation by presenting a more accessible tool that can help to reduce recovery time. Methods Three hundred twenty individuals diagnosed with acute vestibular syndrome (AVS) will be recruited from multiple hospitals in Sweden and the effects of an online vestibular rehabilitation tool, YrselTräning, on vestibular symptoms after acute onset vertigo will be compared to standard care (written instructions leaflet) in a two-armed, evaluator-blinded, multicenter randomized controlled trial. The primary outcome will be the Vertigo Symptom Scale Short Form (VSS-SF) score at 6 weeks after symptom onset. Secondary outcomes include effects of the intervention on activities of daily living, mood and anxiety, vestibular function recovery, mobility measures, health economic effects, and the reliability of the Swedish VSS-SF translation. Discussion Participants using the online vestibular rehabilitation tool are expected to recover earlier and to a greater extent from their symptoms as compared to standard care. Since up to 50% of people with AVS without treatment develop persistent symptoms, effective treatment of AVS will likely lead to a higher quality of life and help reduce the societal costs associated with dizziness and vertigo. Trial registration Clinicaltrials.gov NCT05056324 . Registered on September 24, 2021.

Abstract Background Critically ill COVID-19 patients may develop acute respiratory distress syndrome and the need for respiratory support, including mechanical ventilation in the intensive care unit. Previous observational studies have suggested early tracheotomy to be advantageous. The aim of this parallel, multicentre, single-blinded, randomized controlled trial was to evaluate the optimal timing of tracheotomy. Methods SARS-CoV-2-infected patients within the Region Västra Götaland of Sweden who needed intubation and mechanical respiratory support were included and randomly assigned to early tracheotomy (≤ 7 days after intubation) or late tracheotomy (≥ 10 days after intubation). The primary objective was to compare the total number of mechanical ventilation days between the groups. Results One hundred fifty patients (mean age 65 years, 79% males) were included. Seventy-two patients were assigned to early tracheotomy, and 78 were assigned to late tracheotomy. One hundred two patients (68%) underwent tracheotomy of whom sixty-one underwent tracheotomy according to the protocol. The overall median number of days in mechanical ventilation was 18 (IQR 9; 28), but no significant difference was found between the two treatment regimens in the intention-to-treat analysis (between-group difference: − 1.5 days (95% CI − 5.7 to 2.8); p = 0.5). A significantly reduced number of mechanical ventilation days was found in the early tracheotomy group during the per-protocol analysis (between-group difference: − 8.0 days (95% CI − 13.8 to − 2.27); p = 0.0064). The overall correlation between the timing of tracheotomy and days of mechanical ventilation was significant (Spearman’s correlation: 0.39, p

Abstract Background The generation of human induced pluripotent stem cells (hiPSCs) has opened a world of opportunities for stem cell-based therapies in regenerative medicine. Currently, several human kidney organoid protocols are available that generate organoids containing kidney structures. However, these kidney organoids are relatively small ranging up to 0.13 cm2 and therefore contain a small number of nephrons compared to an adult kidney, thus defying the exploration of future use for therapy. Method We have developed a scalable, easily accessible, and reproducible protocol to increase the size of the organoid up to a nephron sheet of 2.5 cm2 up to a maximum of 12.6 cm2 containing a magnitude of nephrons. Results Confocal microscopy showed that the subunits of the nephrons remain evenly distributed throughout the entire sheet and that these tissue sheets can attain ~ 30,000–40,000 glomerular structures. Upon transplantation in immunodeficient mice, such nephron sheets became vascularized and matured. They also show reuptake of injected low-molecular mass dextran molecules in the tubular structures, indicative of glomerular filtration. Furthermore, we developed a protocol for the cryopreservation of intermediate mesoderm cells during the differentiation and demonstrate that these cells can be successfully thawed and recovered to create such tissue sheets. Conclusion The scalability of the procedures, and the ability to cryopreserve the cells during differentiation are important steps forward in the translation of these differentiation protocols to future clinical applications such as transplantable auxiliary kidney tissue.

Abstract Background Dizziness is a common complaint among older adults and may affect quality of life in a negative way. The aim of this study was to assess health-related quality of life (HRQL), sense of coherence (SOC), self-rated health (SRH) and comorbidity in relation to dizziness, among older persons from an urban population. Methods The study is part of the Gothenburg H70 Birth Cohort Studies (H70). A cross-sectional population-based sample including 662 79-years-olds (404 women, 258 men, 62% response rate) were surveyed with questions regarding dizziness, imbalance, comorbidities and general health. HRQL was assessed using the 36-item Short Form-36 Health Survey (SF-36) and SOC with the 13-items questionnaire Sense of Coherence (SOC-13). Results Half of the participants reported problems with dizziness (54%). Dizziness was negatively associated with HRQL, including after adjusting for comorbidities, especially in the physical domains of SF-36. Having dizziness was also associated with poorer SRH, tiredness and comorbidity among both men and women. SOC (mean total score), however, did not differ between dizzy and non-dizzy participants. Conclusions Dizziness was negatively associated with HRQL, also after adjusting for comorbidities. Identification and treatment of dizziness, when possible, are important because reduction of dizziness symptoms may potentially help to enhance overall well-being in this age group.

This article synthesizes findings from an international virtual conference, funded by the United States National Science Foundation, focused on the home mathematics environment (HME). In light of inconsistencies and gaps in research investigating relations between the HME and children’s outcomes, the purpose of the conference was to discuss actionable steps and considerations for future work. The conference was composed of international researchers with a wide range of expertise and backgrounds. Presentations and discussions during the conference centered broadly on the need to better operationalize and measure the HME as a construct—focusing on issues related to child, family, and community factors, country and cultural factors, and the cognitive and affective characteristics of caregivers and children. Results of the conference and a subsequent writing workshop include a synthesis of core questions and key considerations for the field of research on the HME. Findings highlight the need for the field at large to use multi-method measurement approaches to capture nuances in the HME, and to do so with increased international and interdisciplinary collaboration, open science practices, and communication among scholars.

Abstract Objective This article aimed to investigate older adults visiting a geriatric institution for a bone density measurement who reported dizziness on a daily or weekly basis (but who are not seeking care for dizziness), according to BPPV, vestibular function, walking abilities, and frequency of falls. Methods Patients coming for a bone density measurement answered questions regarding occurrence of dizziness. Patients having dizziness on a daily or weekly basis were considered eligible for the study and invited for investigation at the Ear, Nose, and Throat clinic at Södra Älvsborg Hospital, Sweden. The patients answered questions about history of dizziness, medications, and comorbidities. They were also investigated for BPPV, vestibular deficits using the video head impulse test (vHIT), walking speed, Timed Up and Go test (TUG), and for perception of verticality and horizontality and Romberg test. Results A total of 55 patients with dizziness were included. Fifteen (27%) were diagnosed with BPPV. Forty (73%) patients reported falling during the previous year, including 11 with BPPV. Dizziness when turning in bed was more common among patients with BPPV and increased the risk of BPPV 8‐fold. Conclusion BPPV is common among older adults with dizziness, including among those not seeking medical care. It is important to identify older adults with BPPV and treat the condition since BPPV may contribute to falls. Asking about dizziness when turning in bed can help to distinguish patients with increased risk for BPPV and older adults with dizziness should be investigated for BPPV even when typical history is lacking. Level of evidence: 4

The neural correlates of locomotion impairments observed in people with Parkinson’s disease (PD) are not fully understood. We investigated whether people with PD present distinct brain electrocortical activity during usual walking and the approach phase of obstacle avoidance when compared to healthy individuals. Fifteen people with PD and fourteen older adults walked overground in two conditions: usual walking and obstacle crossing. Scalp electroencephalography (EEG) was recorded using a mobile 64-channel EEG system. Independent components were clustered using a k-means clustering algorithm. Outcome measures included absolute power in several frequency bands and alpha/beta ratio. During the usual walk, people with PD presented a greater alpha/beta ratio in the left sensorimotor cortex than healthy individuals. While approaching obstacles, both groups reduced alpha and beta power in the premotor and right sensorimotor cortices (balance demand) and increased gamma power in the primary visual cortex (visual demand). Only people with PD reduced alpha power and alpha/beta ratio in the left sensorimotor cortex when approaching obstacles. These findings suggest that PD affects the cortical control of usual walking, leading to a greater proportion of low-frequency (alpha) neuronal firing in the sensorimotor cortex. Moreover, the planning for obstacle avoidance changes the electrocortical dynamics associated with increased balance and visual demands. People with PD rely on increased sensorimotor integration to modulate locomotion.

ObjectiveMoringa oleifera possesses multiple biological effects and the 4-[(4′-O-acetyl-α-L- rhamnosyloxy) benzyl] isothiocyanate accounts for them. Based on the original isothiocyanate molecule we obtained a semisynthetic derivative, named 4-[(2′,3′,4′-O-triacetyl-α-L-rhamnosyloxy) N-benzyl] hydrazine carbothioamide (MC-H) which was safe and effective in a temporomandibular joint (TMJ) inflammatory hypernociception in rats. Therefore, considering that there is still a gap in the knowledge concerning the mechanisms of action through which the MC-H effects are mediated, this study aimed to investigate the involvement of the adhesion molecules (ICAM-1, CD55), the pathways heme oxygenase-1 (HO-1) and NO/cGMP/PKG/K+ATP, and the central opioid receptors in the efficacy of the MC-H in a pre-clinical study of TMJ pain.MethodsMolecular docking studies were performed to test the binding performance of MC-H against the ten targets of interest (ICAM-1, CD55, HO-1, iNOS, soluble cGMP, cGMP-dependent protein kinase (PKG), K+ATP channel, mu (μ), kappa (κ), and delta (δ) opioid receptors). In in vivo studies, male Wistar rats were treated with MC-H 1 μg/kg before TMJ formalin injection and nociception was evaluated. Periarticular tissues were removed to assess ICAM-1 and CD55 protein levels by Western blotting. To investigate the role of HO-1 and NO/cGMP/PKG/K+ATP pathways, the inhibitors ZnPP-IX, aminoguanidine, ODQ, KT5823, or glibenclamide were used. To study the involvement of opioid receptors, rats were pre-treated (15 min) with an intrathecal injection of non-selective inhibitor naloxone or with CTOP, naltrindole, or norbinaltorphimine.ResultsAll interactions presented acceptable binding energy values (below −6.0 kcal/mol) which suggest MC-H might strongly bind to its molecular targets. MC-H reduced the protein levels of ICAM-1 and CD55 in periarticular tissues. ZnPP-IX, naloxone, CTOP, and naltrindole reversed the antinociceptive effect of MC-H.ConclusionMC-H demonstrated antinociceptive and anti-inflammatory effects peripherally by the activation of the HO-1 pathway, as well as through inhibition of the protein levels of adhesion molecules, and centrally by μ and δ opioid receptors.

Abstract Background Patients with chronic kidney disease (CKD) and arterial calcification are considered at increased risk of adverse cardiovascular outcomes. However, the optimal site for measurement of arterial calcification has not been determined. The primary aim of this study was to examine the pattern of arterial calcification in different stages of CKD. Methods This was an observational, cross-sectional study that included 580 individuals with CKD stages 1–5 (no dialysis) from the Copenhagen CKD Cohort. Calcification of the carotid, coronary and iliac arteries, thoracic and abdominal aorta was assessed using non-contrast multidetector computed tomography scans and quantified according to the Agatston method. Based on the distribution of Agatston scores in the selected arterial region, the subjects were divided into calcium score categories of 0 (no calcification), 1–100, 101–400 and > 400. Results Participants with CKD stages 3–5 had the highest prevalence of calcification and the highest frequency of calcium scores > 400 in all arterial sites. Calcification in at least one arterial site was present in > 90% of patients with CKD stage 3. In all five CKD stages prevalence of calcification was greatest in both the thoracic and abdominal aorta, and in the iliac arteries. These arterial sites also showed the highest calcium scores. High calcium scores (> 400) in all five arterial regions were independently associated with prevalent cardiovascular disease. In multivariable analyses, after adjusting for cardiovascular risk factors, declining creatinine clearance was associated with increasing calcification of the coronary arteries (p = 0.012) and the thoracic aorta (p = 0.037) only. Conclusions Arterial calcification is highly prevalent throughout all five CKD stages and is most prominent in both the thoracic and abdominal aorta, and in the iliac arteries. Follow-up studies are needed to explore the potential of extracardiac calcification sites in prediction of cardiovascular events in the CKD population.

Background: Clinical and laboratory assessment of people with mild traumatic brain injury (mTBI) indicate impairments in eye movements. These tests are typically done in a static, seated position. Recently, the use of mobile eye-tracking systems has been proposed to quantify subtle deficits in eye movements and visual sampling during different tasks. However, the impact of mTBI on eye movements during functional tasks such as walking remains unknown.Objective: Evaluate differences in eye-tracking measures collected during gait between healthy controls (HC) and patients in the sub-acute stages of mTBI recovery and to determine if there are associations between eye-tracking measures and gait speed.Methods: Thirty-seven HC participants and 67individuals with mTBI were instructed to walk back and forth over 10-m, at a comfortable self-selected speed. A single 1-min trial was performed. Eye-tracking measures were recorded using a mobile eye-tracking system (head-mounted infra-red Tobbii Pro Glasses 2, 100 Hz, Tobii Technology Inc. VA, United States). Eye-tracking measures included saccadic (frequency, mean and peak velocity, duration and distance) and fixation measurements (frequency and duration). Gait was assessed using six inertial sensors (both feet, sternum, right wrist, lumbar vertebrae and the forehead) and gait velocity was selected as the primary outcome. General linear model was used to compare the groups and association between gait and eye-tracking outcomes were explored using partial correlations.Results: Individuals with mTBI showed significantly reduced saccade frequency (p = 0.016), duration (p = 0.028) and peak velocity (p = 0.032) compared to the HC group. No significant differences between groups were observed for the saccade distance, fixation measures and gait velocity (p > 0.05). A positive correlation was observed between saccade duration and gait velocity only for participants with mTBI (p = 0.025).Conclusion: Findings suggest impaired saccadic eye movement, but not fixations, during walking in individuals with mTBI. These findings have implications in real-world function including return to sport for athletes and return to duty for military service members. Future research should investigate whether or not saccade outcomes are influenced by the time after the trauma and rehabilitation.

The aim of this study was to explore self-schemas and attachment style among patients in a methadone or buprenorphine maintenance treatment program of opiate dependence, in relation to treatment outcome (relapse in substance use). The study included 84 patients (21 women and 63 men) in a psychiatric clinic in Malmö, Sweden, providing maintenance treatment of opiate dependence. Three self-report instruments were employed, Young Schema Questionnaire Short version (YSQ-S) and Young Parenting Inventory (YPI) for studying self-schemas and Experiences in Close Relationships–Relationship Structures questionnaire (ECR-RS) for studying attachment style. Demographical data and relapse in substance abuse were registered. The study demonstrated, unsurprisingly, that an insecure attachment style was more common in the group of patients compared to available general population reference data. Significant correlations were found between attachment style and core beliefs about the self (self-schemas). Memories of parenting experiences from childhood (YPI) showed correlations with ongoing self-schemas (YSQ-S). Treatment outcome, defined as relapses in substance abuse, was associated to a minor degree with self-schemas but showed no correlation with attachment style. Patients who did not work or study had more maladaptive self-schemas and insecure attachment style, and a higher incidence of relapse in abuse than patients who were working or studying.

Despite the initial thrust of research aimed at understanding the impact of the COVID-19 pandemic on youth with physical illness and their parents, knowledge gaps in the literature remain, providing the impetus for additional investigation. This study described changes in psychological distress from prior to during the COVID-19 pandemic for parents and youth with physical illness, compared parent-proxy and youth self-reported perceptions of COVID-19-related psychosocial health, and modeled factors associated with psychological and psychosocial distress. There were 147 parent–youth dyads (2–16 years) from MY LIFE—a longitudinal study of youth with physical illness. The Kessler-6 (K6) measured psychological distress for the time before the COVID-19 lockdown (December 19 to March 20) and during the pandemic (December 20 to March 21) among parents and youth. COVID-19-related psychosocial health was measured using the CRISIS. Parents and youth reported increases in K6 scores (d = 0.62 and 0.38). Parent-proxy reports on the K6 were lower vs. youth self-reports prior to and during the pandemic (d = 0.63 for both). In contrast, parents reported lower proxy CRISIS scores for worries (d = 0.38) and effects of social restrictions (d = 0.52). Pandemic parent K6 scores were associated with age, combined in-person and online schooling for youth, COVID-19-related worries, and effects of social restrictions. For youth, only COVID-19-related worries and effects of social restrictions were associated with K6 scores. Parent worries were associated with youth sex, parental stress, family functioning, online and combination learning, and social restrictions. Parental depression and worries were associated with effects of social restrictions. Youth worries were associated with online and combination learning, and social restrictions. Youth disability, online learning, and worries were associated with effects of social restrictions. Few clinical factors are associated with COVID-19-related psychological and psychosocial distress. Instead, parent/family factors and youth learning environment have prominent roles in predicting outcomes and have implications for the health, education, and social services systems.

Cortical bone microstructure deficits may increase fracture risk in individuals with cardiovascular disease and diabetes. High resolution peripheral quantitative computed tomography (HR-pQCT) enables in vivo microstructure characterization but is limited in its ability to visualize important biological features. We conducted histological analyses and HR-pQCT imaging of distal tibia bone samples from 6 donors with cardiovascular disease (CVD) and type 2 diabetes mellitus (T2D). Histology but not HR-pQCT identified previously undocumented morphopathological deficits that may contribute to cortical bone fragility. These observations may provide guidance for improved HR-pQCT microstructural characterization as well as insight into mechanisms of cortical bone degradation.

As mudanças que o contexto educacional vem passando historicamente aparecem na sociedade contemporânea, sobretudo, no dilema da transição dos documentos norteadores da Educação Básica e em suas concepções de ensino, de sujeito e de alfabetização. Assim, este estudo objetivou fazer uma reflexão teórica e comparativa das concepções de alfabetização no documento Diretrizes Curriculares Nacionais da Educação Básica - DCNs (BRASIL, 2013) com o que se pode encontrar no documento que entrou em vigor em 2019, a Base Nacional Comum Curricular – BNCC (BRASIL, 2019a). Para tanto, buscou-se subsídios teóricos em pesquisadores consagrados da educação, como Duarte (2001), Mortatti (2006, 2007, 2010), Sebra e Dias (2011), entre outros autores, que têm estudado os métodos de alfabetização e sua hibridização na então chamada sociedade do conhecimento.

The current Coronavirus Disease 2019 (COVID-19) pandemic has strained hospital systems and training programs across the world. As capacity issues mount and trainees are called upon to provide frontline medical care, programs and institutions have had to rapidly evolve to redefine the trainee experience. To that end, there is a paucity of literature regarding how healthcare training programs should operate during a global pandemic. Here, we aim to describe twelve evidence-based recommendations for coordinating a cohesive, systematic approach to pandemic response planning for Internal Medicine residency training programs. These tips encompass inpatient and outpatient practices, provider safety, resuscitation, virtual education programming and resident wellbeing. Though many of these considerations or recommendations were not described during the COVID-19 pandemic, these tips have been described previously in the literature, are applicable to the current pandemic and could be easily extrapolated to future crises.

As the population ages and the prevalence of heart failure increases, cardiologists and geriatricians can expect to see more elderly patients with heart failure in their everyday practice. With the advancement of medical care and technology, the options for heart failure management have expanded, though current guidelines are based on studies of younger populations, and the evidence in older populations is not as robust. Pharmacologic therapy remains the cornerstone of heart failure management and has improved long-term mortality. Prevention of sudden cardiac death with implantable devices is being more readily utilized in older patients. Advanced therapies have provided more options for end-stage heart failure, though its use is still limited in older patients. In this review, we discuss the current guidelines for medical management of heart failure in older adults, as well as the expanding literature on advanced therapies, such as heart transplantation in older patients with end-stage heart failure. We also discuss the importance of a multidisciplinary care approach including consideration of non-medical co-morbidities such as frailty and cognitive decline.

Abstract Mesenchymal stromal cells (MSC) secrete factors that contribute to organ homeostasis and repair in a tissue specific manner. For instance, kidney perivascular mesenchymal stromal cells (kPSCs) can facilitate renal epithelial repair through secretion of hepatocyte growth factor (HGF) while the secretome of bone marrow MSCs gives rise to immunosuppression. Stromal cells function in a complex 3-dimensional (3D) connective tissue architecture that induces conformational adaptation. Here we tested the hypothesis that surface topography and associated cell adaptations dictate stromal cell function through tuning of the cytokines released. To this end, we cultured human bone marrow and kidney perivascular stromal cells in the TopoWell plate, a custom-fabricated multi-well plate containing 76 unique bioactive surface topographies. Using fluorescent imaging, we observed profound changes in cell shape, accompanied by major quantitative changes in the secretory capacity of the MSCs. The cytokine secretion profile was closely related to cell morphology and was stromal cell type specific. Our data demonstrate that stromal cell function is determined by microenvironment structure and can be manipulated in an engineered setting. Our data also have implications for the clinical manufacturing of mesenchymal stromal cell therapy, where surface topography during bioreactor expansion should be taken into account to preserve therapeutic properties.

Summary: Human pluripotent stem cell (hPSC)-derived kidney organoids may facilitate disease modeling and the generation of tissue for renal replacement. Long-term application, however, will require transferability between hPSC lines and significant improvements in organ maturation. A key question is whether time or a patent vasculature is required for ongoing morphogenesis. Here, we show that hPSC-derived kidney organoids, derived in fully defined medium conditions and in the absence of any exogenous vascular endothelial growth factor, develop host-derived vascularization. In vivo imaging of organoids under the kidney capsule confirms functional glomerular perfusion as well as connection to pre-existing vascular networks in the organoids. Wide-field electron microscopy demonstrates that transplantation results in formation of a glomerular basement membrane, fenestrated endothelial cells, and podocyte foot processes. Furthermore, compared with non-transplanted organoids, polarization and segmental specialization of tubular epithelium are observed. These data demonstrate that functional vascularization is required for progressive morphogenesis of human kidney organoids. : In this article, Van den Berg and colleagues show that PSC-derived kidney organoids contain nephron structures but remain disorganized and immature after prolonged culture. Upon transplantation, the organoids develop host-derived vascularization, functional glomerular perfusion, and connection to pre-existing vascular networks. The authors conclude that patent vasculature is required for ongoing morphogenesis and maturation of these kidney organoids. Keywords: human pluripotent stem cells, directed differentiation, kidney organoids, transplantation, intravital microscopy, vascularization, maturation

In recent years, a variety of circular replicase-encoding single-stranded (CRESS) DNA viruses and unclassified virus-like DNA elements have been discovered in a broad range of animal species and environmental samples. Key questions to be answered concern their presence in the human diet and their potential impact on disease emergence. Especially DNA elements termed bovine meat and milk factors (BMMF) are suspected to act as co-factors in the development of colon and breast cancer. To expand our knowledge on the occurrence of these potential pathogens in human nutrition, a total of 73 sheep and 40 goat milk samples were assayed by combining rolling circle amplification (RCA), PCR and Sanger sequencing. The present study further includes retail milk from the aforementioned species. We recovered 15 single stranded (ss) circular genomes. Of those, nine belong to the family Genomoviridae and six are members of the unclassified group of BMMF. Thus, dairy sheep and goats add to dispersal of CRESS viruses and circular ssDNA elements, which enter the food chain via milk. The presence of these entities is therefore more widespread in Bovidae than initially assumed and seems to be part of the common human nutrition.

Abstract Pro‐inflammatory cytokines are recognized contributors to intervertebral disc (IVD) degeneration and discogenic pain. We have recently reported the anti‐inflammatory effect of pulsed electromagnetic fields (PEMF) on IVD cells in vitro. Whether these potentially therapeutic effects are sufficiently potent to influence disc health in vivo has not been demonstrated. We report here the effect of PEMF on acute inflammation arising from a rat‐tail IVD injury model. Disc degeneration was induced by percutaneously stabbing the Co6‐7, Co7‐8, and Co8‐9 levels using a 20‐gauge needle. Seventy‐two (72) rats were divided into three groups: sham control, needle stab, needle stab+PEMF. Treated rats were exposed to PEMF immediately following surgery and for either 4 or 7 days (4 hr/d). Stab and PEMF effects were evaluated by measuring inflammatory cytokine gene expression (RT‐PCR) and protein levels (ELISA assay), anabolic and catabolic gene expression (RT‐PCR), and histologic changes. We observed in untreated animals that at day 7 after injury, inflammatory cytokines (interleukin [IL]‐6, tumor necrosis factor α, and IL‐1β) were significantly increased at both gene and protein levels (P

Wasps can synthesize chemical compounds called venom whose function is to overcome prey and assist in defense of the colonies. Geographic Parameters such as sex, age, the season of the year, and diet determined the composition of the venom location, genetics, environment. However, studies on the compositional variability of venom are still limited due to the difficulty in obtaining samples and the complexity of these substances. This work describes the use of the Fourier Transform Infrared Photoacoustic Spectroscopy (FTIR-PAS) to investigate inter- and intraspecific variability in the venom chemical composition (VCC) of the social wasp Polybia paulista (Von Ihering 1896) and the ant Ectatomma brunneum (Smith 1858). The results reveal significant differences in VCC among the ant and wasp, even for samples obtained from the same environment. The genetic component, therefore, seemed to be the predominant factor determining the compounds present. The findings also showed that exogenous factors, such as diet, could also be responsible for intraspecific differences, especially in wasps. The FTIR-PAS technique proved to be a reliable way of assessing intra- and interspecific differences in social Hymenoptera VCC.

The integrity of ant colonies depends exclusively on social relationships between their individuals, especially the ability of communication between group members, which is mainly mediated through chemical signals. Another important feature of ant behavior is territory defense, since they need to gather large amounts of food to feed their larvae, males and breeding females. Thereby, ants might display behavioral strategies to defend their territories from intruders. Here we investigated whether Ectatomma brunneum displays the Dear Enemy Phenomenon, what is the relationship between Cuticular Hydrocarbon composition and levels of aggression during their intraspecific interactions and which compounds and/or classes of compounds might be the most important to modulate the level of aggression. To test our hypothesis, we evaluated the levels of aggression through behavioral observations during interactions between 23 pairs of colonies nested in two distinct sites at varied distances. Then, we analyzed the cuticular chemical profile of the individuals involved in the interactions, and compared these results with the levels of aggression displayed between colonies tested. The results allow us to confirm our hypothesis that the DEP occurs in E. brunneum. The higher tolerance between closer colonies can be explained due to their kinship level in addition to sharing the same microhabitats. The results also showed there are significant differences in CHCs profiles, especially between colonies nested at relatively greater distances, and it is likely that differences in content of some branched alkanes are the most important to establish these differences and, therefore, the levels of aggression during the interactions.

There is an increasing prevalence of chronic kidney disease (CKD), which associates with the development of interstitial fibrosis. Pericytes (perivascular fibroblasts) provide a major source of α-SMA-positive myofibroblasts that are responsible for the excessive deposition of extracellular matrix. In order to identify pericyte long non-coding RNAs (lncRNAs) that could serve as a target to decrease myofibroblast formation and counteract the progression of kidney fibrosis we employed two models of experimental kidney injury, one focused on kidney fibrosis (unilateral ureteral obstruction; UUO), and one focused on acute kidney injury that yields kidney fibrosis in the longer term (unilateral ischemia-reperfusion injury; IRI). This was performed in FoxD1-GC;tdTomato stromal cell reporter mice that allowed pericyte fate tracing. Tomato red-positive FoxD1-derivative cells of control and injured kidneys were FACS-sorted and used for lncRNA and mRNA profiling yielding a distinctive transcriptional signature of pericytes and myofibroblasts with 244 and 586 differentially expressed lncRNAs (>twofold, P < 0.05), in the UUO and IRI models, respectively. Next, we selected two differentially expressed and conserved lncRNAs, Rian (RNA imprinted and accumulated in nucleus) and Miat (Myocardial infarction associated transcript), and explored their potential regulatory role in myofibroblast formation through knockdown of their function with gapmers. While Miat was upregulated in myofibroblasts of UUO and IRI in mice, gapmer silencing of Miat attenuated myofibroblast formation as evidenced by decreased expression of α-SMA, col1α1, SMAD2, and SMAD3, as well as decreased α-SMA and pro-collagen-1α1 protein levels. In contrast, silencing Rian, which was found to be downregulated in kidney myofibroblast after IRI and UUO, resulted in increased myofibroblast formation. In addition, we found microRNAs that were previously linked to Miat (miR-150) and Rian (14q32 miRNA cluster), to be dysregulated in the FoxD1-derivative cells, suggesting a possible interaction between miRNAs and these lncRNAs in myofibroblast formation. Taken together, lncRNAs play a regulatory role in myofibroblast formation, possibly through interacting with miRNA regulation, implicating that understanding their biology and their modulation may have the potential to counteract the development of renal fibrosis.