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Abstract Chronic HIV disease is associated with a fivefold increase in albuminuria outside of sub-Saharan Africa. However, very little is known about albuminuria risk among people living with HIV (PLWH) in sub-Saharan Africa. Therefore, we conducted a cross-sectional observational HIV clinic-based study of albuminuria among 1533 adults aged 21 years or older between January 2020 and January 2021 in Gaborone, Botswana. Clinical albuminuria was defined using a sex-based albumin‒creatinine ratio (ACR) of 25–355 mg/g for females and 17–250 mg/g for males. The study population mean age was 48.5 (SD 10.3) years, and 764/1533 (49.7%) were female. The overall prevalence of albuminuria was 20.7% (95% CI 18.7%, 22.8%). A higher proportion of males were more likely to be categorized as having albuminuria than females, 25% (95% CI 22.0, 28.2) versus 16.4% (95% CI 13.8,19.2), P value

Abstract People living with HIV (PLWH) experience increased vulnerability to premature aging and inflammation-associated comorbidities, even when HIV replication is suppressed by antiretroviral therapy (ART). However, the factors associated with this vulnerability remain uncertain. In the general population, alterations in the N-glycans on IgGs trigger inflammation and precede the onset of aging-associated diseases. Here, we investigate the IgG N-glycans in cross-sectional and longitudinal samples from 1214 women and men, living with and without HIV. PLWH exhibit an accelerated accumulation of pro-aging-associated glycan alterations and heightened expression of senescence-associated glycan-degrading enzymes compared to controls. These alterations correlate with elevated markers of inflammation and the severity of comorbidities, potentially preceding the development of such comorbidities. Mechanistically, HIV-specific antibodies glycoengineered with these alterations exhibit a reduced ability to elicit anti-HIV Fc-mediated immune activities. These findings hold potential for the development of biomarkers and tools to identify and prevent premature aging and comorbidities in PLWH.

Objectives: In the United States, a federal emergency program has made SARS-CoV-2 self-test kits available at no cost. It is unclear how widely free tests are preferred. We conducted a survey to estimate the proportion of respondents who do not prefer a free test. We hypothesized that free tests would not be preferred universally, and that a preference for paying would be more common among those with conservative politics than with liberal politics, regardless of income. Design: Observational study design. Methods: A national sample of US adults completed an online survey. To reduce potential enrollment bias, the survey’s focus was not specified beforehand. To prioritize a high-risk group, participation was limited to those who were unvaccinated or were incompletely vaccinated in the primary series against COVID-19. Participants reported their testing preferences and socio-demographic characteristics, including political affiliation. The main outcome assessed if a participant preferred to pay for a self-test or receive a free one (subsidized by the US government). Results: Among 1215 participants, (73%, n = 886) preferred free self-testing, while 27% (n = 329) preferred to pay for the same testing. After adjusting for income, the odds of preferring to pay for self-testing were 66% higher in “strong” Republicans compared to “strong” Democrats (odds ratio = 1.66, 95% confidence interval = 1.07–2.62). Conclusions: More than a quarter of individuals preferred paying for these tests. This preference was more likely among those with right-wing politics. Policy implications are discussed, along with future research directions.

BackgroundWidespread access to testing is critical to public health efforts to control the COVID-19 pandemic. Secondary distribution of COVID-19 self-test kits, where an individual distributes test kits to others in their social networks, is a potential strategy to improve access to testing. In this qualitative study, we identified salient beliefs about distributing and accepting COVID-19 self-test kits within one’s social network, as well as ordering COVID-19 self-test kits from the government.MethodsWe recruited 61 participants from a randomized controlled trial (NCT04797858) in Philadelphia, Pennsylvania to elicit beliefs about (1) distributing COVID-19 self-test kits within one’s social network, (2) receiving test kits from social contacts, and (3) ordering self-test kits from the government. Using validated, open-ended question stems, we identified the most common set of beliefs underlying attitudes, perceived norms (or social referents), and perceived behavioral control (or self-efficacy) toward each of these behaviors.ResultsTwenty-seven out of 30 (90%) of participants who received self-test kits reported distributing the kits to social contacts. These participants described altruistic beliefs about giving others access to testing, and felt approval from family members, friends, and others in their social networks. When receiving test kits from social network contacts, participants described advantages of test kit convenience, but some voiced concern about test kit tampering and confusing instructions. Participants also described perceived logistic barriers to distributing and receiving self-test kits, such as delivering or transporting test kits, or finding time to meet. Participants who ordered test kits from the government also described increased convenience of test access, but described different logistic barriers such as delays in test kit delivery, or not receiving test kits at all.ConclusionIn comparison with government-ordered test kits, the secondary distribution of COVID-19 self-test kits raised unique concerns about test kit quality and instructions, as well as distinctive logistic barriers related to distributing self-test kits to network contacts, which were not raised for test kits ordered from the government. This study demonstrates that beliefs may vary depending on the type of testing behavior, and behavioral interventions may benefit from developing messages tailored to specific testing strategies.

Abstract Objective This study aimed to evaluate safety (infusion‐related reactions [IRRs]) and patient satisfaction (patient‐reported outcomes [PROs]) for at‐home ocrelizumab administration for patients with multiple sclerosis (MS). Methods This open‐label study included adult patients with an MS diagnosis who had completed a ≥ 600‐mg ocrelizumab dose, had a patient‐determined disease steps score of 0 to 6 and had completed PROs. Eligible patients received a 600‐mg ocrelizumab home‐based infusion over 2 h, followed by 24‐h and 2‐week post‐infusion follow‐up calls. IRRs and adverse events (AEs) were documented during infusions and follow‐up calls. PROs were completed before and 2 weeks post infusion. Results Overall, 99 of 100 expected patients were included (mean [SD] age, 42.3 [7.7] years; 72.7% female; 91.9% White). The mean (SD) infusion time was 2.5 (0.6) hours, and 75.8% of patients completed their ocrelizumab infusion between 2 to 2.5 h. The IRR incidence rate was 25.3% (95% CI: 16.7%, 33.8%)—similar to other shorter ocrelizumab infusion studies—and all AEs were mild/moderate. In total, 66.7% of patients experienced AEs, including itch, fatigue, and grogginess. Patients reported significantly increased satisfaction with the at‐home infusion process and confidence in the care provided. Patients also reported a significant preference for at‐home infusion compared with prior infusion center experiences. Interpretation IRRs and AEs occurred at acceptable rates during in‐home infusions of ocrelizumab over a shorter infusion time. Patients reported increased confidence and comfort with the home infusion process. Findings from this study provide evidence of the safety and feasibility of home‐based ocrelizumab infusion over a shorter infusion period.

Introduction To end the HIV epidemic in Philadelphia, implementation of evidence-based practices (EBP) to increase viral suppression and retention in HIV care is critical. Managed problem solving (MAPS), an EBP for antiretroviral therapy adherence, follows a problem-solving approach to empower people living with HIV (PWH) to manage their health. To overcome barriers to care experienced by PWH in Philadelphia, the EBP was adapted to include a focus on care retention and delivery by community health workers (CHWs). The adapted intervention is MAPS+. To maximise the clinical impact and reach of the intervention, evaluation of the effectiveness and implementation of MAPS+ is necessary.Methods and analysis This manuscript describes the protocol for a stepped-wedge cluster-randomised type 2 hybrid effectiveness-implementation trial in 10 clinics in Philadelphia. This research incorporates innovative approaches to accomplish three objectives: (1) to evaluate the effectiveness of the CHW-led MAPS+ intervention to improve viral suppression and retention in care 1 year after the individual implementation period (N=390 participants), (2) to examine the effect of the menu of implementation strategies on reach and implementation cost and (3) to examine processes, mechanisms, and sustainment of the implementation strategies for MAPS+ (N=56 participants). Due to various factors (eg, COVID-19), protocol modifications have occurred.Ethics and dissemination The institutional review board (IRB) at the city of Philadelphia serves as the primary IRB; initial approval was granted on 21 December 2020. The University of Pennsylvania and Northwestern University executed reliance agreements. A safety monitoring committee comprised experts in implementation science, biostatistics and infectious diseases oversee this study. This research will offer insights into achieving the goals to end the HIV epidemic in Philadelphia as well as implementation efforts of MAPS+ and other behavioural interventions aimed at increasing medication adherence and retention in care. Dissemination will include deliverables (eg, peer-reviewed manuscripts and lay publications) to reach multiple constituents.Trial registration number NCT04560621.

Abstract Background Clinical decision support (CDS) is a promising intervention for improving uptake of HIV testing and pre-exposure prophylaxis (PrEP). However, little is known regarding provider perspectives on acceptability, appropriateness, and feasibility of CDS for HIV prevention in pediatric primary care, a key implementation setting. Methods This was a cross-sectional multiple methods study utilizing surveys and in-depth interviews with pediatricians to assess acceptability, appropriateness, and feasibility of CDS for HIV prevention, as well as to identify contextual barriers and facilitators to CDS. Qualitative analysis utilized work domain analysis and a deductive coding approach grounded in the Consolidated Framework of Implementation Research. Quantitative and qualitative data were merged to develop an Implementation Research Logic Model to conceptualize implementation determinants, strategies, mechanisms, and outcomes of potential CDS use. Results Participants (n = 26) were primarily white (92%), female (88%), and physicians (73%). Using CDS to improve HIV testing and PrEP delivery was perceived as highly acceptable (median score 5), IQR [4–5]), appropriate (5, IQR [4–5]), and feasible (4, IQR [3.75–4.75]) using a 5-point Likert scale. Providers identified confidentiality and time constraints as two key barriers to HIV prevention care spanning every workflow step. With respect to desired CDS features, providers sought interventions that were integrated into the primary care workflow, standardized to promote universal testing yet adaptable to the level of a patient’s HIV risk, and addressed providers’ knowledge gaps and bolstered self-efficacy in providing HIV prevention services. Conclusions This multiple methods study indicates that clinical decision support in the pediatric primary care setting may be an acceptable, feasible, and appropriate intervention for improving the reach and equitable delivery of HIV screening and PrEP services. Design considerations for CDS in this setting should include deploying CDS interventions early in the visit workflow and prioritizing standardized but flexible designs.

Abstract Background Treatment management after repeated failure of antiretroviral therapy (ART) is difficult due to resistance and adherence challenges. For people who have failed non-nucleoside reverse transcriptase inhibitor-(NNRTI-) and protease inhibitor-(PI-) based regimens with no or limited resistance, remaining on PI-based ART is an option. Using data from an ART strategy trial (A5288) in low/middle-income countries which included this option, we explored whether predictors can be identified distinguishing those who experienced further virologic failure from those who achieved and maintained virologic suppression. Methods A5288 enrolled people with confirmed HIV-1 RNA ≥ 1000 copies/mL after ≥ 24 weeks of PI-based ART and prior failure on NNRTI-based ART. This analysis focused on the 278 participants with no resistance to the PI being taken and no or limited nucleoside reverse transcriptase inhibitor (NRTI) resistance, who continued their PI with flexibility to change NRTIs. Proportional hazards models were used to evaluate predictors of virologic failure during follow-up (VF: confirmed HIV-1 RNA ≥ 1000 copies/mL at ≥ 24 weeks of follow-up). Results 56% of participants were female. At study entry, median age was 40 years, time on ART 7.8 years, CD4 count 169 cells/mm3, HIV-1 RNA 20,444 copies/mL; and 37% had NRTI resistance. The estimated proportion experiencing VF increased from 39% at week 24 to 60% at week 96. In multivariable analysis, significant predictors at study entry of VF were higher HIV-1 RNA (adjusted hazard ratio: 2.20 for ≥ 10,000 versus

Abstract Introduction The metabolic syndrome, a cluster of inter-related risk factors for cardiovascular diseases is highly prevalent among individuals with obesity and sedentary lifestyle. Chronic psychiatric disorders such as severe mental illness are associated with increased risk for cardiovascular diseases. We aimed to assess the prevalence and correlates of metabolic syndrome among inpatients with severe mental illness in a resource limited setting with high HIV prevalence. Methods This was a cross-sectional study among adult inpatients at a referral psychiatric hospital in Botswana. We used convenience sampling to enrol participants available at the time of the study. The National Cholesterol Education Program Adult Treatment Panel-III (NCEP-ATP III) criteria was used to define the metabolic syndrome. Data were analysed using descriptive statistics as well as multiple logistic regression modelling. Results A total of 137 participants were enrolled. Of these, 119 (87%) had complete data for the main analysis. The overall prevalence of metabolic syndrome was 22.6% (95% CI 15.9, 30.6) and did not differ significantly by gender or HIV status. Age was significantly associated with the risk of having the metabolic syndrome while gender, body mass index, HIV status, and days of moderate physical activity were not. Conclusion There was a moderately high prevalence of metabolic syndrome. Thus, the management of individuals with severe mental illness in resource limited settings should include assessment of cardiovascular risk and target modifiable risk factors in this population. Consideration for the patient’s age should be made when rationalizing the limited resources available for assessing metabolic syndrome among patients with severe mental illness.

Objectives: Although extended-spectrum cephalosporin-resistant Enterobacterales (ESCrE) and carbapenem-resistant Enterobacterales (CRE) are a global challenge, data on these organisms in low- and middle-income countries are limited. In this study, we sought to characterize colonization data critical for greater antibiotic resistance surveillance efforts. Methods: This study was conducted in three hospitals and six clinics in Botswana. We conducted ongoing surveillance of adult patients in hospitals and clinics and adults and children in the community. All participants underwent rectal swab sampling to identify ESCrE and CRE. Results: Enrollment occurred from January 15, 2020, to September 4, 2020, but paused from April 2, 2020, to May 21, 2020, because of a countrywide COVID-19 lockdown. Of 5088 individuals approached, 2469 (49%) participated. ESCrE colonization prevalence was 30.7% overall (43% for hospital participants, 31% for clinic participants, 24% for adult community participants, and 26% for child community participants) (P

Background: The epidemiology of extended-spectrum cephalosporin-resistant Enterobacterales (ESCrE) in hospitalized patients in low- and middle-income countries (LMICs) is poorly described. Although risk factors for ESCrE clinical infection have been studied, little is known of the epidemiology of ESCrE colonization. Identifying risk factors for ESCrE colonization, which can predispose to infection, is therefore critical to inform antibiotic resistance reduction strategies. Methods: This study was conducted in 3 hospitals located in 3 districts in Botswana. In each hospital, we conducted ongoing surveillance in sequential units hospitalwide. All participants had rectal swabs collected which were inoculated onto chromogenic media followed by confirmatory testing using MALDI-TOF MS and VITEK-2. Data were collected via interview and review of the inpatient medical record on demographics, comorbidities, antibiotic use, healthcare exposures, invasive procedures, travel, animal contact, and food consumption. Participants with ESCrE colonization (cases) were compared to noncolonized participants (controls) using bivariable and multivariable analyses to identify risk factors for ESCrE colonization. Results: Enrollment occurred from January 15, 2020, to September 4, 2020, and 469 participants were enrolled. The median age was 42 years (IQR, 31–58) and 320 (68.2%) were female. The median time from hospital admission to date of sampling was 5 days (IQR, 3–12). There were 179 cases and 290 controls (ie, 38.2% of participants were ESCrE colonized). Independent risk factors for ESCrE colonization were a greater number of days on antibiotic, recent healthcare exposure, and tending swine prior to hospitalization. (Table). Conclusions: ESCrE colonization among hospitalized patients was common and was associated with several exposures. Our results suggest prior healthcare exposure may be important in driving ESCrE. The strong link to recent antibiotic use highlights the potential role of antibiotic stewardship interventions for prevention. The association with tending swine suggests that animal husbandry practices may play a role in community exposures, resulting in colonization detected at the time of hospital admission. These findings will help to inform future studies assessing strategies to curb further emergence of hospital ESCrE in LMICs.

Postmodern music therapy is an approach to music therapy that defines itself in diametric opposition to modernist, or medical-model, music therapy. Where modernist music therapy ignores power dynamics and assumes itself to be value neutral, postmodern music therapy is concerned with power dynamics between client, therapist, and broader society. Postmodern music therapy is based on the theories of Bradley Lewis: Lewis (2006) believes that a postmodern psychiatry (or as he calls it, postpsychiatry) will be more aware of politics and social structures in general and will move toward democratization. This article proposes a postmodern music therapy in parallel to postpsychiatric ideas and ideals. Further, the article explores the intersectional nexus between postmodern music therapy and critical race, disability, queer, and feminist studies.

PURPOSEOral cancer is the sixth most common cancer worldwide and is the seventh most common in Botswana. Lack of improvement in oral cancer survival despite the availability of multiple treatment options may be due to the high prevalence of advanced stage at presentation. We identified risk factors for presenting with oral cancer at an advanced stage to facilitate interventions to reduce mortality from oral cancers.METHODSA retrospective cohort analysis was conducted among individuals with biopsy-confirmed oral cancer at Princess Marina Hospital in Gaborone, Botswana, between 2010 and 2020. Data collected included age at diagnosis, sex, place of residence, HIV status, oral cancer stage, and oral subsite. Multivariable analyses were controlled for age, sex, district of residence, and oral subsite.RESULTSOf the 218 records analyzed, 79% were male, 58% were HIV-positive, the median age was 56 years (interquartile range: 47-63), and 67% presented with advanced-stage disease. Cancers from hidden oral sites were more likely to present at an advanced stage with an adjusted odds ratio (OR) of 2.98 (95% CI, 1.29 to 6.89; P = .01). Residence in socioeconomically disadvantaged districts was associated with higher likelihood (OR, 2.36; 95% CI, 1.28 to 4.39; P = .01) of advanced stage presentation compared with other districts. HIV infection was not associated with risk of advanced lesion presentation (OR, 1; 95% CI, 0.61 to 1.61; P = .97).CONCLUSIONHidden oral cancer sites and residence in districts with limited access to care were risk factors for advanced oral cancer at the time of diagnosis in Botswana. These findings support a need to increase efforts to improve access to care and increase oral cancer awareness to decrease the burden of advanced oral cancer.

BackgroundImplementation mapping is a systematic, collaborative, and contextually-attentive method for developing implementation strategies. As an exemplar, we applied this method to strategy development for Managed Problem Solving Plus (MAPS+), an adapted evidence-based intervention for HIV medication adherence and care retention that will be delivered by community health workers and tested in an upcoming trial.MethodsIn Step 1: Conduct Needs Assessment, we interviewed 31 stakeholders to identify determinants of MAPS+ implementation in 13 clinics serving people with HIV in Philadelphia County. In Step 2: Develop Logic Model, we used these determinants as inputs for a working logic model guided by the Consolidated Framework for Implementation Research. In Step 3: Operationalize Implementation Strategies, our team held a virtual stakeholder meeting to confirm determinants. We synthesized stakeholder feedback, then identified implementation strategies that conceptually matched to determinants using the Expert Recommendations for Implementing Change taxonomy. Next, we operationalized implementation strategies with specific examples for clinic settings. We linked strategies to behavior change theories to allow for a mechanistic understanding. We then held a second virtual stakeholder meeting to present the implementation menu for feedback and glean generalizable insights for how these strategies could be operationalized in each stakeholder's clinic. In Step 4: Protocolize Strategies, we incorporated stakeholder feedback and finalized the implementation strategy menu.FindingsImplementation mapping produced a menu of 39 strategies including revise professional roles, identify and prepare champions, use warm handoffs, and change record systems. The process of implementation mapping generated key challenges for implementation strategy development: lack of implementation strategies targeting the outer setting (i.e., sociopolitical context); tension between a one-size-fits-all and individualized approach for all clinics; conceptual confusion between facilitators and strategies; and challenges in translating the implementation science lexicon for partners.ImplicationsThis case exemplar advances both MAPS+ implementation and implementation science methods by furthering our understanding of the use of implementation mapping to develop strategies that enhance uptake of evidence-based interventions. The implementation menu will inform MAPS+ deployment across Philadelphia in an upcoming hybrid trial. We will carry out Step 5: Test Strategies to test the effectiveness and implementation of MAPS+.

PURPOSEPatients who are HIV-positive and have breast cancer have worse overall survival (OS) compared with patients who are HIV-negative. Pathologic complete response (pCR) and relative dose intensity (RDI) of chemotherapy are associated with survival. We assessed whether pCR and RDI rates were lower for patients who are HIV-positive and received neoadjuvant chemotherapy (NACT).METHODSThis was a prospective cohort analysis of patients initiating NACT in Botswana (February 2017 to September 2019). Primary outcomes were pCR and RDI; secondary outcomes were OS and toxicity. HIV status and zidovudine (ZDV) treatment were stratification factors. Multivariable analysis was used to control for confounding.RESULTSIn total, 26 of 110 enrolled individuals were HIV-positive. In univariable analysis, HIV-positive (odds ratio [OR] = 0.2; P = .048) and RDI < 0.85 (OR = 0.30; P = .025) were associated with pCR. In multivariable analysis, the magnitude of association decreased for HIV-positive (OR = 0.28; P = .11), but RDI < 0.85 remained independently associated with pCR (OR = 0.32; P = .035). Patients who are HIV-positive had significantly lower mean RDI, and those on ZDV had significantly lower RDI. Ninety-one (83%) were stage III with 2-year OS significantly worse for patients who are HIV-positive (58% v 74%). Hazard ratio for all-cause mortality was 2.68 (95% CI, 1.17 to 6.13; P = .028) in patients who are HIV-positive compared with patients who are HIV-negative. Toxicity rates were similar despite patients who are HIV-positive receiving significantly lower dose intensity chemotherapy.CONCLUSIONPatients who are HIV-positive and have breast cancer in Botswana have lower pCR rates and also receive lower dose intensity therapy, which may contribute to worse OS. Patients who are HIV-positive on ZDV-containing regimens received even lower dose intensity of NACT. Administering optimal dose intensity in patients who are HIV-positive remains a challenge, and targeted interventions that address modifiable risk factors are needed to improve therapy delivery and outcomes.

Abstract Focal adhesions (FAs) strengthen their link with the actin cytoskeleton to resist force. Talin-vinculin association could reinforce actin anchoring to FAs by controlling actin polymerization. However, the actin polymerization activity of the talin-vinculin complex is not known because it requires the reconstitution of the mechanical and biochemical activation steps that control the association of talin and vinculin. By combining kinetic and binding assays with single actin filament observations in TIRF microscopy, we show that the association of talin and vinculin mutants, mimicking mechanically stretched talin and activated vinculin, triggers a sequential mechanism in which filaments are nucleated, capped and released to elongate. In agreement with these observations, FRAP experiments in cells co-expressing the same constitutive mutants of talin and vinculin revealed accelerated growth of stress fibers. Our findings suggest a versatile mechanism for the regulation of actin assembly in FAs subjected to various combinations of biochemical and mechanical cues.

Abstract Patients with corticosteroid-refractory acute graft-versus-host disease (aGVHD) have a low one-year survival rate. Identification and validation of novel targetable kinases in patients who experience corticosteroid-refractory-aGVHD may help improve outcomes. Kinase-specific proteomics of leukocytes from patients with corticosteroid-refractory-GVHD identified rho kinase type 1 (ROCK1) as the most significantly upregulated kinase. ROCK1/2 inhibition improved survival and histological GVHD severity in mice and was synergistic with JAK1/2 inhibition, without compromising graft-versus-leukemia-effects. ROCK1/2-inhibition in macrophages or dendritic cells prior to transfer reduced GVHD severity. Mechanistically, ROCK1/2 inhibition or ROCK1 knockdown interfered with CD80, CD86, MHC-II expression and IL-6, IL-1β, iNOS and TNF production in myeloid cells. This was accompanied by impaired T cell activation by dendritic cells and inhibition of cytoskeletal rearrangements, thereby reducing macrophage and DC migration. NF-κB signaling was reduced in myeloid cells following ROCK1/2 inhibition. In conclusion, ROCK1/2 inhibition interferes with immune activation at multiple levels and reduces acute GVHD while maintaining GVL-effects, including in corticosteroid-refractory settings.

Abstract Cells rapidly respond to replication stress actively slowing fork progression and inducing fork reversal. How replication fork plasticity is achieved in the context of nuclear organization is currently unknown. Using nuclear actin probes in living and fixed cells, we visualized nuclear actin filaments in unperturbed S phase and observed their rapid extension in number and length upon genotoxic treatments, frequently taking contact with replication factories. Chemically or genetically impairing nuclear actin polymerization shortly before these treatments prevents active fork slowing and abolishes fork reversal. Defective fork remodeling is linked to deregulated chromatin loading of PrimPol, which promotes unrestrained and discontinuous DNA synthesis and limits the recruitment of RAD51 and SMARCAL1 to nascent DNA. Moreover, defective nuclear actin polymerization upon mild replication interference induces chromosomal instability in a PRIMPOL-dependent manner. Hence, by limiting PrimPol activity, nuclear F-actin orchestrates replication fork plasticity and is a key molecular determinant in the rapid cellular response to genotoxic treatments.

Abstract Our current understanding of biomolecular condensate formation is largely based on observing the final near-equilibrium condensate state. Despite expectations from classical nucleation theory, pre-critical protein clusters were recently shown to form under subsaturation conditions in vitro; if similar long-lived clusters comprising more than a few molecules are also present in cells, our understanding of the physical basis of biological phase separation may fundamentally change. Here, we combine fluorescence microscopy with photobleaching analysis to quantify the formation of clusters of NELF proteins in living, stressed cells. We categorise small and large clusters based on their dynamics and their response to p38 kinase inhibition. We find a broad distribution of pre-condensate cluster sizes and show that NELF protein cluster formation can be explained as non-classical nucleation with a surprisingly flat free-energy landscape for a wide range of sizes and an inhibition of condensation in unstressed cells.

Summary: Background: Although immunomodulators have established benefit against the new coronavirus disease (COVID-19) in general, it is uncertain whether such agents improve outcomes without increasing the risk of secondary infections in the specific subgroup of previously immunocompromised patients. We assessed the effect of immunomodulators on outcomes of immunocompromised patients hospitalized for COVID-19. Methods: The protocol was prospectively registered with PROSPERO (CRD42022335397). MEDLINE, Cochrane Central Register of Controlled Trials and references of relevant articles were searched up to 01-06-2022. Authors of potentially eligible randomized controlled trials were contacted to provide data on immunocompromised patients randomized to immunomodulators vs control (i.e., placebo or standard-of-care). Findings: Eleven randomized controlled trials involving 397 immunocompromised patients hospitalized for COVID-19 were included. Ten trials had low risk of bias. There was no difference between immunocompromised patients randomized to immunomodulators vs control regarding mortality [30/182 (16.5%) vs 41/215 (19.1%); RR 0.93, 95% CI 0.61–1.41; p = 0.74], secondary infections (RR 1.00, 95% CI 0.64–1.58; p = 0.99) and change in World Health Organization ordinal scale from baseline to day 15 (weighed mean difference 0.27, 95% CI -0.09–0.63; p = 0.15). In subgroup analyses including only patients with hematologic malignancy, only trials with low risk of bias, only trials administering IL-6 inhibitors, or only trials administering immunosuppressants, there was no difference between comparators regarding mortality. Interpretation: Immunomodulators, compared to control, were not associated with harmful or beneficial outcomes, including mortality, secondary infections, and change in ordinal scale, when administered to immunocompromised patients hospitalized for COVID-19. Funding: Hellenic Foundation for Research and Innovation.

Abstract Artificial intelligence methods including deep neural networks (DNN) can provide rapid molecular classification of tumors from routine histology with accuracy that matches or exceeds human pathologists. Discerning how neural networks make their predictions remains a significant challenge, but explainability tools help provide insights into what models have learned when corresponding histologic features are poorly defined. Here, we present a method for improving explainability of DNN models using synthetic histology generated by a conditional generative adversarial network (cGAN). We show that cGANs generate high-quality synthetic histology images that can be leveraged for explaining DNN models trained to classify molecularly-subtyped tumors, exposing histologic features associated with molecular state. Fine-tuning synthetic histology through class and layer blending illustrates nuanced morphologic differences between tumor subtypes. Finally, we demonstrate the use of synthetic histology for augmenting pathologist-in-training education, showing that these intuitive visualizations can reinforce and improve understanding of histologic manifestations of tumor biology.

Abstract Introduction AO Spine RECODE-DCM was a multi-stakeholder priority setting partnership (PSP) to define the top ten research priorities for degenerative cervical myelopathy (DCM). Priorities were generated and iteratively refined using a series of surveys administered to surgeons, other healthcare professionals (oHCP) and people with DCM (PwDCM). The aim of this work was to utilise word clouds to enable the perspectives of people with the condition to be heard earlier in the PSP process than is traditionally the case. The objective was to evaluate the added value of word clouds in the process of defining research uncertainties in National Institute for Health Research (NIHR) James Lind Alliance (JLA) Priority Setting Partnerships. Methods Patient-generated word clouds were created for the four survey subsections of the AO Spine RECODE-DCM PSP: diagnosis, treatment, long-term management and other issues. These were then evaluated as a nested methodological study. Word-clouds were created and iteratively refined by an online support group of people with DCM, before being curated by the RECODE-DCM management committee and expert healthcare professional representatives. The final word clouds were embedded within the surveys administered at random to 50% of participants. DCM research uncertainties suggested by participants were compared pre- and post-word cloud presentation. Results A total of 215 (50.9%) participants were randomised to the word cloud stream, including 118 (55%) spinal surgeons, 52 (24%) PwDCM and 45 (21%) oHCP. Participants submitted 434 additional uncertainties after word cloud review: word count was lower and more uniform across each survey subsections compared to pre-word cloud uncertainties. Twenty-three (32%) of the final 74 PSP summary questions did not have a post-word cloud contribution and no summary question was formed exclusively on post-word cloud uncertainties. There were differences in mapping of pre- and post-word cloud uncertainties to summary questions, with greater mapping of post-word cloud uncertainties to the number 1 research question priority: raising awareness. Five of the final summary questions were more likely to map to the research uncertainties suggested by participants after having reviewed the word clouds. Conclusions Word clouds may increase the perspective of underrepresented stakeholders in the research question gathering stage of priority setting partnerships. This may help steer the process towards research questions that are of highest priority for people with the condition.

In plants, plasma membrane topologies are predominantly driven by the cell wall. In this study, the authors demonstrate that remorin proteins can take over these functions at specialized, unwalled plasma membranes such as infection droplets associated with symbiotic infection threads.

Abstract Nuclear actin participates in a continuously expanding list of core processes within eukaryotic nuclei, including the maintenance of genomic integrity. In response to DNA damage, nuclear actin polymerises into filaments that are involved in the repair of damaged DNA through incompletely defined mechanisms. We present data to show that the formation of nuclear F-actin in response to genotoxic stress acts as a scaffold for PML NBs and that these filamentous networks are essential for PML NB fission and recruitment of microbodies to DNA lesions. Further to this, we demonstrate that the accumulation of the toxic lamin A precursor prelamin A induces mislocalisation of nuclear actin to the nuclear envelope and prevents the establishment of nucleoplasmic F-actin networks in response to stress. Consequently, PML NB dynamics and recruitment to DNA lesions is ablated, resulting in impaired DNA damage repair. Inhibition of nuclear export of formin mDia2 restores nuclear F-actin formation by augmenting polymerisation of nuclear actin in response to stress and rescues PML NB localisation to sites of DNA repair, leading to reduced levels of DNA damage.

Abstract Vesicle trafficking has emerged as an important process driving tumor progression through various mechanisms. Transforming growth factor beta (TGFβ)‐mediated secretion of Angiopoietin‐like 4 (ANGPTL4) is important for cancer development. Here, Formin‐like 2 (FMNL2) is identified to be necessary for ANGPTL4 trafficking and secretion in response to TGFβ. Protein kinase C (PKC)‐dependent phosphorylation of FMNL2 downstream of TGFβ stimulation is required for cancer cell invasion as well as ANGPTL4 vesicle trafficking and secretion. Moreover, using super resolution microscopy, ANGPTL4 trafficking is actin‐dependent with FMNL2 directly polymerizing actin at ANGPTL4‐containing vesicles, which are associated with Rab8a and myosin Vb. This work uncovers a formin‐controlled mechanism that transiently polymerizes actin directly at intracellular vesicles to facilitate their mobility. This mechanism may be important for the regulation of cancer cell metastasis and tumor progression.