Membraneless organelles (MLOs) formed through liquid-liquid phase separation (LLPS) are becoming increasingly relevant to understanding viral-host interactions, neurodegenerative disease, and cancer. ...The modulation of LLPS involves many parameters and components. To describe these modulators, typical
in vitro
studies require laborious, manual sample preparation of different concentrations and costly biological reagents. Here, we introduce a minimal reagent, microfluidic platform to systematically generate samples of different concentrations and trigger phase separation. We demonstrate the platform's utility by constructing phase diagrams describing the modulation of LLPS using an aqueous two-phase system (ATPS) and an MLO-based phase separating system. We also show on-chip biophysical characterization typical of
in vitro
studies. We expect that this platform will be utilized by scientists to study the growing number of MLOs and inform clinical treatments for pathology related to LLPS.
A microfluidic platform is used to generate phase diagrams for formation of biomolecular condensates without conventional manual sample preparation methods.
Human pluripotent stem cells (hPSCs) provide an invaluable tool for modeling diseases and hold promise for regenerative medicine. For understanding pluripotency and lineage differentiation ...mechanisms, a critical first step involves systematically cataloging essential genes (EGs) that are indispensable for hPSC fitness, defined as cell reproduction in this study. To map essential genetic determinants of hPSC fitness, we performed genome-scale loss-of-function screens in an inducible Cas9 H1 hPSC line cultured on feeder cells and laminin to identify EGs. Among these, we found FOXH1 and VENTX, genes that encode transcription factors previously implicated in stem cell biology, as well as an uncharacterized gene, C22orf43/DRICH1. hPSC EGs are substantially different from other human model cell lines, and EGs in hPSCs are highly context dependent with respect to different growth substrates. Our CRISPR screens establish parameters for genome-wide screens in hPSCs, which will facilitate the characterization of unappreciated genetic regulators of hPSC biology.
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•A robust and flexible platform for genetic screens in hPSCs•Essential gene profiles for hPSCs on two substrates and core hPSC fitness gene sets•Previously unidentified regulators of hPSC fitness (DRICH1 and others)•Confirmation of the TP53 pathway, PAWR, PMAIP1 as negative proliferation regulators
Mair et al. establish a robust, inducible CRISPR screening platform for forward genetics in human pluripotent stem cells (hPSCs). Genome-wide proliferation screens identified core essential genes for hPSCs and revealed context-dependent genetic requirements on different substrates. This underlines hPSC plasticity and helps us to understand the genetic wiring of hPSCs.
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.
Human pluripotent stem cells (hPSCs) have the capacity for self-renewal and differentiation into most cell types and, in contrast to widely used cell lines, are karyotypically normal and ...non-transformed. Hence, hPSCs are considered the gold-standard system for modelling diseases, especially in the field of regenerative medicine. Despite widespread research use of hPSCs and induced pluripotent stem cells (iPSCs), the systematic understanding of pluripotency and lineage differentiation mechanisms are still incomplete. Before tackling the complexities of lineage differentiation with genetic screens, it is critical to catalogue the general genetic requirements for cell fitness and proliferation in the pluripotent state and assess their plasticity under commonly used culture conditions.We describe a method to map essential genetic determinants of hPSC fitness and pluripotency, herein defined as cell reproduction, by genome-scale loss-of-function CRISPR screens in an inducible S. pyogenes Cas9 H1 hPSC line. To address questions of context-dependent gene essentiality, we include protocols for screening hPSCs cultured on feeder cells and laminin, two commonly used growth substrates. This method establishes parameters for genome-wide screens in hPSCs, making human stem cells amenable for functional genomics approaches to facilitate investigation of hPSC biology.
Governments restricted mobility and effectively shuttered much of the global economy in response to the COVID‐19 pandemic. Six San Francisco Bay Area counties were the first region in the United ...States to issue a “shelter‐in‐place” order asking non‐essential workers to stay home. Here we use CO2 observations from 35 Berkeley Environment, Air‐quality and CO2 Network (BEACO2N) nodes and an atmospheric transport model to quantify changes in urban CO2 emissions due to the order. We infer hourly emissions at 900‐m spatial resolution for 6 weeks before and 6 weeks during the order. We observe a 30% decrease in anthropogenic CO2 emissions during the order and show that this decrease is primarily due to changes in traffic (–48%) with pronounced changes to daily and weekly cycles; non‐traffic emissions show small changes (–8%). These findings provide a glimpse into a future with reduced CO2 emissions through electrification of vehicles.
Plain Language Summary
This work uses atmospheric observations to quantify the changes in urban CO2 emissions from different sectors in response to COVID‐19 mobility regulations.
Key Points
A 30% decrease in urban CO2 emissions was observed from the San Francisco Bay Area in response to COVID‐19 mobility restrictions
Changes are primarily driven by a decrease in CO2 emissions from traffic (–48%)
There is a large change to the weekly and diurnal cycle of emissions with reductions in morning rush hour emissions
The adoption of docetaxel for systemic treatment of metastatic prostate cancer (PCa), in both castration-sensitive (mCSPC) and castration-resistant (mCRPC) settings, is poorly understood. This study ...examined the real-world utilization of docetaxel in these patients and their outcomes.
A retrospective population-based study used administrative data from Ontario, Canada, to identify men aged ≥66 years who were diagnosed with de novo mCSPC or mCRPC between 2014 and 2019 and received docetaxel. The study assessed treatment tolerability and toxicity, and survival in both cohorts. Descriptive and comparative statistical analysis were conducted.
The study identified 11.2% (399/3556) and 13.2% (203/1534) patients diagnosed with de novo mCSPC and with mCRPC who received docetaxel respectively. The median age in both cohorts was 72 years (IQR: 68-76). Overall, 43.9% (n = 175) patients with de novo mCSPC and 52.1% (n = 85) with mCRPC completed ≥6 cycles of docetaxel. Over two-fifth also needed dose adjustments in both cohorts. Hospitalization or emergency department visit for febrile neutropenia were noted in 15.8% (n = 63) of de novo mCSPC patients and similarly in 19% (n = 31) of mCRPC cohort. The median survival of PCa patients who completed ≥6 cycles of docetaxel was significantly longer relative to those who completed <4 cycles: 32.7 vs. 23.5 months (p < 0.001) for mCSPC and 20.5 vs. 10.7 (p = 0.012) for mCRPC respectively.
In this population-based study of elderly patients with metastatic PCa, treatment with docetaxel was associated with poor tolerability and higher toxicity compared with clinical trials. Receipt of limited cycles and reduced overall dose of docetaxel were associated with inferior overall survival.
Droplet microfluidics is utilized in a wide range of applications in biomedicine and biology. Applications include rapid biochemical analysis, materials generation, biochemical assays, and ...point-of-care medicine. The integration of aqueous two-phase systems (ATPSs) into droplet microfluidic platforms has potential utility in oil-free biological and biomedical applications, namely, reducing cytotoxicity and preserving the native form and function of costly biomolecular reagents. In this review, we present a design manual for the chemist, biologist, and engineer to design experiments in the context of their biological applications using all-in-water droplet microfluidic systems. We describe the studies achievable using these systems and the corresponding fabrication and stabilization methods. With this information, readers may apply the fundamental principles and recent advancements in ATPS droplet microfluidics to their research. Finally, we propose a development roadmap of opportunities to utilize ATPS droplet microfluidics in applications that remain underexplored.
Despite the wealth of evidence demonstrating the efficacy of treatment intensification beyond androgen-deprivation therapy (ADT) among patients with de novo metastatic castration-sensitive prostate ...cancer (mCSPC), little is known of its real-world use. This study examined the real-world uptake of ADT treatment intensification among older men in a large Canadian province.
We performed a retrospective population-based cohort study using province-wide linked administrative data in Ontario, Canada. Patients 66 years of age and older with de novo mCSPC were included and their treatment with conventional ADT-based regimens, ADT plus next-generation androgen receptor axis-targeted therapy, and ADT plus docetaxel were identified and stratified by time.
From 2014 to 2019, 3556 patients were identified with de novo mCSPC. Most patients (n = 2794 78.6%) were treated with a conventional ADT regimen, whereas 399 (11.2%) patients received ADT intensification with docetaxel and 52 (1.5%) patients received abiraterone acetate plus prednisone. In a time-stratified analysis of ADT intensification before and after the pivotal AA+P trial (LATITUDE), AA+P uptake increased from 0.5% to 3.0%, whereas docetaxel use dropped from 12.0% to 10.0%. The median survival of the study population was 18 months (interquartile range = 10-31).
The majority of patients with de novo mCSPC are treated with ADT alone in the Canadian real-world setting, despite randomized clinical trial evidence of benefit with the use of ADT-intensified regimens. As ADT treatment intensification is substantially underused, better understanding of the barriers to treatment and targeted education to address them are needed.
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.
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.
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.
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 (β = 1.0, p < 0.0001).
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.
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.
•Single line of treatment for men with metastatic castration-resistant prostate cancer.•Most patients received oral androgen-receptor axis-targeted therapies.•Real-world outcomes in Ontario, Canada, ...appear poorer than in clinical trials.
Management of advanced prostate cancer has evolved rapidly with the availability of multiple systemic treatments such as androgen-receptor axis-targeted therapies (ARATs), taxane-based chemotherapy, radium-223, and other approaches. However, limited data exists on real-world treatment selection and clinical outcomes. This study examines the utilization and survival impact of these therapies in men with metastatic castration-resistant prostate cancer (mCRPC) in the real-world setting of Ontario, Canada.
This study was a retrospective, longitudinal, population-based study of administrative claims data between January 2016 and April 2020. Men ≥ 66 years with mCRPC receiving advanced treatment were included. Patients were indexed on the day they initiated mCRPC treatment and followed up until death or end of study period to assess treatment and survival. Multinomial regression was used to model the association between baseline covariates, treatment and survival.
Median age was 75 years among the 944 mCRPC patients who received life-prolonging therapies during this time period. Over 90% of patients used an ARAT as a first-line therapy, and 71.5% received only first-line therapy before death or censoring. Of patients that received two or more lines, over 80% received subsequent therapy with a different mechanism of action. Median overall survival was 18.9 months.
ARATs have become the predominant first-line systemic treatment option for mCRPC patients in recent years. Notably, the majority of patients received only a single line of life-prolonging therapy after developing mCRPC. In keeping with the recognized efficacy-effectiveness gap, real-world outcomes in this cohort appear poorer than in clinical trials.
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