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Polyglutamine (polyQ) sequences undergo repeat-length dependent formation of disease-associated, amyloid-like cross-β core structures with kinetics and aggregate morphologies often influenced by the flanking sequences. In Huntington's disease (HD), the htt NT segment on the polyQ's N-terminal flank enhances aggregation rates by changing amyloid nucleation from a classical homogeneous mechanism to a two-step process requiring an ɑ-helix-rich oligomeric intermediate. A folded, helix-rich htt NT tetrameric structure suggested to be this critical intermediate was recently reported. Here we employ single alanine replacements along the htt NT sequence to assess this proposed structure and refine the mechanistic model. We find that Ala replacement of hydrophobic residues within simple htt NT peptides greatly suppresses helicity, supporting the tetramer model. These same helix-disruptive replacements in the htt NT segment of an exon-1 analog greatly reduce aggregation kinetics, suggesting that an ɑ-helix rich multimer - either the tetramer or a larger multimer - plays an on-pathway role in nucleation. Surprisingly, several other Ala replacements actually enhance helicity and/or amyloid aggregation. The spatial localization of these residues on the tetramer surface suggests a self-association interface responsible for formation of the octomers and higher-order multimers most likely required for polyQ amyloid nucleation. Multimer docking of the tetramer, using the protein-protein docking algorithm ClusPro, predicts this symmetric surface to be a viable tetramer dimerization interface. Intriguingly, octomer formation brings the emerging polyQ chains into closer proximity at this tetramer-tetramer interface. Further supporting the potential importance of tetramer super-assembly, computational docking with a known exon-1 aggregation inhibitor predicts ligand contacts with residues at this interface., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Patients in intensive care units are frequently supported by mechanical ventilation. There is increasing awareness of patient-ventilator dyssynchrony (PVD), a mismatch between patient respiratory effort and assistance provided by the ventilator, as a risk factor for infection, narcotic exposure, lung injury, and adverse neurocognitive effects. One of the most injurious consequences of PVD are double cycled (DC) breaths when two breaths are delivered by the ventilator instead of one. Prior efforts to identify PVD have limited efficacy. An automated method to identify PVD, independent of clinician expertise, acumen, or time, would potentially permit early, targeted treatment to avoid further harm. We performed secondary analyses of data from a clinical trial of children with acute respiratory distress syndrome. Waveforms of ventilator flow, airway pressure and esophageal manometry were annotated to identify DC breaths and underlying PVD subtypes. Spectrograms were generated from those waveforms to train Convolutional Neural Network (CNN) models in detecting DC and underlying PVD subtypes: Reverse Trigger (RT) and Inadequate Support (IS). The DC breath detection model yielded AUROC of 0.980, while the multi-target detection model for underlying dyssynchrony yielded AUROC of 0.980 (RT) and 0.976 (IS). When operating at 75% sensitivity, DC breath detection had a number needed to alert (NNA) 1.3 (99% specificity), while underlying PVD had a NNA 1.6 (98.5% specificity) for RT and NNA 4.0 (98.2% specificity) for IS. CNNs using spectrograms of ventilator waveforms can identify DC breaths and detect the underlying PVD for targeted clinical interventions., Competing Interests: Declaration of interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Purpose: The treatment of nonunion of long bones is difficult particularly in the presence of infection, which often involves staged surgical management. There is limited literature to compare the post operative course and outcomes of patients treated for septic versus aseptic nonunion. Thus, the purpose of this study was to determine if a difference exists between the number of surgical procedures, time to union, and rate of successful union for these two groups., Methods: A retrospective cohort study was performed at a single tertiary care center. Patients suffering nonunion of the humerus, tibia and femur were included. Patient demographic data and characteristics of the post operative course were collected to include number and reason for repeat operations, antibiotic course, time to union, and development of a successful union., Results: About 28 of 122 patients had septic nonunion. After diagnosis of nonunion, the septic group averaged 3.9 surgeries compared to 1.5 in the aseptic group (p < 0.001). There was no difference in the rate of successful union (79.8% versus 85.7%; p = 0.220), though the septic group took 129 days longer on average for successful union. (376 versus 247; p = 0.018)., Conclusion: Septic nonunion of long bones is associated with the need for significantly more operations as well as time to union, though union rates remain similar. The identification of infection is critical for both the appropriate treatment as well as counseling patients on the expected post operative course., (© 2022. The Author(s), under exclusive licence to Springer-Verlag France SAS, part of Springer Nature.)

Intramedullary nailing of humerus fractures has evolved over the past half century and has grown in popularity especially for the polytraumatized patient. The importance of restoring appropriate rotational alignment is equivalent to that of restoring sagittal and coronal alignment to decrease the risk of shoulder degenerative changes and limit range of motion discrepancy from the contralateral limb. This technique is designed to introduce an intraoperative fluoroscopic method to obtain adequate rotational alignment of humeral shaft fractures treated with closed antegrade humeral locked nailing., Competing Interests: Conflicts of Interest and Source of Funding: R.W. is a paid consultant for Stryker and Smith and Nephew. For the remaining authors none were declared., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

Objective: The aim of this work is to evaluate the usefulness of using augmented reality (AR) to train medical professionals in radiological protection (RP) in fluoroscopy., Methods: A Microsoft HoloLens 2 device has been used to simulate a fluoroscopic device. The teaching scenario considers a Philips Azurion able to rotate to pre-defined gantry positions,a dorsal decubitus patient and a ceiling shield. Radiation exposures were simulated using the FLUKA Monte Carlo code. 11 radiologists were asked to reproduce their positioning as per a clinical procedure and to correctly position the ceiling shield. Then, they were presented with the radiation exposure of their choices and were able to further optimise it. After the session, they were asked to complete a questionnaire., Results: Users rated the AR educational approach as Intuitive and relevant to RP education (35%) and inspiring to deepen their knowledge (18%). Nevertheless, a negative aspect was mainly the difficulty in dealing with the system (58%). Although the participants were radiologists, a minority recognised themselves as having accurate knowledge of the RP (18%), indicating a relevant knowledge gap., Conclusion: The usefulness of using AR in RP education for radiologists has been shown. The visual aid of such technology is likely to improve the consolidation of practical knowledge., Advances in Knowledge: The use of interactive teaching techniques has the possibility to both help radiology professionals consolidate their radiation protection training and confidence in their practices.

Background: Wound complications are common after resection of soft tissue sarcomas, with published infection rates ranging from 10% to 35%. Multiple studies have reported on the atypical flora comprising these infections, which are often polymicrobial and contain anaerobic bacteria, and recent studies have noted the high prevalence of anaerobic bacterial infections after soft tissue sarcoma resection [ 26, 35 ]. Based on this, our institution changed clinical practice to include an antibiotic with anaerobic coverage in addition to the standard first-generation cephalosporin for prophylaxis during soft tissue sarcoma resections. The current study was undertaken to evaluate whether this change was associated with a change in major wound complications, and if the change should therefore be adopted for future patients., Questions/purposes: (1) After controlling for potentially confounding variables, was the broadening of the prophylactic antibiotic spectrum to cover anaerobic bacteria associated with a lower odds of major wound complications after soft tissue sarcoma resection? (2) Was the broadening of the prophylactic antibiotic spectrum to cover anaerobic bacteria associated with a lower odds of surgical site infections with polymicrobial or anaerobic infections after soft tissue sarcoma resection? (3) What are the factors associated with major wound complications after soft tissue sarcoma resection?, Methods: We retrospectively identified 623 patients who underwent soft tissue sarcoma resection at a single center between January 2008 and January 2021 using procedural terminology codes. Of these, four (0.6%) pediatric patients were excluded, as were five (0.8%) patients with atypical lipomatous tumors and two (0.3%) patients with primary bone tumors; 5% (33 of 623) who were lost to follow-up, leaving 579 for final analysis. The prophylactic antibiotic regimen given at the resection and whether a wound complication occurred were recorded. Patients received the augmented regimen based on whether they underwent resection after the change in practice in July 2018. A total of 497 patients received a standard antibiotic regimen (usually a first-generation cephalosporin), and 82 patients received an augmented regimen with anaerobic coverage (most often metronidazole). Of the 579 patients, 53% (307) were male (53% [264 of 497] in the standard regimen and 52% [43 of 82] in the augmented regimen), and the mean age was 59 ± 17 years (59 ± 17 and 60 ±17 years in the standard and augmented groups, respectively). Wound complications were defined as any of the following within 120 days of the initial resection: formal wound debridement in the operating room, other interventions such as percutaneous drain placement, readmission for intravenous antibiotics, or deep wound packing for more than 120 days from the resection. Patients were considered to have a surgical site infection if positive cultures resulted from deep tissue cultures taken intraoperatively at the time of debridement. The proportion of patients with major wound complications was 26% (150 of 579); it was 27% (136 of 497) and 17% (14 of 82) in the standard and augmented antibiotic cohorts, respectively (p = 0.049). With the numbers we had, we could not document that the addition of antibiotics with anaerobic coverage was associated with lower odds of anaerobic (4% versus 6%; p = 0.51) or polymicrobial infections (9% versus 14%; p = 0.25). Patient, tumor, and treatment (surgical, radiotherapy, and chemotherapy) variables were collected to evaluate factors associated with overall infection and anaerobic or polymicrobial infection. Patient follow-up was 120 days to capture early wound complications. A multivariable analysis was performed for all variables found to be significant in the univariate analysis. A p value < 0.05 was used as the threshold for statistical significance for all analyses. No patients were found to have an adverse reaction to the augmented regimen, including allergic reactions or Clostridioides (formerly Clostridium) difficile infection., Results: After controlling for other potentially confounding factors such as neoadjuvant radiation, tumor size and anatomic location, as well as patient BMI, anaerobic coverage was associated with smaller odds of wound complications (OR 0.36 [95% confidence interval (CI) 0.18 to 0.68]; p = 0.003). Other factors associated with major wound complications were preoperative radiation (versus no preoperative radiation) (OR 2.66 [95% CI 1.72 to 4.15]; p < 0.001), increasing tumor size (OR 1.04 [95% CI 1.00 to 1.07]; p = 0.03), patient BMI (OR 1.07 [95% CI 1.04 to 1.11]; p < 0.001), and tumor in the distal upper extremity (versus proximal upper extremity, pelvis/groin/hip, and lower extremity) (OR 0.18 [95% CI 0.04 to 0.62]; p = 0.01)., Conclusion: The addition of anaerobic coverage to the standard prophylactic regimen during soft tissue sarcoma resection demonstrated an association with smaller odds of major wound complications and no documented adverse reactions. Treating physicians should consider these findings but note that they are preliminary, and that further work is needed to replicate them in a more controlled study design such as a prospective trial., Level of Evidence: Level III, therapeutic study., Competing Interests: Each author certifies that there are no funding or commercial associations (consultancies, stock ownership, equity interest, patent/licensing arrangements, etc.) that might pose a conflict of interest in connection with the submitted article related to the author or any immediate family members. All ICMJE Conflict of Interest Forms for authors and Clinical Orthopaedics and Related Research® editors and board members are on file with the publication and can be viewed on request., (Copyright © 2022 by the Association of Bone and Joint Surgeons.)

A 74-year-old male with a prior left total knee arthroplasty presented with deformity, loosening, pain, and stiffness associated with multiple raised, erythematous, cutaneous nodules about the anterior knee. Workup was concerning for infection, but the skin nodules were atypical. The patient was sent for biopsy which revealed cutaneous diffuse large B-cell lymphoma. The revision surgery was delayed, and the patient underwent chemotherapy/radiation with complete resolution of his lymphoma. He then underwent a successful aseptic revision total knee arthroplasty. Proper identification and treatment of rare cutaneous skin lesions about a prior surgical site can limit morbidity and result in more desirable outcomes., (© 2022 The Authors.)

Combined hip arthroscopy and periacetabular osteotomy are used for the treatment of concomitant intra-articular hip pathology and acetabular dysplasia or instability. Traditionally, the procedure has been achieved with the use of a traction table or table attachment for the arthroscopic portion and then subsequent transfer of the patient to a fully radiolucent flat bed. In this article, we highlight the technique of a combined hip arthroscopy and periacetabular osteotomy procedure using a single bed attachment system., (© 2022 The Authors.)

Background Aims: Mesenchymal stromal cells (MSCs) are one of the most frequently used cell types in regenerative medicine and cell therapy. Generating sufficient cell numbers for MSC-based therapies is constrained by (i) their low abundance in tissues of origin, which imposes the need for significant ex vivo cell expansion; (ii) donor-specific characteristics, including MSC frequency/quality, that decline with disease state and increasing age; and (iii) cellular senescence, which is promoted by extensive cell expansion and results in decreased therapeutic functionality. The final yield of a manufacturing process is therefore primarily determined by the applied isolation procedure and its efficiency in isolating therapeutically active cells from donor tissue. To date, MSCs are predominantly isolated using media supplemented with either serum or its derivatives, which poses safety and consistency issues., Methods: To overcome these limitations while enabling robust MSC production with constant high yield and quality, the authors developed a chemically defined biomimetic surface coating called isoMATRIX (denovoMATRIX GmbH, Dresden, Germany) and tested its performance during isolation of MSCs., Results: The isoMATRIX facilitates the isolation of significantly higher numbers of MSCs in xenogeneic (xeno)/serum-free and chemically defined conditions. The isolated cells display a smaller cell size and higher proliferation rate than those derived from a serum-containing isolation procedure and a strong immunomodulatory capacity. The high proliferation rates can be maintained up to 5 passages after isolation and cells even benefit from a switch towards a proliferation-specific MSC matrix (myMATRIX MSC) (denovoMATRIX GmbH, Dresden, Germany)., Conclusion: In sum, isoMATRIX promotes enhanced xeno/serum-free and chemically defined isolation of human MSCs and supports consistent and reliable cell performance for improved stem cell-based therapies., (Copyright © 2022 International Society for Cell & Gene Therapy. Published by Elsevier Inc. All rights reserved.)

The rise of the clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein (Cas) system has made it possible to induce double-strand breaks at almost any desired target site in the genome. In plant somatic cells, double-strand breaks are predominantly repaired by the error-prone nonhomologous end-joining pathway, which can lead to mutations at the break site upon repair. So far, it had only been possible to induce genomic changes of up to a few hundred kilobases in plants utilizing this mechanism. However, by combining the highly efficient Staphylococcus aureus Cas9 (SaCas9) with an egg-cell-specific promoter to facilitate heritable mutations, chromosomal rearrangements in the Mb range, such as inversion and translocations, were obtained in Arabidopsis thaliana recently. Here we describe the chromosome-engineering protocol used to generate these heritable chromosomal rearrangements in A. thaliana. The protocol is based on Agrobacterium-mediated transformation of A. thaliana with transfer DNA constructs containing SaCas9, which is driven by an egg-cell-specific promoter, and two guide RNAs that have been preselected based on their cutting efficiency. In the T1 generation, primary transformants are selected and, if required, analyzed by Droplet Digital PCR and propagated. In the following generations, junction-specific PCR screenings are carried out until plants that carry the rearrangement homozygously are identified. Using this protocol, overall rearrangement frequencies range between 0.03% and 0.5%, depending on the type of rearrangement. In total, it takes about 1 year to establish homozygous lines., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

Background: High flow nasal cannula (HFNC) provides noninvasive respiratory support for children who are critically ill who may tolerate it more readily than other noninvasive ventilation (NIV) techniques such as bilevel positive airway pressure and continuous positive airway pressure. Moreover, HFNC may preclude the need for mechanical ventilation (intubation). Nevertheless, NIV or intubation may ultimately be necessary for certain patients. Timely prediction of HFNC failure can provide an indication for increasing respiratory support., Objective: The aim of this study is to develop and compare machine learning (ML) models to predict HFNC failure., Methods: A retrospective study was conducted using the Virtual Pediatric Intensive Care Unit database of electronic medical records of patients admitted to a tertiary pediatric intensive care unit between January 2010 and February 2020. Patients aged <19 years, without apnea, and receiving HFNC treatment were included. A long short-term memory (LSTM) model using 517 variables (vital signs, laboratory data, and other clinical parameters) was trained to generate a continuous prediction of HFNC failure, defined as escalation to NIV or intubation within 24 hours of HFNC initiation. For comparison, 7 other models were trained: a logistic regression (LR) using the same 517 variables, another LR using only 14 variables, and 5 additional LSTM-based models using the same 517 variables as the first LSTM model and incorporating additional ML techniques (transfer learning, input perseveration, and ensembling). Performance was assessed using the area under the receiver operating characteristic (AUROC) curve at various times following HFNC initiation. The sensitivity, specificity, and positive and negative predictive values of predictions at 2 hours after HFNC initiation were also evaluated. These metrics were also computed for a cohort with primarily respiratory diagnoses., Results: A total of 834 HFNC trials (455 [54.6%] training, 173 [20.7%] validation, and 206 [24.7%] test) met the inclusion criteria, of which 175 (21%; training: 103/455, 22.6%; validation: 30/173, 17.3%; test: 42/206, 20.4%) escalated to NIV or intubation. The LSTM models trained with transfer learning generally performed better than the LR models, with the best LSTM model achieving an AUROC of 0.78 versus 0.66 for the 14-variable LR and 0.71 for the 517-variable LR 2 hours after initiation. All models except for the 14-variable LR achieved higher AUROCs in the respiratory cohort than in the general intensive care unit population., Conclusions: ML models trained using electronic medical record data were able to identify children at risk of HFNC failure within 24 hours of initiation. LSTM models that incorporated transfer learning, input data perseveration, and ensembling showed improved performance compared with the LR and standard LSTM models., (©George Pappy, Melissa Aczon, Randall Wetzel, David Ledbetter. Originally published in JMIR Medical Informatics (https://medinform.jmir.org), 03.03.2022.)

Objectives: To compare the number of patients with gunshot wounds presenting to our level 1 trauma center before and during the COVID-19 pandemic with a focus on volume trends after the lifting of stay-at-home directives through August 2020., Design: Retrospective., Setting: Level 1 trauma center., Patients/participants: Seven hundred six gunshot wound patients between 2016 and 2020 (months March to September only)., Intervention: COVID-19 pandemic and resultant stay at home directives., Main Outcome Measurements: Number of patients presenting with gunshot wounds per time period., Results: The number of patients with gunshot wounds presenting to our institution increased by 11.7% in March-April 2020 and by 67% in May-August 2020 when compared to previous years. Length of stay significantly decreased in 2020 compared to 2018 and 2019. In 2020, significantly fewer patients had orthopaedic procedures than in 2018., Conclusions: Patients presenting with gunshot wounds increased during the initial "stay-at-home" portion of the pandemic in March to April and increased significantly more after the restrictions were relaxed during May to August. Level of Evidence: Therapeutic Level III., Competing Interests: The remaining authors certify that he or she, or a member of his or her immediate family, has no funding or commercial associations that might pose a conflict of interest in connection with the submitted article., (Copyright © 2021 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of the Orthopaedic Trauma Association.)