The influence of metal ions on native silk rheology

Whilst flow is the basis for silk fibre formation, subtle changes in a silk feedstocks’ chemical environment may serve to increase both energetic efficiency and control hierarchical structure development during spinning. Despite the role of pH being largely understood, the influence of metal ions is not, only being inferred by correlative work and observations. Through a combination of rheology and microscopy, we provide a causative study of how the most abundant metal ions in the silk feedstock, Ca2+ and K+, affect its flow properties and structure. Our results show that Ca2+ ions increase viscosity and prevent molecular alignment and aggregation, providing ideal storage conditions for unspun silk. In contrast, the addition of K+ ions promotes molecular alignment and aggregation and therefore seems to transfer the silk feedstock into a spinning state which confirms recent ‘sticky reptation’ modelling hypotheses. Additionally, we characterised the influence of the ubiquitous kosmotropic agent Li+, used to prepare regenerated silk solutions, and find that it promotes molecular alignment and prevents aggregation which may permit a range of interesting artificial silk processing techniques to be developed. In summary, our results provide a clearer picture of how metal ions co-ordinate, control and thus contribute towards silk protein self-assembly which in turn can inspire structuring approaches in other biopolymer systems

Extensional flow behaviour and spinnability of native silk

Silk fibres are assembled via flow. While changes in the physiological environment of the gland as well as the shear rheology of silk are largely understood, the effect of extensional flow fields on native silk proteins is almost completely unknown. Here we demonstrate that filament stretching on a conventional tensile tester is a suitable technique to assess silk's extensional flow properties and its ability to form fibres under extensional conditions characteristic of natural spinning. We report that native Bombyx mori silk responds differently to extensional flow fields when compared to synthetic linear polymers, as evidenced by a higher Trouton ratio which we attribute to silk's increased interchain interactions. Finally, we show that native silk proteins can only be spun into stable fibres at low extension rates as a result of dehydration, suggesting that extensional fields alone are unable to induce natural fibre formation

Doppler optical frequency domain reflectometry for remote fiber sensing

Coherent optical frequency domain reflectometry has been widely used to locate static reflectors with high spatial resolution. Here, we present a new type of Doppler optical frequency domain reflectometry that offers simultaneous measurement of the position and speed of moving objects. The system is exploited to track optically levitated "flying" particles inside a hollow-core photonic crystal fiber. As an example, we demonstrate distributed temperature sensing with sub-mm-scale spatial resolution and a standard deviation of similar to 10 degrees C up to 200 degrees C. (C) 2021 Optical Society of America under the terms of the OSA Open Access Publishing Agreemen

Spinning beta silks requires both pH activation and extensional stress

Synthetic silk production has undergone significant technological and commercial advances over the past 5 years, with fibers from most labs and companies now regularly matching the properties of natural silk by one metric or another. Yet the fundamental links between silk protein processing and performance remain largely unresolved and fiber optimization is commonly achieved through non-natural methods. In an effort to address this challenge, data that closes this loop of processing and performance is presented by spinning a native silk feedstock ex vivo into a near-native fiber using just two naturally occurring parameters; pH activation and extensional flow (i.e., spinning rate). This allows us to link previous experimental and modelling hypothesis surrounding silk's pH responsiveness directly to multiscale hierarchical structure development during spinning. Finally, fibers that match, and then exceed, natural silk's mechanical properties are spun and understood by rate of work input. This approach not only provides energetic insights into natural silk spinning and controlled protein denaturation, but is believed will help interpret and improve synthetic silk processing. Ultimately, it is hoped that these results will contribute towards novel bioinspired energy-efficient processing strategies that are driven by work input optimization and where excellent mechanical properties are self-emergent

DNA methylation in human gastric epithelial cells defines regional identity without restricting lineage plasticity

BACKGROUND: Epigenetic modifications in mammalian DNA are commonly manifested by DNA methylation. In the stomach, altered DNA methylation patterns have been observed following chronic Helicobacter pylori infections and in gastric cancer. In the context of epigenetic regulation, the regional nature of the stomach has been rarely considered in detail. RESULTS: Here, we establish gastric mucosa derived primary cell cultures as a reliable source of native human epithelium. We describe the DNA methylation landscape across the phenotypically different regions of the healthy human stomach, i.e., antrum, corpus, fundus together with the corresponding transcriptomes. We show that stable regional DNA methylation differences translate to a limited extent into regulation of the transcriptomic phenotype, indicating a largely permissive epigenetic regulation. We identify a small number of transcription factors with novel region-specific activity and likely epigenetic impact in the stomach, including GATA4, IRX5, IRX2, PDX1 and CDX2. Detailed analysis of the Wnt pathway reveals differential regulation along the craniocaudal axis, which involves non-canonical Wnt signaling in determining cell fate in the proximal stomach. By extending our analysis to pre-neoplastic lesions and gastric cancers, we conclude that epigenetic dysregulation characterizes intestinal metaplasia as a founding basis for functional changes in gastric cancer. We present insights into the dynamics of DNA methylation across anatomical regions of the healthy stomach and patterns of its change in disease. Finally, our study provides a well-defined resource of regional stomach transcription and epigenetics. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s13148-022-01406-4

Predicting base editing outcomes using position-specific sequence determinants.

CRISPR/Cas base editors promise nucleotide-level control over DNA sequences, but the determinants of their activity remain incompletely understood. We measured base editing frequencies in two human cell lines for two cytosine and two adenine base editors at ∼14 000 target sequences and find that base editing activity is sequence-biased, with largest effects from nucleotides flanking the target base. Whether a base is edited depends strongly on the combination of its position in the target and the preceding base, acting to widen or narrow the effective editing window. The impact of features on editing rate depends on the position, with sequence bias efficacy mainly influencing bases away from the center of the window. We use these observations to train a machine learning model to predict editing activity per position, with accuracy ranging from 0.49 to 0.72 between editors, and with better generalization across datasets than existing tools. We demonstrate the usefulness of our model by predicting the efficacy of disease mutation correcting guides, and find that most of them suffer from more unwanted editing than pure outcomes. This work unravels the position-specificity of base editing biases and allows more efficient planning of editing campaigns in experimental and therapeutic contexts

Расчет бокового магнитного сопротивления электромагнитных молотков

Hemolysis is an inevitable side effect of cardiopulmonary bypass resulting in increased plasma free hemoglobin that may impair tissue perfusion by scavenging nitric oxide. Acute kidney injury after on-pump cardiovascular surgery arises from a number of causes and severely affects patient morbidity and mortality. Here, we studied the effect of acute hemolysis on renal injury in 35 patients undergoing on-pump surgical repair of thoracic and thoracoabdominal aortic aneurysms of whom 19 experienced acute kidney injury. During surgery, plasma free hemoglobin increased, as did urinary excretion of the tubular injury marker N-acetyl-β-D-glucosaminidase, in patients with and without acute kidney injury, reaching peak levels at 2 h and 15 min, respectively, after reperfusion. Furthermore, plasma free hemoglobin was independently and significantly correlated with the urine biomarker, which, in turn, was independently and significantly associated with the later postoperative increase in serum creatinine. Importantly, peak plasma free hemoglobin and urine N-acetyl-β-D-glucosaminidase concentrations had significant predictive value for postoperative acute kidney injury. Thus, we found an association between increased plasma free hemoglobin and renal injury casting new light on the pathophysiology of acute kidney injury. Therefore, free hemoglobin is a new therapeutic target to improve clinical outcome after on-pump cardiovascular surgery

In Vivo Gene Essentiality and Metabolism in Bordetella pertussis

Bordetella pertussis is the causative agent of whooping cough, a serious respiratory illness affecting children and adults, associated with prolonged cough and potential mortality. Whooping cough has reemerged in recent years, emphasizing a need for increased knowledge of basic mechanisms of B. pertussis growth and pathogenicity. While previous studies have provided insight into in vitro gene essentiality of this organism, very little is known about in vivo gene essentiality, a critical gap in knowledge, since B. pertussis has no previously identified environmental reservoir and is isolated from human respiratory tract samples. We hypothesize that the metabolic capabilities of B. pertussis are especially tailored to the respiratory tract and that many of the genes involved in B. pertussis metabolism would be required to establish infection in vivo. In this study, we generated a diverse library of transposon mutants and then used it to probe gene essentiality in vivo in a murine model of infection. Using the CON-ARTIST pipeline, 117 genes were identified as conditionally essential at 1 day postinfection, and 169 genes were identified as conditionally essential at 3 days postinfection. Most of the identified genes were associated with metabolism, and we utilized two existing genome-scale metabolic network reconstructions to probe the effects of individual essential genes on biomass synthesis. This analysis suggested a critical role for glucose metabolism and lipooligosaccharide biosynthesis in vivo. This is the first genome-wide evaluation of in vivo gene essentiality in B. pertussis and provides tools for future exploration. IMPORTANCE Our study describes the first in vivo transposon sequencing (Tn-seq) analysis of B. pertussis and identifies genes predicted to be essential for in vivo growth in a murine model of intranasal infection, generating key resources for future investigations into B. pertussis pathogenesis and vaccine design

Role of the CX3C chemokine receptor CX3CR1 in the pathogenesis of atherosclerosis after aortic transplantation

Background: The CX3C chemokine receptor CX3CR1 is expressed on monocytes as well as tissue resident cells, such as smooth muscle cells ( SMCs). Its role in atherosclerotic tissue remodeling of the aorta after transplantation has not been investigated. Methods: We here have orthotopically transplanted infrarenal Cx3cr1(-/-) Apoe(-/-) and Cx3cr1(+/+) Apoe(-/-)aortic segments into Apoe(-/-) mice, as well as Apoe(-/-) aortic segments into Cx3cr1(-/-) Apoe(-/-)mice. The intimal plaque size and cellular plaque composition of the transplanted aortic segment were analyzed after four weeks of atherogenic diet. Results: Transplantation of Cx3cr(-/-) Apoe(-/-) aortic segments into Apoe(-/-) mice resulted in reduced atherosclerotic plaque formation compared to plaque size in Apoe(-/-) or Cx3cr1(-/-) Apoe(-/-) mice after transplantation of Apoe(-/-) aortas. This reduction in lesion formation was associated with reduced numbers of lesional SMCs but not macrophages within the transplanted Cx3cr(-/-) Apoe(-/-) aortic segment. No differences in frequencies of proliferating and apoptotic cells could be observed. Conclusion These results indicate that CX3CR1 on resident vessel wall cells plays a key role in atherosclerotic plaque formation in transplanted aortic grafts. Targeting of vascular CX3CL1/ CX3CR1 may therefore be explored as a therapeutic option in vascular transplantation procedures

Использование жидкостей ГРП на основе дизельного топлива для низкотемпературных газовых пластов

Aim: Stroke and paraplegia are devasting complications of thoracic and thoracoabdominal aortic surgery. The aim of this study was to analyse the value of transcranial Doppler ultrasound (TCD), electroencephalogram (EEG) and motor-evoked potentials (MEP) in preventing neurological complications. Moreover, the principles, technology and surgical protocols are described. Patients and Methods: In 2009, 22 patients (4 females, 18 males) underwent thoracic or thoracoabdominal open aortic repair. We performed 2 arches with descending aortic replacement, 5 arches with TAAA repair, 2 type II, 9 type III, 3 type IV and one type V TAAA aortic repair. In 6 patients, the neuromonitoring included TCD, EEG and MEPs. In 15 patients only MEP monitoring was necessary. In one patient who was operated on in an emergency setting, neuromonitoring was not performed. The surgical approach was a left thoracotomy in 3 and a left thoracolaparotomy in 19 patients. The surgical protocol included cerebrospinal fluid drainage (n=22), moderate (n=19) or deep hypothermia (n=2), and extracorporeal circulation (n=21) with retrograde aortic perfusion and selective cerebral and/or viscerorenal perfusion. Results: In 21 patients, the neuromonitoring could be established successfully. Using TCD and EEG, a relevant cerebral ischaemia during supraaortic clamping was excluded. With a mean distal arterial pressure of 60 mmHg, the MEPs remained adequate in 15 patients (68.2%). Increasing of the blood pressure restored the MEPs in one patient. In 5 patients (22.7%), a reimplantation of segmental arteries (n=4) or of the left subclavian artery (n=1) re-established spinal cord perfusion, as indicated by restored MEPs. We had no absent MEPs at the end of the procedures. Delayed paraparesis developed in 2 patients with a haemodynamic instability during the postoperative course. Paraplegia was not observed. Conclusion: TCD, EEG and MEPs are reliable techniques to unmask cerebral or spinal cord ischaemia during aortic surgery. Immediate operative strategies based on neuromonitoring information prevent neurological complications in aortic surgery