All key chemical transformations in biology are catalysed by linear oligomers. Catalytic properties could be programmed into synthetic oligomers in the same way as they are programmed into proteins, ...and an example of the discovery of emergent catalytic properties in a synthetic oligomer is reported. Dynamic combinatorial chemistry experiments designed to study the templating of a recognition-encoded oligomer by the complementary sequence have uncovered an unexpected imine polymerase activity. Libraries of equilibrating imines were formed by coupling diamine linkers with monomer building blocks composed of dialdehydes functionalised with either a trifluoromethyl phenol (
D
) or phosphine oxide (
A
) H-bond recognition unit. However, addition of the
AAA
trimer to a mixture of the phenol dialdehyde and the diamine linker did not template the formation of the
DDD
oligo-imine. Instead,
AAA
was found to be a catalyst, leading to rapid formation of long oligomers of
D
.
AAA
catalysed a number of different imine formation reactions, but a complementary phenol recognition group on the aldehyde reaction partner is an essential requirement. Competitive inhibition by an unreactive phenol confirmed the role of H-bonding in substrate recognition.
AAA
accelerates the rate of imine formation in toluene by a factor of 20. The kinetic parameters for this enzyme-like catalysis are estimated as 1 × 10
−3
s
−1
for
k
cat
and the dissociation constant for substrate binding is 300 μM. The corresponding
DDD
trimer was found to catalyse oligomerisation the phosphine oxide dialdehyde with the diamine linker, suggesting an important role for the backbone in catalysis. This unexpected imine polymerase activity in a duplex-forming synthetic oligomer suggests that there are many interesting processes to be discovered in the chemistry of synthetic recognition-encoded oligomers that will parallel those found in natural biopolymers.
Enzyme-like catalytic properties have been found in synthetic recognition-encoded oligomers.
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Template-directed synthesis of nucleic acids in the polymerase chain reaction is based on the use of a primer, which is elongated in the replication process. The attachment of a high affinity primer ...to the end of a template chain has been implemented for templating the synthesis of triazole oligomers. A covalent ester base-pair was used to attach a primer to a mixed sequence template. The resulting primed template has phenol recognition units on the template, which can form noncovalent base-pairs with phosphine oxide monomers via H-bonding, and an alkyne group on the primer, which can react with the azide group on a phosphine oxide monomer. Competition reactions between azides bearing phosphine oxide and phenol recognition groups were used to demonstrate a substantial template effect, due to H-bonding interactions between the phenols on the template and phosphine oxides on the azide. The largest rate acceleration was observed when a phosphine oxide 2-mer was used, because this compound binds to the template with a higher affinity than compounds that can only make one H-bond. The 31P NMR spectrum of the product duplex shows that the H-bonds responsible for the template effect are present in the product, and this result indicates that the covalent ester base-pairs and noncovalent H-bonded base-pairs developed here are geometrically compatible. Following the templated reaction, it is possible to regenerate the template and liberate the copy strand by hydrolysis of the ester base-pair used to attach the primer, thus completing a formal replication cycle.
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The presence of amyloid fibrils is a characteristic feature of many diseases, most famously neurodegenerative disease. The supramolecular structure of these fibrils appears to be disease-specific. ...Identifying the unique morphologies of amyloid fibrils could, therefore, form the basis of a diagnostic tool. Here we report a method to characterize the morphology of α-synuclein (αSyn) fibrils based on profiling multiple different ligand binding sites that are present on the surfaces of fibrils. By employing various competition binding assays, seven different types of binding sites were identified on four different morphologies of αSyn fibrils. Similar binding sites on different fibrils were shown to bind ligands with significantly different affinities. We combined this information to construct individual profiles for different αSyn fibrils based on the distribution of binding sites and ligand interactions. These results demonstrate that ligand-based profiling can be used as an analytical method to characterize fibril morphologies with operationally simple fluorescence binding assays.
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The identification of T-bet as a key transcription factor associated with the development of IFNγ-producing CD4
T cells predicted a crucial role for T-bet in cell-mediated immunity and in resistance ...to many intracellular infections. This idea was reinforced by initial reports showing that T-bet-deficient mice were more susceptible to pathogens that survived within the lysosomal system of macrophages. However, subsequent studies revealed IFNγ-dependent, T-bet-independent pathways of resistance to diverse classes of microorganisms that occupy other intracellular niches. Consequently, a more complex picture has emerged of how T-bet and the related transcription factor eomesodermin (EOMES) coordinate many facets of the immune response to bona fide pathogens as well as commensals. This article provides an overview of the discovery and evolutionary relationship between T-bet and EOMES and highlights the studies that have uncovered broader functions of T-bet in innate and adaptive immunity and in the development of the effector and memory T cell populations that mediate long-term resistance to infection.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Size and shape criteria for guest binding inside the cavity of an octanuclear cubic coordination cage in water have been established using a new fluorescence displacement assay to quantify guest ...binding. For aliphatic cyclic ketones of increasing size (from C5 to C11), there is a linear relationship between ΔG for guest binding and the guest’s surface area: the change in ΔG for binding is 0.3 kJ mol–1 Å–2, corresponding to 5 kJ mol–1 for each additional CH2 group in the guest, in good agreement with expectations based on hydrophobic desolvation. The highest association constant is K = 1.2 × 106 M–1 for cycloundecanone, whose volume is approximately 50% of the cavity volume; for larger C12 and C13 cyclic ketones, the association constant progressively decreases as the guests become too large. For a series of C10 aliphatic ketones differing in shape but not size, ΔG for guest binding showed no correlation with surface area. These guests are close to the volume limit of the cavity (cf. Rebek’s 55% rule), so the association constant is sensitive to shape complementarity, with small changes in guest structure resulting in large changes in binding affinity. The most flexible members of this series (linear aliphatic ketones) did not bind, whereas the more preorganized cyclic ketones all have association constants of 104–105 M–1. A crystal structure of the cage·cycloundecanone complex shows that the guest carbonyl oxygen is directed into a binding pocket defined by a convergent set of CH groups, which act as weak hydrogen-bond donors, and also shows close contacts between the exterior surface of the disc-shaped guest and the interior surface of the pseudospherical cage cavity despite the slight mismatch in shape.
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European and North American strains of the parasite Toxoplasma gondii belong to three distinct clonal lineages, type I, type II, and type III, which differ in virulence. Understanding the basis of ...Toxoplasma strain differences and how secreted effectors work to achieve chronic infection is a major goal of current research. Here we show that type I and III infected macrophages, a cell type required for host immunity to Toxoplasma, are alternatively activated, while type II infected macrophages are classically activated. The Toxoplasma rhoptry kinase ROP16, which activates STAT6, is responsible for alternative activation. The Toxoplasma dense granule protein GRA15, which activates NF-κB, promotes classical activation by type II parasites. These effectors antagonistically regulate many of the same genes, and mice infected with type II parasites expressing type I ROP16 are protected against Toxoplasma-induced ileitis. Thus, polymorphisms in determinants that modulate macrophage activation influence the ability of Toxoplasma to establish a chronic infection.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Spinal cord stimulation has been an established treatment for chronic back and leg pain for more than 50 years; however, outcomes are variable and unpredictable, and objective evidence of the ...mechanism of action is needed. A novel spinal cord stimulation system provides the first in vivo, real-time, continuous objective measure of spinal cord activation in response to therapy via recorded evoked compound action potentials (ECAPs) in patients during daily use. These ECAPs are also used to optimise programming and deliver closed-loop spinal cord stimulation by adjusting the stimulation current to maintain activation within patients' therapeutic window. We aimed to examine pain relief and the extent of spinal cord activation with ECAP-controlled closed-loop versus fixed-output, open-loop spinal cord stimulation for the treatment of chronic back and leg pain.
This multicentre, double-blind, parallel-arm, randomised controlled trial was done at 13 specialist clinics, academic centres, and hospitals in the USA. Patients with chronic, intractable pain of the back and legs (Visual Analog Scale VAS pain score ≥60 mm; Oswestry Disability Index ODI score 41–80) who were refractory to conservative therapy, on stable pain medications, had no previous experience with spinal cord stimulation, and were appropriate candidates for a spinal cord stimulation trial were screened. Eligible patients were randomly assigned (1:1) to receive ECAP-controlled closed-loop spinal cord stimulation (investigational group) or fixed-output, open-loop spinal cord stimulation (control group). The randomisation sequence was computer generated with permuted blocks of size 4 and 6 and stratified by site. Patients, investigators, and site staff were masked to the treatment assignment. The primary outcome was the proportion of patients with a reduction of 50% or more in overall back and leg pain with no increase in pain medications. Non-inferiority (δ=10%) followed by superiority were tested in the intention-to-treat population at 3 months (primary analysis) and 12 months (additional prespecified analysis) after the permanent implant. This study is registered with ClinicalTrials.gov, NCT02924129, and is ongoing.
Between Feb 21, 2017, and Feb 20, 2018, 134 patients were enrolled and randomly assigned (67 to each treatment group). The intention-to-treat analysis comprised 125 patients at 3 months (62 in the closed-loop group and 63 in the open-loop group) and 118 patients at 12 months (59 in the closed-loop group and 59 in the open-loop group). The primary outcome was achieved in a greater proportion of patients in the closed-loop group than in the open-loop group at 3 months (51 82·3% of 62 patients vs 38 60·3% of 63 patients; difference 21·9%, 95% CI 6·6–37·3; p=0·0052) and at 12 months (49 83·1% of 59 patients vs 36 61·0% of 59 patients; difference 22·0%, 6·3–37·7; p=0·0060). We observed no differences in safety profiles between the two groups. The most frequently reported study-related adverse events in both groups were lead migration (nine 7% patients), implantable pulse generator pocket pain (five 4%), and muscle spasm or cramp (three 2%).
ECAP-controlled closed-loop stimulation provided significantly greater and more clinically meaningful pain relief up to 12 months than open-loop spinal cord stimulation. Greater spinal cord activation seen in the closed-loop group suggests a mechanistic explanation for the superior results, which aligns with the putative mechanism of action for spinal cord stimulation and warrants further investigation.
Saluda Medical.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
IMPORTANCE: Given the current climate of outcomes-driven quality reporting, it is critical to appropriately risk stratify patients using standardized metrics. OBJECTIVE: To elucidate the risk ...associated with urgent surgery on complications and mortality after general surgical procedures. DESIGN, SETTING, AND PARTICIPANTS: This retrospective review used the American College of Surgeons National Surgery Quality Improvement Program database to capture all general surgery cases performed at 435 hospitals nationwide between January 1, 2013, and December 31, 2013. Data analysis was performed from November 11, 2015, to February 16, 2017. EXPOSURES: Any operations coded as both nonelective and nonemergency were designated into a novel category titled urgent. MAIN OUTCOMES AND MEASURES: The primary outcome was 30-day mortality; secondary outcomes included 30-day rates of complications, reoperation, and readmission in urgent cases compared with both elective and emergency cases. RESULTS: Of 173 643 patients undergoing general surgery (101 632 females and 72 011 males), 130 235 (75.0%) were categorized as elective, 22 592 (13.0%) as emergency, and 20 816 (12.0%) as nonelective and nonemergency. When controlling for standard American College of Surgeons National Surgery Quality Improvement Program preoperative risk factors, with elective surgery as the reference value, the 3 groups had significantly distinct odds ratios (ORs) of experiencing any complication (urgent surgery: OR, 1.38; 95% CI, 1.30-1.45; P < .001; and emergency surgery: OR, 1.65; 95% CI, 1.55-1.76; P < .001) and of mortality (urgent surgery: OR, 2.32; 95% CI, 2.00-2.68; P < .001; and emergency surgery: OR, 2.91; 95% CI, 2.48-3.41; P < .001). Surgical procedures performed urgently had a 12.3% rate of morbidity (n = 2560) and a 2.3% rate of mortality (n = 471). CONCLUSIONS AND RELEVANCE: This study highlights the need for improved risk stratification on the basis of urgency because operations performed urgently have distinct rates of morbidity and mortality compared with procedures performed either electively or emergently. Because we tie quality outcomes to reimbursement, such a category should improve predictive models and more accurately reflect the quality and value of care provided by surgeons who do not have traditional elective practices.
Salt bridges were used to attach polymerisable amidine monomers to an oligomeric benzoic acid template. CuAAC oligomerisation reactions in the presence of a benzoic acid 3-mer template gave the ...amidine 3-mer copy as the major product. Cleavage of ester linkers was used to hydrolyse off the amidine recognition units and convert the product into a benzoic acid 3-mer copy of the original template.
An oligomer equipped with benzoic acid recognition units templates the polymerization of monomers with amidine recognition units, but cleavage of ester linkers in the resulting copy transforms the amidine recognition units into benzoic acids.
Chronic hepatic diseases, such as cirrhosis, hepatocellular carcinoma, and virus‐mediated immunopathogenic infections, affect billions of people worldwide. These diseases commonly initiate with ...fibrosis. Owing to the various side effects of antifibrotic therapy and the difficulty of diagnosing asymptomatic patients, suitable medication remains a major concern. To overcome this drawback, the use of cytokine‐based sustained therapy might be a suitable alternative with minimal side effects. Here, we studied the therapeutic efficacy and potential mechanisms of interleukin (IL)−30 as antifibrosis therapy in murine liver fibrosis models. CCl4 or 3,5‐diethoxycarbonyl‐1,4‐dihydrocollidine (DDC) 0.1% (wt/wt) Purina 5015 Chow (LabDiet, St. Louis, MO) was fed for 3 weeks to induce liver fibrosis. Either control vector (pCtr) or pIL30 was injected hydrodynamically once per week. A significant decrease in collagen deposition and reduced expression of alpha‐smooth muscle actin (α‐SMA) protein indicated that IL‐30‐based gene therapy dramatically reduced bridging fibrosis that was induced by CCl4 or DDC. Immunophenotyping and knockout studies showed that IL‐30 recruits natural‐killer–like T (NKT) cells to the liver to remove activated hepatic stellate cells (HSCs) significantly and ameliorate liver fibrosis. Both flow cytometric and antibody‐mediated neutralization studies showed that liver NKT cells up‐regulate the natural killer group 2, member D (NKG2D) ligand and bind with the NKG2D ligand, retinoic acid early inducible 1 (Rae1), and positively activated HSCs to ameliorate liver fibrosis. Furthermore, adoptive transfer of liver NKT cells in T‐cell‐deficient mice showed reduction of fibrosis upon IL‐30 administration. Conclusions: Highly target‐specific liver NKT cells selectively remove activated HSCs through an NKG2D‐Rae1 interaction to ameliorate liver fibrosis after IL‐30 treatment. (Hepatology 2014;60:2026–2038)
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