Matrix stiffness-dependent regulation of immunomodulatory genes in human MSCs is associated with the lncRNA CYTOR.

Cell-matrix interactions in 3D environments significantly differ from those in 2D cultures. As such, mechanisms of mechanotransduction in 2D cultures are not necessarily applicable to cell-encapsulating hydrogels that resemble features of tissue architecture. Accordingly, the characterization of molecular pathways in 3D matrices is expected to uncover insights into how cells respond to their mechanical environment in physiological contexts, and potentially also inform hydrogel-based strategies in cell therapies. In this study, a bone marrow-mimetic hydrogel was employed to systematically investigate the stiffness-responsive transcriptome of mesenchymal stromal cells. High matrix rigidity impeded integrin-collagen adhesion, resulting in changes in cell morphology characterized by a contractile network of actin proximal to the cell membrane. This resulted in a suppression of extracellular matrix-regulatory genes involved in the remodeling of collagen fibrils, as well as the upregulation of secreted immunomodulatory factors. Moreover, an investigation of long noncoding RNAs revealed that the cytoskeleton regulator RNA (CYTOR) contributes to these 3D stiffness-driven changes in gene expression. Knockdown of CYTOR using antisense oligonucleotides enhanced the expression of numerous mechanoresponsive cytokines and chemokines to levels exceeding those achievable by modulating matrix stiffness alone. Taken together, our findings further our understanding of mechanisms of mechanotransduction that are distinct from canonical mechanotransductive pathways observed in 2D cultures.

Improved efficacy of pembrolizumab combined with soluble EphB4-albumin in HPV-negative EphrinB2 positive head neck squamous cell carcinoma.

OBJECTIVE: Patients with relapsed or metastatic head and neck squamous cell carcinoma (HNSCC) after primary local therapy have low response rates with cetuximab, systemic chemotherapy or check point inhibitor therapy. Novel combination therapies with the potential to improve outcomes for patients with HNSCC is an area of high unmet need. METHODS: This is a phase II single-arm clinical trial of locally advanced or metastatic HNSCC patients treated with a combination of soluble EphB4-human serum albumin (sEphB4-HSA) fusion protein and pembrolizumab after platinum-based chemotherapy with up to 2 prior lines of treatment. The primary endpoints were safety and tolerability and the primary efficacy endpoint was overall response rate (ORR). Secondary endpoints included progression free survival (PFS) and overall survival (OS). HPV status and EphrinB2 expression were evaluated for outcome. RESULTS: Twenty-five patients were enrolled. Median follow up was 40.4 months (range 9.8 - 40.4). There were 6 responders (ORR 24%). There were 5 responders in the 11 HPV-negative and EphrinB2 positive patients, (ORR 45%) with 2 of these patients achieving a complete response (CR). The median PFS in HPV-negative/EphrinB2 positive patients was 3.2 months (95% CI 1.1, 7.3). Median OS in HPV-negative/EphrinB2 positive patients was 10.9 months (95% CI 2.0, 13.7). Hypertension, transaminitis and fatigue were the most common toxicities. DISCUSSION: The combination of sEphB4-HSA and pembrolizumab has a favorable toxicity profile and favorable activity particularly among HPV-negative EphrinB2 positive patients with HNSCC.

High-throughput sensitive screening of small molecule modulators of microexon alternative splicing using dual Nano and Firefly luciferase reporters.

Disruption of alternative splicing frequently causes or contributes to human diseases and disorders. Consequently, there is a need for efficient and sensitive reporter assays capable of screening chemical libraries for compounds with efficacy in modulating important splicing events. Here, we describe a screening workflow employing dual Nano and Firefly luciferase alternative splicing reporters that affords efficient, sensitive, and linear detection of small molecule responses. Applying this system to a screen of ~95,000 small molecules identified compounds that stimulate or repress the splicing of neuronal microexons, a class of alternative exons often disrupted in autism and activated in neuroendocrine cancers. One of these compounds rescues the splicing of several analyzed microexons in the cerebral cortex of an autism mouse model haploinsufficient for Srrm4, a major activator of brain microexons. We thus describe a broadly applicable high-throughput screening system for identifying candidate splicing therapeutics, and a resource of small molecule modulators of microexons with potential for further development in correcting aberrant splicing patterns linked to human disorders and disease.

Conversion Rate of Abstracts Presented at Plastic Surgery The Meeting From 2010 to 2019: A 10-Year Analysis of Factors for Success.

BACKGROUND: Presentations are an important means of knowledge generation. Publication of these studies is important for dissemination of findings beyond meeting attendees. We analyzed a 10-year sample of presented abstracts at Plastic Surgery The Meeting and describe factors that improve rate and speed of conversion to peer-reviewed publication. METHODS: Abstracts presented between 2010 and 2019 at Plastic Surgery The Meeting were sourced from the American Society of Plastic Surgery Abstract Archive. A random sample of 100 abstracts from each year was evaluated. Abstract information and demographics were recorded. The title or author and keywords of each abstract were searched using a standardized workflow to find a corresponding published paper on PubMed, Google Scholar, and Google. Data were analyzed for trends and factors affecting conversion rate. RESULTS: A total of 983 presented abstracts were included. The conversion rate was 54.1%. Residents and fellows constituted the largest proportion of presenters (38.4%). There was a significant increase in medical student and research fellow presenters during the study period (P < 0.001). Conversion rate was not affected by the research rank of a presenter's affiliated institution (ß = 1.001, P = 0.89), geographic location (P = 0.60), or subspecialty tract (P = 0.73). US academics had a higher conversion rate (61.8%) than US nonacademics (32.7%) or international presenters (47.1%) (P < 0.001). Medical students had the highest conversion rate (65.6%); attendings had the lowest (45.0%). Research fellows had the lowest average time to publication (11.6 months, P = 0.007). CONCLUSIONS: Lower levels of training, factors associated with increased institution-level support, and research quality affect rate and time to publication. These findings highlight the success of current models featuring medical student and research fellow-led projects with strong resident and faculty mentorship.

Reactive Oxygen Species Modulation in the Current Landscape of Anticancer Therapies.

Significance: Reactive oxygen species (ROS) are generated during mitochondrial oxidative metabolism, and are tightly controlled through homeostatic mechanisms to maintain intracellular redox, regulating growth and proliferation in healthy cells. However, ROS production is perturbed in cancers where abnormal accumulation of ROS leads to oxidative stress and genomic instability, triggering oncogenic signaling pathways on one hand, while increasing oxidative damage and triggering ROS-dependent death signaling on the other. Recent Advances: Our review illuminates how critical interactions between ROS and oncogenic signaling, the tumor microenvironment, and DNA damage response (DDR) pathways have led to interest in ROS modulation as a means of enhancing existing anticancer strategies and developing new therapeutic opportunities. Critical Issues: ROS equilibrium exists via a delicate balance of pro-oxidant and antioxidant species within cells. "Antioxidant" approaches have been explored mainly in the form of chemoprevention, but there is insufficient evidence to advocate its routine application. More progress has been made via the "pro-oxidant" approach of targeting cancer vulnerabilities and inducing oxidative stress. Various therapeutic modalities have employed this approach, including direct ROS-inducing agents, chemotherapy, targeted therapies, DDR therapies, radiotherapy, and immunotherapy. Finally, emerging delivery systems such as "nanosensitizers" as radiotherapy enhancers are currently in development. Future Directions: While approaches designed to induce ROS have shown considerable promise in selectively targeting cancer cells and dealing with resistance to conventional therapies, most are still in early phases of development and challenges remain. Further research should endeavor to refine treatment strategies, optimize drug combinations, and identify predictive biomarkers of ROS-based cancer therapies.

Programmable RNA-guided DNA endonucleases are widespread in eukaryotes and their viruses.

Programmable RNA-guided DNA nucleases perform numerous roles in prokaryotes, but the extent of their spread outside prokaryotes is unclear. Fanzors, the eukaryotic homolog of prokaryotic TnpB proteins, have been detected in genomes of eukaryotes and large viruses, but their activity and functions in eukaryotes remain unknown. Here, we characterize Fanzors as RNA-programmable DNA endonucleases, using biochemical and cellular evidence. We found diverse Fanzors that frequently associate with various eukaryotic transposases. Reconstruction of Fanzors evolution revealed multiple radiations of RuvC-containing TnpB homologs in eukaryotes. Fanzor genes captured introns and proteins acquired nuclear localization signals, indicating extensive, long-term adaptation to functioning in eukaryotic cells. Fanzor nucleases contain a rearranged catalytic site of the RuvC domain, similar to a distinct subset of TnpBs, and lack collateral cleavage activity. We demonstrate that Fanzors can be harnessed for genome editing in human cells, highlighting the potential of these widespread eukaryotic RNA-guided nucleases for biotechnology applications.

Living Alone, Environmental Hazards, and Falls Among U.S. Older Adults.

Background and Objectives: Physical conditions of living environments can affect the incidence of falls; however, prior work has focused typically on 1 domain at a time-either neighborhood or home, capturing limited environmental boundaries of older adults. We examined how neighborhood together with the home environment affect the incidence of falls over time and whether living arrangement modifies the influence of the environmental risks on falls. Research Design and Methods: Using the 2012-2020 waves of the Health and Retirement Study (HRS; N = 1,893), we fitted logistic regression to estimate the incidence of falls over an 8-year study period. We used the neighborhood and housing data that are collected systematically by trained observers in the HRS to assess environmental hazards. Sidewalk quality, neighborhood disorder, and the presence of green space were measured to capture outdoor environmental hazards. Indoor environmental hazards included the presence of housing decay and poorly maintained stairways. All models were stratified by living arrangement. Results: Neighborhood and housing environment were independently associated with the odds of falls net of demographic characteristics and preexisting health conditions, and effects were significant for people living alone only. The presence of green space and poorly maintained stairways were associated with greater odds of falling, net of covariates during 8 years of follow-up (odds ratios = 2.10 and 2.65, p < .05, respectively). None of the environmental risk factors were significant for those living with others. Discussion and Implications: Falls in old age may be determined in part by a combination of outdoor and indoor risk factors. More research is needed to understand pathways that lead to greater vulnerability among older adults living alone to environmental hazards.

Programmable RNA-guided endonucleases are widespread in eukaryotes and their viruses.

TnpB proteins are RNA-guided nucleases that are broadly associated with IS200/605 family transposons in prokaryotes. TnpB homologs, named Fanzors, have been detected in genomes of some eukaryotes and large viruses, but their activity and functions in eukaryotes remain unknown. We searched genomes of diverse eukaryotes and their viruses for TnpB homologs and identified numerous putative RNA-guided nucleases that are often associated with various transposases, suggesting they are encoded in mobile genetic elements. Reconstruction of the evolution of these nucleases, which we rename Horizontally-transferred Eukaryotic RNA-guided Mobile Element Systems (HERMES), revealed multiple acquisitions of TnpBs by eukaryotes and subsequent diversification. In their adaptation and spread in eukaryotes, HERMES proteins acquired nuclear localization signals, and genes captured introns, indicating extensive, long term adaptation to functioning in eukaryotic cells. Biochemical and cellular evidence show that HERMES employ non-coding RNAs encoded adjacent to the nuclease for RNA-guided cleavage of double-stranded DNA. HERMES nucleases contain a re-arranged catalytic site of the RuvC domain, similar to a distinct subset of TnpBs, and lack collateral cleavage activity. We demonstrate that HERMES can be harnessed for genome editing in human cells, highlighting the potential of these widespread eukaryotic RNA-guided nucleases for biotechnology applications.

Mycobacterium tuberculosis Inhibitors Based on Arylated Quinoline Carboxylic Acid Backbones with Anti-Mtb Gyrase Activity.

New antitubercular agents with either a novel mode of action or novel mode of inhibition are urgently needed to overcome the threat of drug-resistant tuberculosis (TB). The present study profiles new arylated quinoline carboxylic acids (QCAs) having activity against replicating and non-replicating Mycobacterium tuberculosis (Mtb), the causative agent of TB. Thus, the synthesis, characterization, and in vitro screening (MABA and LORA) of 48 QCAs modified with alkyl, aryl, alkoxy, halogens, and nitro groups in the quinoline ring led to the discovery of two QCA derivatives, 7i and 7m, adorned with C-2 2-(naphthalen-2-yl)/C-6 1-butyl and C-2 22-(phenanthren-3-yl)/C-6 isopropyl, respectively, as the best Mtb inhibitors. DNA gyrase inhibition was shown to be exhibited by both, with QCA 7m illustrating better activity up to a 1 µM test concentration. Finally, a docking model for both compounds with Mtb DNA gyrase was developed, and it showed a good correlation with in vitro results.

Arg-tRNA synthetase links inflammatory metabolism to RNA splicing and nuclear trafficking via SRRM2.

Cells respond to perturbations such as inflammation by sensing changes in metabolite levels. Especially prominent is arginine, which has known connections to the inflammatory response. Aminoacyl-tRNA synthetases, enzymes that catalyse the first step of protein synthesis, can also mediate cell signalling. Here we show that depletion of arginine during inflammation decreased levels of nuclear-localized arginyl-tRNA synthetase (ArgRS). Surprisingly, we found that nuclear ArgRS interacts and co-localizes with serine/arginine repetitive matrix protein 2 (SRRM2), a spliceosomal and nuclear speckle protein, and that decreased levels of nuclear ArgRS correlated with changes in condensate-like nuclear trafficking of SRRM2 and splice-site usage in certain genes. These splice-site usage changes cumulated in the synthesis of different protein isoforms that altered cellular metabolism and peptide presentation to immune cells. Our findings uncover a mechanism whereby an aminoacyl-tRNA synthetase cognate to a key amino acid that is metabolically controlled during inflammation modulates the splicing machinery.

Polyphenolic Natural Products Active In Silico Against SARS-CoV-2 Spike Receptor Binding Domains and Non-structural Proteins - A Review.

The ongoing Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) pandemic has been proven to be more severe than the previous coronavirus outbreaks due to the virus' high transmissibility. With the emergence of new variants, this global phenomenon took a more dramatic turn, with many countries recently experiencing higher surges of confirmed cases and deaths. On top of this, the inadequacy of effective treatment options for COVID-19 aggravated the problem. As a way to address the unavailability of target-specific viral therapeutics, computational strategies have been employed to hasten and systematize the search. The objective of this review is to provide initial data highlighting the utility of polyphenols as potential prophylaxis or treatment for COVID-19. In particular, presented here are virtually screened polyphenolic compounds which showed potential as either antagonists to viral entry and host cell recognition through binding with various receptor-binding regions of SARS-CoV-2 spike protein or as inhibitors of viral replication and post-translational modifications through binding with essential SARS-CoV-2 non-structural proteins.

Drag-and-drop genome insertion of large sequences without double-strand DNA cleavage using CRISPR-directed integrases.

Programmable genome integration of large, diverse DNA cargo without DNA repair of exposed DNA double-strand breaks remains an unsolved challenge in genome editing. We present programmable addition via site-specific targeting elements (PASTE), which uses a CRISPR-Cas9 nickase fused to both a reverse transcriptase and serine integrase for targeted genomic recruitment and integration of desired payloads. We demonstrate integration of sequences as large as ~36 kilobases at multiple genomic loci across three human cell lines, primary T cells and non-dividing primary human hepatocytes. To augment PASTE, we discovered 25,614 serine integrases and cognate attachment sites from metagenomes and engineered orthologs with higher activity and shorter recognition sequences for efficient programmable integration. PASTE has editing efficiencies similar to or exceeding those of homology-directed repair and non-homologous end joining-based methods, with activity in non-dividing cells and in vivo with fewer detectable off-target events. PASTE expands the capabilities of genome editing by allowing large, multiplexed gene insertion without reliance on DNA repair pathways.

Piriformis syndrome as a result of intramuscular haematoma mimicking cauda equina effectively treated with piriformis tendon release.

We present a case of piriformis syndrome in a woman in her 30's following low energy trauma, presenting with unilateral lower limb weakness, altered sensation and urinary retention. CT imaging revealed a bulky piriformis muscle which was further clarified on MRI as an intramuscular haematoma within the left piriformis causing compression of the left lumbosacral plexus. Haematoma formation was exacerbated due to use of an antiplatelet medication the patient was taking for Moyamoya disease, which carries an increased risk of cerebrovascular accident. Surgical exploration of the piriformis and sciatic nerve was undertaken and confirmed a haematoma within the piriformis. A full release of the piriformis tendon was undertaken, and the sciatic nerve was inspected, no further abnormality was found. After review in clinic post-discharge, the patient reported normal sensation and normal muscle power in her feet.

Primary total hip replacement using Burch-Schneider cages for acetabular fractures.

INTRODUCTION: Osteoporotic acetabular fractures are common and pose a difficult technical challenge for the trauma surgeon. Acute total hip arthroplasty (THA) using a Burch-Schneider antiprotrusio cage with immediate postoperative weight-bearing is a method to approach these injuries. This case series reports our outcomes of acute THA using Burch-Schneider cages for acetabular fractures from a UK major trauma centre based on length of stay, radiological outcome, complications and outcome scores. METHODS: Data were collected from all patients who underwent acute THA with a Burch-Schneider cage for acetabular fractures between June 2006 and August 2015. Patients were followed up clinically, radiologically, and using Oxford Hip Scores (OHS). RESULTS: 20 patients with a median age of 73 (range 60-90 years) were identified. All patients were independent walkers at follow-up, and had achieved radiological union. There were no dislocations, subsidence, revision or deep infections. Significant complications include 1 perioperative death as a result of complications arising from pre-existing pulmonary fibrosis; 1 deep vein thrombosis; 1 intraoperative arterial injury to the superior gluteal artery; and 1 leg-length discrepancy. Mean length of stay was 10 days. The mean OHS was 37/48 at a mean follow-up of 26 months. CONCLUSIONS: This case series further validates the use of Burch-Schneider cages with primary THA in acute acetabular fractures.

Understanding the Pathophysiology of L5-S1 Loss of Lordosis and Retrolisthesis: An EOS Study of Lumbopelvic Movement Between Standing and Slump Sitting Postures.

OBJECTIVE: To understand the pathophysiology of L5-S1 loss of lordosis and retrolisthesis by comparing 2 commonly assumed physiological weight-bearing postures. METHODS: This was a prospective comparative study of whole-body standing and slump sitting EOS radiographs in clinic patients presenting with back pain or lower limb radicular pain. Patients with prior spinal intervention, malignancy, trauma, inflammatory diseases, transitional lumbosacral vertebra, pregnancy, and L5-S1 retrolisthesis or spondylolisthesis from nondegenerative causes were excluded. C7 sagittal vertical axis, global cervical angle, global thoracic angle, global lumbar angle, thoracolumbar angle, T1-slope, pelvic incidence, pelvic tilt, sacral slope, L5-S1 angle, L5-S1 vertebral translation, L5-S1 disc height, and presence of L4-5 vertebral translation were measured. Univariate and multivariate analyses were performed to identify predictors of L5-S1 lordosis loss and retrolisthesis. RESULTS: L5-S1 loss of lumbar lordosis (7.02 ± 9.90°, P < 0.001), retrolisthesis (0.07 ± 0.411 cm, P < 0.001), and loss of disc height (0.10 ± 0.23 cm, P < 0.001) occurred when changing from standing to slump sitting along with other sagittal profile changes (P < 0.001). Taller L5-S1 disc height (odds ratio [OR] 2.57, P = 0.04), larger lumbar range-of-motion change (OR 3.82, P = 0.012), lower sacral slope on sitting (OR 2.50, P = 0.043), and presence of L4-5 spondylolisthesis (OR 2.75, P = 0.032) were predictive of larger L5-S1 lordosis loss (>7°) on multivariate analysis, while larger lumbar range-of-motion change (OR 2.21, P = 0.050) and presence of L4-5 spondylolisthesis (OR 3.08, P = 0.023) were predictive of greater L5-S1 retrolisthesis (>0.07 cm). CONCLUSIONS: Degenerative L5-S1 loss of lordosis and retrolisthesis likely result from long-standing lower lumbar spine bending forces against the posterior ligamentous complex with slump sitting, predisposed by a negatively sloped sacrum and increased lumbar flexibility.

The Theoretical Framework of the Clinical Pilates Exercise Method in Managing Non-Specific Chronic Low Back Pain: A Narrative Review.

Exercise plays an important role in rehabilitating people with chronic low back pain. Aerobic exercise and resistance training are general exercise strategies to manage chronic low back pain, but these strategies require longer intervention period to achieve clinical outcomes in pain reduction and functional improvements. Directional preference is recognised as an important exercise strategy in managing low back pain. The Clinical Pilates exercise method leverages on the directional preference of an individual to achieve clinical outcomes faster. Clinical Pilates is a hybrid of two of the best exercise interventions for low back pain, which are general Pilates and the McKenzie method. Due to the scarcity of Clinical Pilates literature, a review of its theory and studies was undertaken to provide a structured guide to the technique in managing people with chronic low back pain. Hypothetical algorithms are developed to support translation into clinical practice and future research studies. These algorithms are useful in the management of complex cases involving multiple directional trauma. Although limited, current evidence suggests that the Clinical Pilates exercise method is safe and provides faster functional recovery in the early stage of rehabilitation and similar longer term outcomes as general exercises.

Cattle-Related Trauma: A 5-Year Retrospective Review in a Adult Major Trauma Center.

INTRODUCTION: Bovine injuries are a common and significant cause of trauma, often requiring admission and operative treatment. We review all bovine-related injuries over 5 years, both emergency and general practitioner (GP) referrals at an adult major trauma center in England. METHODS: Retrospective evaluation was undertaken using the keywords through radiology referrals and hospital admissions speciality databases. By searching patient notes, demographics were collected as well as the mechanism and the situation of injury; trauma scores were calculated using: injury severity score (ISS) and probability of survival (Ps19). The results were divided into emergency patients and GP referrals. RESULTS: Sixty-seven patients were identified retrospectively over 5 years, 44 emergency patients (including 23 major traumas), and 23 GP referrals. Combined (emergency and GP) mean age 52 years old; 67% male; and mean ISS 11. Most common combined mechanism of injury, kicked (n = 23). In emergency patients, trampling injuries were the most common. Eighty-six percent of the trampled patients were major traumas and associated with increased ISS (mean 13). Indirect injuries mainly involved farm gates (92%). Seventy-three percent of bull-related injuries were major traumas and had increased ISS scores (mean 17). Orthopaedics was the most common admitting speciality followed by cardiothoracic and neurosurgery. In emergency patients, fractures were the most common primary injury (n = 20), upper limb followed by spine. In GP, soft-tissue injuries were the most common primary injury. Seventy percent of the emergency referrals required admission and 50% operations. Fracture fixation was the most common operative procedure. Only, one GP referral required an operation. There were significant delays in GP patients presenting. Two patients had a Ps19 score <90. There were two mortalities. CONCLUSION: Cattle-related injuries are a significant cause of severe morbidity and mortality. They are under-reported. Patterns of injury are similar to high-velocity road traffic collisions and bull-related injuries or trampling in particular, should alert the clinician to more significant trauma. Farm gates are a frequent cause of trauma associated with cattle. GP referrals with ongoing symptoms for more than 2 weeks seeking medical advice should alert the clinician to a more serious diagnosis.