The mouse embryo is the canonical model for mammalian preimplantation development. Recent advances in single cell profiling allow detailed analysis of embryogenesis in other eutherian species, ...including human, to distinguish conserved from divergent regulatory programs and signalling pathways in the rodent paradigm. Here, we identify and compare transcriptional features of human, marmoset and mouse embryos by single cell RNA-seq. Zygotic genome activation correlates with the presence of polycomb repressive complexes in all three species, while ribosome biogenesis emerges as a predominant attribute in primate embryos, supporting prolonged translation of maternally deposited RNAs. We find that transposable element expression signatures are species, stage and lineage specific. The pluripotency network in the primate epiblast lacks certain regulators that are operative in mouse, but encompasses WNT components and genes associated with trophoblast specification. Sequential activation of GATA6, SOX17 and GATA4 markers of primitive endoderm identity is conserved in primates. Unexpectedly, OTX2 is also associated with primitive endoderm specification in human and non-human primate blastocysts. Our cross-species analysis demarcates both conserved and primate-specific features of preimplantation development, and underscores the molecular adaptability of early mammalian embryogenesis.
Formation of the three primary germ layers during gastrulation is an essential step in the establishment of the vertebrate body plan and is associated with major transcriptional changes
. Global ...epigenetic reprogramming accompanies these changes
, but the role of the epigenome in regulating early cell-fate choice remains unresolved, and the coordination between different molecular layers is unclear. Here we describe a single-cell multi-omics map of chromatin accessibility, DNA methylation and RNA expression during the onset of gastrulation in mouse embryos. The initial exit from pluripotency coincides with the establishment of a global repressive epigenetic landscape, followed by the emergence of lineage-specific epigenetic patterns during gastrulation. Notably, cells committed to mesoderm and endoderm undergo widespread coordinated epigenetic rearrangements at enhancer marks, driven by ten-eleven translocation (TET)-mediated demethylation and a concomitant increase of accessibility. By contrast, the methylation and accessibility landscape of ectodermal cells is already established in the early epiblast. Hence, regulatory elements associated with each germ layer are either epigenetically primed or remodelled before cell-fate decisions, providing the molecular framework for a hierarchical emergence of the primary germ layers.
The mouse inner cell mass (ICM) segregates into the epiblast and primitive endoderm (PrE) lineages coincident with implantation of the embryo. The epiblast subsequently undergoes considerable ...expansion of cell numbers prior to gastrulation. To investigate underlying regulatory principles, we performed systematic single-cell RNA sequencing (seq) of conceptuses from E3.5 to E6.5. The epiblast shows reactivation and subsequent inactivation of the X chromosome, with Zfp57 expression associated with reactivation and inactivation together with other candidate regulators. At E6.5, the transition from epiblast to primitive streak is linked with decreased expression of polycomb subunits, suggesting a key regulatory role. Notably, our analyses suggest elevated transcriptional noise at E3.5 and within the non-committed epiblast at E6.5, coinciding with exit from pluripotency. By contrast, E6.5 primitive streak cells became highly synchronized and exhibit a shortened G1 cell-cycle phase, consistent with accelerated proliferation. Our study systematically charts transcriptional noise and uncovers molecular processes associated with early lineage decisions.
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•A high-resolution scRNA-seq map of mouse from peri-implantation to early gastrulation•Symmetry breaking genes and bivalent chromatin are linked to lineage fate at E4.5•X chromosome inactivation correlates with Rlim and anticorrelates with Dnmt3a and Zfp57•Polycomb targets are repressed in the E6.5 epiblast and activated in the primitive streak
Mohammed et al. chart mouse embryonic development from implantation to early gastrulation at single-cell resolution. They describe regulatory processes associated with lineage commitment. An increased level of transcriptional noise is observed prior to lineage commitment, an observation that provides fresh insights into cell fate decision-making processes.
Rapid immunoprecipitation mass spectrometry of endogenous protein (RIME) is a method that allows the study of protein complexes, in particular chromatin and transcription factor complexes, in a rapid ...and robust manner by mass spectrometry (MS). The method can be used in parallel with chromatin immunoprecipitation-sequencing (ChIP-seq) experiments to provide information on both the cistrome and interactome for a given protein. The method uses formaldehyde fixation to stabilize protein complexes. By using antibodies against the endogenous target, the cross-linked complex is immunoprecipitated, rigorously washed, and then digested into peptides while avoiding antibody contamination (on-bead digestion). By using this method, MS identification of the target protein and several dozen interacting proteins is possible using a 100-min LC-MS/MS run. The protocol does not require substantial proteomics expertise, and it typically takes 2-3 d from the collection of material to results.
Conventional human embryonic stem cells are considered to be primed pluripotent but can be induced to enter a naive state. However, the transcriptional features associated with naive and primed ...pluripotency are still not fully understood. Here we used single-cell RNA sequencing to characterize the differences between these conditions. We observed that both naive and primed populations were mostly homogeneous with no clear lineage-related structure and identified an intermediate subpopulation of naive cells with primed-like expression. We found that the naive-primed pluripotency axis is preserved across species, although the timing of the transition to a primed state is species specific. We also identified markers for distinguishing human naive and primed pluripotency as well as strong co-regulatory relationships between lineage markers and epigenetic regulators that were exclusive to naive cells. Our data provide valuable insights into the transcriptional landscape of human pluripotency at a cellular and genome-wide resolution.
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•A single-cell RNA-seq resource of naive and primed human embryonic stem cells (hESCs)•Naive and primed hESCs are homogeneous except for a naive intermediate subpopulation•Naive and primed pluripotency signatures are conserved between species•Pluripotency and lineage markers correlate with epigenetic machinery in naive hESCs
Messmer et al. demonstrate that the single-cell transcriptomes of naive and primed human embryonic stem cells (hESCs) are mostly homogeneous. The study defines an expression signature that is conserved across species and shows differential epigenetic regulation between naive and primed pluripotency.
Lamina-associated domains (LADs) cover a large part of the human genome and are thought to play a major role in shaping the nuclear architectural landscape. Here, we perform polymer simulations, ...microscopy, and mass spectrometry to dissect the roles played by heterochromatin- and lamina-mediated interactions in nuclear organization. Our model explains the conventional organization of heterochromatin and euchromatin in growing cells and the pathological organization found in oncogene-induced senescence and progeria. We show that the experimentally observed changes in the locality of contacts in senescent and progeroid cells can be explained as arising due to phase transitions in the system. Within our simulations, LADs are highly stochastic, as in experiments. Our model suggests that, once established, the senescent phenotype should be metastable even if lamina-mediated interactions were reinstated. Overall, our simulations uncover a generic physical mechanism that can regulate heterochromatin segregation and LAD formation in a wide range of mammalian nuclei.
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•A 3D polymer model for heterochromatin and lamina interactions is presented•The model captures chromatin organization in growing, senescent, and progeroid cells•The model explains the change in the chromatin contact network between cell states•The model predicts the stochasticity of lamina contacts and stability of senescence
Chiang et al. use polymer simulations to investigate the roles of heterochromatin and lamina interactions on nuclear reorganization in cellular senescence. Their model captures chromatin organization in growing, senescent, and progeroid cells and predicts that a polymeric phase transition underlies the ensuing rearrangements in the chromatin contact network.
Coronaviruses (CoVs) are a family of viruses that are best known as the causative agents of human diseases like the common cold, Middle East Respiratory Syndrome (MERS), Severe Acute Respiratory ...Syndrome (SARS) and COVID-19. CoVs spread by human-to-human transmission via droplets or direct contact. There is, however, concern about potential waterborne transmission of SARS-CoV-2, the virus responsible for COVID-19, as it has been found in wastewater facilities and rivers. To date, little is known about the stability of SARS-CoV-2 or any other free coronavirus in aquatic environments. The inactivation of terrestrial CoVs in seawater is rarely studied. Here, we use a porcine respiratory coronavirus (PRCV) that is commonly found in animal husbandry as a surrogate to study the stability of CoVs in natural water. A series of experiments were conducted in which PRCV (strain 91V44) was added to filtered and unfiltered fresh- and saltwater taken from the river Scheldt and the North Sea. Virus titres were then measured by TCID50-assays using swine testicle cell cultures after various incubation times. The results show that viral inactivation of PRCV in filtered seawater can be rapid, with an observed 99% decline in the viral load after just two days, which may depend on temperature and the total suspended matter concentration. PRCV degraded much slower in filtered water from the river Scheldt, taking over 15 days to decline by 99%, which was somewhat faster than the PBS control treatment (T99 = 19.2 days). Overall, the results suggest that terrestrial CoVs are not likely to accumulate in marine environments. Studies into potential interactions with exudates (proteases, nucleases) from the microbial food web are, however, recommended.
Objectives
The aim of this review was to systematically evaluate the failure rates of miniscrews related to their specific insertion site and explore the insertion site dependent risk factors ...contributing to their failure.
Search methods
An electronic search was conducted in the Cochrane Central Register of Controlled Trials (CENTRAL), Web of Knowledge, Scopus, MEDLINE and PubMed up to October 2017. A comprehensive manual search was also performed.
Eligibility criteria
Randomised clinical trials and prospective non-randomised studies, reporting a minimum of 20 inserted miniscrews in a specific insertion site and reporting the miniscrews’ failure rate in that insertion site, were included.
Data collection and analysis
Study selection, data extraction and quality assessment were performed independently by two reviewers. Studies were sub-grouped according to the insertion site, and the failure rates for every individual insertion site were analysed using a random-effects model with corresponding 95% confidence interval. Sensitivity analyses were performed in order to test the robustness of the reported results.
Results
Overall, 61 studies were included in the quantitative synthesis. Palatal sites had failure rates of 1.3% (95% CI 0.3–6), 4.8% (95% CI 1.6–13.4) and 5.5% (95% CI 2.8–10.7) for the midpalatal, paramedian and parapalatal insertion sites, respectively. The failure rates for the maxillary buccal sites were 9.2% (95% CI 7.4–11.4), 9.7% (95% CI 5.1–17.6) and 16.4% (95% CI 4.9–42.5) for the interradicular miniscrews inserted between maxillary first molars and second premolars and between maxillary canines and lateral incisors, and those inserted in the zygomatic buttress respectively. The failure rates for the mandibular buccal insertion sites were 13.5% (95% CI 7.3–23.6) and 9.9% (95% CI 4.9–19.1) for the interradicular miniscrews inserted between mandibular first molars and second premolars and between mandibular canines and first premolars, respectively. The risk of failure increased when the miniscrews contacted the roots, with a risk ratio of 8.7 (95% CI 5.1–14.7).
Conclusions
Orthodontic miniscrew implants provide acceptable success rates that vary among the explored insertion sites. Very low to low quality of evidence suggests that miniscrews inserted in midpalatal locations have a failure rate of 1.3% and those inserted in the zygomatic buttress have a failure rate of 16.4%. Moderate quality of evidence indicates that root contact significantly contributes to the failure of interradicular miniscrews placed between the first molars and second premolars. Results should be interpreted with caution due to methodological drawbacks in some of the included studies.
Active audiovisual representation of instructions ensures vibrant knowledge acquisition and improves acquaintance needed for self-care with retainer wear. The aim of this trial is to assess the ...impact of audiovisual instructions with additional weekly electronic reminder messages on improving adherence to instructed wear time of Hawley retainer, periodontal outcomes, and participants' experiences. Fifty-two participants (mean age 26.1 y) planned for removable retention, were randomly assigned to two parallel groups to receive either (1) audiovisual instructions with an additional weekly reminder, or (2) verbal instructions alone. Each participant received a Hawley retainer equipped with a TheraMon microsensor and was instructed to wear it for 22 h daily. Participants were monitored for adherence to the wear time after 3 (T1) and 6 months (T2), and had their periodontal health and experiences assessed at T2. Overall, the mean objectively measured daily wear time at T1 was 14.9 (± 4.9 h), and 14.3 (± 5.4 h) at T2. After 3 months, no significant differences were found between the groups (p = 0.065), however, a significant difference favoring better compliance with wear instructions was observed in the audiovisual group after 6 months (p = 0.033). A non-significant difference was observed between both groups regarding the gingival (p = 0.165) and plaque index scores (p = 0.173). Participants' experiences were similar in both groups, except for satisfaction with the way of delivering instructions, being favorably reported in the audiovisual group. Audiovisual instructions with weekly reminders seem to have a significant effect on patient compliance in the longer term.Trial registration: TCTR20230220002.
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