Exacerbated pro‐inflammatory immune response contributes to COVID‐19 pathology. However, despite the mounting evidence about SARS‐CoV‐2 infecting the human gut, little is known about the antiviral ...programs triggered in this organ. To address this gap, we performed single‐cell transcriptomics of SARS‐CoV‐2‐infected intestinal organoids. We identified a subpopulation of enterocytes as the prime target of SARS‐CoV‐2 and, interestingly, found the lack of positive correlation between susceptibility to infection and the expression of ACE2. Infected cells activated strong pro‐inflammatory programs and produced interferon, while expression of interferon‐stimulated genes was limited to bystander cells due to SARS‐CoV‐2 suppressing the autocrine action of interferon. These findings reveal that SARS‐CoV‐2 curtails the immune response and highlights the gut as a pro‐inflammatory reservoir that should be considered to fully understand SARS‐CoV‐2 pathogenesis.
Synopsis
Single cell sequencing and multiplex single‐molecule RNA FISH analyses on SARS‐CoV‐2 infected human intestinal organoids characterize the tropism of SARS‐CoV‐2 and identify strategies developed by the virus to interfere with the host intrinsic innate immune response.
SARS‐CoV‐2 primarily infects the enterocyte lineage.
High expression levels of ACE2 does not correlate with higher infectability of cells by SARS‐CoV‐2.
ACE2 expression is downregulated upon SARS‐CoV‐2 infection of human intestinal epithelial cells.
Infected cells show a high pro‐inflammatory response and little to no interferon‐mediated response as the result of a SARS‐CoV‐2‐mediated inhibition of interferon signaling.
Single cell sequencing and multiplex single‐molecule RNA FISH analyses on SARS‐CoV‐2 infected human intestinal organoids characterize the tropism of SARS‐CoV‐2 and identify strategies developed by the virus to interfere with the host intrinsic innate immune response.
DNA sequence variation has been associated with quantitative changes in molecular phenotypes such as gene expression, but its impact on chromatin states is poorly characterized. To understand the ...interplay between chromatin and genetic control of gene regulation, we quantified allelic variability in transcription factor binding, histone modifications, and gene expression within humans. We found abundant allelic specificity in chromatin and extensive local, short-range, and long-range allelic coordination among the studied molecular phenotypes. We observed genetic influence on most of these phenotypes, with histone modifications exhibiting strong context-dependent behavior. Our results implicate transcription factors as primary mediators of sequence-specific regulation of gene expression programs, with histone modifications frequently reflecting the primary regulatory event.
The transcriptome contains rich information on molecular, cellular and organismal phenotypes. However, experimental and statistical limitations constrain sensitivity and throughput of genetic ...screening with single-cell transcriptomics readout. To overcome these limitations, we introduce targeted Perturb-seq (TAP-seq), a sensitive, inexpensive and platform-independent method focusing single-cell RNA-seq coverage on genes of interest, thereby increasing the sensitivity and scale of genetic screens by orders of magnitude. TAP-seq permits routine analysis of thousands of CRISPR-mediated perturbations within a single experiment, detects weak effects and lowly expressed genes, and decreases sequencing requirements by up to 50-fold. We apply TAP-seq to generate perturbation-based enhancer-target gene maps for 1,778 enhancers within 2.5% of the human genome. We thereby show that enhancer-target association is jointly determined by three-dimensional contact frequency and epigenetic states, allowing accurate prediction of enhancer targets throughout the genome. In addition, we demonstrate that TAP-seq can identify cell subtypes with only 100 sequencing reads per cell.
Medullary thymic epithelial cells (mTECs) play a critical role in central immune tolerance by mediating negative selection of autoreactive T cells through the collective expression of the peripheral ...self-antigen compartment, including tissue-specific antigens (TSAs). Recent work has shown that gene-expression patterns within the mTEC compartment are heterogenous and include multiple differentiated cell states. To further define mTEC development and medullary epithelial lineage relationships, we combined lineage tracing and recovery from transient in vivo mTEC ablation with single-cell RNA-sequencing in
. The combination of bioinformatic and experimental approaches revealed a non-stem transit-amplifying population of cycling mTECs that preceded
expression. We propose a branching model of mTEC development wherein a heterogeneous pool of transit-amplifying cells gives rise to
- and
-expressing mTEC subsets. We further use experimental techniques to show that within the
expressing developmental branch, TSA expression peaked as
expression decreased, implying
expression must be established before TSA expression can occur. Collectively, these data provide a roadmap of mTEC development and demonstrate the power of combinatorial approaches leveraging both in vivo models and high-dimensional datasets.
Copy number variations (CNVs) are a significant source of genetic diversity and commonly found in mammalian genomes. We have generated a genome-wide CNV map for Cynomolgus monkeys (Macaca ...fascicularis). This crab-eating macaque is the closest animal model to humans that is used in biomedical research.
We show that Cynomolgus monkey CNVs are in general much smaller in size than gene loci and are specific to the population of origin. Genome-wide expression data from five vitally important organs demonstrates that CNVs in close proximity to transcription start sites associate strongly with expression changes. Among these eQTL genes we find an overrepresentation of genes involved in metabolism, receptor activity, and transcription.
These results provide evidence that CNVs shape tissue transcriptomes in monkey populations, potentially offering an adaptive advantage. We suggest that this genetic diversity should be taken into account when using Cynomolgus macaques as models.
High-throughput sequencing technologies enable the genome-wide analysis of the impact of genetic variation on molecular phenotypes at unprecedented resolution. However, although powerful, these ...technologies can also introduce unexpected artifacts.
We investigated the impact of library amplification bias on the identification of allele-specific (AS) molecular events from high-throughput sequencing data derived from chromatin immunoprecipitation assays (ChIP-seq). Putative AS DNA binding activity for RNA polymerase II was determined using ChIP-seq data derived from lymphoblastoid cell lines of two parent-daughter trios. We found that, at high-sequencing depth, many significant AS binding sites suffered from an amplification bias, as evidenced by a larger number of clonal reads representing one of the two alleles. To alleviate this bias, we devised an amplification bias detection strategy, which filters out sites with low read complexity and sites featuring a significant excess of clonal reads. This method will be useful for AS analyses involving ChIP-seq and other functional sequencing assays.
The R package abs filter for library clonality simulations and detection of amplification-biased sites is available from http://updepla1srv1.epfl.ch/waszaks/absfilter
Aim
Falls are a leading cause of disability in older people. Here we investigate if daily‐life gait assessments are better than clinical gait assessments at discriminating between older people with ...and without a history of falls.
Methods
A total of 96 independent‐living participants (age 75.5 ± 7.8) underwent sensorimotor, psychological and cognitive assessments, and the Timed Up and Go and 10‐m walk tests. Participants wore a small pendant sensor device for a week in their home environment, from which the new remote assessments of daily‐life gait were determined.
Results
During daily‐life, fallers had significantly lower gait quality (lower gait endurance, higher within‐walk variability and lower between‐walk adaptability), but not reduced gait quantity (total steps) or gait intensity (mean cadence). In the clinic, fallers had slower Timed Up and Go, but not 10‐m walk test times. After adjusting for demographics, only the daily‐life assessments of gait endurance and within‐walk variability remained significant. Reduced daily‐life gait assessments were significantly correlated with older age, higher body mass index, multiple medications, disability, more concern about falling, poor executive function and higher physiological fall risk.
Conclusions
The new daily‐life gait assessments were better than the clinical gait assessments at identifying fall risk in our sample of independent living older people. However, further research is required to validate these findings in other populations or those living in residential aged care. Daily‐life gait was not only associated with demographics and physiological capacity, but also general health, executive function and the ability to undertake a variety of activities of daily living without excessive concern about falling. Geriatr Gerontol Int 2017; 17: 2274–2282.
Accidental falls remain an important problem in older people. The five-times-sit-to-stand (5STS) test is commonly used as a functional test to assess fall risk. Recent advances in sensor technologies ...hold great promise for more objective and accurate assessments.
The aims of this study were: (1) to examine the feasibility of a low-cost and portable Kinect-based 5STS test to discriminate between fallers and nonfallers and (2) to investigate whether this test can be used for supervised clinical, supervised and unsupervised in-home fall risk assessments.
A total of 94 community-dwelling older adults were assessed by the Kinect-based 5STS test in the laboratory and 20 participants were tested in their own homes. An algorithm was developed to automatically calculate timing- and speed-related measurements from the Kinect-based sensor data to discriminate between fallers and nonfallers. The associations of these measurements with standard clinical fall risk tests and the results of supervised and unsupervised in-home assessments were examined.
Fallers were significantly slower than nonfallers on Kinect-based measures. The mean velocity of the sit-to-stand transitions discriminated well between the fallers and nonfallers based on 12-month retrospective fall data. The Kinect-based measures collected in the laboratory correlated strongly with those collected in the supervised (r = 0.704-0.832) and unsupervised (r = 0.775-0.931) in-home assessments.
In summary, we found that the Kinect-based 5STS test discriminated well between the fallers and nonfallers and was feasible to administer in clinical and supervised in-home settings. This test may be useful in clinical settings for identifying high-risk fallers for further intervention or for regular in-home assessments in the future.
Falls remain an important problem in older people. There is strong evidence that falls can be prevented with appropriately designed intervention programs. To start a targeted fall prevention program, ...a first step is to identify those at high risk of falls. Sensor-based tests hold great promise for more frequent and accurate assessment of fall risk in clinical and home settings. The aims of this study were to (a) empirically examine the feasibility of the iStoppFalls (Information and communications technology-based System to Predict & Prevent Falls) assessment, a Kinect and inertial sensor-based test for regular and unsupervised fall risk assessments at home, (b) investigate the experience of older adults with this home-based self-assessment, and (c) make recommendations for future assessments. The iStoppFalls assessment system was installed into the homes of 62 community-living older people in Australia, Germany, and Spain for the duration of 4 months. Participants were asked to perform at least 1 assessment each month. The system use and the user experience were evaluated. To our knowledge, these are the first results on the long-term use of an unsupervised directed routine fall risk assessment system at private homes. In total, 241 assessments were independently performed by the participants. Most participants felt positive about their experience and could see themselves continuing with the assessment on a regular basis. Through the analysis the user motivation, the design and selection of appropriate tests, the user feedback, the reliability and usability of the applied technology, the frequency and duration of the assessment and the safety and support aspects were identified as important characteristics of a home-based self-assessment. The findings demonstrate the feasibility of a sensor-based self-assessment for fall risk but also highlight that further work is necessary. Future research should consider the necessary design requirements identified by this study.
There is good evidence that balance challenging exercises can reduce falls in older people. However, older people often find it difficult to incorporate such programs in their daily life. Videogame ...technology has been proposed to promote enjoyable, balance-challenging exercise. As part of a larger analysis, we compared feasibility and efficacy of two exergame interventions: step-mat-training (SMT) and Microsoft-Kinect® (KIN) exergames.
148 community-dwelling people, aged 65+ years participated in two exergame studies in Sydney, Australia (KIN: n = 57, SMT: n = 91). Both interventions were delivered as unsupervised exercise programs in participants' homes for 16 weeks. Assessment measures included overall physiological fall risk, muscle strength, finger-press reaction time, proprioception, vision, balance and executive functioning.
For participants allocated to the intervention arms, the median time played each week was 17 min (IQR 32) for KIN and 48 min (IQR 94) for SMT. Compared to the control group, SMT participants improved their fall risk score (p = 0.036), proprioception (p = 0.015), reaction time (p = 0.003), sit-to-stand performance (p = 0.011) and executive functioning (p = 0.001), while KIN participants improved their muscle strength (p = 0.032) and vision (p = 0.010), and showed a trend towards improved fall risk scores (p = 0.057).
The findings suggest that it is feasible for older people to conduct an unsupervised exercise program at home using exergames. Both interventions reduced fall risk and SMT additionally improved specific cognitive functions. However, further refinement of the systems is required to improve adherence and maximise the benefits of exergames to deliver fall prevention programs in older people's homes.
ACTRN12613000671763 (Step Mat Training RCT) ACTRN12614000096651 (MS Kinect RCT).
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