Droplet barcoding for single-cell transcriptomics applied to embryonic stem cells.

It has long been the dream of biologists to map gene expression at the single-cell level. With such data one might track heterogeneous cell sub-populations, and infer regulatory relationships between genes and pathways. Recently, RNA sequencing has achieved single-cell resolution. What is limiting is an effective way to routinely isolate and process large numbers of individual cells for quantitative in-depth sequencing. We have developed a high-throughput droplet-microfluidic approach for barcoding the RNA from thousands of individual cells for subsequent analysis by next-generation sequencing. The method shows a surprisingly low noise profile and is readily adaptable to other sequencing-based assays. We analyzed mouse embryonic stem cells, revealing in detail the population structure and the heterogeneous onset of differentiation after leukemia inhibitory factor (LIF) withdrawal. The reproducibility of these high-throughput single-cell data allowed us to deconstruct cell populations and infer gene expression relationships. VIDEO ABSTRACT.

A Randomized Evaluation of MoodFX, a Patient-Centred e-Health Tool to Support Outcome Measurement for Depression: Une évaluation randomisée de MoodFX, un outil de santé en ligne centré sur le patient pour soutenir la mesure du résultat dans la dépression.

BACKGROUND: e-Health tools using validated questionnaires to assess outcomes may facilitate measurement-based care for psychiatric disorders. MoodFX was created as a free online symptom tracker to support patients for outcome measurement in their depression treatment. We conducted a pilot randomized evaluation to examine its usability, and clinical utility. METHODS: Patients presenting with a major depressive episode (within a major depressive or bipolar disorder) were randomly assigned to receive either MoodFX or a health information website as the intervention and control condition, respectively, with follow-up assessment surveys conducted online at baseline, 8 weeks and 6 months. The primary usability outcomes included the percentage of patients with self-reported use of MoodFX 3 or more times during follow up (indicating minimally adequate usage) and usability measures based on the System Usability Scale (SUS). Secondary clinical outcomes included the Quick Inventory of Depressive Symptomatology, Self-Rated (QIDS-SR) and Patient Health Questionnaire (PHQ-9). RESULTS: Forty-nine participants were randomized (24 to MoodFX and 25 to the control condition). Of the 23 participants randomized to MoodFX who completed the user survey, 18 (78%) used MoodFX 3 or more times over the 6 months of the study. The mean SUS score of 72.7 (65th-69th percentile) represents good usability. Compared to the control group, the MoodFX group had significantly better improvement on QIDS-SR and PHQ-9 scores, with large effect sizes and higher response rates at 6 months. There were no differences between conditions on other secondary outcomes such as functioning and quality of life. CONCLUSION: MoodFX demonstrated good usability and was associated with reduction in depressive symptoms. This pilot study supports the use of digital tools in depression treatment.

E-health tools may be useful for measuring and tracking symptoms and other outcomes during treatment for depression. This study is a randomized evaluation of MoodFX, a free web-based app that helps patients track their symptoms using validated questionnaires, and also offers depression information and self-management tips. A total of 49 participants with clinical depression were randomized to using MoodFX or a health information website, for 6 months. In a survey, the participants that used MoodFX found it easy and useful to use. In addition, the participants that used MoodFX had greater improvement in depressive symptoms after 6 months, compared to those who used the health information website. These results suggest that MoodFX may be a useful tool to monitor outcomes and support depression treatment.

Cutting-edge mass spectrometry strategy based on imaged capillary isoelectric focusing (icIEF) technology for characterizing charge heterogeneity of monoclonal antibody.

Imaging capillary isoelectric focusing (icIEF) technology has been becoming the gold criteria of monitoring monoclonal antibody (mAb) charge heterogeneity that is one of the major product-related variants in recombinant biopharmaceuticals, since the first commercial instrument developed twenty years ago. However, the protein identification in icIEF separation is just based on isoelectric point (pI) measurement of protein. Although high resolution mass spectrometry (HRMS) is currently the most powerful means of qualitative protein analysis, traditional icIEF cannot compatibly be used in conjunction with MS due to the use of less volatile reagents. In addition, protein heterogeneity characterization in depth such as peptide mapping by high performance liquid chromatography (HPLC) requires the focused protein bands to be collected as fractions after the icIEF separation, which is a great challenge in biopharmaceutical discovery. In this work, pembrolizumab was employed as targeting mAb (a highly selective anti-PD-1 humanized mAb), an integrated icIEF platform was developed including analytical profiling, MS coupling and fraction collections for charged variant preparation. Multiple operation modes can be rapidly and flexibly switched just by changing customized capillary separation cartridges without more configurations. Main component, four acidic variants (A1-A4) and three basic variants (B1-B3) were baseline separated then directly detected by icIEF-HRMS online coupling for rapid screening of intact protein heterogeneity where reliable and accurate molecular weight of protein charged variants were obtained. Next, by installing preparative capillary separation cartridge, fractions of major charge variants (A2-3 and B1-2) and main component were collected for following LC-MS peptide mapping characterization. The whole workflow of icIEF-based MS strategy for protein heterogeneity is straight forward, reliable and accurate, which provides a comprehensive and revolutionary technology for protein drug quality control (QC) monitoring, MS coupling for fingerprinting intact protein and HPLC-MS peptide mapping in depth.

Imaged capillary isoelectric focusing tandem high-resolution mass spectrometry using nano electrospray ionization (ESI) for protein heterogeneity characterization.

Recombinant monoclonal antibodies (mAbs) have been spurring the rapid growth of commercial biotherapeutics. During production their charge heterogeneity must be assessed as a critical quality attribute to ensure safety, efficacy, and potency. Although imaged capillary isoelectric focusing (icIEF) is a powerful tool for this process, it could be improved further with tandem high-resolution mass spectrometry (HRMS). In this work, a nano-electrospray ionization (nano-ESI) apparatus was constructed to directly couple icIEF to HRMS. The system was evaluated with the standard NISTmAb, as well as more complex mAb, bi-specific antibody, and fusion protein samples. NISTmAb concentrations as low as 0.25 mg/ml demonstrated excellent sensitivity. There were good repeatabilities at 1 mg/ml with 7.58% and 8.01% RSDs for intention time and MS intensity, respectively, and the HRMS signal showed a strong linearity (R = 0.9983) across different concentrations. Meanwhile, the fingerprinting of the complex samples illustrated the versatility and potential of icIEF-HRMS. icIEF-HRMS developed can provide a comprehensive understanding of the underlying structural modifications that impact protein charge heterogeneity. Compared to the traditional ESI, nano-ESI can significantly improve sensitivity while maintaining a reasonable repeatability and throughput. Furthermore, the interface is much easier to connect, and is compatible with many commercial HRMS instruments.

Oral microbiota signatures in post-traumatic stress disorder (PTSD) veterans.

Post-traumatic stress disorder (PTSD) represents a global public health concern, affecting about 1 in 20 individuals. The symptoms of PTSD include intrusiveness (involuntary nightmares or flashbacks), avoidance of traumatic memories, negative alterations in cognition and mood (such as negative beliefs about oneself or social detachment), increased arousal and reactivity with irritable reckless behavior, concentration problems, and sleep disturbances. PTSD is also highly comorbid with anxiety, depression, and substance abuse. To advance the field from subjective, self-reported psychological measurements to objective molecular biomarkers while considering environmental influences, we examined a unique cohort of Israeli veterans who participated in the 1982 Lebanon war. Non-invasive oral 16S RNA sequencing was correlated with psychological phenotyping. Thus, a microbiota signature (i.e., decreased levels of the bacteria sp_HMT_914, 332 and 871 and Noxia) was correlated with PTSD severity, as exemplified by intrusiveness, arousal, and reactivity, as well as additional psychopathological symptoms, including anxiety, hostility, memory difficulties, and idiopathic pain. In contrast, education duration correlated with significantly increased levels of sp_HMT_871 and decreased levels of Bacteroidetes and Firmicutes, and presented an inverted correlation with adverse psychopathological measures. Air pollution was positively correlated with PTSD symptoms, psychopathological symptoms, and microbiota composition. Arousal and reactivity symptoms were correlated with reductions in transaldolase, an enzyme controlling a major cellular energy pathway, that potentially accelerates aging. In conclusion, the newly discovered bacterial signature, whether an outcome or a consequence of PTSD, could allow for objective soldier deployment and stratification according to decreases in sp_HMT_914, 332, 871, and Noxia levels, coupled with increases in Bacteroidetes levels. These findings also raise the possibility of microbiota pathway-related non-intrusive treatments for PTSD.

Blue-Light Therapy for Seasonal and Non-Seasonal Depression: A Systematic Review and Meta-Analysis of Randomized Controlled Trials.

OBJECTIVES: To determine the efficacy and safety of blue-light therapy in seasonal and non-seasonal major depressive disorder (MDD), by comparison to active and inactive control conditions. METHODS: We searched Web of Science, EMBASE, Medline, PsycInfo, and Clinicaltrials.gov through January 17, 2022, for randomized controlled trials (RCTs) using search terms for blue/blue-enhanced, light therapy, and depression/seasonal affective disorder. Two independent reviewers extracted data. The primary outcome was the difference in endpoint scores on the Structured Interview Guide for the Hamilton Depression Rating Scale - Seasonal Affective Disorder (SIGH-SAD) or the Structured Interview Guide for the Hamilton Depression Rating Scale with Atypical Depression Supplement (SIGH-ADS) between blue light and comparison conditions. Secondary outcomes were response (≥ 50% improvement from baseline to endpoint on a depression scale) and remission rates (endpoint score in the remission range). RESULTS: Of 582 articles retrieved, we included nine RCTs (n = 347 participants) assessing blue-light therapy. Seven studies had participants with seasonal MDD and two studies included participants with non-seasonal MDD. Four studies compared blue light to an inactive light condition (efficacy studies), and five studies compared it to an active condition (comparison studies). For the primary outcome, a meta-analysis with random-effects models found no evidence for the efficacy of blue-light conditions compared to inactive conditions (mean difference [MD] = 2.43; 95% confidence interval [CI], -1.28 to 6.14, P = 0.20); however, blue-light also showed no differences compared to active conditions (MD = -0.11; 95% CI, -2.38 to 2.16, P = 0.93). There were no significant differences in response and remission rates between blue-light conditions and inactive or active light conditions. Blue-light therapy was overall well-tolerated. CONCLUSIONS: The efficacy of blue-light therapy in the treatment of seasonal and non-seasonal MDD remains unproven. Future trials should be of longer duration, include larger sample sizes, and attempt to better standardize the parameters of light therapy.

Equality and social determinants of spatial accessibility, availability, and affordability to primary health care in Hong Kong, a descriptive study from the perspective of spatial analysis.

OBJECTIVE: Primary health care (PHC) is widely perceived to be the backbone of health care systems. Since the outbreak of COVID-19, PHC has not only provided primary medical services, but also served as a grassroots network for public health. Our research explored the accessibility, availability, and affordability of primary health care from a spatial perspective, to understand the social determinants affecting access to it in Hong Kong. METHOD: This constitutes a descriptive study from the perspective of spatial analysis. The nearest neighbor method was used to measure the geographic accessibility of PHC based on the road network. The 2SFCA method was used to measure spatial availability and affordability to primary health care, while the SARAR model, Spatial Error model, and Spatial Lag model were then constructed to explain potential factors influencing accessibility and availability of PHC. RESULTS: In terms of accessibility, 95% of residents in Hong Kong can reach a PHC institution within 15 minutes; in terms of availability, 83% of residents can receive PHC service within a month; while in terms of affordability, only 32% of residents can afford PHC services with the support of medical insurance and medical voucher. In Hong Kong, education status and household income show a significant impact on accessibility and availability of PHC. Regions with higher concentrations of residents with post-secondary education receive more PHC resources, while regions with higher concentrations of high-income households show poorer accessibility and poorer availability to PHC. CONCLUSION: The good accessibility and availability of primary health care reflects that the network layout of existing PHC systems in Hong Kong is reasonable and can meet the needs of most residents. No serious gap between social groups further shows equality in resource allocation of PHC in Hong Kong. However, affordability of PHC is not ideal. Indeed, narrowing the gap between availability and affordability is key to fully utilizing the capacity of the PHC system in Hong Kong. The private sector plays an important role in this, but the low coverage of medical insurance in outpatient services exacerbates the crowding of public PHC and underutilization of private PHC. We suggest diverting patients from public to private institutions through medical insurance, medical vouchers, or other ways, to relieve the pressure on the public health system and make full use of existing primary health care in Hong Kong.

A Revisit of Infinite Population Models for Evolutionary Algorithms on Continuous Optimization Problems.

Infinite population models are important tools for studying population dynamics of evolutionary algorithms. They describe how the distributions of populations change between consecutive generations. In general, infinite population models are derived from Markov chains by exploiting symmetries between individuals in the population and analyzing the limit as the population size goes to infinity. In this article, we study the theoretical foundations of infinite population models of evolutionary algorithms on continuous optimization problems. First, we show that the convergence proofs in a widely cited study were in fact problematic and incomplete. We further show that the modeling assumption of exchangeability of individuals cannot yield the transition equation. Then, in order to analyze infinite population models, we build an analytical framework based on convergence in distribution of random elements which take values in the metric space of infinite sequences. The framework is concise and mathematically rigorous. It also provides an infrastructure for studying the convergence of the stacking of operators and of iterating the algorithm which previous studies failed to address. Finally, we use the framework to prove the convergence of infinite population models for the mutation operator and the k-ary recombination operator. We show that these operators can provide accurate predictions for real population dynamics as the population size goes to infinity, provided that the initial population is identically and independently distributed.

Multi-Type Sensor Placements in Gaussian Spatial Fields for Environmental Monitoring.

As citizens are increasingly concerned about the surrounding environment, it is important for modern cities to provide sufficient and accurate environmental information to the public for decision making in the era of smart cities. Due to the limited budget, we often need to optimize the sensor placement in order to maximize the overall information gain according to certain criteria. Existing work is primarily concerned with single-type sensor placement; however, the environment usually requires accurate measurements of multiple types of environmental characteristics. In this paper, we focus on the optimal multi-type sensor placement in Gaussian spatial field for environmental monitoring. We study two representative cases: the one-with-all case when each station is equipped with all types of sensors and the general case when each station is equipped with at least one type of sensor. We propose two greedy algorithms accordingly, each with a provable approximation guarantee. We evaluated the proposed approach via an application in air quality monitoring scenario in Hong Kong and experimental results demonstrate the effectiveness of the proposed approach.

Molecular ties between the cell cycle and differentiation in embryonic stem cells.

Attainment of the differentiated state during the final stages of somatic cell differentiation is closely tied to cell cycle progression. Much less is known about the role of the cell cycle at very early stages of embryonic development. Here, we show that molecular pathways involving the cell cycle can be engineered to strongly affect embryonic stem cell differentiation at early stages in vitro. Strategies based on perturbing these pathways can shorten the rate and simplify the lineage path of ES differentiation. These results make it likely that pathways involving cell proliferation intersect at various points with pathways that regulate cell lineages in embryos and demonstrate that this knowledge can be used profitably to guide the path and effectiveness of cell differentiation of pluripotent cells.

Association between high cumulative dose of benzodiazepine in Chinese patients and risk of dementia: a preliminary retrospective case-control study.

BACKGROUND: Evidence describing the association between high-dose benzodiazepine use and dementia has been conflicting. Most previous studies involved Caucasian populations, with only limited data on Chinese subjects. Possible differences exist between Chinese and Caucasian populations with regard to metabolism and prescription practice. This study aimed to assess the association between high-dose benzodiazepine use and dementia in a Chinese population. METHOD: A retrospective case-control study was carried out in all public hospitals under the Hong Kong Hospital Authority Hong Kong West Cluster between 2000 and 2015. The study recruited 273 Chinese adults (91 cases, 182 controls) aged 75 and over, with at least 6 years of follow-up data. Each dementia case was matched with two controls according to sex, age group, and duration of follow-up. The number of patients with benzodiazepine ever-use and the exposure density based on the prescribed daily doses were assessed. Prescribed daily doses were categorized as either <1096 or ≥1096. Odds ratios and 95% confidence intervals were computed by multivariate analysis. RESULTS: The difference in exposure density between the dementia and control groups was statistically significant between prescribed daily doses <1096 and ≥1096 (P = 0.02). There were two multivariate analyses models; one factored in depression (model 1), and the other (model 2) did not. Model 2 showed a statistically significant association (odds ratio = 1.71, 95% confidence intervals = 1.02-2.89, P = 0.04) between benzodiazepine exposure density and dementia. CONCLUSION: High-dose benzodiazepine use may be associated with dementia in the Chinese population. Prospective studies are required.

Size homeostasis in adherent cells studied by synthetic phase microscopy.

The coupling of the rate of cell growth to the rate of cell division determines cell size, a defining characteristic that is central to cell function and, ultimately, to tissue architecture. The physiology of size homeostasis has fascinated generations of biologists, but the mechanism, challenged by experimental limitations, remains largely unknown. In this paper, we propose a unique optical method that can measure the dry mass of thick live cells as accurately as that for thin cells with high computational efficiency. With this technique, we quantify, with unprecedented accuracy, the asymmetry of division in lymphoblasts and epithelial cells. We can then use the Collins-Richmond model of conservation to compute the relationship between growth rate and cell mass. In attached epithelial cells, we find that due to the asymmetry in cell division and size-dependent growth rate, there is active regulation of cell size. Thus, like nonadherent cells, size homeostasis requires feedback control.

Gap 1 phase length and mouse embryonic stem cell self-renewal.

In somatic cells, the length of the G1 phase of the cell cycle is tightly linked to differentiation, and its elongation can drive differentiation in many cases. Although it has been suggested that the situation is very similar in embryonic stem cells (ESCs), where a rapid cell cycle and a short G1 phase maintain the pluripotent state, evidence has been contradictory. Here we show that, in murine ESCs, elongation of the cell cycle and elongation of G1 are compatible with their pluripotent state. Multiple methods that lengthen the cell cycle and that target cyclin-dependent kinase, retinoblastoma protein, and E2F activity all fail to induce differentiation on their own or even to facilitate differentiation. The resistance of murine ESCs to differentiation induced by lengthening G1 and/or the cell cycle could allow for separate control of these events and provide new opportunities for investigation and application.

Quetiapine attenuates glial activation and proinflammatory cytokines in APP/PS1 transgenic mice via inhibition of nuclear factor-κB pathway.

BACKGROUND: In Alzheimer's disease, growing evidence has shown that uncontrolled glial activation and neuroinflammation may contribute independently to neurodegeneration. Antiinflammatory strategies might provide benefits for this devastating disease. The aims of the present study are to address the issue of whether glial activation and proinflammatory cytokine increases could be modulated by quetiapine in vivo and in vitro and to explore the underlying mechanism. METHODS: Four-month-old amyloid precursor protein (APP) and presenilin 1 (PS1) transgenic and nontransgenic mice were treated with quetiapine (5mg/kg/d) in drinking water for 8 months. Animal behaviors, total Aß levels, and glial activation were evaluated by behavioral tests, enzyme-linked immunosorbent assay, immunohistochemistry, and Western blot accordingly. Inflammatory cytokines and the nuclear factor kappa B pathway were analyzed in vivo and in vitro. RESULTS: Quetiapine improves behavioral performance, marginally affects total Aß40 and Aß42 levels, attenuates glial activation, and reduces proinflammatory cytokines in APP/PS1 mice. Quetiapine suppresses Aß1-42-induced activation of primary microglia by decresing proinflammatory cytokines. Quetiapine inhibits the activation of nuclear factor kappa B p65 pathway in both transgenic mice and primary microglia stimulated by Aß1-42. CONCLUSIONS: The antiinflammatory effects of quetiapine in Alzheimer's disease may be involved in the nuclear factor kappa B pathway. Quetiapine may be an efficacious and promising treatment for Alzheimer's disease targeting on neuroinflammation.

Exploring the relationship between changes in weight and utterances in an online weight loss forum: a content and correlational analysis study.

BACKGROUND: There is increasing interest in the use of online forums as a component of eHealth weight loss interventions. Although the research is mixed on the utility of online forums in general, results suggest that there is promise to this, particularly if the systems can be designed well to support healthful interactions that foster weight loss and continued engagement. OBJECTIVE: The purpose of this study was to examine the relationship between the styles of utterances individuals make on an online weight loss forum and week-to-week fluctuations in weight. This analysis was conducted to generate hypotheses on possible strategies that could be used to improve the overall design of online support groups to facilitate more healthful interactions. METHODS: A convenience sample of individuals using an online weight loss forum (N=4132) included data both on online forum use and weight check-in data. All interactions were coded utilizing the Linguistic Inquiry and Word Count (LIWC) system. Mixed model analyses were conducted to examine the relationship between these LIWC variables and weight over time. RESULTS: Results suggested that increased use of past-tense verbs (P=.05) and motion (P=.02) were associated with lower weekly weights whereas increased use of conjunctions (eg, and, but, whereas; P=.001) and exclusion words (eg, but, without, exclude; P=.07) were both associated with higher weight during the weeks when these utterances were used more. CONCLUSIONS: These results provide some insights on the styles of interactions that appear to be associated with weight fluctuations. Future work should explore the stability of these findings and also explore possibilities for fostering these types of interactions more explicitly within online weight loss forums.

Crystal structure of (S)-5-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)-2-oxazolidinone.

The structure of (S)-5-(3-acetyl-5-chloro-2-ethoxy-6-fluorophenyl)-2-oxazolidinone, C13H13ClFNO4, at 100 K has monoclinic (P21) symmetry. The compound has a polymeric structure propagated by a screw axis parallel to the b axis with N-H⋯O hydrogen bonding. It is of inter-est with respect to efforts in the synthesis of a candidate anti-cancer drug, parsaclisib.

Competition-driven eco-evolutionary feedback reshapes bacteriophage lambda's fitness landscape and enables speciation.

A major challenge in evolutionary biology is explaining how populations navigate rugged fitness landscapes without getting trapped on local optima. One idea illustrated by adaptive dynamics theory is that as populations adapt, their newly enhanced capacities to exploit resources alter fitness payoffs and restructure the landscape in ways that promote speciation by opening new adaptive pathways. While there have been indirect tests of this theory, to our knowledge none have measured how fitness landscapes deform during adaptation, or test whether these shifts promote diversification. Here, we achieve this by studying bacteriophage [Formula: see text], a virus that readily speciates into co-existing receptor specialists under controlled laboratory conditions. We use a high-throughput gene editing-phenotyping technology to measure [Formula: see text]'s fitness landscape in the presence of different evolved-[Formula: see text] competitors and find that the fitness effects of individual mutations, and their epistatic interactions, depend on the competitor. Using these empirical data, we simulate [Formula: see text]'s evolution on an unchanging landscape and one that recapitulates how the landscape deforms during evolution. [Formula: see text] heterogeneity only evolves in the shifting landscape regime. This study provides a test of adaptive dynamics, and, more broadly, shows how fitness landscapes dynamically change during adaptation, potentiating phenomena like speciation by opening new adaptive pathways.

Systematic review on intentional non-medical fentanyl use among people who use drugs.

Objectives: Fentanyl is a highly potent opioid and has, until recently, been considered an unwanted contaminant in the street drug supply among people who use drugs (PWUD). However, it has become a drug of choice for an increasing number of individuals. This systematic review evaluated intentional non-medical fentanyl use among PWUD, specifically by summarizing demographic variance, reasons for use, and resulting patterns of use. Methods: The search strategy was developed with a combination of free text keywords and MeSH and non-MeSH keywords, and adapted with database-specific filters to Ovid MEDLINE, Embase, Web of Science, and PsychINFO. Studies included were human studies with intentional use of non-medical fentanyl or analogues in individuals older than 13. Only peer-reviewed original articles available in English were included. Results: The search resulted in 4437 studies after de-duplication, of which 132 were selected for full-text review. Out of 41 papers included, it was found that individuals who use fentanyl intentionally were more likely to be young, male, and White. They were also more likely to have experienced overdoses, and report injection drug use. There is evidence that fentanyl seeking behaviours are motivated by greater potency, delay of withdrawal, lower cost, and greater availability. Conclusions: Among PWUD, individuals who intentionally use fentanyl have severe substance use patterns, precarious living situations, and extensive overdose history. In response to the increasing number of individuals who use fentanyl, alternative treatment approaches need to be developed for more effective management of withdrawal and opioid use disorder. Systematic review registration: https://www.crd.york.ac.uk/prospero/, identifier CRD42021272111.