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Epidemiology provides a powerful framework for characterizing exposure–disease relationships, but its utility for making causal inferences is limited because epidemiologic data are observational in nature and subject to biases stemming from undetected confounding variables and reverse causation. Mendelian randomization (MR) is an increasingly popular method used to circumvent these limitations. MR uses genetic variants, or instruments, as a natural experiment to proxy an exposure, thus allowing estimation of causal effects upon an outcome that are minimally affected by the usual biases present in epidemiologic studies. Notably, MR relies on three core assumptions related to the selection of the genetic instruments, and adherence to these assumptions must be carefully evaluated to assess the validity of the causal estimates. The goal of this review is to provide readers with a basic understanding of MR studies and how to read and evaluate them. Specifically, we outline the basics of how MR analysis is conducted, the assumptions underlying instrument selection, and how to assess the quality of MR studies.

Background: Despite decades of advocacy for disaggregated data collection and reporting for Asian American, Native Hawaiian, and Pacific Islander (AA and NHPI) people, significant gaps remain in our ability to understand AA and NHPI individuals’ access to care. We assess inequities in access to care measures between non-Hispanic White and AA and NHPI adult Medicaid enrollees. Methods: We used the 2014–15 Nationwide Adult Medicaid Consumer Assessment of Healthcare Providers and Systems, the first-and-only nationally representative sample of Medicaid enrollees. Our main outcomes were access to needed care, access to a personal doctor, timely access to a checkup, and timely access to specialty care. Using multivariable linear probability models, we assessed the relationship between racial/ethnic group and our outcomes, both in the aggregate and disaggregated into ten racial/ethnic groups, and adjusted for enrollee-level sociodemographic characteristics, health status, and state-level Medicaid expansion status. Results: In aggregate, AA and NHPI enrollees reported worse access to care than White enrollees on all four metrics (p< 0.001). The magnitude of disparities varied across the ten AA and NHPI ethnic groups. Disparities relative to White enrollees were particularly large in magnitude, roughly 1.5 to 2 times greater, for Chinese, Korean, and Vietnamese enrollees than for the aggregated AA and NHPI group. Conclusions: Despite comparable insurance coverage, there were inequities in multiple access to care metrics between non-Hispanic White and AA and NHPI Medicaid enrollees. Collection of disaggregated health data on AA and NHPI patients reveals important variation in access to care by ethnic group.

We explored the association between the use of a hospital-based food pantry and subsequent emergency department (ED) utilization among Medicaid patients with diabetes in a large safety-net health system. Leveraging 2015-2019 electronic health record data, we used a staggered difference-in-differences approach to measure changes in ED use before vs after food pantry use. Food pantry use was associated with a 7.3 percentage point decrease per patient per quarter (95% confidence interval, −13.8 to −0.8) in the probability of subsequent ED utilization (P= .03). Addressing food insecurity through hospital-based food pantries may be one mechanism for reducing ED use among low-income patients with diabetes.

Osteochondral lesions of the talus are a common sequelae of trauma and are often associated with ankle sprains and ankle fractures. Because the surface of the talus is composed primarily of hyaline cartilage, the regenerative capacity of these injuries is limited. Therefore, several open and arthroscopic techniques have been described to treat osteochondral injuries of the talus and underlying bone marrow lesions. Throughout this review, these treatment options are discussed along with their indications and currently reported outcomes. A commentary on the authors’ preferences among these techniques is also provided.

Prescription opioids are the gold standard for treating moderate to severe pain despite their well-documented adverse effects. Of all prescription medications, opioids are abused most widely, and fatal overdoses have reached epidemic levels. One strategy for improving the margin of safety of opioids is combining them with non-opioid drugs to decrease the opioid dose needed for pain relief, thereby reducing adverse effects that occur with larger doses. The N-methyl-D-aspartate receptor antagonist ketamine has been used safely as an analgesic but only under a very limited range of conditions. The current studies characterized the antinociceptive, behavioral suppressant, and gastrointestinal effects of morphine and ketamine alone and in mixtures to determine their interaction in 24 adult male Sprague–Dawley rats (n = 8 per assay). Given alone, both morphine and ketamine produced antinociception, decreased responding for food, and reduced gastrointestinal transit (i.e. produced constipation). The effects of morphine:ketamine mixtures generally were additive, except for the antinociceptive effects of 1:1 mixtures for which the difference in slope (i.e. non-parallel shift) between the observed and predicted effects suggested synergy at smaller doses and additivity at larger doses. The potency of morphine to produce constipation was not enhanced by administration of morphine:ketamine mixtures with antinociceptive effects. The nature of the interaction between morphine and ketamine for adverse effects such as dependence, withdrawal, abuse, or respiratory depression remains unknown but also might be related to the ratio of each drug in mixtures. It will be important to identify conditions that produce the largest potential therapeutic window in humans.

Mendelian randomization (MR) has increasingly been utilized as a tool for establishing causal relations between modifiable exposures and osteoarthritis (OA). The goal of this review was to summarize available MR studies of OA that evaluate the causal role of modifiable risk factors on OA. This review was performed following the Preferred Reporting Items for Systematic Reviews and Meta‐Analysis (PRISMA) Extension for Scoping Reviews model. We performed a literature search for relevant studies published before December 2021 across multiple databases using the search terms “osteoarthritis” and (“Mendelian randomization” or “polygenic risk score”). We reported the MR estimates of causal associations between exposures and OA and then assessed methodologic quality of abstracted studies according to their efforts to validate the three key MR assumptions. Our search identified 45 studies reporting on 141 exposure‐association analyses. All studies performed a formal instrumental variable analysis to estimate the causal effect of exposure on OA. Causal associations (P< 0.05) were reported in 60 of these analyses representing 36 unique publications, and MR‐Egger sensitivity analyses were performed in 45 of these analyses. MR studies provided support for causal associations of OA with increased levels of adiposity, coffee consumption, bone mineral density, and sleep disturbance, and decreased levels of serum calcium and low‐density lipoprotein cholesterol. These results highlight the potential benefits of weight reduction and improvement of sleep quality to reduce the risk of OA and call for a better understanding of the relations of coffee consumption and serum calcium to OA risk.

Human immunodeficiency virus (HIV) persistence during antiretroviral therapy (ART) is associated with heightened plasma interleukin-10 (IL-10) levels and PD-1 expression. We hypothesized that IL-10 and PD-1 blockade would lead to control of viral rebound following analytical treatment interruption (ATI). Twenty-eight ART-treated, simian immunodeficiency virus (SIV)mac239-infected rhesus macaques (RMs) were treated with anti-IL-10, anti-IL-10 plus anti-PD-1 (combo) or vehicle. ART was interrupted 12 weeks after introduction of immunotherapy. Durable control of viral rebound was observed in nine out of ten combo-treated RMs for >24 weeks post-ATI. Induction of inflammatory cytokines, proliferation of effector CD8+T cells in lymph nodes and reduced expression of BCL-2 in CD4+T cells pre-ATI predicted control of viral rebound. Twenty-four weeks post-ATI, lower viral load was associated with higher frequencies of memory T cells expressing TCF-1 and of SIV-specific CD4+and CD8+T cells in blood and lymph nodes of combo-treated RMs. These results map a path to achieve long-lasting control of HIV and/or SIV following discontinuation of ART.

Introduction: In a previously published randomized controlled trial, automated self-association training (ASAT), a novel digital intervention, was found to extend the rapid antidepressant effect of a single infusion of ketamine for at least 30 days. In this secondary analysis, we aimed to understand the potential role of implicit self-esteem in the combined antidepressant effect of ketamine and ASAT training, by investigating the novel synergistic treatment’s effects on implicit self-associations and their relation to symptom improvement. Methods: A total of 154 adults (ages 18–60) with treatment-resistant unipolar depression and lower-than-normative explicit self-esteem were randomized in a double-blind, parallel-arm design to receive one of three treatment allocations: an active/active treatment combination consisting of one infusion of ketamine (0.5 mg/kg) followed by four days of ASAT ( ~ 30–40 min/day), or one of two control arms that lacked either the active drug or the active behavioral component. The Implicit Association Test (IAT) was used to behaviorally assess the strength of association between self-related stimuli and negative concepts. Linear regression models were used to test the relationship between group assignment, IAT scores acquired immediately post-treatment, and both acute and extended clinical outcomes (% change in Montgomery-Asberg Depression Rating Scale scores, relative to pre-treatment baseline) in the trial. Results: The group assigned to ketamine + ASAT intervention, compared to the other groups, had a pattern of IAT scores indicating more positive self-associations immediately after treatment relative to the control arms (F(1, 131) = 3.979; p= 0.048). In regression models, IAT scores tracked with concurrent (acute post-treatment) % change in MADRS scores across all treatment arms (p= 0.001), and mediated more extended (Day 30) depression improvements specifically for the ketamine+ASAT arm (group * IAT interaction term: β = –0.201; p= 0.049). Discussion: Our findings suggest that changing implicit self-worth during a post-ketamine ‘plasticity window’ is one key mechanism whereby the novel ketamine+ASAT treatment combination exerts its antidepressant benefit, confirming the intended treatment target at the level of implicit cognition. Future studies should seek to further enhance the reliability of the biobehavioral intervention’s impact on implicit cognition, as this mechanism appears linked to the intervention’s enduring clinical benefits.

Human immunodeficiency virus (HIV) cure efforts are increasingly focused on harnessing CD8+T cell functions, which requires a deeper understanding of CD8+T cells promoting HIV control. Here we identifiy an antigen-responsive TOXhiTCF1+CD39+CD8+T cell population with high expression of inhibitory receptors and low expression of canonical cytolytic molecules. Transcriptional analysis of simian immunodeficiency virus (SIV)-specific CD8+T cells and proteomic analysis of purified CD8+T cell subsets identified TOXhiTCF1+CD39+CD8+T cells as intermediate effectors that retained stem-like features with a lineage relationship with terminal effector T cells. TOXhiTCF1+CD39+CD8+T cells were found at higher frequency than TCF1−CD39+CD8+T cells in follicular microenvironments and were preferentially located in proximity of SIV-RNA+cells. Their frequency was associated with reduced plasma viremia and lower SIV reservoir size. Highly similar TOXhiTCF1+CD39+CD8+T cells were detected in lymph nodes from antiretroviral therapy-naive and antiretroviral therapy-suppressed people living with HIV, suggesting this population of CD8+T cells contributes to limiting SIV and HIV persistence.

Measurable residual disease (MRD) assessed by multiparametric flow cytometry (MFC) has gained importance in clinical decision‐making for acute myeloid leukemia (AML) patients. However, complying with the recent In Vitro Diagnostic Regulations (IVDR) in Europe and Food and Drug Administration (FDA) guidance in the United States requires rigorous validation prior to their use in investigational clinical trials and diagnostics. Validating AML MRD‐MFC assays poses challenges due to the unique underlying disease biology and paucity of patient specimens. In this study, we describe an experimental framework for validation that meets regulatory expectations. Our validation efforts focused on evaluating assay accuracy, analytical specificity, analytical and functional sensitivity (limit of blank (LoB), detection (LLoD) and quantitation (LLoQ)), precision, linearity, sample/reagent stability and establishing the assay background frequencies. Correlation between different MFC methods was highly significant (r= 0.99 for %blasts and r= 0.93 for %LAIPs). The analysis of LAIP specificity accurately discriminated from negative control cells. The assay demonstrated a LoB of 0.03, LLoD of 0.04, and LLoQ of 0.1%. Precision experiments yielded highly reproducible results (Coefficient of Variation <20%). Stability experiments demonstrated reliable measurement of samples up to 96 h from collection. Furthermore, the reference range of LAIP frequencies in non‐AML patients was below 0.1%, ranging from 0.0% to 0.04%. In this manuscript, we present the validation of an AML MFC‐MRD assay using BM/PB patient specimens, adhering to best practices. Our approach is expected to assist other laboratories in expediting their validation activities to fulfill recent health authority guidelines.

Natural science datasets frequently violate assumptions of independence. Samples may be clustered (e.g., by study site, subject, or experimental batch), leading to spurious associations, poor model fitting, and confounded analyses. While largely unaddressed in deep learning, this problem has been handled in the statistics community through mixed effects models, which separate cluster-invariant fixed effects from cluster-specific random effects. We propose a general-purpose framework for Adversarially-Regularized Mixed Effects Deep learning (ARMED) models through non-intrusive additions to existing neural networks: 1) an adversarial classifier constraining the original model to learn only cluster-invariant features, 2) a random effects subnetwork capturing cluster-specific features, and 3) an approach to apply random effects to clusters unseen during training. We apply ARMED to dense, convolutional, and autoencoder neural networks on 4 datasets including simulated nonlinear data, dementia prognosis and diagnosis, and live-cell image analysis. Compared to prior techniques, ARMED models better distinguish confounded from true associations in simulations and learn more biologically plausible features in clinical applications. They can also quantify inter-cluster variance and visualize cluster effects in data. Finally, ARMED matches or improves performance on data from clusters seen during training (5-28% relative improvement) and generalization to unseen clusters (2-9% relative improvement) versus conventional models.

Objectives: Toward common methods for system monitoring and evaluation, we proposed a key performance indicator framework and discussed lessons learned while implementing a statewide exposure notification (EN) system in California during the COVID-19 epidemic.Materials and Methods: California deployed the Google Apple Exposure Notification framework, branded CA Notify, on December 10, 2020, to supplement traditional COVID-19 contact tracing programs. For system evaluation, we defined 6 key performance indicators: adoption, retention, sharing of unique codes, identification of potential contacts, behavior change, and impact. We aggregated and analyzed data from December 10, 2020, to July 1, 2021, in compliance with the CA Notify privacy policy.Results: We estimated CA Notify adoption at nearly 11 million smartphone activations during the study period. Among 1 654 201 CA Notify users who received a positive test result for SARS-CoV-2, 446 634 (27%) shared their unique code, leading to ENs for other CA Notify users who were in close proximity to the SARS-CoV-2–positive individual. We identified at least 122 970 CA Notify users as contacts through this process. Contact identification occurred a median of 4 days after symptom onset or specimen collection date of the user who received a positive test result for SARS-CoV-2.Practice Implications: Smartphone-based EN systems are promising new tools to supplement traditional contact tracing and public health interventions, particularly when efficient scaling is not feasible for other approaches. Methods to collect and interpret appropriate measures of system performance must be refined while maintaining trust and privacy.

AimsWhile interdisciplinary protocols and expedited surgical treatment improve the management of hip fractures in the elderly, the impact of such interventions on patients specifically undergoing arthroplasty for a femoral neck fracture is not clear. We sought to evaluate the efficacy of an interdisciplinary protocol for the management of patients with a femoral neck fracture who are treated with an arthroplasty.MethodsIn 2017, our institution introduced a standardized interdisciplinary hip fracture protocol. We retrospectively reviewed adult patients who underwent hemiarthroplasty (HA) or total hip arthroplasty (THA) for femoral neck fracture between July 2012 and March 2020, and compared patient characteristics and outcomes between those treated before and after the introduction of the protocol.ResultsA total of 157 patients were treated before the introduction of the protocol (35 (22.3%) with a THA), and 114 patients were treated after its introduction (37 (32.5%) with a THA). The demographic details and medical comorbidities were similar in the two groups. Patients treated after the introduction of the protocol had a significantly reduced median time between admission and surgery (22.8 hours (interquartile range (IQR) 18.8 to 27.7) compared with 24.8 hours (IQR 18.4 to 43.3) (p = 0.042), and a trend towards a reduced mean time to surgery (24.1 hours (SD 10.7) compared with 46.5 hours (SD 165.0); p = 0.150), indicating reduction in outliers. Patients treated after the introduction of the protocol had a significantly decreased rate of major complications (4.4% vs 17.2%; p = 0.005), decreased median hospital length of stay in hospital (4.0 days vs 4.8 days; p = 0.008), increased rate of discharge home (26.3% vs 14.7%; p = 0.030), and decreased one-year mortality (14.7% vs 26.3%; p = 0.049). The 90-day readmission rate (18.2% vs 21.7%; p = 0.528) and 30-day mortality (3.7% vs 5.1%; p = 0.767) did not significantly differ. Patients who underwent HA were significantly older than those who underwent THA (82.1 years (SD 10.4) vs 71.1 years (SD 9.5); p < 0.001), more medically complex (mean Charlson Comorbidity Index 6.4 (SD 2.6) vs 4.1 (SD 2.2); p < 0.001), and more likely to develop delirium (8.5% vs 0%; p = 0.024).ConclusionThe introduction of an interdisciplinary protocol for the management of elderly patients with a femoral neck fracture was associated with reduced time to surgery, length of stay, complications, and one-year mortality. Such interventions are critical in improving outcomes and reducing costs for an ageing population.Cite this article: Bone Joint J2021;103-B(7 Supple B):3–8.

Exceptional clinical responses produced by the first chimeric antigen receptor T [CAR‐T] cell therapies, and their entry into commercial markets prompted a logarithmic increase in the number of next generation CAR‐T clinical trials. As a result, there is a growing interest in understanding the analytical approaches utilized for reliable monitoring of these “living” drugs, and the challenges encountered during their clinical development. Multiparametric flow cytometry (MFC) assays have played a crucial role in understanding the phenotype and function of first approved CAR‐T therapies. Herein, three main areas for monitoring CAR‐T therapies in clinical trials are discussed: (1) analytical considerations critical for development of MFC assays for the reliable enumeration of CAR‐T levels, (2) operational challenges associated with clinical trial sampling and transportation, and (3) differential cellular kinetics observed by MFC and qPCR analyses and their relationship with efficacy (measurable residual disease levels). Initial experiences described here may enable design of fit‐for‐purpose tools and help to more rapidly advance the development of next generation CAR‐T therapies.

Background: There is controversy regarding the effectiveness of postoperative antibiotics to prevent wound infection. Some surgeons still use a routine postoperative oral antibiotic regimen. The purpose of this study was to review a series of cases and document statistically any difference in infection rates and whether routine postoperative antibiotics in foot and ankle surgery are justified.Methods: A retrospective chart review of 649 patients was performed who underwent elective foot and ankle surgery. Six hundred thirty-one patient charts were included in the final analysis. Evaluated were patients who did and did not receive postoperative oral antibiotics in order to identify whether a difference in infection rate or wound healing occurred. The study also evaluated risk factors for developing infection following foot and ankle surgery.Results: The number of infections in patients receiving postoperative oral antibiotics was 6 (3%), while the number of infections in those who did not receive postoperative oral antibiotics was 10 (2%) (P= .597). The difference of deep versus superficial infections and delays in wound healing between the 2 groups was not statistically significant. Patients who developed infections were older and had a higher prevalence of hypertension, a history of neoplasm, and a greater American Society of Anesthesiologists Classification of Physical Health.Conclusion: This study suggests that routine use of postoperative antibiotics in foot and ankle surgery does not affect wound complications or infection rates. Additionally, patients who are older and those with multiple medical problems may be at higher risk for developing postoperative infection following foot and ankle surgeries.Level of Evidence: Level III, retrospective comparative series.

Recent research has supported the use of student-centered teaching practices, such as active learning, because of its effectiveness in improving student learning and retention when compared with traditional, lecture-based teaching practices. Despite evidence supporting the effectiveness of active learning in improving STEM undergraduate education, the adoption of active learning by instructors has been slow for reasons, including negative student response to active learning. In this systematic literature review, we examine students’negative responses to active learning and reasons for the negative responses noted in 57 published STEM studies. Our findings identify three types of negative responses: affect, engagement, and evaluation. The reasons behind negative response represented six overarching categories based on student feedback: limited value, lack of time, difficulty and increased workload, lack of guidance, logistical difficulties, unfamiliarity with active learning, lack of preparation, and confidence. We leverage different theoretical perspectives to explain the reasons behind negative responses and offer insights for lowering the barrier for instructors to adopt active learning in STEM classrooms.

The primary human immunodeficiency virus (HIV) reservoir is composed of resting memory CD4+T cells, which often express the immune checkpoint receptors programmed cell death protein 1 (PD-1) and cytotoxic T lymphocyte-associated protein 4 (CTLA-4), which limit T cell activation via synergistic mechanisms. Using simian immunodeficiency virus (SIV)-infected, long-term antiretroviral therapy (ART)-treated rhesus macaques, we demonstrate that PD-1, CTLA-4 and dual CTLA-4/PD-1 immune checkpoint blockade using monoclonal antibodies is well tolerated, with evidence of bioactivity in blood and lymph nodes. Dual blockade was remarkably more effective than PD-1 blockade alone in enhancing T cell cycling and differentiation, expanding effector-memory T cells and inducing robust viral reactivation in plasma and peripheral blood mononuclear cells. In lymph nodes, dual CTLA-4/PD-1 blockade, but not PD-1 alone, decreased the total and intact SIV-DNA in CD4+T cells, and SIV-DNA and SIV-RNA in B cell follicles, a major site of viral persistence during ART. None of the tested interventions enhanced SIV-specific CD8+T cell responses during ART or viral control after ART interruption. Thus, despite CTLA-4/PD-1 blockade inducing robust latency reversal and reducing total levels of integrated virus, the degree of reservoir clearance was still insufficient to achieve viral control. These results suggest that immune checkpoint blockade regimens targeting PD-1 and/or CTLA-4, if performed in people living with HIV with sustained aviremia, are unlikely to induce HIV remission in the absence of additional interventions.

Targeted therapeutics are needed for triple-negative breast cancer (TNBC). In this study, we investigated the activation of Src family of cytoplasmic tyrosine kinases (SFKs) and two SFK substrates—CUB-domain containing protein 1 (CDCP1) and protein kinase C δ (PKCδ)—in 56 formalin-fixed, paraffin-embedded (FFPE) TNBCs. Expression of SFK phosphorylated at Y416 (SFK_pY416+) in tumor cells was strongly associated with phosphorylation of CDCP1 and PKCδ (CDCP1_ pY743+and PKCδ_pY311+), as assessed by immunohistochemistry, indicating increased SFK activity in situ. To enable biochemical analysis, protein extraction from FFPE tissue was optimized. Cleaved CDCP1 isoform (70 kDa) was expressed to a varying degree in all samples but only phosphorylated in TNBC tumor cells that expressed SFK_pY416. Interestingly, active SFK was found to be biphosphorylated (SFK_pY416+/pY527+). Biphosphorylated active SFK was observed more frequently in forkhead box protein A1 (FOXA1)−TNBCs. In addition, in SFK_pY416−samples, FOXA1+TNBC tended to be SFK_pY527+(classic inactive SFK), and FOXA1−TNBC tended to be SFK_pY527−(SFK poised for activation). Strong SFK_pY416 staining was also observed in tumor-infiltrating lymphocytes in a subset of TNBCs with high tumor-infiltrating lymphocyte content. This report will facilitate protein biochemical analysis of FFPE tumor samples and justifies the development of therapies targeting the SFK/CDCP1/PKCδ pathway for TNBC treatment.