PLSCR1 is a cell-autonomous defence factor against SARS-CoV-2 infection.

Understanding protective immunity to COVID-19 facilitates preparedness for future pandemics and combats new SARS-CoV-2 variants emerging in the human population. Neutralizing antibodies have been widely studied; however, on the basis of large-scale exome sequencing of protected versus severely ill patients with COVID-19, local cell-autonomous defence is also crucial1-4. Here we identify phospholipid scramblase 1 (PLSCR1) as a potent cell-autonomous restriction factor against live SARS-CoV-2 infection in parallel genome-wide CRISPR-Cas9 screens of human lung epithelia and hepatocytes before and after stimulation with interferon-γ (IFNγ). IFNγ-induced PLSCR1 not only restricted SARS-CoV-2 USA-WA1/2020, but was also effective against the Delta B.1.617.2 and Omicron BA.1 lineages. Its robust activity extended to other highly pathogenic coronaviruses, was functionally conserved in bats and mice, and interfered with the uptake of SARS-CoV-2 in both the endocytic and the TMPRSS2-dependent fusion routes. Whole-cell 4Pi single-molecule switching nanoscopy together with bipartite nano-reporter assays found that PLSCR1 directly targeted SARS-CoV-2-containing vesicles to prevent spike-mediated fusion and viral escape. A PLSCR1 C-terminal ß-barrel domain-but not lipid scramblase activity-was essential for this fusogenic blockade. Our mechanistic studies, together with reports that COVID-associated PLSCR1 mutations are found in some susceptible people3,4, identify an anti-coronavirus protein that interferes at a late entry step before viral RNA is released into the host-cell cytosol.

Supramolecular organization and dynamics of mannosylated phosphatidylinositol lipids in the mycobacterial plasma membrane.

Mycobacterium tuberculosis (Mtb) is the causative agent of tuberculosis (TB), a disease that claims ~1.6 million lives annually. The current treatment regime is long and expensive, and missed doses contribute to drug resistance. Therefore, development of new anti-TB drugs remains one of the highest public health priorities. Mtb has evolved a complex cell envelope that represents a formidable barrier to antibiotics. The Mtb cell envelop consists of four distinct layers enriched for Mtb specific lipids and glycans. Although the outer membrane, comprised of mycolic acid esters, has been extensively studied, less is known about the plasma membrane, which also plays a critical role in impacting antibiotic efficacy. The Mtb plasma membrane has a unique lipid composition, with mannosylated phosphatidylinositol lipids (phosphatidyl-myoinositol mannosides, PIMs) comprising more than 50% of the lipids. However, the role of PIMs in the structure and function of the membrane remains elusive. Here, we used multiscale molecular dynamics (MD) simulations to understand the structure-function relationship of the PIM lipid family and decipher how they self-organize to shape the biophysical properties of mycobacterial plasma membranes. We assess both symmetric and asymmetric assemblies of the Mtb plasma membrane and compare this with residue distributions of Mtb integral membrane protein structures. To further validate the model, we tested known anti-TB drugs and demonstrated that our models agree with experimental results. Thus, our work sheds new light on the organization of the mycobacterial plasma membrane. This paves the way for future studies on antibiotic development and understanding Mtb membrane protein function.

Psychosocial Distress Screening Among Interprofessional Palliative Care Teams: A Narrative Review.

With increased need for palliative care and limited staffing resources, non-social workers are increasingly responsible for screening for urgent psychosocial distress. The National Consensus Project guidelines call for all palliative care team members to be competent in screening across domains. Yet, in contrast to an abundance of evidence-informed tools for palliative social work assessments, standardization for interprofessional psychosocial screening is lacking. This lack of standardized practice may lead to harmful disparities in care delivery. The purpose of this narrative review is to examine current literature on evidence-informed practices for psychosocial screening within palliative care. Google Scholar, a university Summon library search engine, and prominent palliative care journals were searched using the same phrases to locate articles for inclusion. Each article was reviewed and synthesized across common themes. Although an abundance of validated screening tools exists for outpatient oncology-specific settings, there is minimal guidance on psychosocial screening tools intended for specialty palliative care. The most oft-cited tools have been met with concern for validity across diverse palliative care populations and settings. Additional research is needed to operationalize and measure brief psychosocial screening tools that can be validated for use by interprofessional palliative care teams, a stepping-stone for increased equity in palliative care practice.

Amyloidosis and Considerations for the Hand Surgeon.

Amyloidosis can lead to cardiac, renal, and other multiorgan failure. New treatments have become available that can prolong survival but rely on early diagnosis. Manifestations of amyloidosis in hand surgery include carpal tunnel syndrome, trigger finger, peripheral neuropathy, and spontaneous distal biceps rupture. Often, these can predate systemic amyloidosis, offering hand surgeons an opportunity to diagnose patients with amyloidosis before systemic disease, refer them for treatment, and potentially alter disease course and prolong survival. In this review, we describe the pathophysiology and two most common subtypes of amyloidosis seen by hand surgeons. We provide guidance on biopsy practices and referral for patients with amyloidosis. Lastly, we provide a brief overview of the treatments for amyloidosis and their effect on disease course.

Missed opportunities for diagnosis of hepatitis B and C in individuals diagnosed with decompensated cirrhosis or hepatocellular carcinoma.

BACKGROUND AND AIM: This study aimed to assess utilization of health-care services in people with decompensated cirrhosis (DC) or hepatocellular carcinoma (HCC) and a "late diagnosis" of hepatitis B or hepatitis C. METHODS: Hepatitis B and C cases during 1997-2016 in Victoria, Australia, were linked with hospitalizations, deaths, liver cancer diagnoses, and medical services. A late diagnosis was defined as hepatitis B or hepatitis C notification occurring after, at the same time, or within 2 years preceding an HCC/DC diagnosis. Services provided during the 10-year period before HCC/DC diagnosis were assessed, including general practitioner (GP) or specialist visits, emergency department presentations, hospital admissions, and blood tests. RESULTS: Of the 25 766 notified cases of hepatitis B, 751 (2.9%) were diagnosed with HCC/DC, and hepatitis B was diagnosed late in 385 (51.3%). Of 44 317 cases of hepatitis C, 2576 (5.8%) were diagnosed with HCC/DC, and hepatitis C was diagnosed late in 857 (33.3%). Although late diagnosis dropped over time, missed opportunities for timely diagnosis were observed. Most people diagnosed late had visited a GP (97.4% for hepatitis B, 98.9% for hepatitis C) or had a blood test (90.9% for hepatitis B, 88.6% for hepatitis C) during the 10 years before HCC/DC diagnosis. The median number of GP visits was 24 and 32, and blood tests 7 and 8, for hepatitis B and C, respectively. CONCLUSIONS: Late diagnosis of viral hepatitis remains a concern, with the majority having frequent health-care service provision in the preceding period, indicating missed opportunities for diagnosis.

Trauma-Informed Care for Inpatient Palliative Care Social Work: Applying Existing Models at the Bedside.

Coexisting serious illness and posttraumatic stress place hospitalized individuals at risk for complex pain, anxiety, and retraumatization. Hospital palliative care social workers increasingly recognize the value of trauma-informed care (TIC) for reducing harm in the inpatient setting. Despite this recognition, there is limited operationalization of TIC principles for inpatient interventions. This paper integrates each TIC principle with inpatient psychosocial interventions to advance trauma-informed competencies among inpatient palliative care social workers and to provide a foundation for future TIC implementation research.

Reducing Sialylation Enhances Electrotaxis of Corneal Epithelial Cells.

Corneal wound healing is a complex biological process that integrates a host of different signals to coordinate cell behavior. Upon wounding, there is the generation of an endogenous wound electric field that serves as a powerful cue to guide cell migration. Concurrently, the corneal epithelium reduces sialylated glycoforms, suggesting that sialylation plays an important role during electrotaxis. Here, we show that pretreating human telomerase-immortalized corneal epithelial (hTCEpi) cells with a sialyltransferase inhibitor, P-3FAX-Neu5Ac (3F-Neu5Ac), improves electrotaxis by enhancing directionality, but not speed. This was recapitulated using Kifunensine, which inhibits cleavage of mannoses and therefore precludes sialylation on N-glycans. We also identified that 3F-Neu5Ac enhanced the responsiveness of the hTCEpi cell population to the electric field and that pretreated hTCEpi cells showed increased directionality even at low voltages. Furthermore, when we increased sialylation using N-azidoacetylmannosamine-tetraacylated (Ac4ManNAz), hTCEpi cells showed a decrease in both speed and directionality. Importantly, pretreating enucleated eyes with 3F-Neu5Ac significantly improved re-epithelialization in an ex vivo model of a corneal injury. Finally, we show that in hTCEpi cells, sialylation is increased by growth factor deprivation and reduced by PDGF-BB. Taken together, our results suggest that during corneal wound healing, reduced sialylated glycoforms enhance electrotaxis and re-epithelialization, potentially opening new avenues to promote corneal wound healing.

Structural basis of trehalose recognition by the mycobacterial LpqY-SugABC transporter.

The Mycobacterium tuberculosis (Mtb) LpqY-SugABC ATP-binding cassette transporter is a recycling system that imports trehalose released during remodeling of the Mtb cell-envelope. As this process is essential for the virulence of the Mtb pathogen, it may represent an important target for tuberculosis drug and diagnostic development, but the transporter specificity and molecular determinants of substrate recognition are unknown. To address this, we have determined the structural and biochemical basis of how mycobacteria transport trehalose using a combination of crystallography, saturation transfer difference NMR, molecular dynamics, site-directed mutagenesis, biochemical/biophysical assays, and the synthesis of trehalose analogs. This analysis pinpoints key residues of the LpqY substrate binding lipoprotein that dictate substrate-specific recognition and has revealed which disaccharide modifications are tolerated. These findings provide critical insights into how the essential Mtb LpqY-SugABC transporter reuses trehalose and modified analogs and specifies a framework that can be exploited for the design of new antitubercular agents and/or diagnostic tools.

Outcomes of treatment in men with prostate cancer at the cancer centre Bahamas.

PURPOSE: This is a first report from The Bahamas of management and long-term outcomes in men with non-metastatic prostate cancer treated with radiotherapy, with or without androgen deprivation therapy, from 2004 to 2016. METHODS: Patients were characterized by baseline factors, stratified by risk groups using tumor stage (clinical T-stage), prostate-specific antigen (PSA) test result and Gleason grade, and sorted by treatment combinations (by radiation volume and use of androgen deprivation). RESULTS: Overall, 205/216 men were Afro-Caribbean. Median age was 66. There were 18 low-, 77 intermediate-, and 121 high-risk patients, treated with prostate-only versus pelvis plus prostate radiotherapy, many receiving 2 years of androgen suppression. Time to commence radiation was about 6 months from initial diagnosis. In those not relapsing, global PSA nadir was reached in 4 years and was under 1.0, reduced from a mean at baseline of 31. At 10 years, disease-free experience (32 relapses) was 68% and overall survival was 87%, although only 2/12 deaths were related to prostate cancer. This experience compares favorably with recently published outcomes from other countries using very similar treatments. CONCLUSIONS: This study establishes benchmark statistics from diagnosis to long-term follow-up. Outcomes in Bahamian men are consistent with expectations from risk-stratified guidelines followed in developed countries.

Injuries and Illnesses Occurring on a Recreational Bicycle Tour: The Great Ohio Bicycle Adventure.

OBJECTIVE: Recreational multiday cycling events have grown popular in the United States; however, little has been published regarding the injuries and illnesses that occur during these events. The objective of this study was to describe injuries and illnesses that occur in the Great Ohio Bicycle Adventure (GOBA) and to examine associated risk factors. METHODS: Injury and illness data collected from an on-site medical clinic during the 2013 and 2014 GOBA tours were merged with rider registration data for analysis. Diagnoses were classified as acute injury, overuse injury, or medical illness. The odds ratios of sustaining at least 1 injury/illness, as well as sustaining an acute injury, were assessed adjusting for riders' demographics. RESULTS: A total of 4005 (2172 in 2013 and 1833 in 2014) cyclists participated in GOBA, with an an age of 50.7±17.6 (2-86) years (mean±SD [range]), of whom 59.8% were male. During the tours, 143 (3.6%) riders reported at least 1 injury/illness, which resulted in 220 clinical diagnoses, including 114 (51.8%) acute injuries, 27 (12.3 %) overuse injuries, and 79 (35.9%) medical illnesses. The lower extremities were the most commonly injured body site (n=71, 50.4%). "Superficial/Abrasions/Contusions" was the most common injury/illness type (n=68, 30.9%). Riders who had no previous tour experience or who were at least 50 years old had a greater risk of injury/illness than their counterparts. CONCLUSIONS: The prevalence of injury and illness is relatively low in multiday recreational cycling events. Our findings provide important data for planning and preparing for medical coverage at mass recreational cycling events.

Lifestyle, medication and socio-demographic determinants of mental and physical health-related quality of life in people with multiple sclerosis.

BACKGROUND: Health-related quality of life (QOL) is a key outcome for people with multiple sclerosis (MS). While modifiable lifestyle factors, like smoking, physical activity and vitamin D, have strong associations with development and progression of MS, few studies have examined such associations with QOL. METHODS: Using patient-reported data from 2312 people with MS from 54 countries, regression models explored associations of socio-demographic, therapeutic and lifestyle factors with QOL, using the Multiple Sclerosis Quality of Life-54 (MSQOL-54). RESULTS: Participants were on average 45.6 years old, 82.4% women, mostly partnered (74.1%), with a university degree (59.5%). Controlling for socio-demographic factors and disability, factors associated with better physical health composite (PHC) (on a 100 point scale) were: moderate and high physical activity compared to low (5.9 [95% confidence interval: 4.2, 7.6] and 9.9 [CI: 8.1, 11.6] points higher score respectively); non-smoking compared to current smoking (4.6 points [CI: 2.4, 6.7]); better diet (per 10 points on the 100 point Diet Habits Questionnaire scale (DHQ) 1.6 points [CI: 1.0, 2.2] points); normal body mass index (BMI) versus overweight or obese (2.1 points [CI: 0.4, 3.7] and 2.4 points [CI: 0.5, 4.3]); fewer comorbidities (4.4 points [CI: 3.9, 4.9]); and not taking a disease-modifying drug (DMD) (2.1 points [CI: 0.7, 3.4]). Better mental health composite (MHC) determinants were: moderate and high physical activity compared to low (4.0 points [CI: 2.0, 6.0] and 5.7 points [CI: 3.5, 8.0]); non-smoking compared to current (6.7 points [CI: 4.1, 9.3]); better diet (2.8 points [CI: 1.9, 3.5]); normal BMI versus overweight or obese (3.1 points [CI: 1.1, 5.1] and 3.5 points [CI: 1.3, 5.7]); meditating regularly (2.2 points [CI: 0.2, 4.2]); and no DMD use (2.9 points [CI: 1.3, 4.6]). CONCLUSIONS: While causality cannot be concluded from cross-sectional data, the associations between modifiable lifestyle factors and QOL suggest significant potential for secondary prevention of the known deterioration of QOL for people with MS through lifestyle risk factor modification.

Development of functional trait biomarkers for trace metal exposure in freshwater clams (Musculium spp.).

Exposure to trace metals typically causes oxidative stress; these consequences are better-characterized in estuarine and marine species than in freshwater species. How cellular-level responses to metal pollution influence whole-organism and population-level traits is poorly understood. We tested whether exposure to single metals (zinc and cadmium) and to metal mixtures (water in equilibrium with sediment from a highly polluted lake) alters two ecologically-relevant traits in freshwater clams, locomotion and reproduction. Fingernail clams (Musculium spp.) from unimpacted habitats were exposed to single metals and the metal mixture for up to 49days. The single metal doses (≤5mg/L Zn and ≤20µg/L Cd) were not toxicologically meaningful as clam survival, burial, and climbing activity did not differ across treatments. Water in equilibrium with the lake sediment contained cadmium, copper, lead, and zinc. Clams exposed to this metal mixture had decreased climbing activity but no change in burial activity. Metal-exposed clams had lower fecundity (number of shelled juveniles extruded by adult clams) and patterns in metal accumulation corresponded with lake sediment dose and clam activity. In contrast to the functional traits, stress protein expression and whole-clam glycogen content did not vary across treatment groups. These results indicate that fingernail clams of the genus Musculium are appropriate for development as sentinel species for metal pollution and can serve as a model for determining how metal pollution alters metabolic allocation patterns in freshwater organisms.

Kinetic isotope effect of proton-coupled electron transfer in a hydrogen bonded phenol-pyrrolidino[60]fullerene.

Proton-coupled electron transfer (PCET) plays a central role in photosynthesis and potentially in solar-to-fuel systems. We report a spectroscopy study on a phenol-pyrrolidino[60]fullerene. Quenching of the singlet excited state from 1 ns to 250 ps is assigned to PCET. A H/D exchange study reveals a kinetic isotope effect (KIE) of 3.0, consistent with a concerted PCET mechanism.

A de novo designed 2[4Fe-4S] ferredoxin mimic mediates electron transfer.

[Fe-S] clusters, nature's modular electron transfer units, are often arranged in chains that support long-range electron transfer. Despite considerable interest, the design of biomimetic artificial systems emulating multicluster-binding proteins, with the final goal of integrating them in man-made oxidoreductases, remains elusive. Here, we report a novel bis-[4Fe-4S] cluster binding protein, DSD-Fdm, in which the two clusters are positioned within a distance of 12 Å, compatible with the electronic coupling necessary for efficient electron transfer. The design exploits the structural repeat of coiled coils as well as the symmetry of the starting scaffold, a homodimeric helical protein (DSD). In total, eight hydrophobic residues in the core of DSD were replaced by eight cysteine residues that serve as ligands to the [4Fe-4S] clusters. Incorporation of two [4Fe-4S] clusters proceeds with high yield. The two [4Fe-4S] clusters are located in the hydrophobic core of the helical bundle as characterized by various biophysical techniques. The secondary structure of the apo and holo proteins is conserved; further, the incorporation of clusters results in stabilization of the protein with respect to chemical denaturation. Most importantly, this de novo designed protein can mimic the function of natural ferredoxins: we show here that reduced DSD-Fdm transfers electrons to cytochrome c, thus generating the reduced cyt c stoichiometrically.

A new method for generating high purity motoneurons from mouse embryonic stem cells.

A common problem with using embryonic stem (ES) cells as a source for analysis of gene expression, drug toxicity, or functional characterization studies is the heterogeneity that results from many differentiation protocols. The ability to generate large numbers of high purity differentiated cells from pluripotent stem cells could greatly enhance their utility for in vitro characterization studies and transplantation in pre-clinical injury models. Population heterogeneity is particularly troublesome for post-mitotic neurons, including motoneurons, because they do not proliferate and are quickly diluted in culture by proliferative phenotypes, such as glia. Studies of motoneuron biology and disease, in particular amyotrophic lateral sclerosis, can benefit from high purity motoneuron cultures. In this study, we engineered a transgenic-ES cell line where highly conserved enhancer elements for the motoneuron transcription factor Hb9 were used to drive puromycin N-acetyltransferase expression in ES cell-derived motoneurons. Antibiotic selection with puromycin was then used to obtain high purity motoneuron cultures following differentiation of mouse ES cells. Purity was maintained during maturation allowing the production of consistent, uniform populations of cholinergic ES cell-derived motoneurons. Appropriate functional properties of purified motoneurons were verified by acetylcholinesterase activity and electrophysiology. Antibiotic selection, therefore, can provide an inexpensive alternative to current methods for isolating ES cell-derived motoneurons at high purity that does not require specialized laboratory equipment and provides a unique platform for studies in motoneuron development and degeneration.

Modeling membranes in situ.

Molecular dynamics simulations of cellular membranes have come a long way-from simple model lipid bilayers to multicomponent systems capturing the crowded and complex nature of real cell membranes. In this opinionated minireview, we discuss the current challenge to simulate the dynamics of membranes in their native environment, in situ, with the prospect of reaching the level of whole cells and cell organelles using an integrative modeling framework.

Neurodivergent patient experience in a tertiary children's hospital-a qualitative analysis.

Introduction: Sensory processing challenges are commonly encountered in pediatric patients, particularly in those who are neurodivergent. We previously developed a novel clinical pathway (named "Sensory Pathway") which aimed at improving patient care for those with sensory barriers via staff training, provision of sensory toolkits and early integration of families throughout the hospital stay. We hypothesized that utilization of this pathway will result in improved patient experience and provide valuable feedback to improve care. Methods: A voluntary survey was made available to all patients who utilized this resource as part of our hospital wide patient satisfaction survey. Qualitative data was coded using open coding as part of the constant comparison method data using NVivo 12 for Windows software for analysis. Software was used to create word clouds and clusters for visualization, which confirmed the themes and patterns that were noted from initial open coding. Results: Between 2021 and 2022, surveys were obtained from 160 patients who utilized the Sensory Pathway. More than 50% reported that the most helpful components of the pathway were the approach by the staff and sensory tools. The three major themes identified from the survey were (1) Tools and techniques that benefited their children; (2) Positive interactions and communication with the hospital staff, and (3) Suggestions for future improvement. Conclusion: The survey results highlight the importance of having tools readily available to aid with sensory regulation and comfort of patients during healthcare encounters, the value of a positive patient and staff encounter, as well as opportunities for improvement.

Diabetic Ocular Surface Has Defects in Oxygen Uptake Revealed by Optic Fiber Microsensor.

Purpose: Diabetes mellitus causes diabetic keratopathy (DK). This and other ocular surface disorders are underdiagnosed and problematic for affected patients as well as recipients of diabetic donor corneas. Thus, it is important to find noninvasive means to facilitate determination of the potentially vision-threatening DK. It has been reported that diabetic corneas uptake significantly less oxygen (O2) than healthy controls. However, an integral assessment of the ocular surface is missing. Methods: Using an optic-fiber O2 micro-sensor (optrode) we demonstrated recently that the healthy ocular surface displays a unique spatiotemporal map of O2 consumption. We hypothesize that diabetes impairs the spatiotemporal profile of O2 uptake at the ocular surface. Results: Using streptozotocin (STZ)-induced diabetic mice, we found diminished O2 uptake and loss of the unique pattern across the ocular surface. A diabetic cornea consumes significantly less O2 at the bulbar conjunctiva and limbus, but not the central and peripheral cornea, compared to controls. Further, we show that, contrary to the healthy cornea, the diabetic cornea does not increase the O2 consumption at the limbus in the evening as the normal control. Conclusions: Altogether, our measurements reveal a previously unknown impairment in O2 uptake at the diabetic cornea, making it a potential tool to diagnose ocular surface abnormalities and suggesting a new etiology mechanism.

Considerations for Hand Surgery in Patients With Scleroderma.

Systemic sclerosis (scleroderma, SSc) is an autoimmune disease that causes significant dysfunction to multiple organ systems, including the musculoskeletal system. It poses significant challenges to the hand surgeon, including calcinosis, ischemic changes, Raynaud phenomenon, tendinopathies, synovitis, and joint contractures. Patients with SSc also suffer from multiorgan dysfunction, which makes them high-risk surgical patients. The hand surgeon must understand the pathophysiology, treatment strategies, and special operative considerations required in this population to avoid complications and help maintain or improve hand function.