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Morphogenesis and pattern formation are vital processes in any organism, whether unicellular or multicellular. But in contrast to the developmental biology of plants and animals, the principles of morphogenesis and pattern formation in single cells remain

Ribosomes can read through stop codons in a regulated manner, elongating rather than terminating the nascent peptide. Stop codon readthrough is essential to diverse viruses, and phylogenetically predicted to occur in a few hundred genes in Drosophila melan

Rationale: Studies have shown that reducing sedation of critically ill patients shortens time on the ventilator and in the intensive care unit (ICU). Little is known, however, of how such strategies affect long-term cognitive, psychological, and functional outcomes.Objectives: To determine the long-term effects of a wake up and breathe protocol that interrupts and reduces sedative exposure in the ICU.Methods: In this a priori planned substudy conducted at one tertiary care hospital during the Awakening and Breathing Controlled Trial, a multicenter randomized controlled trial, we assessed cognitive, psychological, and functional/quality-of-life outcomes 3 and 12 months postdischarge among 180 medical ICU patients randomized to paired daily spontaneous awakening trials with spontaneous breathing trials (SBTs) or to sedation per usual care plus daily SBTs.Measurements and Main Results: Cognitive impairment was less common in the intervention group at 3-month follow-up (absolute risk reduction, 20.2%; 95% confidence interval, 1.5-36.1%; P = 0.03) but not at 12-month follow-up (absolute risk reduction, -1.9%; 95% CI, -21.3 to 27.1%; P = 0.89). Composite cognitive scores, alternatively, were similar in the two groups at 3-month and 12-month follow-up (P = 0.80 and 0.61, respectively), as were symptoms of depression (P = 0.59 and 0.82) and posttraumatic stress disorder (P = 0.59 and 0.97). Activities of daily living, functional status, and mental and physical quality of life were similar between groups throughout follow-up.Conclusions: In this trial, management of mechanically ventilated medical ICU patients with a wake up and breathe protocol resulted in similar cognitive, psychological, and functional outcomes among patients tested 3 and 12 months post-ICU. The proven benefits of this protocol, including improved 1-year survival, were not offset by adverse long-term outcomes. Clinical trial registered with www.clinicaltrials.gov (NCT 00097630). [ABSTRACT FROM AUTHOR]

Purpose: To assess the biomechanical performance of different stitching methods using a suturing device by comparing the elongation, stiffness, failure load, and time to stitch completion in cadaveric semitendinosus tendons (STs) and quadriceps tendons (QTs)., Methods: A total of 24 STs and 16 QTs were harvested from cadaveric knee specimens (N = 40). Samples were randomly divided into 2 groups: whipstitch (WS) and whip-lock (WL) stitch. Both tendon ends were clamped to a graft preparation stand, and a 2-part needle was used to place 5 stitches, each 0.5 cm apart. Stitching time was recorded. Samples were preconditioned and then underwent cyclic loading from 50 to 200 N at 1 Hz for 500 cycles, followed by load-to-failure testing at 20 mm/min. Stiffness (in newtons per millimeter), ultimate failure load (in newtons), peak-to-peak displacement (in millimeters), elongation (in millimeters), and failure displacement (in millimeters) were recorded., Results: Completion of the WS was significantly faster than the WL stitch in the ST ( P < .001) and QT ( P  = .004). For the ST, the WL stitch exhibited higher ultimate failure loads and construct stiffness than the WS. Regarding the QT, the WL stitch showed higher stiffness and displacement than the WS; however, the ultimate failure load was higher for the WS in the QT. The ultimate failure load in the QT was higher than that in the ST for both stitches. In the ST, only 25% of WSs and 100% of WL stitches failed due to suture breakage. In the QT, suture breakage led to the failure of 100% of both the WL stitches and WSs., Conclusions: In the ST, the WL stitch resulted in improved biomechanical performance through higher ultimate load and fewer failures from tissue damage compared with the WS. In the QT, both the WS and the WL stitch showed similar biomechanical performance with ultimate failure loads above established clinical failure thresholds., Clinical Relevance: Various types of ligament and tendon injuries require suturing to enable repair or reconstruction. The success of ligament or tendon surgery often relies on soft-tissue quality. It is important to investigate the biomechanical properties of stitching techniques that help preserve soft-tissue quality as a step to determining their clinical suitability., Competing Interests: The authors report the following potential conflicts of interest or sources of funding: This study was funded by Winter Innovations and was supported by the 10.13039/100000001National Science Foundation (under grant No. 2112103). M.A.D. receives a research grant from Winter Innovations (grant No. 2112103, paid directly to the Foundation for Orthopaedic Research and Education). In addition, the Foundation for Orthopaedic Research and Education has received a research grant from DePuy Mitek. A.B. receives support from Sanara MedTech and ZIMVIE, outside the submitted work. S.E.J. receives support from Arthrex, CGG Medical, Vericel, GE Healthcare, BREG, and SI-BONE, outside the submitted work. All other authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this article. Full ICMJE author disclosure forms are available for this article online, as supplementary material., (© 2023 The Authors.)

One method of preventing surgical-site infection is lowering intraoperative environmental contamination. The authors sought to evaluate their hospital's operating room (OR) contamination rate and compare it with the remainder of the hospital. They tested environmental contamination in preoperative, intraoperative, and postoperative settings for a total joint arthroplasty patient. A total of 190 air settle plates composed of trypsin soy agar were placed in 19 settings within the hospital. Locations included the OR with light and heavy traffic, with and without masks, jackets, and shoe covers; the substerile room; OR hallways; the sterile equipment processing center; preoperative areas; post-anesthesia care units; orthopedic floors; the emergency department; OR locker rooms and restrooms; a resident's home; and controls. The trypsin soy agar plates were incubated at 36 °C for 48 hours. Colony counts were performed for each plate. Average colony-forming units (CFUs) were calculated in each setting. The highest CFUs were in the OR locker room, at 28 CFUs per plate per hour. Preoperative and post-anesthesia care unit holding areas were 7.4 CFUs and 9.6 CFUs, respectively. The main orthopedic surgical ward had 10.0 CFUs per plate per hour, whereas the VIP hospital ward had 17.0 CFUs per plate per hour. All OR environments had low CFUs. A live OR had slightly higher CFUs than settings without OR personnel. In comparison with the local community household, the OR locker room, restrooms, hospital orthopedic wards, emergency department, preoperative holding, post-anesthesia care unit, and OR hallway all had higher airborne contamination. On the basis of these results, the authors recommend environmental sampling as a simple, fast, inexpensive tool for monitoring airborne contamination. [ Orthopedics . 2021;44(3):e414-e416.].

Machine learning provides researchers a unique opportunity to make metabolic engineering more predictable. In this review, we offer an introduction to this discipline in terms that are relatable to metabolic engineers, as well as providing in-depth illustrative examples leveraging omics data and improving production. We also include practical advice for the practitioner in terms of data management, algorithm libraries, computational resources, and important non-technical issues. A variety of applications ranging from pathway construction and optimization, to genetic editing optimization, cell factory testing, and production scale-up are discussed. Moreover, the promising relationship between machine learning and mechanistic models is thoroughly reviewed. Finally, the future perspectives and most promising directions for this combination of disciplines are examined., (Copyright © 2020. Published by Elsevier Inc.)

We are just beginning to understand how translational control affects tumour initiation and malignancy. Here we use an epidermis-specific, in vivo ribosome profiling strategy to investigate the translational landscape during the transition from normal homeostasis to malignancy. Using a mouse model of inducible SOX2, which is broadly expressed in oncogenic RAS-associated cancers, we show that despite widespread reductions in translation and protein synthesis, certain oncogenic mRNAs are spared. During tumour initiation, the translational apparatus is redirected towards unconventional upstream initiation sites, enhancing the translational efficiency of oncogenic mRNAs. An in vivo RNA interference screen of translational regulators revealed that depletion of conventional eIF2 complexes has adverse effects on normal but not oncogenic growth. Conversely, the alternative initiation factor eIF2A is essential for cancer progression, during which it mediates initiation at these upstream sites, differentially skewing translation and protein expression. Our findings unveil a role for the translation of 5' untranslated regions in cancer, and expose new targets for therapeutic intervention., Competing Interests: The authors declare no competing financial interests. Readers are welcome to comment on the online version of the paper.

Background: Next-generation sequencing (NGS) informs many biological questions with unprecedented depth and nucleotide resolution. These assays have created a need for analytical tools that enable users to manipulate data nucleotide-by-nucleotide robustly and easily. Furthermore, because many NGS assays encode information jointly within multiple properties of read alignments - for example, in ribosome profiling, the locations of ribosomes are jointly encoded in alignment coordinates and length - analytical tools are often required to extract the biological meaning from the alignments before analysis. Many assay-specific pipelines exist for this purpose, but there remains a need for user-friendly, generalized, nucleotide-resolution tools that are not limited to specific experimental regimes or analytical workflows., Results: Plastid is a Python library designed specifically for nucleotide-resolution analysis of genomics and NGS data. As such, Plastid is designed to extract assay-specific information from read alignments while retaining generality and extensibility to novel NGS assays. Plastid represents NGS and other biological data as arrays of values associated with genomic or transcriptomic positions, and contains configurable tools to convert data from a variety of sources to such arrays. Plastid also includes numerous tools to manipulate even discontinuous genomic features, such as spliced transcripts, with nucleotide precision. Plastid automatically handles conversion between genomic and feature-centric coordinates, accounting for splicing and strand, freeing users of burdensome accounting. Finally, Plastid's data models use consistent and familiar biological idioms, enabling even beginners to develop sophisticated analytical workflows with minimal effort., Conclusions: Plastid is a versatile toolkit that has been used to analyze data from multiple NGS assays, including RNA-seq, ribosome profiling, and DMS-seq. It forms the genomic engine of our ORF annotation tool, ORF-RATER, and is readily adapted to novel NGS assays. Examples, tutorials, and extensive documentation can be found at https://plastid.readthedocs.io .

Morphogenesis and pattern formation are vital processes in any organism, whether unicellular or multicellular. But in contrast to the developmental biology of plants and animals, the principles of morphogenesis and pattern formation in single cells remain largely unknown. Although all cells develop patterns, they are most obvious in ciliates; hence, we have turned to a classical unicellular model system, the giant ciliate Stentor coeruleus. Here we show that the RNA interference (RNAi) machinery is conserved in Stentor. Using RNAi, we identify the kinase coactivator Mob1--with conserved functions in cell division and morphogenesis from plants to humans-as an asymmetrically localized patterning protein required for global patterning during development and regeneration in Stentor. Our studies reopen the door for Stentor as a model regeneration system., Competing Interests: The authors have declared that no competing interests exist.

Ribosomes can read through stop codons in a regulated manner, elongating rather than terminating the nascent peptide. Stop codon readthrough is essential to diverse viruses, and phylogenetically predicted to occur in a few hundred genes in Drosophila melanogaster, but the importance of regulated readthrough in eukaryotes remains largely unexplored. Here, we present a ribosome profiling assay (deep sequencing of ribosome-protected mRNA fragments) for Drosophila melanogaster, and provide the first genome-wide experimental analysis of readthrough. Readthrough is far more pervasive than expected: the vast majority of readthrough events evolved within D. melanogaster and were not predicted phylogenetically. The resulting C-terminal protein extensions show evidence of selection, contain functional subcellular localization signals, and their readthrough is regulated, arguing for their importance. We further demonstrate that readthrough occurs in yeast and humans. Readthrough thus provides general mechanisms both to regulate gene expression and function, and to add plasticity to the proteome during evolution. DOI: http://dx.doi.org/10.7554/eLife.01179.001.

The conserved transcriptional regulator heat shock factor 1 (Hsf1) is a key sensor of proteotoxic and other stress in the eukaryotic cytosol. We surveyed Hsf1 activity in a genome-wide loss-of-function library in Saccaromyces cerevisiae as well as ~78,000 double mutants and found Hsf1 activity to be modulated by highly diverse stresses. These included disruption of a ribosome-bound complex we named the Ribosome Quality Control Complex (RQC) comprising the Ltn1 E3 ubiquitin ligase, two highly conserved but poorly characterized proteins (Tae2 and Rqc1), and Cdc48 and its cofactors. Electron microscopy and biochemical analyses revealed that the RQC forms a stable complex with 60S ribosomal subunits containing stalled polypeptides and triggers their degradation. A negative feedback loop regulates the RQC, and Hsf1 senses an RQC-mediated translation-stress signal distinctly from other stresses. Our work reveals the range of stresses Hsf1 monitors and elucidates a conserved cotranslational protein quality control mechanism., (Copyright © 2012 Elsevier Inc. All rights reserved.)

The purpose of this study was to determine whether developmental differences exist in childrens' use of self-regulatory strategies while learning supra-memory-span movement sequences. Grade 1 and Grade 4 children were asked to study two movement sequences long enough to be absolutely certain that they would be able to recall the entire sequences correctly. The older group displayed superior recall readiness in comparison to the younger group. Older children used a greater variety of strategies and also displayed significantly greater numbers of strategy orchestration patterns. In addition, Grade 4 children more frequently used overt encoding and self-checking strategies. Further, evidence suggests that Grade 4 children used language to encode movement components or to regulate their own learning more frequently than Grade 1 children.

A microwave oven has been specifically modified to permit rapid thawing of fresh-frozen plasma (FFP) by using a rotating disc with a temperature sensor to hold the plasma bag. This modification makes it possible to mix the FFP continuously during thawing, and automatically shuts the oven off when the plasma reaches 21 degrees C. Comparisons were made between FFP thawed in the modified microwave oven and FFP thawed conventionally in a 37 degrees C waterbath. The following tests were done: total protein, albumin, and immunoglobulin concentrations; plasma fibrinogen, factor VIII, and factor IX activities; protein electrophoresis, albumin aggregation, hemolytic complement activity, and plasma particle count and size. In no case was there a significant difference between plasma thawed in the microwave oven compared with that thawed in the waterbath. Further, microwave thawing was reliable and rapid; all units of FFP thawed in less than 6 minutes, and the thawed plasma did not vary by more than 6 degrees C from the preselected final temperature of 21 degrees C. Thus, it appears that controlled thawing of FFP in a microwave oven specifically designed for this purpose is an effective and reliable method and has many advantages over conventional thawing of FFP.