Rapid Activation Research Articles

Central and peripheral neuromuscular fatigue following ramp and rapid maximal voluntary isometric contractions.

Maximal voluntary isometric contractions (MVICs) as a fatiguing modality have been widely studied, but little attention has been given to the influence of the rate of torque development. Given the established differences in motor command and neuromuscular activation between ramp and rapid MIVCs, it is likely performance fatigue differs as well as the underlying physiological mechanisms. To compare responses for rapid and maximal torque following ramp and rapid MVICs, and the corresponding neuromuscular and corticospinal alterations. On separate visits, twelve healthy males (22.8 ± 2.5years) performed fatiguing intermittent MVICs of the knee extensors with either 2s (RAMP) or explosive (RAPID) ramp-ups until a 50% reduction in peak torque was achieved. Before and after each condition, maximal and rapid torque measures were determined from an MVIC. Additionally, peripheral (twitch parameters) and central (voluntary activation) fatigue, as well as rapid muscle activation, and cortical-evoked twitch and electromyographic responses were recorded. Maximal and late-phase rapid torque measures (p ≤ 0.001; = 0.635-0.904) were reduced similarly, but early rapid torque capacity (0-50ms) (p = 0.003; d = 1.11 vs. p = 0.054; d = 0.62) and rapid muscle activation (p = 0.008; d = 1.07 vs. p = 0.875; d = 0.06) decreased more after RAMP. Changes in peripheral fatigue, as indicated by singlet and doublet contractile parameters (p < 0.001 for all; = 0.752-0.859), and nerve-evoked voluntary activation (p < 0.001; 0.660) were similar between conditions. Corticospinal inhibition (via silent period) was only increased after RAPID (p = 0.007; d = 0.94 vs. p = 0.753; d = 0.09), whereas corticospinal voluntary activation and excitability were unchanged. Ramp, fatiguing MVICs impaired early rapid torque capacity more than rapid MVICs, and this was accompanied by decrements in rapid muscle activation. Responses for peripheral and central fatigue (nerve and cortical stimulation) were largely similar between conditions, except that rapid MVICs increased corticospinal inhibition.

The antimicrobial peptide Temporin-L induces vesicle formation and reduces the virulence in S. aureus

The evolution of methicillin-resistant Staphylococcus aureus (MRSA) has required the development of new antimicrobial agents and new approaches to prevent and overcome drug resistance. AntiMicrobial Peptides (AMPs) represent promising alternatives due to their rapid bactericidal activity and their broad-spectrum of action against a wide range of microorganisms. The amphibian Temporins constitute a well-known family of AMPs with high antibacterial properties against both Gram-positive and Gram-negative bacteria. In this paper, we evaluated the in vivo effect of Temp-L on S. aureus performing morphological studies using Transmission Electron Microscopy (TEM) that revealed the occurrence of protrusions from the cell surface. The formation of vesicle-like structure was confirmed by Dynamic Light Scattering (DLS). The global effect of Temp-L on Staphylococcus aureus (S. aureus) was deeply investigated by differential proteomics leading to the identification of up-regulated proteins involved in the synthesis of the cell membrane and fatty acids, and down-regulated virulence factors. GC-MS analysis suggested a possible protective response mechanism implemented by the bacterium after treatment with Temp-L, as the synthesis of fatty acids was increased. Adhesion and invasion assays on eukaryotic cells confirmed a reduced virulence of S. aureus following treatment with Temp-L. These results suggested the targeting of virulence factors as novel strategy to replace traditional antimicrobial agents that can be used to treat infections, especially infections caused by the resistant pathogen S. aureus.

Probing PAC1 receptor activation across species with an engineered sensor.

Class-B1 G-protein-coupled receptors (GPCRs) are an important family of clinically relevant drug targets that remain difficult to investigate via high-throughput screening and in animal models. Here, we engineered PAClight1P78A, a novel genetically encoded sensor based on a class-B1 GPCR (the human PAC1 receptor, hmPAC1R) endowed with high dynamic range (ΔF/F0 = 1100%), excellent ligand selectivity, and rapid activation kinetics (τON = 1.15 s). To showcase the utility of this tool for in vitro applications, we thoroughly characterized and compared its expression, brightness and performance between PAClight1P78A-transfected and stably expressing cells. Demonstrating its use in animal models, we show robust expression and fluorescence responses upon exogenous ligand application ex vivo and in vivo in mice, as well as in living zebrafish larvae. Thus, the new GPCR-based sensor can be used for a wide range of applications across the life sciences empowering both basic research and drug development efforts.

The Lysine Acetyltransferase PCAF Functionally Interacts with Estrogen Receptor Alpha in the Hippocampus of Gonadally Intact Male-But Not Female-Rats to Enhance Short-Term Memory.

Acetylation of histone proteins by histone acetyltransferases (HATs), and the resultant change in gene expression, is a well-established mechanism necessary for long-term memory (LTM) consolidation, which is not required for short-term memory (STM). However, we previously demonstrated that the HAT p300/CBP-associated factor (PCAF) also influences hippocampus (HPC)-dependent STM in male rats. In addition to their epigenetic activity, HATs acetylate nonhistone proteins involved in nongenomic cellular processes, such as estrogen receptors (ERs). Given that ERs have rapid, nongenomic effects on HPC-dependent STM, we investigated the potential interaction between ERs and PCAF for STM mediated by the dorsal hippocampus (dHPC). Using a series of pharmacological agents administered directly into the dHPC, we reveal a functional interaction between PCAF and ERα in the facilitation of short-term object-in-place memory in male but not female rats. This interaction was specific to ERα, while ERβ agonism did not enhance STM. It was further specific to dHPC STM, as the effect was not present in the dHPC for LTM or in the perirhinal cortex. Further, while STM required local (i.e., dHPC) estrogen synthesis, the facilitatory interaction effect appeared independent of estrogens. Finally, western blot analyses demonstrated that PCAF activation in the dHPC rapidly (5 min) activated downstream estrogen-related cell signaling kinases (c-Jun N-terminal kinase and extracellular signal-related kinase). Collectively, these findings indicate that PCAF, which is typically implicated in LTM through epigenetic processes, also influences STM in the dHPC, possibly via nongenomic ER activity. Critically, this novel PCAF-ER interaction might exist as a male-specific mechanism supporting STM.

Brain region-specific action of ketamine as a rapid antidepressant.

Ketamine has been found to have rapid and potent antidepressant activity. However, despite the ubiquitous brain expression of its molecular target, the N-methyl-d-aspartate receptor (NMDAR), it was not clear whether there is a selective, primary site for ketamine's antidepressant action. We found that ketamine injection in depressive-like mice specifically blocks NMDARs in lateral habenular (LHb) neurons, but not in hippocampal pyramidal neurons. This regional specificity depended on the use-dependent nature of ketamine as a channel blocker, local neural activity, and the extrasynaptic reservoir pool size of NMDARs. Activating hippocampal or inactivating LHb neurons swapped their ketamine sensitivity. Conditional knockout of NMDARs in the LHb occluded ketamine's antidepressant effects and blocked the systemic ketamine-induced elevation of serotonin and brain-derived neurotrophic factor in the hippocampus. This distinction of the primary versus secondary brain target(s) of ketamine should help with the design of more precise and efficient antidepressant treatments.

Uncovering Endolysins against Methicillin-Resistant Staphylococcus aureus Using a Microbial Single-Cell Genome Database.

Endolysins, peptidoglycan hydrolases derived from bacteriophages (phages), are being developed as a promising alternative to conventional antibiotics. To obtain highly active endolysins, a diverse library of these endolysins is vital. We propose here microbial single-cell genome sequencing as an efficient tool to discover dozens of previously unknown endolysins, owing to its culture-independent sequencing method. As a proof of concept, we analyzed and recovered endolysin genes within prophage regions of Staphylococcus single-amplified genomes in human skin microbiome samples. We constructed a library of chimeric endolysins by shuffling domains of the natural endolysins and performed high-throughput screening against Staphylococcus aureus. One of the lead endolysins, bbst1027, exhibited desirable antimicrobial properties, such as rapid bactericidal activity, no detectable resistance development, and in vivo efficacy. We foresee that this endolysin discovery pipeline is in principle applicable to any bacterial target and boost the development of novel antimicrobial agents.

Robust peroxidase from Bacillus mojavensis TH309: Immobilization on walnut shell hydrochar and evaluation of its potential in dye decolorization

Peroxidases have received considerable attention as a cost-effective and environmentally friendly catalyst for bioremediation. Their rapid activity loss under harsh environmental conditions and inability to be used repetitively limit their exploitation in real-world wastewater treatment. First, a peroxidase was produced extracellularly by Bacillus mojavensis TH309 and purified 8.12-fold with a final yield of 47.10 % using Sephadex G-100 superfine resin. The pure peroxidase (BmPer) possessed a relatively low molecular weight of ∼21 kDa and was active against L-DOPA on acrylamide gel after electrophoresis. BmPer was immobilized by adsorption functionalized walnut shell hydrochar (WsH) with 61.99 ± 1.34 % efficiency and 37.07 ± 4.16 % activity loss. BmPer and its immobilized form (WsH-BmPer) exhibited maximum activity at 50 °C and pH 9. WsH-BmPer exhibited 3.23-, 2.37-, 1.65-, and 2.25-fold longer half-life than BmPer at 50, 60, 70, and 80 °C, respectively. Immobilization significantly enhanced the stability of the enzyme under acidic conditions. BmPer and WsH-BmPer showed maximal activity in the presence of 1 % salt and retained more than 85 % of their activity even after pre-incubation with 2.5 M salt for 60 min at 50 °C. Their catalytic efficiency was significantly stimulated by pre-incubation with Triton X-100 (1 mM), Tween20 (1 mM), and Mg2+ (1 and 10 mM). Immobilization strongly reduced the loss of activity caused by inhibitors including Ba2+, Hg2+, and Cu2+. Moreover, both forms of the enzyme were compatible with solvents. The Michaelis constant (Km) values of BmPer and WsH-BmPer were 0.88 and 2.66 mM for 2,4 DCP, respectively. WsH-BmPer peroxidase maintained about 82 % and 85 % of its activity when stored at 4 °C for 30 days and reused for up to 10 cycles, respectively. Furthermore, it decolorized Cibacron red (CR), Poly R-478 (PR), Remazol Brilliant Blue R (RBBR), and Methyl red (MR) dyes by 60.13 %, 91.34 %, 86.41 %, and 50.51 % within 60 min, respectively.

Valsalva maneuver pressure recovery time is prolonged following spinal cord injury with correlations to autonomically-influenced secondary complications.

This work's purpose was to quantify rapid sympathetic activation in individuals with spinal cord injury (SCI), and to identify associated correlations with symptoms of orthostatic hypotension and common autonomically mediated secondary medical complications. This work was a cross-sectional study of individuals with SCI and uninjured individuals. Symptoms of orthostatic hypotension were recorded using the Composite Autonomic Symptom Score (COMPASS)-31 and Autonomic Dysfunction following SCI (ADFSCI) survey. Histories of secondary complications of SCI were gathered. Rapid sympathetic activation was assessed using pressure recovery time of Valsalva maneuver. Stepwise multiple linear regression models identified contributions to secondary medical complication burden. In total, 48 individuals (24 with SCI, 24 uninjured) underwent testing, with symptoms of orthostatic hypotension higher in those with SCI (COMPASS-31, 3.3 versus 0.6, p < 0.01; ADFSCI, 21.2 versus. 3.2, p < 0.01). Pressure recovery time was prolonged after SCI (7.0s versus. 1.7s, p < 0.01), though poorly correlated with orthostatic symptom severity. Neurological level of injury after SCI influenced pressure recovery time, with higher injury levels associated with more prolonged time. Stepwise multiple linear regression models identified pressure recovery time as the primary explanation for variance in number of urinary tract infections (34%), histories of hospitalizations (12%), and cumulative secondary medical complication burden (24%). In all conditions except time for bowel program, pressure recovery time outperformed current clinical tools for assessing such risk. SCI is associated with impaired rapid sympathetic activation, demonstrated here by prolonged pressure recovery time. Prolonged pressure recovery time after SCI predicts higher risk for autonomically mediated secondary complications, serving as a viable index for more "autonomically complete" injury.

Characterization of the small Arabidopsis thaliana GTPase and ADP-ribosylation factor-like 2 protein TITAN 5.

Small GTPases switch between GDP- and GTP-bound states during cell signaling. The ADP-ribosylation factor (ARF) family of small GTPases is involved in vesicle trafficking. Although evolutionarily well conserved, little is known about ARF and ARF-like GTPases in plants. We characterized biochemical properties and cellular localization of the essential small ARF-like GTPase TITAN 5 (TTN5; also known as HALLIMASCH, ARL2 and ARLC1) from Arabidopsis thaliana, and two TTN5 proteins with point mutants in conserved residues, TTN5T30N and TTN5Q70L, that were expected to be unable to perform nucleotide exchange and GTP hydrolysis, respectively. TTN5 exhibited very rapid intrinsic nucleotide exchange and remarkably low GTP hydrolysis activity, functioning as a non-classical small GTPase being likely present in a GTP-loaded active form. We analyzed signals from YFP-TTN5 and HA3-TTN5 by in situ immunolocalization in Arabidopsis seedlings and through use of a transient expression system. Colocalization with endomembrane markers and pharmacological treatments suggests that TTN5 can be present at the plasma membrane and that it dynamically associates with membranes of vesicles, Golgi stacks and multivesicular bodies. Although TTN5Q70L mirrored wild-type TTN5 behavior, the TTN5T30N mutant differed in some aspects. Hence, the unusual rapid nucleotide exchange activity of TTN5 is linked with its membrane dynamics, and TTN5 likely has a role in vesicle transport within the endomembrane system.

Millisecond-scale behaviours of plankton quantified invitro and insitu using the Event-based Vision Sensor.

The Event-based Vision Sensor (EVS) is a bio-inspired sensor that captures detailed motions of objects, aiming to become the 'eyes' of machines like self-driving cars. Compared to conventional frame-based image sensors, the EVS has an extremely fast motion capture equivalent to 10,000-fps even with standard optical settings, plus high dynamic ranges for brightness and also lower consumption of memory and energy. Here, we developed 22 characteristic features for analysing the motions of aquatic particles from the EVS raw data and tested the applicability of the EVS in analysing plankton behaviour. Laboratory cultures of six species of zooplankton and phytoplankton were observed, confirming species-specific motion periodicities up to 41 Hz. We applied machine learning to automatically classify particles into four categories of zooplankton and passive particles, achieving an accuracy up to 86%. At the insitu deployment of the EVS at the bottom of Lake Biwa, several particles exhibiting distinct cumulative trajectory with periodicities in their motion (up to 16 Hz) were identified, suggesting that they were living organisms with rhythmic behaviour. We also used the EVS in the deep sea, observing particles with active motion and periodicities over 40 Hz. Our application of the EVS, especially focusing on its millisecond-scale temporal resolution and wide dynamic range, provides a new avenue to investigate organismal behaviour characterised by rapid and periodical motions. The EVS will likely be applicable in the near future for the automated monitoring of plankton behaviour by edge computing on autonomous floats, as well as quantifying rapid cellular-level activities under microscopy.

Cyclin-dependent kinase 4 drives cystic kidney disease in the absence of mTORC1 signaling activity

Progression of cystic kidney disease has been linked to activation of the mTORC1 signaling pathway. Yet the utility of mTORC1 inhibitors to treat patients with polycystic kidney disease remains controversial despite promising preclinical data. To define the cell intrinsic role of mTORC1 for cyst development, the mTORC1 subunit gene Raptor was selectively inactivated in kidney tubular cells lacking cilia due to simultaneous deletion of the kinesin family member gene Kif3A. In contrast to a rapid onset of cyst formation and kidney failure in mice with defective ciliogenesis, both kidney function, cyst formation discerned by magnetic resonance imaging and overall survival were strikingly improved in mice additionally lacking Raptor. However, these mice eventually succumbed to cystic kidney disease despite mTORC1 inactivation. In-depth transcriptome analysis revealed the rapid activation of other growth-promoting signaling pathways, overriding the effects of mTORC1 deletion and identified cyclin-dependent kinase (CDK) 4 as an alternate driver of cyst growth. Additional inhibition of CDK4-dependent signaling by the CDK4/6 inhibitor Palbociclib markedly slowed disease progression in mice and human organoid models of polycystic kidney disease and potentiated the effects of mTORC1 deletion/inhibition. Our findings indicate that cystic kidneys rapidly adopt bypass mechanisms typically observed in drug resistant cancers. Thus, future clinical trials need to consider combinatorial or sequential therapies to improve therapeutic efficacy in patients with cystic kidney disease.

A rapid analysis of aftershock processes after a moderate magnitude earthquake with ML methods

SUMMARY Moderate magnitude earthquakes and seismic sequences frequently develop on fault systems, but whether they are linked to future major ruptures is always ambiguous. In this study, we investigated a seismic sequence that has developed within a portion of the stretching region of the Apennines in Italy where moderate to large earthquakes are likely to occur. We captured a total of 2039 aftershocks of the 2023 September 18, Mw 4.9 earthquake occurred during the first week, by using machine-learning (ML) based algorithms. Aftershocks align on two 5–7 km long parallel faults, from a length that exceeds what is expected from the main shock magnitude. The segments are ramping at about 6 km depth on closely spaced N100 striking 70 N dipping planes, at a distance of some kilometres from the main shock hypocentre. Our results indicate that even moderate magnitude events trigger seismicity on a spread set of fault segments around the main shock hypocentre, revealing processes of interaction within the crustal layer. The possibility that larger earthquakes develop during seismicity spread is favoured by pore pressure diffusion, in relation with the closeness to criticality of fault segments. Based on the very rapid activation of seismicity on the entire system and a back-front signal from the hypocentre of the main event, we infer that fluid pressure, initially high within the crustal layer, rapidly dropped after the main shock. Our study reinforces the importance of timely extracting information on fault geometry and seismicity distribution on faults. ML-based methods represent a viable tool for semi-real-time application, yielding constraints on short-time forecasts.

Event-triggered semi-active TLCD for ground motion-induced vibration control

One potential drawback of tuned liquid column dampers (TLCDs) is their relatively low control efficiency during the initial stage of structural vibration caused by external excitations. This is because satisfactory control effects can only be achieved when the liquid inside TLCD is fully oscillating, which is not the case during the initial stage. To solve this problem, in this study, an event-triggered semi-active technique is creatively proposed to improve the vibration reduction efficiency of TLCDs during the initial stage. The fundamental idea of the proposed approach is to provide an initial displacement to the liquid column via baffles, and then release the constraints on the initial liquid displacement at an appropriate time to achieve the rapid activation of TLCDs. A strategy from the standpoint of phase difference between liquid column motion and structural motion is proposed to determine the triggering conditions (i.e. when to release the constraints). The effectiveness of the proposed semi-active system is examined under both harmonic and stochastic excitations. The results show that the proposed strategy successfully improves the vibration suppression performance of TLCDs in the early stage of structural vibration.

Ca2+ transients on the T cell surface trigger rapid integrin activation in a timescale of seconds

One question in lymphocyte homing is how integrins are rapidly activated to enable immediate arrest of fast rolling lymphocytes upon encountering chemokines at target vascular beds given the slow chemokine-induced integrin inside-out activation. Herein we demonstrate that chemokine CCL25-triggered Ca2+ influx induces T cell membrane-proximal external Ca2+ concentration ([Ca2+]ex) drop in 6 s from physiological concentration 1.2 mM to 0.3 mM, a critical extracellular Ca2+ threshold for inducing αLβ2 activation, triggering rapid αLβ2 activation and T cell arrest before occurrence of αLβ2 inside-out activation. Talin knockdown inhibits the slow inside-out activation of αLβ2 but not [Ca2+]ex drop-triggered αLβ2 quick activation. Blocking Ca2+ influx significantly suppresses T cell rolling-to-arrest transition and homing to skin lesions in a mouse psoriasis model, thus alleviating skin inflammation. [Ca2+]ex decrease-triggered rapid integrin activation bridges the gap between initial chemokine stimulation and slow integrin inside-out activation, ensuring immediate lymphocyte arrest and subsequent diapedesis on the right location.

Validation of a rapid collagenase activity detection technique based on fluorescent quenched gelatin with synovial fluid samples

BackgroundMeasuring collagenase activity is crucial in the field of joint health and disease management. Collagenases, enzymes responsible for collagen degradation, play a vital role in maintaining the balance between collagen synthesis and breakdown in joints. Dysregulation of collagenase activity leads to joint tissue degradation and diseases such as rheumatoid arthritis and osteoarthritis. The development of methods to measure collagenase activity is essential for diagnosis, disease severity assessment, treatment monitoring, and identification of therapeutic targets.ResultsThis study aimed to validate a rapid collagenase activity detection technique using synovial fluid samples. Antibody microarray analysis was initially performed to quantify the levels of matrix metalloproteinase-9 (MMP-9), a major collagenase in joints. Subsequently, the developed gelatin-based test utilizing fluorescence measurement was used to determine collagenase activity. There was a significant correlation between the presence of MMP-9 and collagenase activity. In addition, Lower Limit of Detection and Upper Limit of Detection can be preliminary estimated as 8 ng/mL and 48 ng/mL respectively.ConclusionsThe developed technique offers a potential point-of-care assessment of collagenase activity, providing real-time information for clinicians and researchers. By accurately quantifying collagenase activity, healthcare professionals can optimize patient care, improve treatment outcomes, and contribute to the understanding and management of joint-related disorders. Further research and validation are necessary to establish the full potential of this rapid collagenase activity detection method in clinical practice.

In situ fabrication of oxygen deficient Bi2MoO6/InVO4/CeVO4 dual S-scheme ternary heterostructure for robust photocatalytic H2 and H2O2 production

Mild in situ synthesis of ternary dual S-scheme heterostructure photocatalyst with high redox ability of photo-excitons is a pragmatic approach for achieving rapid activation of tenacious atmospheric molecules. In the present study, a series of Bi2MoO6/InVO4/CeVO4 ternary heterostructures were constructed by in situ deposition of Bi2MoO6 nanoplates over one pot synthesised InVO4/CeVO4 using a facile oil bath heating method. The as-synthesised heterostructure materials were comprehensively characterised to understand their structural, optical, electrochemical and morphological properties. The ternary heterostructure displayed a distinct morphology consisting of Bi2MoO6 nanoplates, CeVO4 nanosheets and InVO4 nanorods. The significant intergrowth among the constituent phases through the in situ coupling strategy also led to the construction of tight interfacial microscopic junctions. The important attributes of the ternary heterostructures included intense photon absorption in whole UV–visible region, drastic decrease in photogenerated charge recombination and higher excited state lifetime. Both InVO4 and CeVO4 individually as well as the ternary heterostructure contained surface oxygen vacancies that further promoted efficient charge separation and activation of atmospheric molecules. The optimised ternary photocatalyst displayed 2314 μmol/g/h H2 generation and 1700 μM/g/h H2O2 production which are 12–86 and 11–27 times higher than the pristine semiconductors, respectively. Detailed band position assessment and radical entrapment analysis suggested the occurrence of a dual S-scheme charge transfer mechanism that rationally accounted for the remarkable enhancement in the photocatalytic performance.

Coupling Cu Coordination Polymers with CdS Forming an S-Scheme Heterojunction for Rapid Charge Separation and High Photocatalytic Activity.

Energy and the environment are significant impacting factors for the future development of humankind. In order to improve the corrosion resistance of CdS and decrease the recombination of photogenerated carriers, a novel Cu-CPs@CdS heterojunction with high efficiency mesopores was constructed by a simple hydrothermal method. The effective interfacial contact formation between nano-CdS and Cu-CPs promotes the transfer of photogenerated carriers while exhibiting a high spatial separation rate of charges. The photocatalytic performance of the heterojunction was evaluated by the photocatalytic degradation of organic pollutants and photocatalytic hydrogen generation. The photocatalytic degradation of ciprofloxacin (CIP) could reach 90.34%, and the hydrogen generation was high as 9227.82 μmol·g-1 under simulated sunlight irradiation. The boosted photocatalytic activity of Cu-CPs@CdS results from (i) the formation of coordination bonds, which not only enhanced the stability of heterojunctions but also provided a path for photogenerated carrier migration, (ii) integrating Cu-CPs, which provided more active sites, and (iii) the matched energy band structure between CdS and Cu-CPs that promoted speedy S-scheme interfacial charge-transfer pathways, culminating in efficient photogenerated charge separation and transfer. This research offered a fresh tactic to restrict photocorrosion and enhance the production of photocatalytic H2 over CdS-based catalysts.

Rapid functional activation of horizontally transferred eukaryotic intron-containing genes in the bacterial recipient.

Horizontal gene transfer has occurred across all domains of life and contributed substantially to the evolution of both prokaryotes and eukaryotes. Previous studies suggest that many horizontally transferred eukaryotic genes conferred selective advantages to bacterial recipients, but how these eukaryotic genes evolved into functional bacterial genes remained unclear, particularly how bacteria overcome the expressional barrier posed by eukaryotic introns. Here, we first confirmed that the presence of intron would inactivate the horizontally transferred gene in Escherichiacoli even if this gene could be efficiently transcribed. Subsequent large-scale genetic screens for activation of gene function revealed that activation events could rapidly occur within several days of selective cultivation. Molecular analysis of activation events uncovered two distinct mechanisms how bacteria overcome the intron barrier: (i) intron was partially deleted and the resulting stop codon-removed mutation led to one intact foreign protein or (ii) intron was intactly retained but it mediated the translation initiation and the interaction of two split small proteins (derived from coding sequences up- and downstream of intron, respectively) to restore gene function. Our findings underscore the likelihood that horizontally transferred eukaryotic intron-containing genes could rapidly acquire functionality if they confer a selective advantage to the prokaryotic recipient.

Exploring Reduced Graphene Oxide Sheets Stabilized by Cu(II) and Cu(I) Cations in Ethanol

In this study, ultrasound treatment was used to exfoliate commercially available graphite flakes into reduced graphene oxide (rGO) dispersed in ethanol. After centrifugation, solid copper chloride trihydrate was added, resulting in a green liquor containing Cu(II), Cu(I), and rGO. These liquors exhibited good and rapid photocatalytic activity in the degradation of eosin and bromophenol blue dyes (elimination in a few seconds) under visible-light irradiation. UV–visible spectroscopy confirmed the presence of rGO and Cu species. The size and morphology of the rGO sheets were investigated by several methods (SAXS, wide-angle XRD, SEM, and TEM). Negative UV peaks indicated light emission, which was independently verified by fluorescence. Intense plasmon peaks, with absorbances greater than 10, were observed after adding copper chloride salt. These plasmons were eliminated by a high dilution before the described catalytic tests were performed.

Intermittent cyclic stretch of engineered ligaments drives hierarchical collagen fiber maturation in a dose- and organizational-dependent manner

Hierarchical collagen fibers are the primary source of strength in tendons and ligaments; however, these fibers largely do not regenerate after injury or with repair, resulting in limited treatment options. We previously developed a static culture system that guides ACL fibroblasts to produce native-sized fibers and early fascicles by 6 weeks. These constructs are promising ligament replacements, but further maturation is needed. Mechanical cues are critical for development in vivo and in engineered tissues; however, the effect on larger fiber and fascicle formation is largely unknown. Our objective was to investigate whether intermittent cyclic stretch, mimicking rapid muscle activity, drives further maturation in our system to create stronger engineered replacements and to explore whether cyclic loading has differential effects on cells at different degrees of collagen organization to better inform engineered tissue maturation protocols. Constructs were loaded with an established intermittent cyclic loading regime at 5 or 10 % strain for up to 6 weeks and compared to static controls. Cyclic loading drove cells to increase hierarchical collagen organization, collagen crimp, and tissue tensile properties, ultimately producing constructs that matched or exceeded immature ACL properties. Further, the effect of loading on cells varied depending on degree of organization. Specifically, 10 % load drove early improvements in tensile properties and composition, while 5 % load was more beneficial later in culture, suggesting a shift in mechanotransduction. This study provides new insight into how cyclic loading affects cell-driven hierarchical fiber formation and maturation, which will help to develop better rehabilitation protocols and engineer stronger replacements. Statement of significanceCollagen fibers are the primary source of strength and function in tendons and ligaments throughout the body. These fibers have limited regenerate after injury, with repair, and in engineered replacements, reducing treatment options. Cyclic load has been shown to improve fibril level alignment, but its effect at the larger fiber and fascicle length-scale is largely unknown. Here, we demonstrate intermittent cyclic loading increases cell-driven hierarchical fiber formation and tissue mechanics, producing engineered replacements with similar organization and mechanics as immature ACLs. This study provides new insight into how cyclic loading affects cell-driven fiber maturation. A better understanding of how mechanical cues regulate fiber formation will help to develop better engineered replacements and rehabilitation protocols to drive repair after injury.