Altered Immunoglobulin Repertoire and Decreased IgA Somatic Hypermutation in the Gut during Chronic HIV-1 Infection.

Humoral immune perturbations contribute to pathogenic outcomes in persons with HIV-1 infection (PWH). Gut barrier dysfunction in PWH is associated with microbial translocation and alterations in microbial communities (dysbiosis), and IgA, the most abundant immunoglobulin (Ig) isotype in the gut, is involved in gut homeostasis by interacting with the microbiome. We determined the impact of HIV-1 infection on the antibody repertoire in the gastrointestinal tract by comparing Ig gene utilization and somatic hypermutation (SHM) in colon biopsies from PWH (n = 19) versus age and sex-matched controls (n = 13). We correlated these Ig parameters with clinical, immunological, microbiome and virological data. Gene signatures of enhanced B cell activation were accompanied by skewed frequencies of multiple Ig Variable genes in PWH. PWH showed decreased frequencies of SHM in IgA and possibly IgG, with a substantial loss of highly mutated IgA sequences. The decline in IgA SHM in PWH correlated with gut CD4+ T cell loss and inversely correlated with mucosal inflammation and microbial translocation. Diminished gut IgA SHM in PWH was driven by transversion mutations at A or T deoxynucleotides, suggesting a defect not at the AID/APOBEC3 deamination step but at later stages of IgA SHM. These results expand our understanding of humoral immune perturbations in PWH that could have important implications in understanding mucosal immune defects in individuals with chronic HIV-1 infection. IMPORTANCE The gut is a major site of early HIV-1 replication and pathogenesis. Extensive CD4+ T cell depletion in this compartment results in a compromised epithelial barrier that facilitates the translocation of microbes into the underlying lamina propria and systemic circulation, resulting in chronic immune activation. To date, the consequences of microbial translocation on the mucosal humoral immune response (or vice versa) remains poorly integrated into the panoply of mucosal immune defects in PWH. We utilized next-generation sequencing approaches to profile the Ab repertoire and ascertain frequencies of somatic hypermutation in colon biopsies from antiretroviral therapy-naive PWH versus controls. Our findings identify perturbations in the Ab repertoire of PWH that could contribute to development or maintenance of dysbiosis. Moreover, IgA mutations significantly decreased in PWH and this was associated with adverse clinical outcomes. These data may provide insight into the mechanisms underlying impaired Ab-dependent gut homeostasis during chronic HIV-1 infection.

Qualitative Differences Between the IFNα subtypes and IFNß Influence Chronic Mucosal HIV-1 Pathogenesis.

The Type I Interferons (IFN-Is) are innate antiviral cytokines that include 12 different IFNα subtypes and IFNß that signal through the IFN-I receptor (IFNAR), inducing hundreds of IFN-stimulated genes (ISGs) that comprise the 'interferome'. Quantitative differences in IFNAR binding correlate with antiviral activity, but whether IFN-Is exhibit qualitative differences remains controversial. Moreover, the IFN-I response is protective during acute HIV-1 infection, but likely pathogenic during the chronic stages. To gain a deeper understanding of the IFN-I response, we compared the interferomes of IFNα subtypes dominantly-expressed in HIV-1-exposed plasmacytoid dendritic cells (1, 2, 5, 8 and 14) and IFNß in the earliest cellular targets of HIV-1 infection. Primary gut CD4 T cells from 3 donors were treated for 18 hours ex vivo with individual IFN-Is normalized for IFNAR signaling strength. Of 1,969 IFN-regulated genes, 246 'core ISGs' were induced by all IFN-Is tested. However, many IFN-regulated genes were not shared between the IFNα subtypes despite similar induction of canonical antiviral ISGs such as ISG15, RSAD2 and MX1, formally demonstrating qualitative differences between the IFNα subtypes. Notably, IFNß induced a broader interferome than the individual IFNα subtypes. Since IFNß, and not IFNα, is upregulated during chronic HIV-1 infection in the gut, we compared core ISGs and IFNß-specific ISGs from colon pinch biopsies of HIV-1-uninfected (n = 13) versus age- and gender-matched, antiretroviral-therapy naïve persons with HIV-1 (PWH; n = 19). Core ISGs linked to inflammation, T cell activation and immune exhaustion were elevated in PWH, positively correlated with plasma lipopolysaccharide (LPS) levels and gut IFNß levels, and negatively correlated with gut CD4 T cell frequencies. In sharp contrast, IFNß-specific ISGs linked to protein translation and anti-inflammatory responses were significantly downregulated in PWH, negatively correlated with gut IFNß and LPS, and positively correlated with plasma IL6 and gut CD4 T cell frequencies. Our findings reveal qualitative differences in interferome induction by diverse IFN-Is and suggest potential mechanisms for how IFNß may drive HIV-1 pathogenesis in the gut.

tidyMicro: a pipeline for microbiome data analysis and visualization using the tidyverse in R.

BACKGROUND: The drive to understand how microbial communities interact with their environments has inspired innovations across many fields. The data generated from sequence-based analyses of microbial communities typically are of high dimensionality and can involve multiple data tables consisting of taxonomic or functional gene/pathway counts. Merging multiple high dimensional tables with study-related metadata can be challenging. Existing microbiome pipelines available in R have created their own data structures to manage this problem. However, these data structures may be unfamiliar to analysts new to microbiome data or R and do not allow for deviations from internal workflows. Existing analysis tools also focus primarily on community-level analyses and exploratory visualizations, as opposed to analyses of individual taxa. RESULTS: We developed the R package "tidyMicro" to serve as a more complete microbiome analysis pipeline. This open source software provides all of the essential tools available in other popular packages (e.g., management of sequence count tables, standard exploratory visualizations, and diversity inference tools) supplemented with multiple options for regression modelling (e.g., negative binomial, beta binomial, and/or rank based testing) and novel visualizations to improve interpretability (e.g., Rocky Mountain plots, longitudinal ordination plots). This comprehensive pipeline for microbiome analysis also maintains data structures familiar to R users to improve analysts' control over workflow. A complete vignette is provided to aid new users in analysis workflow. CONCLUSIONS: tidyMicro provides a reliable alternative to popular microbiome analysis packages in R. We provide standard tools as well as novel extensions on standard analyses to improve interpretability results while maintaining object malleability to encourage open source collaboration. The simple examples and full workflow from the package are reproducible and applicable to external data sets.

Integrating the Practical Robust Implementation and Sustainability Model With Best Practices in Clinical Decision Support Design: Implementation Science Approach.

BACKGROUND: Clinical decision support (CDS) design best practices are intended to provide a narrative representation of factors that influence the success of CDS tools. However, they provide incomplete direction on evidence-based implementation principles. OBJECTIVE: This study aims to describe an integrated approach toward applying an existing implementation science (IS) framework with CDS design best practices to improve the effectiveness, sustainability, and reproducibility of CDS implementations. METHODS: We selected the Practical Robust Implementation and Sustainability Model (PRISM) IS framework. We identified areas where PRISM and CDS design best practices complemented each other and defined methods to address each. Lessons learned from applying these methods were then used to further refine the integrated approach. RESULTS: Our integrated approach to applying PRISM with CDS design best practices consists of 5 key phases that iteratively interact and inform each other: multilevel stakeholder engagement, designing the CDS, design and usability testing, thoughtful deployment, and performance evaluation and maintenance. The approach is led by a dedicated implementation team that includes clinical informatics and analyst builder expertise. CONCLUSIONS: Integrating PRISM with CDS design best practices extends user-centered design and accounts for the multilevel, interacting, and dynamic factors that influence CDS implementation in health care. Integrating PRISM with CDS design best practices synthesizes the many known contextual factors that can influence the success of CDS tools, thereby enhancing the reproducibility and sustainability of CDS implementations. Others can adapt this approach to their situation to maximize and sustain CDS implementation success.

A longitudinal intervention to improve young children's liking and consumption of new foods: findings from the Colorado LEAP study.

BACKGROUND: Many interventions have been conducted to improve young children's liking and consumption of new foods however their impacts on children's consumption have been limited. Consistent evidence supports the use of repeated exposure to improve liking for new foods however longitudinal effects lasting greater than 6 months often have not been demonstrated. Here we report the eating-related findings of the Colorado Longitudinal Eating And Physical Activity (LEAP) Study, a multi-component intervention, delivered primarily in the school setting, which aimed to improve children's liking and consumption of a target food via repeated exposure and positive experiential learning. METHODS: Four sites in rural Colorado, each housing Head Start preschool programs, matched on state vital statistics for childhood obesity rates, (2 intervention and 2 control sites) took part in a quasi-experimental study design which included 4 time points (baseline, post-intervention, one-year [Y1] and two- year [Y2] follow ups). A total of 250 children and families were enrolled (n = 143 intervention and n = 107 control; 41% Hispanic and 69% low-income). A 12-week intervention, Food Friends - Fun With New Foods®, delivered by trained preschool teachers and which focuses on positive and repeated experiences with new foods, and a 5-month (1 unit/month) social marketing "booster program" was delivered in kindergarten (one-year follow up) and 1st grade (two-year follow up). Main outcome measures included change in children's liking for new foods, analyzed by ordinal regression using generalized estimating equations, and change in weighed consumption of new foods over time, analyzed using a hierarchical mixed effects model. RESULTS: The intervention was delivered with good fidelity (87%). Both intervention and control groups demonstrated an increase in liking for the target food over time (p = 0.0001). The pattern of consumption of the target food was different, over time, for intervention and control groups (p < 0.005). In particular the change in intake between baseline and post-intervention was significantly greater in the intervention compared to the control group (p < 0.0001) though this pattern of change did not hold between baseline and Y2 follow up (p = 0.1144). Children in the intervention group who liked the target food consumed nearly double their baseline consumption at post-intervention (p < 0.0001;) and maintained this increase at Y2 follow up (p < 0.0001). CONCLUSIONS: The Food Friends intervention, which utilized positive, repeated experiences with new foods, and was delivered with good fidelity by trained preschool teachers, found that larger improvements were observed in children's eating behaviors than would be expected with developmentally-based changes in eating behaviors. TRIAL REGISTRATION NUMBER: This trial is registered at ClinicalTrials.gov : NCT01937481. Date registered: 09/09/2013; Retrospectively registered. Date first participant registered: 09/15/2010.

Factors influencing the acceptance of referrals for clinical pharmacist managed disease states in primary care.

OBJECTIVE: Clinical pharmacists use population health methods to generate chronic disease management referrals for patients with uncontrolled chronic conditions. The purpose of this study was to compare primary care providers' (PCPs) referral responses for 4 pharmacist-managed indications and to identify provider and patient characteristics that are predictive of PCP response. DESIGN: Retrospective cohort study. SETTING: This study occurred in an academic internal medicine clinic. PARTICIPANTS: Clinical pharmacy referrals generated through a population health approach between 2012 and 2016 for hypertension, chronic pain, depression, and benzodiazepine management were included. MAIN OUTCOME MEASURES: Proportion of referrals accepted, left pending, or rejected and influencing provider and patient characteristics. RESULTS: Of 1769 referrals generated, PCPs accepted 869 (49%), left pending 300 (17%), and rejected 600 (34%). Compared with referrals for hypertension, benzodiazepine management, and depression, chronic pain referrals had the lowest likelihood of rejection (odds ratio [OR] 0.31; 95% CI 0.19-0.49). Depression referrals had an equal likelihood of being accepted or rejected (OR 1.04; 95% CI 0.66-1.64). Provider characteristics were not significantly associated with referral response, but residents were more likely to accept referrals. Patient characteristics associated with lower referral rejection included black race (OR 0.39; 95% CI 0.18-0.87), higher systolic blood pressure (OR 0.98; 95% CI 0.97-0.99), and missed visits (OR 0.24; 95% CI 0.07-0.81). CONCLUSION: The majority of referrals for clinical pharmacists in primary care settings were responded to, varying mostly between acceptance and rejection. There was variability in referral acceptance across indications, and some patient characteristics were associated with increased referral acceptance.

Comparative Immune Responses to Licensed Herpes Zoster Vaccines.

Background: Live attenuated (ZV) and recombinant adjuvanted (HZ/su) zoster vaccines differ with respect to efficacy, effect of age, and persistence of protection. We compared cell-mediated immunity (CMI responses to ZV and HZ/su. Methods: This was a randomized, double-blind, placebo-controlled trial stratified by age (50-59 and 70-85 years) and by HZ vaccination status (received ZV ≥5 years before entry or not). Varicella zoster virus (VZV)- and glycoprotein E (gE)-specific CMI were analyzed by interleukin 2 (IL-2) and interferon gamma (IFN-γ) FluoroSpot and flow cytometry at study days 0, 30, 90, and 365. Results: Responses to ZV peaked on day 30 and to HZ/su (administered in 2 doses separated by 60 days) peaked on day 90. Age and vaccination status did not affect peak responses, but higher baseline CMI correlated with higher peak responses. HZ/su generated significantly higher VZV-specific IL-2+ and gE-specific IL-2+, IFN-γ+, and IL-2+/IFN-γ+ peak and 1-year baseline-adjusted responses compared with ZV. VZV-specific IFN-γ+ and IL-2+/IFN-γ+ did not differ between vaccines. HZ/su generated higher memory and effector-memory CD4+ peak responses and ZV generated higher effector CD4+ responses . Conclusions: The higher IL-2 and other memory responses generated by HZ/su compared with ZV may contribute to its superior efficacy. Clinical Trials Registration: NCT02114333.

Examining the role of unmeasured confounding in mediation analysis with genetic and genomic applications.

BACKGROUND: In mediation analysis if unmeasured confounding is present, the estimates for the direct and mediated effects may be over or under estimated. Most methods for the sensitivity analysis of unmeasured confounding in mediation have focused on the mediator-outcome relationship. RESULTS: The Umediation R package enables the user to simulate unmeasured confounding of the exposure-mediator, exposure-outcome, and mediator-outcome relationships in order to see how the results of the mediation analysis would change in the presence of unmeasured confounding. We apply the Umediation package to the Genetic Epidemiology of Chronic Obstructive Pulmonary Disease (COPDGene) study to examine the role of unmeasured confounding due to population stratification on the effect of a single nucleotide polymorphism (SNP) in the CHRNA5/3/B4 locus on pulmonary function decline as mediated by cigarette smoking. CONCLUSIONS: Umediation is a flexible R package that examines the role of unmeasured confounding in mediation analysis allowing for normally distributed or Bernoulli distributed exposures, outcomes, mediators, measured confounders, and unmeasured confounders. Umediation also accommodates multiple measured confounders, multiple unmeasured confounders, and allows for a mediator-exposure interaction on the outcome. Umediation is available as an R package at https://github.com/SharonLutz/Umediation A tutorial on how to install and use the Umediation package is available in the Additional file 1.

Anchoring in Destination-Therapy Left Ventricular Assist Device Decision Making: A Mechanical Turk Survey.

BACKGROUND: People with end-stage heart failure may have to decide about destination-therapy left ventricular assist device (DT-LVAD). Individuals facing difficult decisions often rely on heuristics, such as anchoring, which predictably bias decision outcomes. We aimed to investigate whether showing a larger historical Heartmate XVE creates an anchoring effect, making the smaller Heartmate II (HMII) appear more favorable. METHODS: With the use of Amazon Mechanical Turk, participants watched videos asking them to imagine themselves dying of end-stage heart failure, then were presented the option of LVAD as potentially life-prolonging therapy. Participants were randomized to a control group who were only shown the HMII device, and the intervention group who saw the XVE device before the HMII. Participants then completed surveys. RESULTS: A total of 487 participants completed the survey (control = 252; intervention = 235); 79% were <40 years of age, 84% were white, and 55% were male. The intervention group was not more likely to accept the LVAD therapy (68% vs 61%; P = .37). However, participants in the intervention group were more likely (51% vs 17%; P < .01) to agree or strongly agree with the statement that the HMII was "smaller than expected." Participants in the intervention group were also more likely to rate the size of the device as "important" or "very important" in their decision (61% vs 46%; P < .01). CONCLUSIONS: Although the XVE anchor did not affect likelihood of accepting the LVAD, it did affect device perception. This article highlights an important point with clinical implications: factors such as anchoring have the potential to inappropriately influence perceptions and decisions and should be carefully considered in research and practice.

Sugar intake is associated with progression from islet autoimmunity to type 1 diabetes: the Diabetes Autoimmunity Study in the Young.

AIMS/HYPOTHESIS: Dietary sugar intake may increase insulin production, stress the beta cells and increase the risk for islet autoimmunity (IA) and subsequent type 1 diabetes. METHODS: Since 1993, the Diabetes Autoimmunity Study in the Young (DAISY) has followed children at increased genetic risk for type 1 diabetes for the development of IA (autoantibodies to insulin, GAD or protein tyrosine phosphatase-like protein [IA2] twice or more in succession) and progression to type 1 diabetes. Information on intake of fructose, sucrose, total sugars, sugar-sweetened beverages, beverages with non-nutritive sweetener and juice was collected prospectively throughout childhood via food frequency questionnaires (FFQs). We examined diet records for 1,893 children (mean age at last follow-up 10.2 years); 142 developed IA and 42 progressed to type 1 diabetes. HLA genotype was dichotomised as high risk (HLA-DR3/4,DQB1*0302) or not. All Cox regression models were adjusted for total energy, FFQ type, type 1 diabetes family history, HLA genotype and ethnicity. RESULTS: In children with IA, progression to type 1 diabetes was significantly associated with intake of total sugars (HR 1.75, 95% CI 1.07-2.85). Progression to type 1 diabetes was also associated with increased intake of sugar-sweetened beverages in those with the high-risk HLA genotype (HR 1.84, 95% CI 1.25-2.71), but not in children without it (interaction p value = 0.02). No sugar variables were associated with IA risk. CONCLUSIONS/INTERPRETATION: Sugar intake may exacerbate the later stage of type 1 diabetes development; sugar-sweetened beverages may be especially detrimental to children with the highest genetic risk of developing type 1 diabetes.

Timing of solid food introduction is associated with urinary F2-isoprostane concentrations in childhood.

BACKGROUND: Timing of solid food introduction in infancy has been associated with several chronic diseases. To explore potential mechanisms, we investigated the relationship between timing of solid food introduction and F2-isoprostanes-a marker of oxidative stress. METHODS: Urinary F2-isoprostanes were assessed in 336 healthy children aged less than 11.5 y with 1,266 clinic visits (mean = 3.8 visits per child) in the Diabetes Autoimmunity Study in the Young. We analyzed the association between F2-isoprostane concentrations and infant diet exposures using linear mixed models adjusted for age, age(2), HLA-DR3/4,DQB1*0302 genotype, first-degree relative with type 1 diabetes, maternal age, maternal education, sex, and exposure to in utero cigarette smoke. RESULTS: Later solid food introduction was associated with lower F2-isoprostane concentrations in childhood (on average, 0.10 ng/mg per month of age at introduction; estimate: -0.10 (95% confidence interval (CI): -0.18, -0.02) P value = 0.02). Moreover, childhood F2-isoprostane concentrations were, on average, 0.24 ng/mg lower in individuals breastfed at solid food introduction (estimate: -0.24 (95% CI: -0.47, -0.01) P value = 0.04) compared with those who were not. Associations remained significant after limiting analyses to F2-isoprostanes after 2 y of age. CONCLUSION: Our results suggest a long-term protective effect of later solid food introduction and breastfeeding at solid food introduction against increased F2-isoprostane concentrations throughout childhood.

The effect of childhood cow's milk intake and HLA-DR genotype on risk of islet autoimmunity and type 1 diabetes: the Diabetes Autoimmunity Study in the Young.

BACKGROUND: Cow's milk intake has been inconsistently associated with islet autoimmunity (IA) and type 1 diabetes (T1D) development. Genetic and environmental factors may modify the effect of cow's milk on IA and T1D risk. METHODS: The Diabetes Autoimmunity Study in the Young (DAISY) follows children at increased T1D risk of IA (presence of autoantibodies to insulin, GAD65, or IA-2 twice in succession) and T1D development. We examined 1835 DAISY children with data on cow's milk intake: 143 developed IA, 40 subsequently developed T1D. Cow's milk protein and lactose intake were calculated from prospectively collected parent- and self-reported food frequency questionnaires (FFQ). High risk HLA-DR genotype: HLA-DR3/4,DQB1*0302; low/moderate risk: all other genotypes. We examined interactions between cow's milk intake, age at cow's milk introduction, and HLA-DR genotype in IA and T1D development. Interaction models contained the base terms (e.g., cow's milk protein and HLA-DR genotype) and an interaction term (e.g., cow's milk protein*HLA-DR genotype). RESULTS: In survival models adjusted for total calories, FFQ type, T1D family history, and ethnicity, greater cow's milk protein intake was associated with increased IA risk in children with low/moderate risk HLA-DR genotypes [hazard ratio (HR): 1.41, 95% confidence interval (CI): 1.08-1.84], but not in children with high risk HLA-DR genotypes. Cow's milk protein intake was associated with progression to T1D (HR: 1.59, CI: 1.13-2.25) in children with IA. CONCLUSIONS: Greater cow's milk intake may increase risk of IA and progression to T1D. Early in the T1D disease process, cow's milk intake may be more influential in children with low/moderate genetic T1D risk.

Erythrocyte membrane docosapentaenoic acid levels are associated with islet autoimmunity: the Diabetes Autoimmunity Study in the Young.

AIMS/HYPOTHESES: We previously reported that lower n-3 fatty acid intake and levels in erythrocyte membranes were associated with increased risk of islet autoimmunity (IA) but not progression to type 1 diabetes in children at increased risk for diabetes. We hypothesise that specific n-3 fatty acids and genetic markers contribute synergistically to this increased risk of IA in the Diabetes Autoimmunity Study in the Young (DAISY). METHODS: DAISY is following 2,547 children at increased risk for type 1 diabetes for the development of IA, defined as being positive for glutamic acid decarboxylase (GAD)65, IA-2 or insulin autoantibodies on two consecutive visits. Using a case-cohort design, erythrocyte membrane fatty acids and dietary intake were measured prospectively in 58 IA-positive children and 299 IA-negative children. RESULTS: Lower membrane levels of the n-3 fatty acid, docosapentaenoic acid (DPA), were predictive of IA (HR 0.23; 95% CI 0.09, 0.55), while α-linolenic acid (ALA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were not, adjusting for HLA and diabetes family history. We examined whether the effect of dietary intake of the n-3 fatty acid ALA on IA risk was modified by fatty acid elongation and desaturation genes. Adjusting for HLA, diabetes family history, ethnicity, energy intake and questionnaire type, ALA intake was significantly more protective for IA in the presence of an increasing number of minor alleles at FADS1 rs174556 (pinteraction = 0.017), at FADS2 rs174570 (pinteraction = 0.016) and at FADS2 rs174583 (pinteraction = 0.045). CONCLUSIONS/INTERPRETATION: The putative protective effect of n-3 fatty acids on IA may result from a complex interaction between intake and genetically controlled fatty acid desaturation.

Evidence of stage- and age-related heterogeneity of non-HLA SNPs and risk of islet autoimmunity and type 1 diabetes: the diabetes autoimmunity study in the young.

Previously, we examined 20 non-HLA SNPs for association with islet autoimmunity (IA) and/or progression to type 1 diabetes (T1D). Our objective was to investigate fourteen additional non-HLA T1D candidate SNPs for stage- and age-related heterogeneity in the etiology of T1D. Of 1634 non-Hispanic white DAISY children genotyped, 132 developed IA (positive for GAD, insulin, or IA-2 autoantibodies at two or more consecutive visits); 50 IA positive children progressed to T1D. Cox regression was used to analyze risk of IA and progression to T1D in IA positive children. Restricted cubic splines were used to model SNPs when there was evidence that risk was not constant with age. C1QTNF6 (rs229541) predicted increased IA risk (HR: 1.57, CI: 1.20-2.05) but not progression to T1D (HR: 1.13, CI: 0.75-1.71). SNP (rs10517086) appears to exhibit an age-related effect on risk of IA, with increased risk before age 2 years (age 2 HR: 1.67, CI: 1.08-2.56) but not older ages (age 4 HR: 0.84, CI: 0.43-1.62). C1QTNF6 (rs229541), SNP (rs10517086), and UBASH3A (rs3788013) were associated with development of T1D. This prospective investigation of non-HLA T1D candidate loci shows that some SNPs may exhibit stage- and age-related heterogeneity in the etiology of T1D.

Approaches to analyzing binary data for large-scale A/B testing.

An industry-academic collaboration was established to evaluate the choice of statistical test and study design for A/B testing in larger-scale industry experiments. Specifically, the standard approach at the industry partner was to apply a t-test for all outcomes, both continuous and binary, and to apply naïve interim monitoring strategies that had not evaluated the potential implications on operating characteristics such as power and type I error rates. Although many papers have summarized the robustness of the t-test, its performance for the A/B testing context of large-scale proportion data, with or without interim analyses, is needed. Investigating the effect of interim analyses on the robustness of the t-test is important, because interim analyses rely on a fraction of the total sample size and one should ensure that desired properties are maintained when a t-test is implemented not just at the end of the study, but for making interim decisions. Through simulation studies, the performance of the t-test, Chi-squared test, and Chi-squared test with Yate's correction when applied to binary outcomes data is evaluated. Further, interim monitoring through a naïve approach with no correction for multiple testing versus the O'Brien-Fleming boundary are considered in designs that allow early termination for futility, difference, or both. Results indicate that the t-test achieves similar power and type I error rates for binary outcomes data with the large sample sizes used in industrial A/B tests with and without interim monitoring, and naïve interim monitoring without corrections leads to poorly performing studies.

Examination of Sex Differences in a Chronic Rhinosinusitis Surgical Cohort.

OBJECTIVE: Sex discrepancies have been reported in chronic rhinosinusitis (CRS), but limited data exist exploring sex-specific biological processes and sinonasal quality of life. STUDY DESIGN: Prospective cohort. SETTING: Academic medical center. METHODS: Demographics, clinical data, and sinonasal mucus were collected from patients with CRS presenting for surgical consideration over a 5-year period. A random forest model and linear regression were used to assess predictor variables for the 22-item Sino-Nasal Outcome Test (SNOT-22) and subdomains. Enzyme-linked immunosorbent assays were used to measure substance P and tryptase in a subset of mucus samples to explore biological differences by sex. RESULTS: In total, 520 patients were studied (mean age 48.3 years, 50.9% female). Males were older (50.1 vs 46.6 years, P = .008), had more polyp disease (48.2% vs 35.5%, P = .004), and had higher mean Lund-Mackay CT score (11.3 vs 9.5, P = .004). Females had a higher overall mean SNOT-22 (40.9 vs 46.9, P = .001) and higher scores in ear/facial, psychological, and sleep domains (P < .01). Age, objective disease measures, and sex were top predictors for total SNOT-22. Neither mucus substance P or tryptase, alone or paired with sex, correlated with total SNOT-22. Analysis of mucus biomarkers by sex revealed correlation between mucus tryptase in females with the extranasal subdomain (P = .01). CONCLUSION: Sex differences exist in CRS disease manifestations and presentation for surgical consideration. Detection of mucus (substance P and tryptase) was reliable, but in this exploratory study, we were not able to establish neurogenic or allergic inflammatory processes as a large source of differential disease features between sexes.

Association of Sputum Neutrophil Extracellular Trap Subsets With IgA Anti-Citrullinated Protein Antibodies in Subjects at Risk for Rheumatoid Arthritis.

OBJECTIVE: Mechanisms leading to anti-citrullinated protein antibody (ACPA) generation in rheumatoid arthritis (RA) are hypothesized to originate in the lung. We undertook this study to understand associations between neutrophil extracellular trap (NET) formation in the lung and local ACPA generation in subjects at risk of developing RA. METHODS: Induced sputum was collected from 49 subjects at risk of developing RA, 12 patients with RA, and 18 controls. Sputum neutrophils were tested for ex vivo NET formation, and sputum-induced NET formation of control neutrophils was measured using immunofluorescence imaging. Sputum macrophages were tested for ex vivo endocytosis of apoptotic and opsonized cells. Levels of ACPA, NET remnants, and inflammatory proteins were quantified in sputum supernatant. RESULTS: Spontaneous citrullinated histone H3 (Cit-H3)-expressing NET formation was higher in sputum neutrophils from at-risk subjects and RA patients compared to controls (median 12%, 22%, and 0%, respectively; P < 0.01). In at-risk subjects, sputum IgA ACPA correlated with the percentage of neutrophils that underwent Cit-H3+ NET formation (r = 0.49, P = 0.002) and levels of Cit-H3+ NET remnants (r = 0.70, P < 0.001). Reduced endocytic capacity of sputum macrophages was found in at-risk subjects and RA patients compared to controls. Using a mediation model, we found that sputum inflammatory proteins were associated with sputum IgA ACPA through a pathway mediated by Cit-H3+ NET remnants. Sputum-induced Cit-H3+ NET formation also correlated with sputum levels of interleukin-1ß (IL-1ß), IL-6, and tumor necrosis factor in at-risk subjects, suggesting a causal relationship. CONCLUSION: These data support a potential mechanism for mucosal ACPA generation in subjects at risk of developing RA, whereby inflammation leads to increased citrullinated protein-expressing NETs that promote local ACPA generation.

Depression-related stigma among primary care providers.

INTRODUCTION: Depression is one of the most common mental illnesses in the United States and is often treated in primary care settings. Despite its prevalence, depression remains underdiagnosed and undertreated for a variety of reasons, including stigma. This may result in suboptimal management of depression. Studies evaluating stigma in US primary care providers (PCP) are scarce. The main objective of this study was to describe stigma in a cohort of PCPs. METHODS: We utilized a validated questionnaire to measure stigma (score range 15 to 75 with lower scores indicating lower stigma levels). PCPs in 2 academic internal medicine clinics were sent an electronic questionnaire and received a small monetary incentive for responding. In addition to the stigma survey, we collected demographic data, including age, provider type, gender, and other data related to social proximity to mental illness. To describe stigma, differences in stigma between provider characteristics were evaluated using t tests and ANOVA tests as appropriate. RESULTS: Of 107 PCPs, 71 responded (66.4% response rate). Male responders displayed higher stigma scores than females (31.8 vs 27.4, P = .0021). Medical residents displayed higher stigma scores than nonresidents (31.3 vs 27.2, P = .0045). Providers with personal exposure to mental illness and those who reported they frequently treated depression had less stigma. DISCUSSION: Overall, a range of stigma was present among PCPs surveyed. Higher levels of stigma were found in men, medical residents, those without personal exposure to mental illness, younger PCPs, and those who reported treating depression less frequently. Future studies should utilize larger sample sizes and focus on the impact of stigma on quality of care.

Patient Outreaches for Clinical Pharmacy Services: A Population Health Management Program Assessment.

BACKGROUND: Pharmacists in ambulatory care can utilize population health approaches to identify patients needing disease management and improve outcomes. However, population health is only effective when identified patients are successfully outreached and show to appointments. OBJECTIVE: Describe a population health approach utilized by pharmacists in primary care, report outcomes of outreach attempts and scheduled appointments, and determine whether patient and referral characteristics predict no-show appointments. METHODS: Retrospective cohort study of patients referred for pharmacist management of hypertension or chronic pain through population health between 2013-2016. Outreach attempt and appointments outcomes were collected. Patient and referral characteristics were analyzed to determine whether predictive of no-show appointments using logistic regression. RESULTS: Of 450 outreach attempts, 250 (56%) patients were not reached, 164 (36%) scheduled appointments, and 36 (8%) were reached but declined an appointment. Of 164 patients with appointments, 71 (43%) no-showed. Patients with higher systolic blood pressure were more likely to no-show (OR: 1.02, 95% CI: 1.00-1.04). Other characteristics were not predictive of no-show appointments. CONCLUSION: Successful outreach and showed appointments are essential components of successful population health programs. Using multiple outreach modalities and further identifying factors predictive of no-show appointments to refine the current approach may lead to increased efficiency.