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The nuclear long non-coding RNA LUCAT1 has previously been identified as a negative feedback regulator of type I interferon and inflammatory cytokine expression in human myeloid cells. Here, we define the mechanistic basis for the suppression of inflammatory gene expression by LUCAT1. Using comprehensive identification of RNA-binding proteins by mass spectrometry as well as RNA immunoprecipitation, we identified proteins important in processing and alternative splicing of mRNAs as LUCAT1-binding proteins. These included heterogeneous nuclear ribonucleoprotein C, M, and A2B1. Consistent with this finding, cells lacking LUCAT1 have altered splicing of selected immune genes. In particular, upon lipopolysaccharide stimulation, the splicing of the nuclear receptor 4A2 (NR4A2) gene was particularly affected. As a consequence, expression of NR4A2 was reduced and delayed in cells lacking LUCAT1. NR4A2-deficient cells had elevated expression of immune genes. These observations suggest that LUCAT1 is induced to control the splicing and stability of NR4A2, which is in part responsible for the anti-inflammatory effect of LUCAT1. Furthermore, we analyzed a large cohort of patients with inflammatory bowel disease as well as asthma and chronic obstructive pulmonary disease. In these patients, LUCAT1 levels were elevated and in both diseases, positively correlated with disease severity. Collectively, these studies define a key molecular mechanism of LUCAT1-dependent immune regulation through post-transcriptional regulation of mRNAs highlighting its role in the regulation of inflammatory disease.

Both stemmed and stemless designs for total shoulder arthroplasty (TSA) have demonstrated efficacious outcomes for the surgical treatment of primary glenohumeral joint osteoarthritis. The purpose of this systematic review and meta-analysis was to compare the clinical outcomes of stemmed versus stemless TSA in randomized controlled trials. We hypothesized that there would be no differences in Constant Score (CS), range of motion, or adverse events, such as periprosthetic fracture and/or revision surgery.Using the Preferred Reporting Items for Systematic Reviews and Meta-analyses guidelines, a systematic review of the literature was done using MEDLINE, SPORTDiscus, Cumulative Index to Nursing and Allied Health Literature, Cochrane Central Registry of Controlled Trials, Embase, and Web of Science databases. Outcomes of interest included CS, range of motion, and adverse events (periprosthetic fracture and revision). Summary effect estimates of the mean difference between stemmed and stemless TSA for each outcome were estimated in random effects models.The search yielded 301 articles with 4 appropriate for qualitative analysis, including the results of 229 stemmed and 358 stemless TSAs. No significant difference was observed in postoperative CS (P = 0.36), forward flexion (P = 0.93), abduction (P = 0.30), or external rotation (P = 0.34) between stemmed and stemless TSA. No significant difference was observed in change in CS (P = 0.27), forward flexion (P = 0.25), or external rotation (P = 0.74). A change in abduction was significantly different between stemmed and stemless TSA (standardized mean difference = -0.64; 95% confidence interval, -1.20 to -0.08) in favor of stemmed TSA (P = 0.02), attributed to preoperative differences. No significant difference was observed in periprosthetic fractures (P = 0.07) or revision (P = 0.90).TSA with stemless versus stemmed humeral components was not associated with notable differences in functional and clinical outcomes. No difference was observed between stemmed and stemless designs in postoperative forward flexion, abduction, or external rotation. Similarly, there was no difference in change in forward flexion or external rotation. A markedly greater improvement in abduction was observed with stemmed TSA, likely due to the lower preoperative motion in the stemmed cohort in one of the studies. No differences were observed between stemmed and stemless designs in the rate of humeral fracture or risk of revision.Level II; systematic review and meta-analysis of prospective randomized controlled trials.

Exhausted CD8 T cells (TEX) are a distinct state of T cell differentiation associated with failure to clear chronic viruses and cancer. Immunotherapies such as PD-1 blockade can reinvigorate TEX cells, but reinvigoration is not durable. A major unanswered question is whether TEX cells differentiate into functional durable memory T cells (TMEM) upon antigen clearance. Here, using a mouse model, we found that upon eliminating chronic antigenic stimulation, TEX cells partially (re)acquire phenotypic and transcriptional features of TMEM cells. These ‘recovering’ TEX cells originated from the T cell factor (TCF-1+) TEX progenitor subset. Nevertheless, the recall capacity of these recovering TEX cells remained compromised as compared to TMEM cells. Chromatin-accessibility profiling revealed a failure to recover core memory epigenetic circuits and maintenance of a largely exhausted open chromatin landscape. Thus, despite some phenotypic and transcriptional recovery upon antigen clearance, exhaustion leaves durable epigenetic scars constraining future immune responses. These results support epigenetic remodeling interventions for TEX cell–targeted immunotherapies. Wherry and colleagues examine whether exhausted T cells (TEX) can differentiate into functional memory T cells (TMEM) when chronic antigen is withdrawn. Using the chronic LCMV infection mouse model, they show that ‘recovering’ TEX cells (REC-TEX) only partially recover immunophenotypic and functional characteristics of TMEM cells. The epigenomic status of REC-TEX cells more closely resembles that of TEX cells, and, upon rechallenge, the REC-TEX cells were still compromised in their ability to respond to virus.

Background: Anaphylaxis is a life-threatening allergic reaction that is often inadequately treated in the hospital setting, leading to adverse outcomes. We hypothesize that a brief educational intervention will enhance knowledge of community-based medical professionals evaluated by pre- and post-questionnaires, leading to improved recognition and management of anaphylaxis. Methods: An initial questionnaire consisting of eight multiple-choice questions and two fill in response pertaining to anaphylaxis identification, management, and treatment was completed by 189 University Hospitals Regional Hospitals personnel, including faculty, nurses, student, residents, and Emergency Medical Services (EMS). The participants were then offered an educational intervention, including a 10-slide, 20-minute PowerPoint presentation on anaphylaxis, and review of the pre-educational intervention questionnaire responses, followed by a post-educational intervention questionnaire similar to the initial questionnaire. Seventy-seven participants completed the same questionnaire at a six-month follow-up to assess retention. Results: Participant scores improved from 62% to 94%, from the initial questionnaire to the immediate post- educational intervention questionnaire. The six-month post-educational intervention questionnaire revealed a return to near baseline (65%) medical knowledge regarding anaphylaxis Conclusion: Healthcare personnel demonstrate a knowledge deficit of identification and management of anaphylaxis. In the short-term, a brief, educational intervention did improve knowledge of anaphylaxis (p

Background A 4-month regimen containing rifapentine and moxifloxacin has noninferior efficacy compared to the standard 6-month regimen for drug-sensitive tuberculosis. We evaluated the effect of regimens containing daily, high-dose rifapentine on efavirenz pharmacokinetics and viral suppression in patients with human immunodeficiency virus (HIV)-associated tuberculosis (TB). Methods In the context of a Phase 3 randomized controlled trial, HIV-positive individuals already virally suppressed on efavirenz--containing antiretroviral therapy (ART) (EFV1), or newly initiating efavirenz (EFV2) received TB treatment containing rifapentine (1200 mg), isoniazid, pyrazinamide, and either ethambutol or moxifloxacin. Mid-interval efavirenz concentrations were measured (a) during ART and TB cotreatment (Weeks 4, 8, 12, and 17, different by EFV group) and (b) when ART was taken alone (pre- or post-TB treatment, Weeks 0 and 22). Apparent oral clearance (CL/F) was estimated and compared. Target mid-interval efavirenz concentrations were > 1 mg/L. Co-treatment was considered acceptable if > 80% of participants had mid-interval efavirenz concentrations meeting this target. Results EFV1 and EFV2 included 70 and 41 evaluable participants, respectively. The geometric mean ratio comparing efavirenz CL/F with vs without TB drugs was 0.79 (90% confidence interval [CI] .72–.85) in EFV1 and 0.84 [90% CI .69–.97] in EFV2. The percent of participants with mid-interval efavirenz concentrations > 1mg/L in EFV1 at Weeks 0, 4, 8, and 17 was 96%, 96%, 88%, and 89%, respectively. In EFV2, at approximately 4 and 8 weeks post efavirenz initiation, the value was 98%. Conclusions TB treatment containing high-dose daily rifapentine modestly decreased (rather than increased) efavirenz clearance and therapeutic targets were met supporting the use of efavirenz with these regimens, without dose adjustment. Clinical Trials Registration NCT 02410772.

BACKGROUND: Tuberculosis (TB) Trials Consortium Study 31/AIDS Clinical Trials Group A5349, an international randomized open-label phase 3 noninferiority trial showed that a 4-month daily regimen substituting rifapentine for rifampin and moxifloxacin for ethambutol had noninferior efficacy and was safe for the treatment of drug-susceptible pulmonary TB (DS-PTB) compared with the standard 6-month regimen. We explored results among the prespecified subgroup of people with human immunodeficiency virus (HIV) (PWH). METHODS: PWH and CD4+ counts ≥100 cells/μL were eligible if they were receiving or about to initiate efavirenz-based antiretroviral therapy (ART). Primary endpoints of TB disease-free survival 12 months after randomization (efficacy) and ≥ grade 3 adverse events (AEs) on treatment (safety) were compared, using a 6.6% noninferiority margin for efficacy. Randomization was stratified by site, pulmonary cavitation, and HIV status. PWH were enrolled in a staged fashion to support cautious evaluation of drug–drug interactions between rifapentine and efavirenz. RESULTS: A total of 2516 participants from 13 countries in sub-Saharan Africa, Asia, and the Americas were enrolled. Among 194 (8%) microbiologically eligible PWH, the median CD4+ count was 344 cells/μL (interquartile range: 223–455). The rifapentine-moxifloxacin regimen was noninferior to control (absolute difference in unfavorable outcomes −7.4%; 95% confidence interval [CI] −20.8% to 6.0%); the rifapentine regimen was not noninferior to control (+7.5% [95% CI, −7.3% to +22.4%]). Fewer AEs were reported in rifapentine-based regimens (15%) than the control regimen (21%). CONCLUSIONS: In people with HIV-associated DS-PTB with CD4+ counts ≥100 cells/μL on efavirenz-based ART, the 4-month daily rifapentine-moxifloxacin regimen was noninferior to the 6-month control regimen and was safe. CLINICAL TRIALS REGISTRATION: NCT02410772.

Background: Low education levels and language barriers present challenges in obtaining informed consent for clinical research. Objective: To describe and correlate the association between the level of education and the participant’s preferred language of consent. Design: Descriptive-analytical cross-sectional study. Participants: Adults being consented for participation in tuberculosis(TB) research studies in an East African community with varying levels of education. Procedures: We analyzed data on demographic and educational characteristics collected from adults being consented for participation in TB studies .Only participants who could understand and speak Luganda (the main local language) or English ( the official language of Uganda) were included in this analysis. Results: A total of 523 participants were consented between April 2015 and December 2017 and included in this analysis; 250 below Senior four (< 11yrs of education), 114 senior four (at 11yrs of education),73 senior five-senior six (12-13yrs of education) and 86 beyond senior six (> 13yrs of education). We noted that the preference for English rises with the rising levels of education and peaked at beyond senior six (83%Vs17%,OR=49,95%CI:22.8-106.3,p

Background The cost of health care in the United States accounts for 18% of the nation's gross domestic product and is expected to reach 20% by 2020. Physicians are responsible for 60%-80% of decisions resulting in health care expenditures. Rotator cuff repairs account for $1.2-$1.6 billion in US health care expenditures annually. The purpose of this study is to assess surgeons' cost awareness in the setting of rotator cuff repairs. The hypothesis is that practice environment and training affect cost consciousness and incentivization will lead to more cost-effective choices. Methods In this cross-sectional study, a 21-item survey was distributed via the email list services of the American Shoulder and Elbow Surgeons and Arthroscopy Association of North America. Data collected included demographics, variables regarding rotator cuff repair (technique, number of companies used, procedures per month), and knowledge of costs. Results Responses from 345 surgeons in 23 countries were obtained with the majority (89%) being from the United States. Most surgeons were “cost-conscious” (275, 70.7%). Of these surgeons, 62.9% are willing to switch suture anchors brands to reduce overall costs if incentivized. Cost-conscious surgeons were more likely to be fellowship trained in shoulder and elbow (51.81% vs. 38.57%, P = .048), be paid based on productivity (73.53% vs. 61.43%, P = .047), and receive shared profits (85.4% vs. 75%, P = .02). Conclusion The majority of orthopedic surgeons are both cost-conscious and willing to change their practice to reduce costs if incentivized to do so. A better understanding of implant costs combined with incentives may help reduce health care expenditure.

Mycobacterium bovis bacille Calmette-Guérin (BCG) has been used for 100 years and prevents disseminated tuberculosis and death in young children. However, it shows only partial efficacy against pulmonary tuberculosis (TB) in adults, so new vaccines are urgently needed. The protective efficacy of BCG depends on T cells, which are typically activated by pathogen-derived protein antigens that bind to highly polymorphic major histocompatibility complex (MHC) molecules. Some T cells recognize non-protein antigens via antigen presenting systems that are independent of genetic background, leading to their designation as donor-unrestricted T (DURT) cells. Whether live whole cell vaccines, like BCG, can induce durable expansions of DURT cells in humans is not known. We used combinatorial tetramer staining, multi-parameter flow cytometry, and immunosequencing to comprehensively characterize the effect of BCG on activation and expansion of DURT cell subsets. We examined peripheral blood mononuclear cells (PBMC) derived from a Phase I study of South African adults in which samples were archived at baseline, 3 weeks, and 52 weeks post-BCG revaccination. We did not observe a change in the frequency of total mucosal-associated invariant T (MAIT) cells, invariant natural killer T (iNKT) cells, germline encoded mycolyl-reactive (GEM) T cells, or γδ T cells at 52 weeks post-BCG. However, immunosequencing revealed a set of TCR-δ clonotypes that were expanded at 52 weeks post-BCG revaccination. These expanded clones expressed the Vδ2 gene segment and could be further defined on the basis of biochemical similarity into several ‘meta-clonotypes’ that likely recognize similar epitopes. Our data reveal that BCG vaccination leads to durable expansion of DURT cell clonotypes despite a limited effect on total circulating frequencies in the blood and have implications for defining the immunogenicity of candidate whole cell TB vaccines.

The identification of sensitive, specific, and reliable biomarkers that can be quantified in the early phases of tuberculosis treatment and predictive of long-term outcome is key for the development of an effective short-course treatment regimen. Time to positivity (TTP), a biomarker of treatment outcome against Mycobacterium tuberculosis, measures longitudinal bacterial growth in mycobacterial growth indicator tube broth culture and may be predictive of standard time to stable culture conversion (TSCC). In two randomized phase 2b trials investigating dose-ranging rifapentine (Studies 29 and 29X), 662 participants had sputum collected over 6 months where TTP, TSCC, and time to culture conversion were quantified. The goals of this post hoc study were to characterize longitudinal TTP profiles and to identify individual patient characteristics associated with delayed time to culture conversion. In order to do so, a nonlinear mixed-effects model describing longitudinal TTP was built. Independent variables associated with increased bacterial clearance (increased TTP), assessed by subject-specific and population-level trajectories, were higher rifapentine exposure, lower baseline grade of sputum acid-fast bacillus smear, absence of productive cough, and lower extent of lung infiltrates on radiographs. Importantly, sensitivity analysis revealed that major learning milestones in phase 2b trials, such as significant exposure-response and covariate relationships, could be detected using truncated TTP data as early as 6 weeks from start of treatment, suggesting alternative phase 2b study designs. The TTP model built depicts a novel phase 2b surrogate endpoint that can inform early assessment of experimental treatment efficacy and treatment failure or relapse in patients treated with shorter and novel TB treatment regimens, improving efficiency of phase 2 clinical trials. (The studies discussed in this paper have been registered at ClinicalTrials.gov under identifiers NCT00694629 and NCT01043575.).

Key Points MAIT cells comprise half the CD8 T cell IFN-γ response to BCG in blood. Frequencies of MAIT cells were not sustainably modulated by BCG vaccination. Innate cytokines mediate BCG-induced MAIT cell responses in whole blood., Tuberculosis (TB) is the leading cause of mortality from a single infectious agent, Mycobacterium tuberculosis. Relevant immune targets of the partially efficacious TB vaccine bacille Calmette–Guérin (BCG) remain poorly defined. Mucosal-associated invariant T (MAIT) cells are MHC-related protein 1 (MR1)–restricted T cells, which are reactive against M. tuberculosis, and underexplored as potential TB vaccine targets. We sought to determine whether BCG vaccination activated mycobacteria-specific MAIT cell responses in humans. We analyzed whole blood samples from M. tuberculosis–infected South African adults who were revaccinated with BCG after a six-month course of isoniazid preventative therapy. In vitro BCG stimulation potently induced IFN-γ expression by phenotypic (CD8+CD26+CD161+) MAIT cells, which constituted the majority (75%) of BCG-reactive IFN-γ–producing CD8+ T cells. BCG revaccination transiently expanded peripheral blood frequencies of BCG-reactive IFN-γ+ MAIT cells, which returned to baseline frequencies a year following vaccination. In another cohort of healthy adults who received BCG at birth, 53% of mycobacteria-reactive–activated CD8 T cells expressed CDR3α TCRs, previously reported as MAIT TCRs, expressing the canonical TRAV1-2-TRAJ33 MAIT TCRα rearrangement. CD26 and CD161 coexpression correlated with TRAV1-2+CD161+ phenotype more accurately in CD8+ than CD4−CD8− MAIT cells. Interestingly, BCG-induced IFN-γ expression by MAIT cells in vitro was mediated by the innate cytokines IL-12 and IL-18 more than MR1-induced TCR signaling, suggesting TCR-independent activation. Collectively, the data suggest that activation of blood MAIT cells by innate inflammatory cytokines is a major mechanism of responsiveness to vaccination with whole cell vaccines against TB or in vitro stimulation with mycobacteria (Clinical trial registration: NCT01119521).

Introduction When conservative therapy for hallux rigidus fails, surgical options such as arthrodesis and interposition arthroplasty can be considered. Although arthrodesis of MTP joint is the gold standard treatment. However patients desiring MTP joint movement may opt for either interposition arthroplasty or implant arthroplasty to avoid the movement restrictions of arthrodesis. The purpose of this systematic review was to investigate clinical outcomes and complications following interposition arthroplasty for moderate to severe hallux rigidus, for patietns who would prefer to maintain range of motion in the MTP joint. Methods A systematic search on MEDLINE, EMBASE and Cochrane library database was performed during February 2018. Demographics, surgical techniques, clinical outcomes, radiological outcomes and complications were recorded from each included study. Pooled statistics performed for variables with homogenous data across the studies. A linear regression model used to compare the clinical outcomes between autogenous vs allogenous material interposition arthroplasty. Results Fifteen articles were included in the systematic review. Mean AOFAS scores improved from preoperative 41.35 to postoperative 83.17. Mean pain, function, and alignment score improved from preoperative values of 14.9, 24.9, and 10 to postoperative values of 33.3, 35.8, and 14.5. Mean dorsiflexion increased from 21.27° (5–30) to 42.03° (25–71). Mean ROM improved from 21.06° to 46.43°. Joint space increased from 0.8 mm to 2.5 mm. The most common postoperative complications included metatarsalgia (13.9%), loss of ground contact (9.7%), osteonecrosis (5.4%), great toe weakness (4.8%), hypoesthesia (4.2%), decreased push off power (4.2%), and callous formation (4.2%). Conclusion Interposition arthroplasty is an effective treatment option with acceptable clinical outcomes in patients with moderate-severe hallux rigidus who prefer to maintain range of motion and accept the risk of future complications. Level of Evidence: IV.

Whether conventional dendritic cells (cDC) acquire subset identity under direction of Wnt family glycoproteins is unknown. We demonstrate that Wnt4, a β-catenin–independent Wnt ligand, is produced by both hematopoietic and nonhematopoietic cells and is both necessary and sufficient for preconventional DC1/cDC1 maintenance. Whereas bone marrow cDC precursors undergo phosphoJNK/c-Jun activation upon Wnt4 treatment, loss of cDC Wnt4 in CD11cCreWnt4flox/flox mice impaired differentiation of CD24+, Clec9A+, CD103+ cDC1 compared with CD11cCre controls. Conversely, single-cell RNA sequencing analysis of bone marrow revealed a 2-fold increase in cDC2 gene signature genes, and flow cytometry demonstrated increased numbers of SIRP-α+ cDC2 amid lack of Wnt4. Increased cDC2 numbers due to CD11c-restricted Wnt4 deficiency increased IL-5 production, group 2 innate lymphoid cell expansion, and host resistance to the hookworm parasite Nippostrongylus brasiliensis. Collectively, these data uncover a novel and unexpected role for Wnt4 in cDC subset differentiation and type 2 immunity.

Background Iatrogenic or traumatic injury to the spinal accessory nerve is a rare but debilitating injury. An effective treatment, known as the Eden-Lange modification triple-tendon transfer procedure, involves the transfer of the rhomboid major (RM), rhomboid minor (Rm), and levator scapulae (LS). Careful detachment of their insertions is necessary to avoid injury of the dorsal scapular nerve (DSN). This study evaluated the surgical anatomy and safety of the DSN relative to this procedure. Methods The study used 12 cadavers (22 shoulders). The RM, Rm, and LS were detached from their insertions, and the DSN was dissected. Measurements were taken to evaluate the anatomy of each relative to the triple-tendon transfer procedure. Additional measurements were taken to identify “danger zones” for DSN injury, regarding detachment of RM, Rm, and LS from their respective insertions. Results Measurements of the 22 shoulders included in the study showed wide variation in anatomy. The minimum distance between the scapula and the DSN at the vertebral scapular border was 0.7 cm, suggesting that care and precision are needed to perform this technique. The region where the DSN crosses the superior border of the Rm was shown to be the greatest “danger zone” of this technique, with a mean distance to the scapula of 1.61 ± 0.53 cm Conclusions This study provides insight into the surgical anatomy of the DSN relative to a rare but successful procedure used to treat trapezius paralysis. The results of this study can inform the surgeon regarding potential anatomic considerations when performing the triple-tendon transfer.

BackgroundRifapentine-based regimens have potent antimycobacterial activity that may allow for a shorter course in patients with drug-susceptible pulmonary tuberculosis.MethodsIn an open-label, phase 3, randomized, controlled trial involving persons with newly diagnosed pulmonary tuberculosis from 13 countries, we compared two 4-month rifapentine-based regimens with a standard 6-month regimen consisting of rifampin, isoniazid, pyrazinamide, and ethambutol (control) using a noninferiority margin of 6.6 percentage points. In one 4-month regimen, rifampin was replaced with rifapentine; in the other, rifampin was replaced with rifapentine and ethambutol with moxifloxacin. The primary efficacy outcome was survival free of tuberculosis at 12 months.ResultsAmong 2516 participants who had undergone randomization, 2343 had a culture positive for Mycobacterium tuberculosis that was not resistant to isoniazid, rifampin, or fluoroquinolones (microbiologically eligible population; 768 in the control group, 791 in the rifapentine-moxifloxacin group, and 784 in the rifapentine group), of whom 194 were coinfected with human immunodeficiency virus and 1703 had cavitation on chest radiography. A total of 2234 participants could be assessed for the primary outcome (assessable population; 726 in the control group, 756 in the rifapentine-moxifloxacin group, and 752 in the rifapentine group). Rifapentine with moxifloxacin was noninferior to the control in the microbiologically eligible population (15.5% vs. 14.6% had an unfavorable outcome; difference, 1.0 percentage point; 95% confidence interval [CI], -2.6 to 4.5) and in the assessable population (11.6% vs. 9.6%; difference, 2.0 percentage points; 95% CI, -1.1 to 5.1). Noninferiority was shown in the secondary and sensitivity analyses. Rifapentine without moxifloxacin was not shown to be noninferior to the control in either population (17.7% vs. 14.6% with an unfavorable outcome in the microbiologically eligible population; difference, 3.0 percentage points [95% CI, -0.6 to 6.6]; and 14.2% vs. 9.6% in the assessable population; difference, 4.4 percentage points [95% CI, 1.2 to 7.7]). Adverse events of grade 3 or higher occurred during the on-treatment period in 19.3% of participants in the control group, 18.8% in the rifapentine-moxifloxacin group, and 14.3% in the rifapentine group.ConclusionsThe efficacy of a 4-month rifapentine-based regimen containing moxifloxacin was noninferior to the standard 6-month regimen in the treatment of tuberculosis. (Funded by the Centers for Disease Control and Prevention and others; Study 31/A5349 ClinicalTrials.gov number, NCT02410772.).

SummaryAntigen classes other than proteins can be presented to T cells by near-monomorphic antigen-presenting molecules such as CD1, MR1, and butyrophilin 3A1. We sought to define the roles of donor unrestricted T (DURT) cells, including MR1-reactive MAIT cells, CD1b-reactive glucose monomycolate (GMM)-specific T cells, CD1d-reactive NKT cells, and γδ T cells, in vaccination against Mycobacterium tuberculosis. We characterized DURT cells following primary bacille Calmette-Guerin (BCG) vaccination in infants or BCG-revaccination in adults. BCG (re)vaccination did not modulate peripheral blood frequencies, T cell activation or memory profiles of MAIT cells, CD1b-restricted GMM-specific and germline-encoded mycolyl-reactive (GEM) cells or CD1d- restricted NKT cells. By contrast, BCG vaccination was associated with increased frequencies of γδ T cells as well as a novel subset of IFN-γ-expressing CD4+ T cells with a CD26+CD161+TRAV1-2− phenotype in infants. More studies are required to understand the full potential of DURT cells in new TB vaccine strategies.

Adjuvants are critical for improving the quality and magnitude of adaptive immune responses to vaccination. Lipid nanoparticle (LNP)-encapsulated nucleoside-modified mRNA vaccines have shown great efficacy against SARS-CoV-2, but the mechanism of action of this vaccine platform is not well-characterized. Using influenza virus and SARS-CoV-2 mRNA and protein subunit vaccines, we demonstrated that our LNP formulation has intrinsic adjuvant activity that promotes the induction of strong T follicular helper cell, germinal center B cell, long-lived plasma cell and memory B cell responses that are associated with durable and protective antibodies in mice. Comparative experiments demonstrated that this LNP outperformed a widely used MF59-like adjuvant, AddaVax™. The adjuvant activity of the LNP relies on the ionizable lipid component and on IL-6 cytokine induction, but not on MyD88- or MAVS-dependent sensing of LNPs. Our study identified LNPs as a versatile adjuvant that enhances the efficacy of both traditional and next-generation vaccine platforms., Graphical Abstract, The mechanism of action of nucleoside-modified mRNA-LNP vaccines is unknown. Alameh, Tombácz, Bettini et al. demonstrate that LNPs can possess adjuvant activity and promote a robust induction of Tfh cell, B cell and humoral responses when utilized in mRNA and protein subunit vaccines in mice. IL-6 induction and the ionizable lipid component are critical for the adjuvant activity of LNPs.

Background Rifapentine exposure is associated with bactericidal activity against Mycobacterium tuberculosis, but high interindividual variation in plasma concentrations is encountered. Objectives To investigate a genomic association with interindividual variation of rifapentine exposure, SNPs of six human genes involving rifamycin metabolism (AADAC, CES2), drug transport (SLCO1B1, SLCO1B3) and gene regulation (HNF4A, PXR) were evaluated. Methods We characterized these genes in 173 adult participants in treatment trials of the Tuberculosis Trials Consortium. Participants were stratified by self-identified race (black or non-black), and rifapentine AUC from 0 to 24 h (AUC0–24) was adjusted by analysis of covariance for SNPs, rifapentine dose, sex, food and HIV coinfection. This study was registered at ClinicalTrials.gov under identifier NCT01043575. Results The effect on rifapentine least squares mean AUC0–24 in black participants overall decreased by –10.2% for AADAC rs1803155 G versus A allele (Wald test: P = 0.03; false discovery rate, 0.10). Black participants with one G allele in AADAC rs1803155 were three times as likely to have below target bactericidal rifapentine exposure than black participants with the A allele (OR, 2.97; 95% CI: 1.16, 7.58). With two G alleles, the OR was greater. In non-black participants, AADAC rs1803155 SNP was not associated with rifapentine exposure. In both black and non-black participants, other evaluated genes were not associated with rifapentine exposure (P > 0.05; false discovery rate > 0.10). Conclusions Rifapentine exposure in black participants varied with AADAC rs1803155 genotype and the G allele was more likely to be associated with below bactericidal target rifapentine exposure. Further pharmacogenomic study is needed to characterize the association of the AADAC rs1803155 with inadequate rifapentine exposure in different patient groups.

Study Objective Determine near-optimal dose, safety, and efficacy of nerindocianine in pelvic ureter detection with near-infrared fluorescence imaging in women undergoing minimally invasive pelvic surgery with 3 Food and Drug Administration–cleared imaging systems. Design Open label, phase 1/2a study. Setting University of Alabama at Birmingham. Patients Forty-one female subjects undergoing minimally invasive gynecologic surgery. Interventions Subjects received a single dose of nerindocianine sodium, starting at 0.06-mg/kg body weight and increased/decreased until the near-optimal dose was determined (part A). Examine the degree of concordance between endoscopic and robotic devices (part B). Measurements and Main Results In part A, composite scores were collected every 10 minutes for 30 minutes and then every 15 minutes through 90 minutes using a scale measuring the anatomy/laterality of ureter visualization. In part B (paired imaging system efficacy), 2 cohorts of 8 subjects each received the near-optimal dose. Composite scores for visualization of the ureter were collected at 10 and 30 minutes postinfusion with the Firefly Imaging System and either the PINPOINT or 1588 AIM endoscope. Composite scores were compared to examine the degree of concordance between devices. Part A comprised 25 total subjects enrolled in dosing groups 1, 2, and 3 (0.06-, 0.12-, and 0.045-mg/kg, respectively). Median time to first ureter visualization was 10 minutes (all groups). The nerindocianine 0.06-mg/kg and 0.12-mg/kg groups had longer length of time of visualization than the 0.045-mg/kg group, resulting in the selection of 0.06 mg/kg as the near-optimal dose. Part B enrolled 16 total subjects in 2 groups dosed at 0.06 mg/kg. Efficacy analysis showed no statistically significant difference in composite scores with Firefly versus PINPOINT or 1588 AIM. Conclusion Nerindocianine was well tolerated with visualization of the ureter demonstrated in 88.9% of the subjects through 90 minutes postdosing. No meaningful visualization differences were observed among the Food and Drug Administration–cleared surgical imaging systems used.

Tuberculosis kills more people than any other infectious disease. Three pivotal trials testing 4-month regimens failed to meet non-inferiority margins; however, approximately four-fifths of participants were cured. Through a pooled analysis of patient-level data with external validation, we identify populations eligible for 4-month treatment, define phenotypes that are hard to treat and evaluate the impact of adherence and dosing strategy on outcomes. In 3,405 participants included in analyses, baseline smear grade of 3+ relative to 6 months to cure all. Regimen duration can be selected in order to improve outcomes, providing a stratified medicine approach as an alternative to the 'one-size-fits-all' treatment currently used worldwide.

Sintering behavior of nanoscale WC-Co was investigated based on master sintering curve model. Densification and grain growth were mapped for grain size assessment and optimization. Parameters for model fitting were acquired by experimental approaches. Using the sensitivity analysis with respect to processing parameters, effects of key conditions on final properties were investigated. Based on the derived objective function, an optimization algorithm to minimize grain size under the given target density was developed. For example using our nanoscale WC-Co, grain size of 332 nm was the minimum size when the compaction pressure of 565 MPa and 97% of sintered density were the processing constraints. Using the constructed mapping function, the achievable minimum grain size under constrained processing condition is predicted and the further requirements for the powder are suggested to achieve the specific target grain size. In this study, we use the master sintering curve model and optimization methods to identify process conditions, such as compaction pressure and sintering temperature, to tailor specified tolerances of the fabricated components.

Retroviral (RV) expression of genes of interest (GOIs) is an invaluable tool and has formed the foundation of cellular engineering for adoptive cell therapy in cancer and other diseases. However, monitoring of transduced T cells long term (weeks to months) in vivo remains challenging because of the low frequency and often poor durability of transduced T cells over time when transferred without enrichment. Traditional methods often require additional overnight in vitro culture after transduction. Moreover, in vitro-generated effector CD8+ T cells enriched by sorting often have reduced viability, making it difficult to monitor the fate of transferred cells in vivo. Here, we describe an optimized mouse CD8+ T-cell RV transduction protocol that uses simple and rapid Percoll density centrifugation to enrich RV-susceptible activated CD8+ T cells. Percoll density centrifugation is simple, can be done on the day of transduction, requires minimal time, has low reagent costs and improves cell recovery (up to 60%), as well as the frequency of RV-transduced cells (~sixfold over several weeks in vivo as compared with traditional methods). We have used this protocol to assess the long-term stability of CD8+ T cells after RV transduction by comparing the durability of T cells transduced with retroviruses expressing each of six commonly used RV reporter genes. Thus, we provide an optimized enrichment and transduction approach that allows long-term in vivo assessment of RV-transduced T cells. The overall procedure from T-cell isolation to RV transduction takes 2 d, and enrichment of activated T cells can be done in 1 h.

The sputum smear-positive, culture-negative state poses a challenge for clinicians. Previous studies have shown that most samples with positive smears during the later stages of treatment are culture-negative. Earlier studies generally used solid culture media, which tend to be less sensitive than current liquid culture systems. We examined the smear-positive, culture-negative state in the era of MGIT™ 960™ liquid cultures. We found that the smear-positive, culture-negative state occurred less frequently with MGIT culture, and that the majority of the samples with late positive smears were culture-negative, regardless of media type.

TCF-1 is a key transcription factor in progenitor exhausted CD8 T cells (Tex). Moreover, this Tex cell subset mediates responses to PD-1 checkpoint pathway blockade. However, the role of the transcription factor TCF-1 in early fate decisions and initial generation of Tex cells is unclear. Single cell RNA-sequencing (scRNA-seq) and lineage tracing identified a TCF-1(+)Ly108(+)PD-1(+) CD8 T cell population early during chronic infection that seeds development of mature Tex cells. TCF-1 mediated the bifurcation between divergent fates, repressing development of terminal KLRG1(Hi) effectors while fostering KLRG1(Lo) Tex precursor cells, and PD-1 stabilized this TCF-1(+) Tex precursor cell pool. TCF-1 mediated a T-bet to Eomes transcription factor transition in Tex precursors by promoting Eomes expression and drove c-Myb expression that controlled Bcl-2 and survival. These data define a role for TCF-1 in early fate bifurcation driving Tex precursor cells, and also identify PD-1 as a protector of this early TCF-1 subset.

Biomarkers for TB treatment response and outcome are needed. This study characterize changes in immune profiles during TB treatment, define biosignatures associated with treatment outcomes, and explore the feasibility of predictive models for relapse. Seventy-two markers were measured by multiplex cytokine array in serum samples from 78 cured, 12 relapsed and 15 failed treatment patients from South Africa before and during therapy for pulmonary TB. Promising biosignatures were evaluated in a second cohort from Uganda/Brazil consisting of 17 relapse and 23 cured patients. Thirty markers changed significantly with different response patterns during TB treatment in cured patients. The serum biosignature distinguished cured from relapse patients and a combination of two clinical (time to positivity in liquid culture and BMI) and four immunological parameters (TNF-β, sIL-6R, IL-12p40 and IP-10) at diagnosis predicted relapse with a 75% sensitivity (95%CI 0.38-1) and 85% specificity (95%CI 0.75-0.93). This biosignature was validated in an independent Uganda/Brazil cohort correctly classifying relapse patients with 83% (95%CI 0.58-1) sensitivity and 61% (95%CI 0.39-0.83) specificity. A characteristic biosignature with value as predictor of TB relapse was identified. The repeatability and robustness of these biomarkers require further validation in well-characterized cohorts.

Therapeutic drug monitoring (TDM) of moxifloxacin is recommended to improve the response to tuberculosis treatment and reduce acquired drug resistance. Limited sampling strategies (LSSs) are able to reduce the burden of TDM by using a small number of appropriately timed samples to estimate the parameter of interest, the area under the concentration-time curve. This study aimed to develop LSSs for moxifloxacin alone (MFX) and together with rifampin (MFX+RIF) in tuberculosis (TB) patients. Population pharmacokinetic (popPK) models were developed for MFX (n = 77) and MFX+RIF (n = 24). In addition, LSSs using Bayesian approach and multiple linear regression were developed. Jackknife analysis was used for internal validation of the popPK models and multiple linear regression LSSs. Clinically feasible LSSs (one to three samples, 6-h timespan postdose, and 1-h interval) were tested. Moxifloxacin exposure was slightly underestimated in the one-compartment models of MFX (mean -5.1%, standard error [SE] 0.8%) and MFX+RIF (mean -10%, SE 2.5%). The Bayesian LSSs for MFX and MFX+RIF (both 0 and 6 h) slightly underestimated drug exposure (MFX mean -4.8%, SE 1.3%; MFX+RIF mean -5.5%, SE 3.1%). The multiple linear regression LSS for MFX (0 and 4 h) and MFX+RIF (1 and 6 h), showed mean overestimations of 0.2% (SE 1.3%) and 0.9% (SE 2.1%), respectively. LSSs were successfully developed using the Bayesian approach (MFX and MFX+RIF; 0 and 6 h) and multiple linear regression (MFX, 0 and 4h; MFX+RIF, 1 and 6h). These LSSs can be implemented in clinical practice to facilitate TDM of moxifloxacin in TB patients.

The evaluation of new tuberculosis (TB) therapies is limited by the paucity of biomarkers to monitor treatment response. Previous work detected an uncharacterized urine metabolite with a molecular mass of 874.3547 Da that showed promise as a biomarker for successful TB treatment. Using mass spectrometry combined with enzymatic digestions, the metabolite was structurally characterized as a seryl-leucine core 1 O-glycosylated peptide (SLC1G) of human origin. Examination of SLC1G in urine revealed a significant abundance increase in individuals with active TB versus their household contacts and healthy controls. Moreover, differential decreases in SLC1G levels were observed by week one in TB patients during successful treatment versus those that failed treatment. The SLC1G levels were also associated with clinical parameters used to measure bacterial burden (GeneXpert) and inflammation (positron emission tomography-computed tomography (PET-CT)). These results demonstrate the importance of metabolite identification and provide strong evidence for applying SLC1G as a biomarker of TB treatment response.

First tarsometatarsal (TMT) joint fusion is effective for treatment of arthritis and some first ray deformities. To prepare the articular surfaces, cartilage should be carefully but completely denuded. Inadequate preparation may result in non-union, while excessive preparation may cause ray shortening and consequential transfer metatarsalgia. Preparation can be performed with an osteotome or a saw. The purpose of this study was to investigate whether utilization of an osteotome or saw would minimize shortening of the first ray in TMT arthrodesis.Ten fresh-frozen cadaver specimens were randomly assigned to undergo joint preparation using either an osteotome (n=5) or saw (n=5). Sample size was determined by cadaver availability. Fusion was performed using a cross-screw construct through the dorsal aspect of the proximal phalanx and the medial cuneiform. Pre- and post-operative X-rays were taken with a radiopaque ruler in the field, and changes in length in the first metatarsal and first cuneiform were compared between osteotome and sawblade groups.The average change in metatarsal length was significantly smaller in the osteotome group (1.6mm) as compared to the saw group (4.4mm) (p=0.031). The average percent change in metatarsal length was also significantly smaller in the osteotome group (3.0%) compared to the saw group (8.4%) (p=0.025). There was no significant difference between the two groups with respect to change in cuneiform length. The osteotome group demonstrated a significantly smaller average measured change (3.0mm vs. 6.9mm, p=0.001) and percent change (4.1% vs. 9.3%, p0.001) in total length (cuneiform plus metatarsal) in comparison to the saw group.In first TMT fusion, joint preparation with an osteotome may prevent over-shortening of the first ray in comparison to preparation with a saw.

Background Complications following orthopedic surgeries are undesirable and costly. A potential method to reduce these costs is to perform traditionally inpatient surgical procedures in the outpatient setting. The purpose of this study is to compare outcomes between inpatient and outpatient settings for elective foot and ankle surgeries using the National Surgical Quality Improvement Program (NSQIP) database. Methods Patients with Current Procedural Terminology (CPT) codes specific to orthopedic foot and ankle surgery were identified from the 2011-2015 American College of Surgeons NSQIP database. Demographics, comorbidities, and complications were compared between patients undergoing inpatient and outpatient procedures. Results Patients receiving inpatient surgery were significantly older and more frequently male. Black patients were significantly more likely to undergo inpatient surgery than outpatient surgery while white patients were significantly more likely to undergo outpatient surgery. Outpatients had a significantly higher mean body mass index (BMI) than inpatients. Smokers were at a significantly greater risk of undergoing inpatient surgery than outpatient surgery. Outpatients had significantly longer operative times, were more likely to receive general anesthesia, had a lower American Society of Anesthesiologists (ASA) class, were more likely to be functionally independent, and were less likely to expire postoperatively. Patients who received surgery as an inpatient were significantly more likely to have comorbidities as compared to outpatients. The overall risk of surgical complications was significant between groups with 8.6% in the inpatient group and 2.0% in the outpatient group. The overall risk of medical complications was 16.9% in the inpatient group and 1.7% in the outpatient group. Similar to the surgical complications, inpatients were significantly more likely to sustain each of the individual medical complications except for stroke/CVA and venous thromboembolism. Conclusions Outpatient management is associated with decreased postoperative complications in select patients. Performing more operations in the outpatient setting in select patients may be beneficial for cost reduction and patient satisfaction.

Tuberculosis kills more people than any other infectious disease. Three pivotal trials testing 4-month regimens failed to meet non-inferiority margins; however, approximately four-fifths of participants were cured. Through a pooled analysis of patient-level data with external validation, we identify populations eligible for 4-month treatment, define phenotypes that are hard to treat and evaluate the impact of adherence and dosing strategy on outcomes. In 3,405 participants included in analyses, baseline smear grade of 3+ relative to2+, HIV seropositivity and adherence of ≤90% were significant risk factors for unfavorable outcome. Four-month regimens were non-inferior in participants with minimal disease defined by2+ sputum smear grade or non-cavitary disease. A hard-to-treat phenotype, defined by high smear grades and cavitation, may require durations6 months to cure all. Regimen duration can be selected in order to improve outcomes, providing a stratified medicine approach as an alternative to the 'one-size-fits-all' treatment currently used worldwide.

One third of the global population is estimated to be latently infected with Mycobacterium tuberculosis. We performed a phase I randomized controlled trial of isoniazid preventive therapy (IPT) before revaccination with bacillus Calmette–Guérin (BCG) in healthy, tuberculin skin test–positive (≥15-mm induration), HIV-negative South African adults. We hypothesized that preclearance of latent bacilli with IPT modulates BCG immunogenicity following revaccination. Frequencies and coexpression of IFN-γ, TNF-α, IL-2, IL-17, and/or IL-22 in CD4 T cells and IFN-γ–expressing CD8 T, γδ T, CD3+CD56+ NKT-like, and NK cells in response to BCG were measured using whole blood intracellular cytokine staining and flow cytometry. We analyzed 72 participants who were revaccinated with BCG after IPT (n = 33) or without prior IPT (n = 39). IPT had little effect on frequencies or cytokine coexpression patterns of M. tuberculosis– or BCG-specific responses. Revaccination transiently boosted BCG-specific Th1 cytokine-expressing CD4, CD8, and γδ T cells. Despite high frequencies of IFN-γ–expressing BCG-reactive CD3+CD56+ NKT-like cells and CD3−CD56dim and CD3−CD56hi NK cells at baseline, BCG revaccination boosted these responses, which remained elevated up to 1 y after revaccination. Such BCG-reactive memory NK cells were induced by BCG vaccination in infants, whereas in vitro IFN-γ expression by NK cells upon BCG stimulation was dependent on IL-12 and IL-18. Our data suggest that isoniazid preclearance of M. tuberculosis bacilli has little effect on the magnitude, persistence, or functional attributes of lymphocyte responses boosted by BCG revaccination. Our study highlights the surprising durability of BCG-boosted memory NKT-like and NK cells expressing antimycobacterial effector molecules, which may be novel targets for tuberculosis vaccines.

Although considerable experience exists with sintering on Earth, the behavior under reduced gravity conditions is poorly understood. This study analyzes replica microgravity liquid phase sintering data for seven tungsten alloys (35 to 88 wt pct tungsten) sintered for three hold times (1, 180, or 600 minutes) at 1773 K (1500 °C) using 0.002 pct of standard gravity. Equivalent sintering is performed on Earth using the same heating cycles. Microgravity sintering results in a lower density and more shape distortion. For Earth-based sintering, minimized distortion is associated with low liquid contents to avoid solid settling and slumping. Distortion in microgravity sintering involves viscous spreading of the component at points of contact with the containment crucible. Distortion in microgravity is minimized by short hold times; long hold times allow progressive component reshaping toward a spherical shape. Microgravity sintering also exhibits pore coalescence into large, stable voids that cause component swelling. The microgravity sintering results show good replication in terms of mass change and sintered density. Distortion is scattered but statistically similar between the replica microgravity runs. However, subtle factors, not typically of concern on Earth, emerge to influence microgravity sintering, such that ground experiments do not provide a basis to predict microgravity behavior.

B cell subsets expressing the transcription factor T-bet are associated with humoral immune responses and autoimmunity. Here we examined the anatomic distribution, clonal relationships, and functional properties of T-bet(+) and T-bet(−) memory B cells (MBCs) in the context of the influenza-specific immune response. In mice, both T-bet(−) and T-bet(+) hemagglutinin-specific B cells arose in germinal centers, acquired memory B cell markers, and persisted indefinitely. Lineage tracing and IgH repertoire analyses revealed minimal interconversion between T-bet(−) and T-bet(+) MBCs, and parabionts showed differential tissue residency and recirculation properties. T-bet(+) MBCs could be subdivided into recirculating T-bet(lo) MBCs and spleen-resident T-bet(hi) MBCs. Human MBCs displayed similar features. Conditional gene deletion studies revealed that T-bet expression in B cells was required for nearly all HA stalk-specific IgG2c antibodies and for durable neutralizing titers to influenza. Thus, T-bet expression distinguishes MBC subsets that have profoundly different homing, residency, and functional properties, and mediate distinct aspects of humoral immune memory.