Development of an activity-based therapy tracking tool: Item generation and reduction using the Delphi method.

CONTEXT/OBJECTIVE: Activity-based therapies (ABT) are increasingly used in rehabilitation after spinal cord injury or disease (SCI/D). However, the absence of standardized tools to track the details of an ABT program hinders the collection of data needed for client-tailored programming and resource allocation. The objective of this study is to determine the content to include in an ABT tracking tool for people living with SCI/D. DESIGN: Cross-sectional e-survey. SETTING: Community. PARTICIPANTS: The 60 participants from Canada and the United States who had knowledge and/or experience with ABT included: individuals with SCI/D; hospital clinicians (i.e. physical and occupational therapists/assistants); community-based clinicians; hospital or community clinic administrators; researchers; and funders, advocates and policy makers. INTERVENTIONS: None. OUTCOME MEASURES: A Delphi e-survey comprised 16 types of ABT (e.g. treadmill training) and 4 types of technology (e.g. virtual reality). Participants rated the importance of including each item on a tracking tool and the feasibility to track each item using a 9-point Likert scale. RESULTS: After two survey rounds, nine types of ABT and one technology were identified as important to include in a tracking tool. All items rated as important were considered feasible for clinicians and people with SCI/D to track, except crawling. CONCLUSION: This study identified the types of ABT and technology to include in an ABT tracking tool. Such a tool may provide details of an ABT program that can support decision-making at the individual, program and health system levels and aid the development of best practice guidelines.

Identifying human pre-mRNA cleavage and polyadenylation factors by genome-wide CRISPR screens using a dual fluorescence readthrough reporter.

Messenger RNA precursors (pre-mRNA) generally undergo 3' end processing by cleavage and polyadenylation (CPA), which is specified by a polyadenylation site (PAS) and adjacent RNA sequences and regulated by a large variety of core and auxiliary CPA factors. To date, most of the human CPA factors have been discovered through biochemical and proteomic studies. However, genetic identification of the human CPA factors has been hampered by the lack of a reliable genome-wide screening method. We describe here a dual fluorescence readthrough reporter system with a PAS inserted between two fluorescent reporters. This system enables measurement of the efficiency of 3' end processing in living cells. Using this system in combination with a human genome-wide CRISPR/Cas9 library, we conducted a screen for CPA factors. The screens identified most components of the known core CPA complexes and other known CPA factors. The screens also identified CCNK/CDK12 as a potential core CPA factor, and RPRD1B as a CPA factor that binds RNA and regulates the release of RNA polymerase II at the 3' ends of genes. Thus, this dual fluorescence reporter coupled with CRISPR/Cas9 screens reliably identifies bona fide CPA factors and provides a platform for investigating the requirements for CPA in various contexts.

A multi-site cultural and linguistic adaptation of a hypospadias decision aid for Latinx communities.

INTRODUCTION: Latinx, Spanish-speaking (LSS) patients are more likely to experience decisional conflict and regret about healthcare decisions than non-Hispanic, white, English-speaking patients. OBJECTIVE: To adapt the Hypospadias Hub (Hub), a rigorously developed and tested web-based decision aid (DA), for LSS parents. METHODS: Guided by the Ecological Validity Model (EVM), a heuristic framework was followed to culturally adapt the Hub (see Extended Summary Figure). In stage 1, recommendations were obtained from a focus group with members of the institution's Latinx Community Review Board (Latinx CRB) and semi-structured interviews with pediatricians with Latinx-focused practices. In stage 2, preliminary cultural modifications were made, the Hub was translated into Spanish, and a second focus group with the Latinx CRB was convened to review the revised Hub. In stage 3, semi-structured interviews with LSS parents of healthy boys (i.e., without hypospadias) ≤ 5 years old were conducted to identify any cultural adaptations and/or usability issues regarding the revised Hub. In stage 4, based on parents' feedback, final revisions to the Hub were made. The focus groups and parent interviews were conducted in Spanish; then, the recordings were professionally transcribed in Spanish and translated into English. Interviews with pediatricians were conducted in English; then, the recordings were professionally transcribed. Three coders conducted a qualitative content analysis to identify areas for revision. Changes were applied iteratively. RESULTS: Participants included 3 Latinx CRB members (2 women, 1 did not disclose gender; mean age = 48.3, SD = 21.2), 3 non-Latinx pediatricians (2 women, 1 man; mean age = 49.6, SD = 9.1), and 5 Latinx mothers (mean age = 34.0, SD = 1.26). Participants recommended: 1) featuring video testimonials from Latinx families or including Spanish voice-overs/subtitles; 2) diversifying racial/ethnic/geographic representation and including extended families in photographs/illustrations; 3) adding information about health insurance coverage and circumcision, 4) reassuring parents that the condition is not their fault, 5) considering cultural values (e.g., reliance on expert advice), and 6) clarifying medical terminology. Feedback related to seven EVM dimensions: concepts, content, context, goals, language, metaphors, and methods. DISCUSSION: Participants perceived the Hub to be informative in guiding parents' treatment decisions. Revisions were reasonable and acceptable for a linguistic and cultural adaptation for LSS parents. CONCLUSIONS: We identified and implemented preliminary cultural modifications to the Hub and applied user-centered design methods to test and revise the website. The product is a culturally appropriate DA for LSS parents. Next, English and Spanish-versions of the Hub will be tested in a randomized controlled trial.

Mapping of DDX11 genetic interactions defines sister chromatid cohesion as the major dependency.

DDX11/Chl1R is a conserved DNA helicase with roles in genome maintenance, DNA replication, and chromatid cohesion. Loss of DDX11 in humans leads to the rare cohesinopathy Warsaw breakage syndrome. DDX11 has also been implicated in human cancer where it has been proposed to have an oncogenic role and possibly to constitute a therapeutic target. Given the multiple roles of DDX11 in genome stability and its potential as an anticancer target, we set out to define a complete genetic interaction profile of DDX11 loss in human cell lines. Screening the human genome with clustered regularly interspaced short palindromic repeats (CRISPR) guide RNA drop out screens in DDX11-wildtype (WT) or DDX11-deficient cells revealed a strong enrichment of genes with functions related to sister chromatid cohesion. We confirm synthetic lethal relationships between DDX11 and the tumor suppressor cohesin subunit STAG2, which is frequently mutated in several cancer types and the kinase HASPIN. This screen highlights the importance of cohesion in cells lacking DDX11 and suggests DDX11 may be a therapeutic target for tumors with mutations in STAG2.

A scalable platform for efficient CRISPR-Cas9 chemical-genetic screens of DNA damage-inducing compounds.

Current approaches to define chemical-genetic interactions (CGIs) in human cell lines are resource-intensive. We designed a scalable chemical-genetic screening platform by generating a DNA damage response (DDR)-focused custom sgRNA library targeting 1011 genes with 3033 sgRNAs. We performed five proof-of-principle compound screens and found that the compounds' known modes-of-action (MoA) were enriched among the compounds' CGIs. These scalable screens recapitulated expected CGIs at a comparable signal-to-noise ratio (SNR) relative to genome-wide screens. Furthermore, time-resolved CGIs, captured by sequencing screens at various time points, suggested an unexpected, late interstrand-crosslinking (ICL) repair pathway response to camptothecin-induced DNA damage. Our approach can facilitate screening compounds at scale with 20-fold fewer resources than commonly used genome-wide libraries and produce biologically informative CGI profiles.

Tracking activity-based therapy for people living with spinal cord injury or disease: insights gained through focus group interviews with key stakeholders.

PURPOSE: The development of a tool to track participation in activity-based therapy (ABT) for people with spinal cord injury or disease (SCI/D) was identified as a priority of the Canadian ABT Community of Practice. The objective of this study was to understand multi-stakeholder perspectives on tracking ABT participation across the continuum of care. MATERIALS AND METHODS: Forty-eight individuals from six stakeholder groups (persons living with SCI/D; hospital therapists; community trainers; administrators; researchers; and funders, advocates and policy experts) were recruited to participate in focus group interviews. Participants were asked open-ended questions concerning the importance of and parameters around tracking ABT. Transcripts were analyzed using conventional content analysis. RESULTS: Themes reflected the Who, What, Where, When, Why and How of tracking ABT. Participants described the importance of involving hospital therapists, community trainers and individuals with SCI/D in tracking ABT to capture both subjective and objective parameters across the continuum of care and injury trajectory. Digital tracking tools were favoured, although paper-based versions were regarded as a necessity in some circumstances. CONCLUSIONS: Findings highlighted the importance of tracking ABT participation for individuals with SCI/D. The information may guide the development of ABT practice guidelines and support the implementation of ABT in Canada.

Tracking the details of activity-based therapy (ABT) sessions and programs across the continuum of care and injury trajectory may provide important information to support the development of ABT practice guidelines and implementation strategies.Tracking objective and subjective parameters are needed to provide a comprehensive description of an ABT session and program.Clinicians and individuals with spinal cord injury or disease (SCI/D) should both be able to track ABT to accommodate all settings and types of data.Digital tracking tools, such as an app, may provide an accessible, versatile and efficient way of tracking ABT.

Novel Methionine Aminopeptidase 2 Inhibitor M8891 Synergizes with VEGF Receptor Inhibitors to Inhibit Tumor Growth of Renal Cell Carcinoma Models.

N-terminal processing by methionine aminopeptidases (MetAP) is a crucial step in the maturation of proteins during protein biosynthesis. Small-molecule inhibitors of MetAP2 have antiangiogenic and antitumoral activity. Herein, we characterize the structurally novel MetAP2 inhibitor M8891. M8891 is a potent, selective, reversible small-molecule inhibitor blocking the growth of human endothelial cells and differentially inhibiting cancer cell growth. A CRISPR genome-wide screen identified the tumor suppressor p53 and MetAP1/MetAP2 as determinants of resistance and sensitivity to pharmacologic MetAP2 inhibition. A newly identified substrate of MetAP2, translation elongation factor 1-alpha-1 (EF1a-1), served as a pharmacodynamic biomarker to follow target inhibition in cell and mouse studies. Robust angiogenesis and tumor growth inhibition was observed with M8891 monotherapy. In combination with VEGF receptor inhibitors, tumor stasis and regression occurred in patient-derived xenograft renal cell carcinoma models, particularly those that were p53 wild-type, had Von Hippel-Landau gene (VHL) loss-of-function mutations, and a mid/high MetAP1/2 expression score.

Topoisomerase 1 Inhibition in MYC-Driven Cancer Promotes Aberrant R-Loop Accumulation to Induce Synthetic Lethality.

MYC is a central regulator of gene transcription and is frequently dysregulated in human cancers. As targeting MYC directly is challenging, an alternative strategy is to identify specific proteins or processes required for MYC to function as a potent cancer driver that can be targeted to result in synthetic lethality. To identify potential targets in MYC-driven cancers, we performed a genome-wide CRISPR knockout screen using an isogenic pair of breast cancer cell lines in which MYC dysregulation is the switch from benign to transformed tumor growth. Proteins that regulate R-loops were identified as a potential class of synthetic lethal targets. Dysregulated MYC elevated global transcription and coincident R-loop accumulation. Topoisomerase 1 (TOP1), a regulator of R-loops by DNA topology, was validated to be a vulnerability in cells with high MYC activity. Genetic knockdown of TOP1 in MYC-transformed cells resulted in reduced colony formation compared with control cells, demonstrating synthetic lethality. Overexpression of RNaseH1, a riboendonuclease that specifically degrades R-loops, rescued the reduction in clonogenicity induced by TOP1 deficiency, demonstrating that this vulnerability is driven by aberrant R-loop accumulation. Genetic and pharmacologic TOP1 inhibition selectively reduced the fitness of MYC-transformed tumors in vivo. Finally, drug response to TOP1 inhibitors (i.e., topotecan) significantly correlated with MYC levels and activity across panels of breast cancer cell lines and patient-derived organoids. Together, these results highlight TOP1 as a promising target for MYC-driven cancers. SIGNIFICANCE: CRISPR screening reveals topoisomerase 1 as an immediately actionable vulnerability in cancers harboring MYC as a driver oncoprotein that can be targeted with clinically approved inhibitors.

N-terminal acetylation shields proteins from degradation and promotes age-dependent motility and longevity.

Most eukaryotic proteins are N-terminally acetylated, but the functional impact on a global scale has remained obscure. Using genome-wide CRISPR knockout screens in human cells, we reveal a strong genetic dependency between a major N-terminal acetyltransferase and specific ubiquitin ligases. Biochemical analyses uncover that both the ubiquitin ligase complex UBR4-KCMF1 and the acetyltransferase NatC recognize proteins bearing an unacetylated N-terminal methionine followed by a hydrophobic residue. NatC KO-induced protein degradation and phenotypes are reversed by UBR knockdown, demonstrating the central cellular role of this interplay. We reveal that loss of Drosophila NatC is associated with male sterility, reduced longevity, and age-dependent loss of motility due to developmental muscle defects. Remarkably, muscle-specific overexpression of UbcE2M, one of the proteins targeted for NatC KO-mediated degradation, suppresses defects of NatC deletion. In conclusion, NatC-mediated N-terminal acetylation acts as a protective mechanism against protein degradation, which is relevant for increased longevity and motility.

The experience of falls and fall risk during the subacute phase of spinal cord injury: a mixed methods study.

Purpose: To understand the circumstances, causes and consequences of falls experienced by individuals with subacute SCI, and to explore their perspectives on how falls/fall risk impacted their transition to community living.Materials and methods: Sixty adults with subacute SCI participated. A sequential explanatory mixed methods design was adopted. In Phase I, falls were monitored for six months post-inpatient rehabilitation discharge through a survey. In Phase II, a qualitative focus group (n = 5) was held to discuss participants' perspectives on Phase I results and falls/fall risk. Descriptive statistics and thematic analysis were used to analyze Phase I and II data, respectively.Results: Falls commonly occurred in the daytime, at home and about half resulted in minor injury. Three themes reflecting participants' perspectives were identified in Phase II. 1) Lack of preparedness to manage fall risk upon returning home from inpatient rehabilitation. 2) Adjusting to increased fall risk following discharge from inpatient rehabilitation. 3) Psychological impact of the transition to living at home with an increased fall risk.Conclusions: The findings highlight the need for fall prevention initiatives during subacute SCI, when individuals are learning to manage their increased fall risk.

Falls are common in the subacute phase of spinal cord injury (SCI), with falls commonly occurring in the daytime at home while walking or changing positions and resulting in minor injury.Individuals living with SCI feel unprepared to manage the increased fall risk experienced after discharge from inpatient rehabilitation, and the possibility of falling can cause anxiety and fear.Following the transition from inpatient rehabilitation to living at home, individuals with SCI would like continued support from health professionals and/or peers to prevent falls and adjust to living independently with SCI.

Quality of reporting for qualitative studies in pediatric urology-A scoping review.

INTRODUCTION: Qualitative research has gained popularity in pediatric urology due to rich data and insights about quantitative results. To date, there has been no study evaluating the comprehensiveness of the reporting of these studies based on established guidelines. OBJECTIVE: The objective of this study is to perform a scoping review of the quality of reporting in recent qualitative studies in pediatric urology based on a predominant checklist, the 21-item Standards of Reporting Qualitative Research (SRQR) and identify areas for improvement. STUDY DESIGN: In accordance with the Preferred Reported Items for Systematic Reviews and Meta-Analyses extension for Scoping Reviews, we performed a systematic literature search to identify qualitative studies on pediatric urology topics published from 2015 to 2021. We used clustering technology to eliminate articles with unrelated keywords. Articles not in English and those published prior to 2015 were excluded. Two reviewers performed title/abstract screening and full text review and resolved discrepancies by consensus. We reported the median and interquartile range of total SRQR scores (maximum: 21). SRQR-reported items were summarized; overall proportion of reported items for each article was estimated. Bivariate analyses examined the association between study characteristics and SRQR tertile. Simple linear regression was performed to examine the relationship between year and SRQR score. RESULTS: Of the 2562 titles/abstracts screened, 26 studies were included. The most common topics were hypospadias and congenital adrenal hyperplasia (Summary Figure). The median total score was 18.0 of 21 possible items (interquartile range: 3). All studies reported an abstract, problem formulation, purpose/context of the study, data collection methods, integration with prior work, limitations, and ethics review board approval. Most (25/26; 96.2%) reported sampling strategy, data analysis, synthesis/interpretation of findings and links to empirical data. Less fulfilled items included: a title identifying the study as qualitative (11/26, 42.3%), qualitative approach & research paradigm (11/26, 42.3%) and researcher characteristics & reflexivity (9/26, 34.6%). There was no association between study characteristics and SRQR score. There was a statistically significant increase in the SRQR score during the study period (ß = 1.0, p < 0.0001). DISCUSSION: Studies fulfilled most SRQR checklist items. There was significant improvement in quality during the study period. Limitations include possible recency bias and exclusion of articles due to inconsistent categorizations in Pub Med. CONCLUSION: The quality and trajectory of qualitative study reporting in pediatric urology is encouraging. SRQR standards should be implemented by journals to continue improving the robustness and transparency of future qualitative manuscripts in pediatric urology.

Identifying priorities for balance interventions through a participatory co-design approach with end-users.

BACKGROUND: Most individuals living with spinal cord injuries/diseases (SCI/D) or stroke experience at least one fall each year; hence, the development of interventions and technologies that target balance control is needed. The purpose of this study was to identify and explore the priorities for balance-focused interventions and technologies from the perspectives of end-users to assist with the design of an intervention that combines functional electrical stimulation (FES) with visual feedback training for standing balance. METHODS: Two individuals with SCI/D, one individual with stroke, two physical therapists (PT) and one hospital administrator were recruited. Participants attended three focus group meetings that followed a participatory co-design approach. A semi-structured interview guide, developed from the FAME (Feasibility, Appropriateness, Meaningfulness, Effectiveness, Economic Evidence) framework, was used to lead the discussion, querying participants' experiences with balance deficits and interventions, and FES. Meetings were audio-recorded and transcribed verbatim. An iterative and reflexive inductive thematic analysis was applied to the transcripts by three researchers. RESULTS: Four themes were identified: (1) Balance is meaningful for daily life and rehabilitation. Participants acknowledged various factors influencing balance control and how balance deficits interfered with participation in activities. End-users stressed the importance of continuing to work on one's balance after discharge from hospital-based rehabilitation. (2) Desired characteristics of balance interventions. Participants explained that balance interventions should be tailored to an individual's unique needs and goals, relevant to their lives, balance their safety and risk, and be engaging. (3) Prior experiences with FES to inform future therapeutic use. Participants with stroke or SCI/D described initial apprehension with FES, but experienced numerous benefits that motivated them to continue with FES. Challenges with FES were mentioned, including wires, cost, and time of set up. (4) Potential role of FES in balance interventions. Participants felt that FES would complement balance interventions; however, they had not experienced this combination of therapies previously. CONCLUSIONS: End-users described how their experiences with balance deficits, rehabilitation, and FES informed their priorities for balance interventions. The findings inform the design and implementation of future balance interventions for individuals with SCI/D or stroke, including an intervention involving FES and visual feedback training.

Interrater reliability of the Standing and Walking Assessment Tool for spinal cord injury.

STUDY DESIGN: Psychometric study. OBJECTIVES: The Standing and Walking Assessment Tool (SWAT) is a standardized approach to the evaluation of standing and walking capacity following traumatic spinal cord injury (tSCI) in Canada. The SWAT classifies individuals with a tSCI into 12 stages of standing and walking capacity that are paired with well-established outcome measures, such as the Berg Balance Scale and 10-m Walk Test. Prior research has demonstrated the validity and responsiveness of the SWAT stages; however, the reliability remains unknown. The objective of this study was to evaluate the interrater reliability of the SWAT stages. SETTING: Inpatient units of two Canadian rehabilitation hospitals. METHODS: Adults with sub-acute tSCI were recruited. SWAT stage was evaluated for each participant by two physical therapists separately. The two therapists aimed to complete the evaluations within one day of each other. To evaluate interrater reliability, the percentage agreement between the SWAT stages rated by the two physical therapists was calculated, along with a linear weighted kappa statistic with a 95% CI. RESULTS: Forty-five individuals with sub-acute tSCI (36 males, 9 females, mean (SD) age of 54.8 (17.9) years) participated. The percentage agreement in SWAT stages between the two physical therapists was 75.6%. A kappa statistic of 0.93 with a 95% CI, 0.81-1.05 was obtained. In cases where therapists disagreed (18% of participants), therapists differed by 1-2 stages only. CONCLUSIONS: The SWAT stages have high interrater reliability, providing further support for the use of the SWAT in rehabilitation practice in Canada.

Quality over quantity: How qualitative research informs and improves quantitative research.

We highlight five recent, high-quality, representative, qualitative articles about Spina Bifida and DSD care that contextualize their findings within the scope of a larger envisioned clinical project. Qualitative research is uniquely poised to address issues such as how to define treatment success and optimize fertility-related education and surgical experiences in DSD care. This approach is also well-suited to explore caregiver burden and financial toxicity in spina bifida care and identify areas for improvement. We encourage researchers to add a qualitative approach to their quantitative research to provide a more holistic, patient-centered view of their subject matter.

Spatial characteristics of reactive stepping among people living with chronic incomplete spinal cord injury.

Objective: Compare the spatial characteristics of reactive stepping between individuals with chronic motor incomplete spinal cord injuries (iSCI) and able-bodied (AB) individuals.Design: Cross sectional.Setting: Lyndhurst Centre.Participants: Twelve individuals with iSCI (3 males, 53.6 ± 15.2 years old) and 11 age- and sex-matched AB individuals (3 males, 54.8 ± 14.0 years old).Interventions: The Lean-and-Release test was used to elicit reactive stepping. A horizontal cable, attached at waist height, was released when 8-12% body weight was supported in a forward lean position. Participants underwent up to 10 Lean-and-Release trials in a session. Kinematic and kinetic data were recorded.Outcome measures: The length, width and height of the first reactive step of each trial were calculated. Standard deviation between trials was calculated to represent the variability in step length, width and height within a participant. Among participants with iSCI, correlation coefficients were used to explore the relationship between step length and width variability and (1) Lean-and-Release test behavioral responses, (2) 3-month fall history, and (3) lower extremity strength.Results: Step length (P = 0.94), width (P = 0.52) and height (P = 0.97), normalized for participant height, did not differ between groups. Participants with iSCI showed greater variability in step length (P = 0.02) and width (P = 0.01), but not height (P = 0.32). No correlation was found between step length or width variability and behavioral responses, 3-month fall history, or lower extremity strength.Conclusions: Individuals with iSCI showed increased variability in length and width of reactive stepping compared to AB individuals, which may contribute to their impaired ability to execute single-step reactive responses.Trial registration: ClinicalTrials.gov identifier: NCT02960178.

Stepping responses for reactive balance for individuals with incomplete spinal cord injury.

Incomplete spinal cord injury (iSCI) causes impairment of reactive balance control, leading to higher fall risk. In our previous work, we found that individuals with iSCI were more likely to exhibit multiple-step response during the lean-and-release (LR) test, where the participant leaned forward while a tether supported 8-12% of the body weight and received a sudden release, inducing reactive steps. Here we investigated the foot placement of people with iSCI during the LR test using margin-of-stability (MOS). Twenty-one individuals with iSCI (age: 56.1 ± 16.1 years old; mass: 72.5 ± 19.0 kg; height: 166 ± 12 cm), and fifteen age- and sex-matched able-bodied (AB) individuals (age: 56.1 ± 12.9 years old; mass: 57.4 ± 10.9 kg; height: 164 ± 8 cm) participated in the study. The participants performed ten trials of the LR test and also completed clinical assessment of balance and strengths, including the Mini-Balance Evaluations Systems Test, the Community Balance and Mobility Scale, gait speed, and lower extremity manual muscle testing. MOS was significantly smaller during multiple-step responses than during single-step responses for both individuals with iSCI and AB counterparts. Using binary logistic regression and receiver operating characteristic analyses, we demonstrated that MOS can distinguish single- and multiple-step responses. In addition, individuals with iSCI demonstrated significantly larger intra-subject variability of MOS compared to AB individuals at first foot contact. Further, we found that MOS correlated with clinical measures of balance including one for reactive balance. We conclude that individuals with iSCI were less likely to demonstrate foot placement with sufficiently large MOS, which may increase the tendency to exhibit multiple-step responses.

A multi-site pilot study of a parent-centered tool to promote shared decision-making in hypospadias care.

BACKGROUND: Using a user-centered design approach, we conducted a two-site pilot study to evaluate a decision aid (DA) website, the Hypospadias Hub, for parents of hypospadias patients. OBJECTIVES: The objectives were to assess the Hub's acceptability, remote usability, and feasibility of study procedures, and to evaluate its preliminary efficacy. METHODS: From June 2021-February 2022, we recruited English-speaking parents (≥18 years old) of hypospadias patients (≤5 years) and delivered the Hub electronically ≤2 months before their hypospadias consultation. We collected website analytic data using an ad tracker plug-in. We inquired about treatment preference, hypospadias knowledge, and decisional conflict (Decisional Conflict Scale) at baseline, after viewing the Hub (pre-consultation), and post-consultation. We administered the Decision Aid Acceptability Questionnaire (DAAQ) and the Preparation for Decision-Making Scale (PrepDM) which assessed how well the Hub prepared parents for decision-making with the urologist. Post-consultation, we assessed participants' perception of involvement in decision-making with the Shared Decision-making Questionnaire (SDM-Q-9) and the Decision Regret Scale (DRS). A bivariate analysis compared participants' baseline and pre/post-consultation hypospadias knowledge, decisional conflict, and treatment preference. Using a thematic analysis, we analyzed our semi-structured interviews to uncover how the Hub impacted the consultation and what influenced participants' decisions. RESULTS: Of 148 parents contacted, 134 were eligible and 65/134 (48.5%) enrolled: mean age 29.2, 96.9% female, 76.6% White (Extended Summary Figure). Pre/post-viewing the Hub, there was a statistically significant increase in hypospadias knowledge (54.3 vs. 75.6, p < 0.001) and decrease in decisional conflict (36.0 vs. 21.9, p < 0.001). Most participants (83.3%) thought Hub's length and amount of information (70.4%) was "about right", and 93.0% found most or everything was clear. Pre/post-consultation, there was a statistically significant decrease in decisional conflict (21.9 vs. 8.8, p < 0.001). PrepDM's mean score was 82.6/100 (SD = 14.1); SDM-Q-9's mean score was 82.5/100 (SD = 16.7). DCS's mean score was 25.0/100 (SD = 47.03). Each participant spent an average of 25.75 min reviewing the Hub. Based on thematic analysis, the Hub helped participants feel prepared for the consultation. DISCUSSION: Participants engaged extensively with the Hub and demonstrated improved hypospadias knowledge and decision quality. They felt prepared for the consultation and perceived a high degree of involvement in decision-making. CONCLUSION: As the first pilot test of a pediatric urology DA, the Hub was acceptable and study procedures were feasible. We plan to conduct a randomized controlled trial of the Hub versus usual care to test its efficacy to improve the quality of shared decision-making and reduce long-term decisional regret.

Survival-based CRISPR genetic screens across a panel of permissive cell lines identify common and cell-specific SARS-CoV-2 host factors.

SARS-CoV-2 depends on host cell components for infection and replication. Identification of virus-host dependencies offers an effective way to elucidate mechanisms involved in viral infection and replication. If druggable, host factor dependencies may present an attractive strategy for anti-viral therapy. In this study, we performed genome wide CRISPR knockout screens in Vero E6 cells and four human cell lines including Calu-3, UM-UC-4, HEK-293 and HuH-7 to identify genetic regulators of SARS-CoV-2 infection. Our findings identified only ACE2, the cognate SARS-CoV-2 entry receptor, as a common host dependency factor across all cell lines, while other host genes identified were largely cell line specific, including known factors TMPRSS2 and CTSL. Several of the discovered host-dependency factors converged on pathways involved in cell signalling, immune-related pathways, and chromatin modification. Notably, the chromatin modifier gene KMT2C in Calu-3 cells had the strongest impact in preventing SARS-CoV-2 infection when perturbed.

Microfluidically-generated Encapsulated Spheroids (µ-GELS): An All-Aqueous Droplet Microfluidics Platform for Multicellular Spheroids Generation.

Spheroids are three-dimensional clusters of cells that serve as in vitro tumor models to recapitulate in vivo morphology. A limitation of many existing on-chip platforms for spheroid formation is the use of cytotoxic organic solvents as the continuous phase in droplet generation processes. All-aqueous methods do not contain cytotoxic organic solvents but have so far been unable to achieve complete hydrogel gelation on chip. Here, we describe an enhanced droplet microfluidic platform that achieves on-chip gelation of all-aqueous hydrogel multicellular spheroids (MCSs). Specifically, we generate dextran-alginate droplets containing MCF-7 breast cancer cells, surrounded by polyethylene glycol, at a flow-focusing junction. Droplets then travel to a second flow-focusing junction where they interact with calcium chloride and gel on chip to form hydrogel MCSs. On-chip gelation of the MCSs is possible here because of an embedded capillary at the second junction that delays the droplet gelation, which prevents channel clogging problems that would otherwise exist. In drug-free experiments, we demonstrate that MCSs remain viable for 6 days. We also confirm the applicability of this system for cancer drug testing by observing that dose-dependent cell death is achievable using doxorubicin.