Implementation Science to Achieve Equity in Heart Failure Care: A Scientific Statement From the American Heart Association.

Guideline-directed medical therapies and guideline-directed nonpharmacological therapies improve quality of life and survival in patients with heart failure (HF), but eligible patients, particularly women and individuals from underrepresented racial and ethnic groups, are often not treated with these therapies. Implementation science uses evidence-based theories and frameworks to identify strategies that facilitate uptake of evidence to improve health. In this scientific statement, we provide an overview of implementation trials in HF, assess their use of conceptual frameworks and health equity principles, and provide pragmatic guidance for equity in HF. Overall, behavioral nudges, multidisciplinary care, and digital health strategies increased uptake of therapies in HF effectively but did not include equity goals. Few HF studies focused on achieving equity in HF by engaging stakeholders, quantifying barriers and facilitators to HF therapies, developing strategies for equity informed by theory or frameworks, evaluating implementation measures for equity, and titrating strategies for equity. Among these HF equity studies, feasibility was established in using various educational strategies to promote organizational change and equitable care. A couple include ongoing randomized controlled pragmatic trials for HF equity. There is great need for additional HF implementation trials designed to promote delivery of equitable guideline-directed therapy.

Genetic risk impacts the association of menopausal hormone therapy with colorectal cancer risk.

BACKGROUND: Menopausal hormone therapy (MHT), a common treatment to relieve symptoms of menopause, is associated with a lower risk of colorectal cancer (CRC). To inform CRC risk prediction and MHT risk-benefit assessment, we aimed to evaluate the joint association of a polygenic risk score (PRS) for CRC and MHT on CRC risk. METHODS: We used data from 28,486 postmenopausal women (11,519 cases and 16,967 controls) of European descent. A PRS based on 141 CRC-associated genetic variants was modeled as a categorical variable in quartiles. Multiplicative interaction between PRS and MHT use was evaluated using logistic regression. Additive interaction was measured using the relative excess risk due to interaction (RERI). 30-year cumulative risks of CRC for 50-year-old women according to MHT use and PRS were calculated. RESULTS: The reduction in odds ratios by MHT use was larger in women within the highest quartile of PRS compared to that in women within the lowest quartile of PRS (p-value = 2.7 × 10-8). At the highest quartile of PRS, the 30-year CRC risk was statistically significantly lower for women taking any MHT than for women not taking any MHT, 3.7% (3.3%-4.0%) vs 6.1% (5.7%-6.5%) (difference 2.4%, P-value = 1.83 × 10-14); these differences were also statistically significant but smaller in magnitude in the lowest PRS quartile, 1.6% (1.4%-1.8%) vs 2.2% (1.9%-2.4%) (difference 0.6%, P-value = 1.01 × 10-3), indicating 4 times greater reduction in absolute risk associated with any MHT use in the highest compared to the lowest quartile of genetic CRC risk. CONCLUSIONS: MHT use has a greater impact on the reduction of CRC risk for women at higher genetic risk. These findings have implications for the development of risk prediction models for CRC and potentially for the consideration of genetic information in the risk-benefit assessment of MHT use.

Structural basis for Retriever-SNX17 assembly and endosomal sorting.

During endosomal recycling, Sorting Nexin 17 (SNX17) facilitates the transport of numerous membrane cargo proteins by tethering them to the Retriever complex. Despite its importance, the mechanisms underlying this interaction have remained elusive. Here, we report the structure of the Retriever-SNX17 complex determined using cryogenic electron microscopy (cryo-EM). Our structure reveals that the C-terminal tail of SNX17 engages with a highly conserved interface between the VPS35L and VPS26C subunits of Retriever. Through comprehensive biochemical, cellular, and proteomic analyses, we demonstrate that disrupting this interface impairs the Retriever-SNX17 interaction, subsequently affecting the recycling of SNX17-dependent cargos and altering the composition of the plasma membrane proteome. Intriguingly, we find that the SNX17-binding pocket on Retriever can be utilized by other ligands that share a consensus acidic C-terminal tail motif. By showing how SNX17 is linked to Retriever, our findings uncover a fundamental mechanism underlying endosomal trafficking of critical cargo proteins and reveal a mechanism by which Retriever can engage with other regulatory factors.

Stakeholder perspectives of mental healthcare services in Bangladesh, its challenges and opportunities: a qualitative study.

This study explores Bangladesh's mental health services from an individual- and system-level perspective and provides insights and recommendations for strengthening it's mental health system. We conducted 13 in-depth interviews and 2 focus group discussions. Thirty-one participants were recruited using a combination of purposive and snowball sampling methods. All interviews and group discussions were audio-recorded and transcribed, and key findings were translated from Bengali to English. Data were coded manually and analysed using a thematic and narrative analysis approach. Stakeholders perceived scarcity of service availability at the peripheral level, shortage of professionals, weak referral systems, lack of policy implementation and regulatory mechanisms were significant challenges to the mental health system in Bangladesh. At the population level, low levels of mental health literacy, high societal stigma, and treatment costs were barriers to accessing mental healthcare. Key recommendations included increasing the number of mental health workers and capacity building, strengthening regulatory mechanisms to enhance the quality of care within the health systems, and raising awareness about mental health. Introducing measures that relate to tackling stigma, mental health literacy as well as building the capacity of the health workforce and governance systems will help ensure universal mental health coverage.

Karyotypic changes and diversification time in Epinephelidae groupers (Perciformes). Implications on reproductive isolation.

Groupers (Epinephelidae and Serranidae) have attracted special attention to fish farming, and their species offer good opportunities for successful hybridizations. Cytogenetic data allow a better understanding of the role of karyotypic diversification in the acquisition of post-zygotic reproductive isolation (RI). Thus, chromosomal analyses were performed on E. striatus (Caribbean Sea), E. coioides and E. tauvina (Indo-Pacific Region), using standard procedures and mapping of six repetitive DNA classes by the in situ hybridization. The three species have 2n=48 chromosomes. The karyotypes of E. coioides and E. striatus are composed only of acrocentric chromosomes (FN=48), while E. tauvina has 8 submetacentric chromosomes (FN=56). Heterochromatin has a preferential centromeric distribution, and the microsatellite repeats are dispersed throughout the chromosomes of all species. The 18S and 5S rDNA sites are unique but show a colocalization arrangement in E. tauvina and E. striatus. The chromosomal organization suggests that the three species still maintain a significant amount of syntenic regions. The range of the karyotype divergence and the RI levels showed low, but goes turn proportionally greater in relation to the divergence time between the parental species. The slow acquisition of postzygotic RI is consistent with the high karyotype homogeneity presented by Epinephelidae family.

Yeast TLDc domain proteins regulate assembly state and subcellular localization of the V-ATPase.

Yeast vacuoles perform crucial cellular functions as acidic degradative organelles, storage compartments, and signaling hubs. These functions are mediated by important protein complexes, including the vacuolar-type H+-ATPase (V-ATPase), responsible for organelle acidification. To gain a more detailed understanding of vacuole function, we performed cross-linking mass spectrometry on isolated vacuoles, detecting many known as well as novel protein-protein interactions. Among these, we identified the uncharacterized TLDc-domain-containing protein Rtc5 as a novel interactor of the V-ATPase. We further analyzed the influence of Rtc5 and of Oxr1, the only other yeast TLDc-domain-containing protein, on V-ATPase function. We find that both Rtc5 and Oxr1 promote the disassembly of the vacuolar V-ATPase in vivo, counteracting the role of the RAVE complex, a V-ATPase assembly chaperone. Furthermore, Oxr1 is necessary for the retention of a Golgi-specific subunit of the V-ATPase in this compartment. Collectively, our results shed light on the in vivo roles of yeast TLDc-domain proteins as regulators of the V-ATPase, highlighting the multifaceted regulation of this crucial protein complex.

Direct Probe of Vibrational Fingerprint and Combination Band Coupling.

Vibrational fingerprints and combination bands are a direct measure of couplings that control molecular properties. However, most combination bands possess small transition dipoles. Here we use multiple, ultrafast coherent infrared pulses to resolve vibrational coupling between CH3CN fingerprint modes at 918 and 1039 cm-1 and combination bands in the 2750-6100 cm-1 region via doubly vibrationally enhanced (DOVE) coherent multidimensional spectroscopy (CMDS). This approach provides a direct probe of vibrational coupling between fingerprint modes and near-infrared combination bands of large and small transition dipoles in a molecular system over a large frequency range.

Triboelectric Nanogenerators for the Masses: A Low-Cost Do-It-Yourself Pulsed Ion Source for Sample-Limited Applications.

Triboelectric nanogenerators (TENG) are useful devices for converting mechanical motion into electric current using readily available materials. Though the applications for these devices span across many fields, TENG can be leveraged for mass spectrometry (MS) as inexpensive and effective power supplies for pulsed nanoelectrospray ionization (nESI). The inherently discontinuous spray provided by TENG is particularly useful in scenarios where high sample economy is imperative, as in the case of ultraprecious samples. Previous work has shown the utility of TENG MS as a highly sensitive technique capable of yielding quality spectra from only a few microliters of sample at low micromolar concentrations. As the field of miniaturized, fieldable mass spectrometers grows, it remains critical to develop advanced ion sources with similarly small power requirements and footprints. Here, we present a redesigned TENG ion source with a sub-1000 USD material cost, lower power consumption, reduced footprint, and improved capabilities. We validate the performance of this new device for a diverse set of applications, including lipid double bond localization and native protein analysis.

Estrogen Receptor, Progesterone Receptor, and Human Epidermal Growth Factor Receptor 2 Expression Rates in Invasive Breast Carcinoma: A Study of 21 Institutions.

CONTEXT.­: Laboratories performing predictive marker testing for breast carcinoma are encouraged to compare patient results to published benchmarks. OBJECTIVE.­: To collect expression rates for estrogen receptor (ER), progesterone receptor (PgR), and human epidermal growth factor receptor 2 (HER2) in invasive breast carcinoma from multiple laboratories. DESIGN.­: Participants submitted data from up to 50 primary cases during the study period. Participants reported ER, PgR, and HER2 results in addition to demographic and histologic information. Participants also provided annual institution-level expression rates. RESULTS.­: A total of 21 institutions submitted data for 687 cases. Aggregate positivity rates for ER and PgR were 85.6% and 75.1%, respectively. Receptor positivity rates were higher in well-differentiated (grade 1) tumors (ER, 97.4%; PgR, 88.0%) compared with moderately differentiated (grade 2) tumors (ER, 92.4%; PgR, 84.0%) and poorly differentiated (grade 3) tumors (ER, 61.8%; PgR, 48.0%). Expression rates were higher in postmenopausal women (ER, 87.2%) than premenopausal women (ER, 79.6%) and higher in lobular carcinomas (ER, 98.7%; PgR, 85.3%) than ductal carcinomas (ER, 84.1%; PgR, 74.5%). The aggregate HER2 positivity (score 3+) rate was 9.0%. The aggregate HER2 equivocal (score 2+) rate was 14.5%. Of 81 equivocal (score 2+) cases, 70 (86.4%) were nonamplified. CONCLUSIONS.­: The data from this study provide multi-institutional benchmark data to assist laboratories performing periodic comparisons as part of a quality management program. Overall expression rates were generally similar to those of other published reports, with the exception of the ER-negative and HER2-positive rates, both of which were somewhat lower.

The loss of the PDIM/PGL virulence lipids causes differential secretion of ESX-1 substrates in Mycobacterium marinum.

The mycobacterial cell envelope is a major virulence determinant in pathogenic mycobacteria. Specific outer lipids play roles in pathogenesis, modulating the immune system and promoting the secretion of virulence factors. ESX-1 (ESAT-6 system-1) is a conserved protein secretion system required for mycobacterial pathogenesis. Previous studies revealed that mycobacterial strains lacking the outer lipid PDIM have impaired ESX-1 function during laboratory growth and infection. The mechanisms underlying changes in ESX-1 function are unknown. We used a proteo-genetic approach to measure phthiocerol dimycocerosate (PDIM)- and phenolic glycolipid (PGL)-dependent protein secretion in M. marinum, a non-tubercular mycobacterial pathogen that causes tuberculosis-like disease in ectothermic animals. Importantly, M. marinum is a well-established model for mycobacterial pathogenesis. Our findings showed that M. marinum strains without PDIM and PGL showed specific, significant reductions in protein secretion compared to the WT and complemented strains. We recently established a hierarchy for the secretion of ESX-1 substrates in four (I-IV) groups. Loss of PDIM differentially impacted secretion of Group III and IV ESX-1 substrates, which are likely the effectors of pathogenesis. Our data suggest that the altered secretion of specific ESX-1 substrates is responsible for the observed ESX-1-related effects in PDIM-deficient strains.IMPORTANCEMycobacterium tuberculosis, the cause of human tuberculosis, killed an estimated 1.3 million people in 2022. Non-tubercular mycobacterial species cause acute and chronic human infections. Understanding how these bacteria cause disease is critical. Lipids in the cell envelope are essential for mycobacteria to interact with the host and promote disease. Strains lacking outer lipids are attenuated for infection, but the reasons are unclear. Our research aims to identify a mechanism for attenuation of mycobacterial strains without the PDIM and PGL outer lipids in M. marinum. These findings will enhance our understanding of the importance of lipids in pathogenesis and how these lipids contribute to other established virulence mechanisms.

Anti-tumor activity of a novel LAIR1 antagonist in combination with anti-PD-1 to treat collagen-rich solid tumors.

We recently reported that resistance to PD-1-blockade in a refractory lung cancer-derived model involved increased collagen deposition and the collagen-binding inhibitory receptor leukocyte-associated immunoglobulin-like receptor 1 (LAIR1), and thus we hypothesized that LAIR1 and collagen cooperated to suppress therapeutic response. Here, we report LAIR1 is associated with tumor stroma and is highly expressed by intratumoral myeloid cells in both human tumors and mouse models of cancer. Stroma-associated myeloid cells exhibit a suppressive phenotype and correlate with LAIR1 expression in human cancer. NGM438, a novel humanized LAIR1 antagonist monoclonal antibody, elicits myeloid inflammation and allogeneic T cell responses by binding to LAIR1 and blocking collagen engagement. Further, a mouse-reactive NGM438 surrogate antibody sensitized refractory KP mouse lung tumors to anti-PD-1 therapy and resulted in increased intratumoral CD8+ T cell content and inflammatory gene expression. These data place LAIR1 at the intersection of stroma and suppressive myeloid cells and support the notion that blockade of the LAIR1/collagen axis can potentially address resistance to checkpoint inhibitor therapy in the clinic.

Statine-based peptidomimetic compounds as inhibitors for SARS-CoV-2 main protease (SARS-CoV­2 Mpro).

COVID-19 is a multisystemic disease caused by the SARS-CoV-2 airborne virus, a member of the Coronaviridae family. It has a positive sense single-stranded RNA genome and encodes two non-structural proteins through viral cysteine-proteases processing. Blocking this step is crucial to control virus replication. In this work, we reported the synthesis of 23 statine-based peptidomimetics to determine their ability to inhibit the main protease (Mpro) activity of SARS-CoV-2. Among the 23 peptidomimetics, 15 compounds effectively inhibited Mpro activity by 50% or more, while three compounds (7d, 8e, and 9g) exhibited maximum inhibition above 70% and IC50 < 1 µM. Compounds 7d, 8e, and 9g inhibited roughly 80% of SARS-CoV-2 replication and proved no cytotoxicity. Molecular docking simulations show putative hydrogen bond and hydrophobic interactions between specific amino acids and these inhibitors. Molecular dynamics simulations further confirmed the stability and persisting interactions in Mpro's subsites, exhibiting favorable free energy binding (ΔGbind) values. These findings suggest the statine-based peptidomimetics as potential therapeutic agents against SARS-CoV-2 by targeting Mpro.

Organization of spontaneous spatial behaviors under dark conditions is unaffected in adult male and female long-Evans rats after moderate prenatal alcohol exposure.

Prenatal alcohol exposure can produce disruptions in a wide range of cognitive functions, but it is especially detrimental to spatial navigation. In open environments, rodents organize their spatial behaviors around centralized locations, termed home bases, from which they make circuitous and slow locomotor trips (progressions) into the rest of the environment. Open-field behaviors are organized even under darkened test conditions, suggesting a role for self-motion cues (vestibular, motor, etc.). The impact of moderate prenatal alcohol exposure (mPAE) on the organization of spontaneous open-field behaviors under darkened conditions has not been investigated. Here we tested adult female and male rats with mPAE or saccharin control exposure in a circular open field for 30 min in a testing room that was made completely dark. While general locomotion, as measured by reductions in travel distance and increased stop duration, decreased across the test session, the organization of these behaviors, as measured by stop duration, home base establishment, home base stability, progression accuracy, and scaling of peak speeds with progression length, did not differ between mPAE and saccharin control rats. Together, the findings strongly suggest that spontaneous movement organization in relation to self-motion cues remains intact in adult mPAE rats. (PsycInfo Database Record (c) 2024 APA, all rights reserved).

Requirements for efficient endosomal escape by designed mini-proteins.

ZF5.3 is a compact, rationally designed mini-protein that escapes efficiently from the endosomes of multiple cell types. Despite its small size (27 amino acids), ZF5.3 can be isolated intact from the cytosol of treated cells and guides multiple classes of proteins into the cytosol and/or nucleus. In the best cases, delivery efficiencies reach or exceed 50% to establish nuclear or cytosolic concentrations of 500 nM or higher. But other than the requirement for unfoldable cargo and an intact HOPS complex, there is little known about how ZF5.3 traverses the limiting endocytic membrane. Here we delineate the attributes of ZF5.3 that enable efficient endosomal escape. We confirm that ZF5.3 is stable at pH values between 5.5 and 7.5, with no evidence of unfolding even at temperatures as high as 95 °C. The high-resolution NMR structure of ZF5.3 at pH 5.5, also reported here, shows a canonical p zinc-finger fold with the penta-arg motif integrated seamlessly into the C-terminal α-helix. At lower pH, ZF5.3 unfolds cooperatively as judged by both circular dichroism and high-resolution NMR. Unfolding occurs upon protonation of a single Zn(II)-binding His side chain whose pKa corresponds almost exactly to that of the late endosomal lumen. pH-induced unfolding is essential for endosomal escape, as a ZF5.3 analog that remains folded at pH 4.5 fails to efficiently reach the cytosol, despite high overall uptake. Finally, using reconstituted liposomes, we identify a high-affinity interaction of ZF5.3 with a specific lipid-BMP-that is selectively enriched in the inner leaflet of late endosomal membranes. This interaction is 10-fold stronger at low pH than neutral pH, providing a molecular picture for why escape occurs preferentially and in a HOPS-dependent manner from late endosomal compartments. The requirements for programmed endosomal escape identified here should aid and inform the design of proteins, peptidomimetics, and other macromolecules that reach cytosolic or nuclear targets intact and at therapeutically relevant concentrations.

Cas9 RNP Physiochemical Analysis for Enhanced CRISPR-AuNP Assembly and Function.

CRISPR therapy for hematological disease has proven effective for transplant dependent beta thalassemia and sickle cell anemia, with additional disease targets in sight. The success of these therapies relies on high rates of CRISPR-induced double strand DNA breaks in hematopoietic stem and progenitor cells (HSPC). To achieve these levels, CRISPR complexes are typically delivered by electroporation ex vivo which is toxic to HSPCs. HSPCs are then cultured in stimulating conditions that promote error-prone DNA repair, requiring conditioning with chemotherapy to facilitate engraftment after reinfusion. In vivo delivery by nanocarriers of CRISPR gene editing tools has the potential to mitigate this complexity and toxicity and make this revolutionary therapy globally available. To achieve in vivo delivery, the inherent restriction factors against oligonucleotide delivery into HSPCs, that make ex vivo manipulation including electroporation and stimulation essential, must be overcome. To this end, our group developed a CRISPR carrying gold nanoparticle (CRISPR-AuNP) capable of delivering either Cas9 or Cas12a CRISPRs as ribonucleoprotein complexes (RNP) without compromising HSPC fitness. However, the most commonly used CRISPR, Cas9, demonstrated inconsistent activity in this delivery system, with lower activity relative to Cas12a. Investigation of Cas9 RNP biophysics relative to Cas12a revealed duplex RNA instability during the initial loading onto Au cores, resulting in undetectable Cas9 loading to the particle surface. Here we demonstrate preformation of RNP before loading, coupled with optimization of the loading chemistry and conditions, resulted in 39.6 ± 7.0 Cas9 RNP/AuNP without compromising RNP activity in both in vitro assays and primary human HSPC. The same alterations improved Cas12a RNP/AuNP loading 10-fold over previously reported levels. To achieve particle stability, the reported polyethyleneimine outer coating was altered to include PEGylation and the resulting 2nd generation CRISPR-AuNP demonstrates favorable nanoformulation characteristics for in vivo administration, with a hydrophilic, more neutral nanoparticle surface. Direct treatment of HSPC in vitro showed 72.5 ± 7.37% uptake of 2nd generation CRISPR-AuNP in primary human HSPC, but with endosomal accumulation and low rates of gene editing consistent with low levels of endosomal escape.

Metabologenomic characterization uncovers a clinically aggressive IDH mutant glioma subtype.

Mutations in the pivotal metabolic isocitrate dehydrogenase (IDH) enzymes are recognized to drive the molecular footprint of diffuse gliomas, and patients with IDH mutant gliomas have overall favorable outcomes compared to patients with IDH wild-type tumors. However, survival still varies widely among patients with IDH mutated tumors. Here, we aimed to characterize molecular signatures that explain the range of IDH mutant gliomas. By integrating matched epigenome-wide methylome, transcriptome, and global metabolome data in 154 patients with gliomas, we identified a group of IDH mutant gliomas with globally altered metabolism that resembled IDH wild-type tumors. IDH-mutant gliomas with altered metabolism have significantly shorter overall survival from their IDH mutant counterparts that is not fully accounted for by recognized molecular prognostic markers of CDKN2A/B loss and glioma CpG Island Methylator Phenotype (GCIMP) status. IDH-mutant tumors with dysregulated metabolism harbored distinct epigenetic alterations that converged to drive proliferative and stem-like transcriptional profiles, providing a window to target novel dependencies in gliomas.

The top-cited military relevant spine articles.

Introduction: Historically musculoskeletal injury has substantially affected United States (US) service members. Lumbosacral spine injuries are among the most common sites of injury for service members across all US military branches and usually presents with pain in the lower back and extremities. The aim of this study is to identify and describe the 50 most-cited articles relevant to military medicine on the subject of the spine. Methods: In April 2020 Web of Science was used to search the key words: spinal cord injury, spine, thoracic spine, lumbar spine, cervical spine, sacrum, sacral, cervical fusion, lumbar fusion, sacral fracture, combat, back pain, neck pain, and military. Articles published from 1900 to 2020 were evaluated for relevance to military spine orthopaedics and ranked based on citation number. The 50 most-cited articles were characterized based on country of origin, journal of publication, affiliated institution, topic, military branch, and conflict. Results: 1900 articles met search criteria. The 50 most-cited articles were cited 24 to 119 times and published between 1993 and 2017. 30 articles (60%) originated in the United States. Aviation, Space, and Environmental Medicine accounted for the most frequent (n = 10) destination journal followed by Spine (n = 8). 37 institutions contributed to the top 50 most-cited articles. The most common article type was clinically focused retrospective analysis 36% (n = 18), clinically focused cohort study 10% (n = 5), and clinically focused cohort questionnaire, cross-sectional analysis, and randomized study 8% each (n = 4). 90% of articles were non-surgical (n = 45). The US Army had the greatest number of associated articles. Operation Iraqi Freedom and Operation Enduring Freedom were the most-cited conflicts. Conclusion: The 50 most-cited articles relevant to military spine orthopaedics are predominantly clinically focused, arising from the US, and published in Aviation, Space, and Environmental Medicine, Spine, and The Spine Journal.

Non-Operative Treatment of Humeral Shaft Fractures with Immediate Functional Bracing versus Coaptation Splinting and Delayed Functional Bracing: A Retrospective Study.

OBJECTIVES: To compare radiographic and clinical outcomes in non-operative management of humeral shaft fractures treated initially with coaptation splinting (CS) followed by delayed functional bracing (FB) versus treatment with immediate FB. METHODS: Design: Retrospective cohort study. SETTING: Academic Level 1 Trauma Center. PATIENT SELECTION CRITERIA: Patients with closed humeral shaft fractures managed non-operatively with initial CS followed by delayed FB or with immediate FB from 2016 to 2022. Patients younger than 18 years and/or with less than 3 months follow-up were excluded.Outcome Measures and Comparisons: The primary outcome was coronal and sagittal radiographic alignment assessed at final follow-up. Secondary outcomes included rate of failure of non-operative management (defined as surgical conversion and/or fracture nonunion), fracture union, and skin complications secondary to splint/brace wear. RESULTS: Ninety-seven patients were managed non-operatively with delayed FB (n=58) or immediate FB (n=39). Overall, the mean age was 49.9 years (range 18-94), and 64 (66%) patients were female. The immediate FB group had less smokers (p=0.003) and lower incidence of radial nerve palsy (p=0.025), with more proximal third humeral shaft fractures (p=0.001). There were no other significant differences in demographic or clinical characteristics (p>0.05). There were no significant differences in coronal (p=0.144) or sagittal (p=0.763) radiographic alignment between groups. In total, 33 (34.0%) humeral shaft fractures failed non-operative management, with 11 (28.2%) in the immediate FB group, and 22 (37.9%) in the delayed FB group (p=0.322). There were no significant differences in fracture union (p=0.074) or skin complications (p=0.259) between groups. CONCLUSIONS: This study demonstrated that non-operative treatment of humeral shaft fractures with immediate functional bracing did not result in significantly different radiographic or clinical outcomes compared to treatment with coaptation splinting followed by delayed functional bracing. Future prospective studies assessing patient reported outcomes (PROs) will further guide clinical decision making. LEVEL OF EVIDENCE: Therapeutic Level III. See Instructions for Authors for a complete description of levels of evidence.