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Translocations involving FGFR2 gene fusions are common in cholangiocarcinoma and predict response to FGFR kinase inhibitors. However, response rates and durability are limited due to the emergence of resistance, typically involving FGFR2 kinase domain mutations, and to sub-optimal dosing, relating to drug adverse effects. Here, we develop biparatopic antibodies targeting the FGFR2 extracellular domain (ECD), as candidate therapeutics. Biparatopic antibodies can overcome drawbacks of bivalent monospecific antibodies, which often show poor inhibitory or even agonist activity against oncogenic receptors. We show that oncogenic transformation by FGFR2 fusions requires an intact ECD. Moreover, by systematically generating biparatopic antibodies targeting distinct epitope pairs in FGFR2 ECD, we identified antibodies that effectively block signaling and malignant growth driven by FGFR2-fusions. Importantly, these antibodies demonstrate efficacy in vivo, synergy with FGFR inhibitors, and activity against FGFR2 fusions harboring kinase domain mutations. Thus, biparatopic antibodies may serve as an innovative treatment option for patients with FGFR2-altered cholangiocarcinoma.

This paper presents the results of a study on the formation of nanostructures of electrochemical titanium oxide for neuromorphic applications. Three anodization synthesis techniques were considered to allow the formation of structures with different sizes and productivity: nanodot, lateral, and imprint. The mathematical model allowed us to calculate the processes of oxygen ion transfer to the reaction zone; the growth of the nanostructure due to the oxidation of the titanium film; and the formation of TiO, Ti 2 O 3 , and TiO 2 oxides in the volume of the growing nanostructure and the redistribution of oxygen vacancies and conduction channel. Modeling of the nanodot structure synthesis process showed that at the initial stages of growth, a conductivity channel was formed, connecting the top and bottom of the nanostructure, which became thinner over time; at approximately 640 ms, this channel broke into upper and lower nuclei, after which the upper part disappeared. Modeling of the lateral nanostructure synthesis process showed that at the initial stages of growth, a conductivity channel was also formed, which quickly disappeared and left a nucleus that moved after the moving AFM tip. The simulation of the imprint nanostructure synthesis process showed the formation of two conductivity channels at a distance corresponding to the dimensions of the template tip. After about 460 ms, both channels broke, leaving behind embryos. The nanodot, lateral, and imprint nanostructure XPS spectra confirmed the theoretical calculations presented earlier: in the near-surface layers, the TiO 2 oxide was observed, with the subsequent titanium oxide nanostructure surface etching proportion of TiO 2 decreasing, and proportions of Ti 2 O 3 and TiO oxides increasing. All nanodot, lateral, and imprint nanostructures showed reproducible resistive switching over 1000 switching cycles and holding their state for 10,000 s at read operation.

Approximately 3.4 million patients worldwide are diagnosed each year with cancers that have pathogenic mutations in one of three RAS proto-oncogenes (KRAS, NRAS and HRAS) 1,2 . These mutations impair the GTPase activity of RAS, leading to activation of downstream signalling and proliferation 3-6 . Long-standing efforts to restore the hydrolase activity of RAS mutants have been unsuccessful, extinguishing any consideration towards a viable therapeutic strategy 7 . Here we show that tri-complex inhibitors-that is, molecular glues with the ability to recruit cyclophilin A (CYPA) to the active state of RAS-have a dual mechanism of action: not only do they prevent activated RAS from binding to its effectors, but they also stimulate GTP hydrolysis. Drug-bound CYPA complexes modulate residues in the switch II motif of RAS to coordinate the nucleophilic attack on the γ-phosphate of GTP in a mutation-specific manner. RAS mutants that were most sensitive to stimulation of GTPase activity were more susceptible to treatment than mutants in which the hydrolysis could not be enhanced, suggesting that pharmacological stimulation of hydrolysis potentiates the therapeutic effects of tri-complex inhibitors for specific RAS mutants. This study lays the foundation for developing a class of therapeutics that inhibit cancer growth by stimulating mutant GTPase activity., Competing Interests: Competing interests: P.L. is listed as an inventor on patents filed by MSKCC regarding treatment of KRAS- or BRAF-mutant cancers; reports grants to his institution from Revolution Medicines, Amgen, Mirati and Boehringer Ingelheim; and reports consulting fees or honoraria from Black Diamond Therapeutics, AmMax, OrbiMed, PAQ-Tx, Repare Therapeutics, Boehringer Ingelheim, Menarini Group and Revolution Medicines, as well as membership on the scientific advisory board of Frontier Medicines, Ikena, Biotheryx and PAQ-Tx (consulting fees and equity in each). The other authors declare no competing interests., (© 2024. The Author(s).)

Gestational diabetes mellitus (GDM) affects around 10% of pregnancies in the United States and has been linked to neurodevelopmental sequelae in children. However, there is a paucity of studies investigating early-life neural markers in GDM-exposed infants. This study examined the association of GDM with relative EEG power among healthy term-age neonates collected during natural sleep. Participants included a diverse cohort of 101 mothers (45% multiracial, 25% Black, and 69% Hispanic or Latina) and their infants (gestational age at birth M age = 39.0 ± 0.95; 46.5% female). We did not observe the main effect of GDM on infant relative EEG power. Our post hoc analyses revealed a significant interaction effect between GDM and infant birth weight on relative EEG power in active sleep. Among GDM-exposed neonates, increased birth weight was associated with increased relative theta EEG power and decreased relative beta and gamma EEG power across multiple electrode regions. Among non-GDM-exposed infants, increased birth weight was associated with decreased relative theta EEG power and increased relative beta and gamma EEG power across multiple electrode regions. Our findings suggest that alterations in fetal growth may serve as either an indirect marker or pathway through which GDM influences the developing fetal brain., (© 2024 Wiley Periodicals LLC.)

Designs for two automated atomic layer deposition (ALD) flow reactors are presented, and their capabilities for coating additively manufactured (AM) metal prints are described. One instrument allows the coating of several AM parts in batches, while the other is useful for single part experiments. To demonstrate reactor capabilities, alumina (Al2O3) was deposited onto AM 316L stainless steel by dosing with water (H2O) vapor and trimethylaluminum (TMA) and purging with nitrogen gas (N2). Both instruments are controlled by custom-programmed LabVIEW software that enables in situ logging of temperature, total pressure, and film thickness using a quartz crystal microbalance. An initial result shows that 150 ALD cycles led to a film thickness of ∼55 nm, which was verified with Rutherford backscattering spectroscopy. This indicates that the reactors were indeed depositing single atomic layers of Al2O3 per ALD cycle, as intended., (© 2024 Author(s). All article content, except where otherwise noted, is licensed under a Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).)

Translocations involving FGFR2 gene fusions are common in cholangiocarcinoma and predict response to FGFR kinase inhibitors. However, the rate and durability of response are limited due to the emergence of resistance, typically involving acquired FGFR2 kinase domain mutations, and to sub-optimal dosing, relating to drug adverse effects. Here, we report the development of biparatopic antibodies targeting the FGFR2 extracellular domain (ECD), as candidate therapeutics. Biparatopic antibodies can overcome drawbacks of standard bivalent monoparatopic antibodies, which often show poor inhibitory or even agonist activity against oncogenic receptors. We show that oncogenic transformation by FGFR2 fusions requires an intact ECD. Moreover, by systematically generating biparatopic antibodies that target distinct epitope pairs along the FGFR2 ECD, we identified antibodies that effectively block signaling and malignant growth driven by FGFR2-fusions. Importantly, these antibodies demonstrate efficacy in vivo, synergy with FGFR inhibitors, and activity against FGFR2 fusions harboring kinase domain mutations. Thus, biparatopic antibodies may serve as new treatment options for patients with FGFR2-altered cholangiocarcinoma., Summary: We identify biparatopic FGFR2 antibodies that are effective against FGFR2 fusion driven cholangiocarcinoma.

Lightning strikes, a prominent meteorological event, pose a significant risk of triggering technological disruptions within the process industry. To better understand this phenomenon, an analysis focused on past lightning-triggered events was carried out, examining open-source industrial-accident databases to compile a new NaTech-driven dataset of 689 records. First, an overall quantitative analysis revealed that over 80 % of these events involved incidents or loss of containment. Notably, 83.3 % of them occurred during the spring and summer, indicating a seasonal pattern. Based on the frequency of functional attributes, the chemical and petrochemical macro-sector was the most vulnerable, followed by storage and warehousing. About 40 % of all classifiable events happened on storage equipment, while 21 % happened on electric and electronic devices. Given the lack of valuable information for the principal source of data (NRC), the technological scenarios triggered were characterized using a refined subset of 127 observations, obtained considering the "other sources" of data. Fire scenarios predominated at 56 %; coincidentally, roughly 70 % of all scenarios involved hazardous substances classified as physical hazards. Estimated losses for the available information underscored the adverse consequences of lightning-triggered NaTech events, highlighting their major impact on both safety and the environment. An analysis of the event tree showed the logical path from the lightning strike to the final ignition scenarios (considering a subset of 107 records). This path accounted for 36 % of the classifiable records that directly affected the structure, while more than 50 % of them did not. Bayesian network structures made it possible to get conditional probabilities from the event tree and improved the model by adding attributes for vulnerable equipment and macro-sectors. In order to deal with the uncertain data, algorithms were used to generalize the models that were obtained from smaller subsets of data with more accurate information to the whole dataset. It provides an important additional view of unclassifiable data that otherwise remained in the dark. This novel insight contributes to increase the vulnerability awareness of industrial assets against lightning strikes., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)

The paradigm of cancer-targeted therapies has focused largely on inhibition of critical pathways in cancer. Conversely, conditional activation of signaling pathways as a new source of selective cancer vulnerabilities has not been deeply characterized. In this study, we sought to systematically identify context-specific gene-activation-induced lethalities in cancer. To this end, we developed a method for gain-of-function genetic perturbations simultaneously across ~500 barcoded cancer cell lines. Using this approach, we queried the pan-cancer vulnerability landscape upon activating ten key pathway nodes, revealing selective activation dependencies of MAPK and PI3K pathways associated with specific biomarkers. Notably, we discovered new pathway hyperactivation dependencies in subsets of APC-mutant colorectal cancers where further activation of the WNT pathway by APC knockdown or direct β-catenin overexpression led to robust antitumor effects in xenograft and patient-derived organoid models. Together, this study reveals a new class of conditional gene-activation dependencies in cancer., (© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Introduction: Esophageal 24-hour pH/impedance testing is routinely performed to diagnose gastroesophageal reflux disease. Interpretation of these studies is time-intensive for expert physicians and has high inter-reader variability. There are no commercially available machine learning tools to assist with automated identification of reflux events in these studies., Methods: A machine learning system to identify reflux events in 24-hour pH/impedance studies was developed, which included an initial signal processing step and a machine learning model. Gold-standard reflux events were defined by a group of expert physicians. Performance metrics were computed to compare the machine learning system, current automated detection software (Reflux Reader v6.1), and an expert physician reader., Results: The study cohort included 45 patients (20/5/20 patients in the training/validation/test sets, respectively). The mean age was 51 (standard deviation 14.5) years, 47% of patients were male, and 78% of studies were performed off proton-pump inhibitor. Comparing the machine learning system vs current automated software vs expert physician reader, area under the curve was 0.87 (95% confidence interval [CI] 0.85-0.89) vs 0.40 (95% CI 0.37-0.42) vs 0.83 (95% CI 0.81-0.86), respectively; sensitivity was 68.7% vs 61.1% vs 79.4%, respectively; and specificity was 80.8% vs 18.6% vs 87.3%, respectively., Discussion: We trained and validated a novel machine learning system to successfully identify reflux events in 24-hour pH/impedance studies. Our model performance was superior to that of existing software and comparable to that of a human reader. Machine learning tools could significantly improve automated interpretation of pH/impedance studies., (Copyright © 2023 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of The American College of Gastroenterology.)

Chemical solution deposition (CSD) methods involving the thermal decomposition of 5.0 M Er(NO 3 ) 3 ·5H 2 O and Y(NO 3 ) 3 ·6H 2 O precursor solutions were employed to fabricate protective erbia and yttria coatings onto stainless steel (SS304/SS316) coupons. The two techniques tested were dip and spray coating, which were then compared to a commercial yttria spray (ZYP Coatings). It was determined that solution concentration, solvent choice, injection of Er 2 O 3 and Y 2 O 3 micropowder, and the annealing temperature/ramp profile were critical to the coatings' physical properties. For dip coatings, thicknesses were 1-20 μm after two dipping/annealing cycles, and adhesion strength was ∼1000 psi, increasing up to ∼1300 psi if the SS coupons had preliminary sandblasting. Spray coatings from precursor solutions were reported to have thicknesses of 20-80 μm and adhesion strength less than 500 psi (regardless of the coupon surface finish). Cross-sectional views of the coatings confirmed subsurface porosity, and XRD results indicated that the coatings were polycrystalline, with patterns typical to that of cubic Er 2 O 3 and Y 2 O 3 .

Absolute kinetics for reactions of C2H2 with a series of ∼60 individual carbon nanoparticles (NPs) from graphite, graphene, graphene oxide, carbon black, diamond, and nano-onion feedstocks were measured for temperatures (TNP) ranging from 1200 to 1700 K. All the NPs were observed to gain mass by carbon addition under conditions that varied with feedstock but with large variations in initial growth rate. Long reaction periods were studied to allow the evolution of growth rates over time to be observed. Diamond NPs were found to passivate against C2H2 addition if heated above ∼1400 K, and the highly variable initial reactivity for carbon nano-onions was found to depend on the presence of non-onion-structure surface carbon. For graphitic and carbon black NPs, three distinct growth modes were observed, correlated with the initial NP mass (Minitial). Smallest graphitic and carbon black NPs, with masses <∼25 MDa, initially grew rapidly but also passivated quickly after adding <4% of Minitial. NPs in the 20-50 MDa range also passivated but only after multiple waves of fast growth separated by periods of low reactivity, with up to ∼11% total mass gain before passivation. The largest carbon black and graphitic NPs, with Minitial >50 MDa, grew rapidly and continuously, adding up to ∼300% of Minitial with no sign of rate slowing as long as C2H2 was present. The efficiencies for C2H2 addition and etching by O2 are strongly correlated, but the correlation changes as the NPs passivate. Growth and passivation mechanisms are discussed., (© 2023 Author(s). Published under an exclusive license by AIP Publishing.)

Combinatorial CRISPR technologies have emerged as a transformative approach to systematically probe genetic interactions and dependencies of redundant gene pairs. However, the performance of different functional genomic tools for multiplexing sgRNAs vary widely. Here, we generate and benchmark ten distinct pooled combinatorial CRISPR libraries targeting paralog pairs to optimize digenic knockout screens. Libraries composed of dual Streptococcus pyogenes Cas9 (spCas9), orthogonal spCas9 and Staphylococcus aureus (saCas9), and enhanced Cas12a from Acidaminococcus were evaluated. We demonstrate a combination of alternative tracrRNA sequences from spCas9 consistently show superior effect size and positional balance between the sgRNAs as a robust combinatorial approach to profile genetic interactions of multiple genes., (© 2022. The Author(s).)

O 2 oxidation and sublimation kinetics for >30 individual nanoparticles (NPs) of five different feedstocks (graphite, graphene oxide, carbon black, diamond, and nano-onion) were measured using single-NP mass spectrometry at temperatures ( T NP ) in the 1100-2900 K range. It was found that oxidation, studied in the 1200-1600 K range, is highly sensitive to the NP surface structure, with etching efficiencies (EE O 2 ) varying by up to 4 orders of magnitude, whereas sublimation rates, significant only for T NP ≥ ∼1700 K, varied by only a factor of ∼3. Its sensitivity to the NP surface structure makes O 2 etching a good real-time structure probe, which was used to follow the evolution of the NP surface structures over time as they were either etched or annealed at high T NP . All types of carbon NPs were found to have initial EE O 2 values in the range near 10 -3 Da/O 2 collision, and all eventually evolved to become essentially inert to O 2 (EE O 2 < 10 -6 Da/O 2 collision); however, the dependence of EE O 2 on time and mass loss was very different for NPs from different feedstocks. For example, diamond NPs evolved rapidly and monotonically toward inertness, and evolution occurred in both oxidizing and inert atmospheres. In contrast, graphite NPs evolved only under oxidizing conditions and were etched with complex time dependence, with multiple waves of fast but non-monotonic etching separated by periods of near-inertness. Possible mechanisms to account for the complex etching behavior are proposed.

Although single-gene perturbation screens have revealed a number of new targets, vulnerabilities specific to frequently altered drivers have not been uncovered. An important question is whether the compensatory relationship between functionally redundant genes masks potential therapeutic targets in single-gene perturbation studies. To identify digenic dependencies, we developed a CRISPR paralog targeting library to investigate the viability effects of disrupting 3,284 genes, 5,065 paralog pairs and 815 paralog families. We identified that dual inactivation of DUSP4 and DUSP6 selectively impairs growth in NRAS and BRAF mutant cells through the hyperactivation of MAPK signaling. Furthermore, cells resistant to MAPK pathway therapeutics become cross-sensitized to DUSP4 and DUSP6 perturbations such that the mechanisms of resistance to the inhibitors reinforce this mechanism of vulnerability. Together, multigene perturbation technologies unveil previously unrecognized digenic vulnerabilities that may be leveraged as new therapeutic targets in cancer., (© 2021. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Background: The prevalence of gestational diabetes mellitus (GDM) has rapidly increased, yet few prior studies have investigated parameters of early brain development in infants born to gestational diabetic mothers. The present study assessed visual evoked potentials (VEPs) in healthy infants born to gestational diabetic mothers and matched controls., Methods: After exclusions, in this prospective study we examined VEPs in 73 neonates between 37 weeks and 41 weeks gestation at birth (n = 37 infants of gestational diabetic mothers). Stroboscopic flashes were presented through closed eyelids during passive electroencephalography (EEG) recording to derive VEP waveforms during natural sleep., Results: There was a statistically significant moderate correlation between gestational age at birth and P2 latency of the flash VEP where P2 latency significantly decreased with increasing gestational age (Pearson's R(73) = -0.32, p < .01). There was also a significant moderate correlation between postnatal age (hours of life) and P2 latency of the flash VEP where P2 latency significantly decreased with increasing postnatal age (Pearson's R(73) = -0.23, p < .05). When controlling for gestational age at birth, postnatal age, and sex, there was a significant effect of group (GDM-exposed vs. control) on P2 latency of the flash VEP (p < .05). Infants of gestational diabetic mothers had a significantly longer P2 latency (M: 215.29 ± SD: 2.58 ms) than controls (M: 206.41 ± SD: 2.62 ms)., Conclusion: Our findings suggest P2 flash VEP latency is a potential measure of cortical maturation and marker of immature development in infants of gestational diabetic mothers., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Remdesivir is an antiviral nucleoside phosphoramidate with activity against multiple viruses, including SARS-CoV-2. To enable studies of viral polymerases with RNA containing remdesivir, we report an efficient synthesis of a phosphoramidite of GS-441524, the nucleoside precursor of remdesivir, and its incorporation into RNA using automated solid-phase RNA synthesis., Competing Interests: The authors declare no conflicts of interest., (This journal is © The Royal Society of Chemistry.)

PRMT5 is an essential arginine methyltransferase and a therapeutic target in MTAP-null cancers. PRMT5 uses adaptor proteins for substrate recruitment through a previously undefined mechanism. Here, we identify an evolutionarily conserved peptide sequence shared among the three known substrate adaptors (CLNS1A, RIOK1, and COPR5) and show that it is necessary and sufficient for interaction with PRMT5. We demonstrate that PRMT5 uses modular adaptor proteins containing a common binding motif for substrate recruitment, comparable with other enzyme classes such as kinases and E3 ligases. We structurally resolve the interface with PRMT5 and show via genetic perturbation that it is required for methylation of adaptor-recruited substrates including the spliceosome, histones, and ribosomal complexes. Furthermore, disruption of this site affects Sm spliceosome activity, leading to intron retention. Genetic disruption of the PRMT5-substrate adaptor interface impairs growth of MTAP-null tumor cells and is thus a site for development of therapeutic inhibitors of PRMT5., Competing Interests: Declaration of interests Materials related to this article are in a provisional patent application by the Broad Institute. W.R.S. is a board or scientific advisory board (SAB) member and holds equity in Peloton Therapeutics, Ideaya Biosciences, Civetta Therapeutics, and Bluebird bio; has consulted for Array, Astex, Dynamo Therapeutics, Ipsen, Merck Pharmaceuticals, PearlRiver Therapeutics, Sanofi, Scorpion Therapeutics, and Servier; and receives research funding from Pfizer Pharmaceuticals, Merck Pharmaceuticals, Ideaya Biosciences, and Deerfield Management. B.J.M. is an employee and equity holder of Tango Therapeutics. D. Porter is an employee and equity holder of Cedilla Therapeutics. A.J.A. has consulted for Oncorus Inc., Arrakis Therapeutics, and Merck & Company and has research funding from Mirati Therapeutics and Deerfield Management., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Multiple copies of WW domains and PPXY motif sequences are often reciprocally presented by regulatory proteins that interact at crucial regulatory steps in the cell life cycle. While biophysical studies of single WW domain-single PPXY motif complexes abound in the literature, the molecular mechanisms of multivalent WW domain-PPXY assemblies are still poorly understood. By way of investigating such assemblies, we characterized the multivalent association of the entire cognate binding domains, two WW sequences and five PPXY motifs respectively, of the Yorkie transcription coactivator and the Warts tumor suppressor. Isothermal titration calorimetry, sedimentation velocity, size-exclusion chromatography coupled to multi-angle light scattering and native-state mass spectrometry of Yorkie WW domains interactions with the full-length Warts PPXY domain, and numerous PPXY motif variants of Warts show that the two proteins assemble via binding of tandem WW domains to adjacent PPXY pairs to produce an ensemble of interconverting complexes of variable stoichiometries, binding energetics and WW domain occupancy. Apparently, the Yorkie tandem WW domains first target the two adjacent PPXY motifs at the C-terminus of the Warts polypeptide and additional WW domains bind unoccupied motifs. Similar ensembles of interconverting conformers may be common in multivalent WW domain-PPXY interactions to promote the adaptability and versatility of WW domain-PPXY mediated cellular processes., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

A single nanoparticle (NP) mass spectrometry method was used to measure sublimation rates as a function of nanoparticle temperature (T NP ) for sets of individual graphite and graphene NPs. Initially, the NP sublimation rates were ∼400 times faster than those for bulk graphite, and there were large NP-to-NP variations. Over time, the rates slowed substantially, though they remained well above the bulk rate. The initial activation energies ( E a values) were correspondingly low and doubled, as a few monolayers worth of material was sublimed from the surfaces. The high initial rates and low E a values are attributed to large numbers of edge, defect, and other low coordination sites on the NP surfaces, and the changes are attributed to atomic-scale "smoothing" of the surface by preferential sublimation of the less stable sites. The emissivity of the NPs also changed after heating, more frequently increasing. The emissivity and sublimation rates were anticorrelated, leading to the conclusion that high densities of low-coordination sites on the NP surfaces enhance sublimation but suppress emissivity.

The second WW domain (WW2) of the kidney and brain scaffolding protein, KIBRA, has an isoleucine (Ile-81) rather than a second conserved tryptophan and is primarily unstructured. However, it adopts the canonical triple-stranded antiparallel β-sheet structure of WW domains when bound to a two-PP X Y motif peptide of the synaptic protein Dendrin. Here, using a series of biophysical experiments, we demonstrate that the WW2 domain remains largely disordered when bound to a 69-residue two-PP X Y motif polypeptide of the synaptic and podocyte protein synaptopodin (SYNPO). Isothermal titration calorimetry and CD experiments revealed that the interactions of the disordered WW2 domain with SYNPO are significantly weaker than SYNPO's interactions with the well-folded WW1 domain and that an I81W substitution in the WW2 domain neither enhances binding affinity nor induces substantial WW2 domain folding. In the tandem polypeptide, the two WW domains synergized, enhancing the overall binding affinity with the I81W variant tandem polypeptide 2-fold compared with the WT polypeptide. Solution NMR results showed that SYNPO binding induces small but definite chemical shift perturbations in the WW2 domain, confirming the disordered state of the WW2 domain in this complex. These analyses also disclosed that SYNPO binds the tandem WW domain polypeptide in an antiparallel manner, that is, the WW1 domain binds the second PP X Y motif of SYNPO. We propose a binding model consisting of a bipartite interaction mode in which the largely disordered WW2 forms a "fuzzy" complex with SYNPO. This binding mode may be important for specific cellular functions., (© 2019 Kwok et al.)

We discuss the design of an optical system that allows measurement of 600-1650 nm emission spectra for individual nanoparticles (NPs), laser-heated in an electrodynamic trap in controlled atmospheres. An approach to calibration of absolute intensity versus wavelength for very low emission intensities is discussed, and examples of NP graphite and carbon black spectra are used to illustrate the methodology.

Fluorescent mRNA molecules offer a wide range of applications for studying capping/decapping reactions, translation, and other biophysical studies. Furthermore, fluorescent tags prove invaluable for tracking RNA molecules in cells. Here, we describe an efficient synthesis of a fluorescent cap analog, anthranioyl-GTP, its purification, and in vitro cap labeling of transcribed mRNA catalyzed by the recombinant vaccinia capping enzyme to produce anthranioyl-m(7)GpppG-capped RNA.

Atherosclerosis is a chronic inflammatory disease characterized by the accumulation of lipid-loaded macrophages in the arterial wall. We demonstrate that macrophage lipid body formation can be induced by modified lipoproteins or by inflammatory Toll-like receptor agonists. We used an unbiased approach to study the overlap in these pathways to identify regulators that control foam cell formation and atherogenesis. An analysis method integrating epigenomic and transcriptomic datasets with a transcription factor (TF) binding site prediction algorithm suggested that the TF ATF3 may regulate macrophage foam cell formation. Indeed, we found that deletion of this TF results in increased lipid body accumulation, and that ATF3 directly regulates transcription of the gene encoding cholesterol 25-hydroxylase. We further showed that production of 25-hydroxycholesterol (25-HC) promotes macrophage foam cell formation. Finally, deletion of ATF3 in Apoe(-/-) mice led to in vivo increases in foam cell formation, aortic 25-HC levels, and disease progression. These results define a previously unknown role for ATF3 in controlling macrophage lipid metabolism and demonstrate that ATF3 is a key intersection point for lipid metabolic and inflammatory pathways in these cells.

Introduction: Pacemaker and defibrillator infections are an uncommon, but catastrophic complication of device implantation. The present study examined the prevalence of device-related infections, the patterns of antibiotic resistance, and the presence of methicillin resistant staphylococcus aureus (MRSA) nares colonization in device implant recipients., Methods: Two protocols were employed using a retrospective and a prospective analysis. A retrospective chart review of 218 patients with suspected device infection from 1/2000 to 1/2011 was performed. Demographics, infection rates, and patterns of antibiotic resistance were compared. The prospective analysis enrolled one hundred eighty two patients undergoing device implantations or generator replacements. The nares were swabbed and analyzed for the presence of staphylococcus aureus, and tested for methicillin sensitivity., Results: Over a period of ten years, 12,771 device implants/generator changes/system revisions were performed, with an infection rate of 1.2%. Methicillin resistance (MR) was identified in 98/218 (44.9%) of patients. Those with MR infection had more diabetes and cardiomyopathy. There was no significant increase in methicillin resistance over time (p=0.30). Our prospective analysis included 110 men. A total of 32 patients (17.6%) had positive cultures for SA: 6.6% with MRSA. Patients positive for MRSA nares colonization had a statistically significant greater length of hospital stay 8.5 days (mean) versus 4.4 days (P=0.049)., Conclusions: Methicillin resistant organisms appear to be emerging and persistent pathogens in device implants. The screening of MRSA colonization may identify new populations at risk. Further studies and analysis are needed to determine the cost effectiveness of a screening protocol.

We describe a microfluidic immunoassay device that permits sensitive and quantitative multiplexed protein measurements on nano-liter-scale samples. The device exploits the combined power of integrated microfluidics and optically encoded microspheres to create an array of approximately 100-microm(2) sensors functionalized with capture antibodies directed against distinct targets. This strategy overcomes the need for performing biochemical coupling of affinity reagents to the device substrate, permits multiple proteins to be detected in a nano-liter-scale sample, is scalable to large numbers of samples, and has the required sensitivity to measure the abundance of proteins derived from single mammalian cells. The sensitivity of the device is sufficient to detect 1000 copies of tumor necrosis factor (TNF) in a volume of 4.7nl.

Although iron is required to sustain life, its free concentration and metabolism have to be tightly regulated. This is achieved through a variety of iron-binding proteins including transferrin and ferritin. During infection, bacteria acquire much of their iron from the host by synthesizing siderophores that scavenge iron and transport it into the pathogen. We recently demonstrated that enterochelin, a bacterial catecholate siderophore, binds to the host protein lipocalin 2 (ref. 5). Here, we show that this event is pivotal in the innate immune response to bacterial infection. Upon encountering invading bacteria the Toll-like receptors on immune cells stimulate the transcription, translation and secretion of lipocalin 2; secreted lipocalin 2 then limits bacterial growth by sequestrating the iron-laden siderophore. Our finding represents a new component of the innate immune system and the acute phase response to infection.

The spontaneous occurrence of resistance to the herbicide glyphosate in weed species has been an extremely infrequent event, despite over 20 years of extensive use. Recently, a glyphosate-resistant biotype of goosegrass (Eleusine indica) was identified in Malaysia exhibiting an LD(50) value approximately 2- to 4-fold greater than the sensitive biotype collected from the same region. A comparison of the inhibition of 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) activity by glyphosate in extracts prepared from the resistant (R) and sensitive (S) biotypes revealed an approximately 5-fold higher IC(50)(glyphosate) for the (R) biotype. Sequence comparisons of the predicted EPSPS mature protein coding regions from both biotypes revealed four single-nucleotide differences, two of which result in amino acid changes. One of these changes, a proline to serine substitution at position 106 in the (R) biotype, corresponds to a substitution previously identified in a glyphosate-insensitive EPSPS enzyme from Salmonella typhimurium. Kinetic data generated for the recombinant enzymes suggests that the second substitution identified in the (R) EPSPS does not contribute significantly to its reduced glyphosate sensitivity. Escherichia coli aroA- (EPSPS deficient) strains expressing the mature EPSPS enzyme from the (R) biotype exhibited an approximately 3-fold increase in glyphosate tolerance relative to strains expressing the mature EPSPS from the (S) biotype. These results provide the first evidence for an altered EPSPS enzyme as an underlying component of evolved glyphosate resistance in any plant species.

Poly(hydroxyalkanoates) are natural polymers with thermoplastic properties. One polymer of this class with commercial applicability, poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) can be produced by bacterial fermentation, but the process is not economically competitive with polymer production from petrochemicals. Poly(hydroxyalkanoate) production in green plants promises much lower costs, but producing copolymer with the appropriate monomer composition is problematic. In this study, we have engineered Arabidopsis and Brassica to produce PHBV in leaves and seeds, respectively, by redirecting the metabolic flow of intermediates from fatty acid and amino acid biosynthesis. We present a pathway for the biosynthesis of PHBV in plant plastids, and also report copolymer production, metabolic intermediate analyses, and pathway dynamics.

Objective: To design and implement the Lovelace Diabetes EPISODES OF CARE program in a managed care setting. This program is intended to address the complex needs of patients with type 2 diabetes mellitus by using specific physician-provider and patient interventions., Design: Observational study., Setting: Lovelace Health Systems, the second-largest and most fully integrated health care delivery system in New Mexico. The main facility is located in Albuquerque., Participants: Lovelace Health Plan members with type 2 diabetes., Interventions: Physician-provider interventions included practice guidelines medical profile screens, and provider support reports. Patients interventions included diabetes education; improved access to care, with focused diabetes clinic visits and "Diabetes Days"; and reminder systems., Main Outcome Measures: Glycohemoglobin values, dilated eye examination rates, and access to education., Results: Significant lowering of glycohemoglobin values, dilated eye examination rates exceeding benchmark measures, and increases in educational access rates have occurred since the Lovelace Diabetes EPISODES OF CARE program was implemented., Conclusions: An integrated health care delivery system with a comprehensive, diabetes disease management program can substantially improve outcomes.

Time delays in density dependence have been associated with regular and sustained fluctuations in population size. Mathematical models have found that delays that are long enough can produce limit cycles. In experimental populations with preadult and adult phases in the life cycle and density regulation in any of the phases, it has been found that the time delay represented by maturation time can produce fluctuations in adult size. This has been corroborated by mathematical models. The present study develops mathematical models of growth in this type of population. The analysis of these models indicate that when preadult cohorts do not overlap in space, and preadult mortality is affected by the density of preadults at the present moment, the time delay does not destabilize the population. However, with spatial overlap of preadult cohorts and the same mechanism of density dependence, time delays can be destabilizing. It is concluded that time delays in density effects during preadulthood are not necessarily destabilizing, and that fluctuations in population size can also be due to delays with density effects in preadulthood and adulthood, or exogenous agents. Copyright 1998 Academic Press Limited

The metabolic response to trauma, including neurotrauma in general, has been studied extensively, but the acute metabolic response to spinal cord injury (SCI) has not. Therefore, 12 patients with SCI are presented in whom intensive nutrition assessment and management were instituted immediately after injury. Nitrogen balance (NB), predicted energy expenditure (PEE), and actual energy expenditure (MEE) were calculated or measured in each patient. A persistent negative NB was observed in all but one of the 12 patients. The single patient who did not exhibit persistent negative NB (no positive NB from week 2 to week 4 in the face of appropriate feeding) had an incomplete myelopathy, thus implying that the degree of motor dysfunction correlates with the obligatory nature of the negative NB. The negative NB observed in several of the patients did not occur until the second or third post-injury week. In addition, calculations of PEE by successively multiplying the Harris-Benedict equation by an activity factor of 1.2 and then by a stress factor of 1.6, resulted in excessive feeding (as assessed by metabolic cart measurements; ie indirect calorimetry) in the majority of the patients. In all of the 11 patients with persistent negative NBs, protein administration in the amount of 2 g/k of ideal body weight and aggressive caloric delivery did not alter the negative pattern of the NBs. Therefore, it is concluded that the negative NB following SCI is obligatory. Furthermore, the extent of SCI (extent of myelopathy or of neurological injury) correlates with the obligatory nature of the negative NB. In addition, the results from using the above method for estimating caloric requirements and the delayed manifestation of the negative NB may cause an additional tendency to acutely overfeed SCI patients. Therefore, eliminating the activity factor of 1.2 (due to the diminished activity arising from paralysis) and a diminution of the stress factor is recommended for initial PEE calculations. Serial metabolic cart (indirect calorimetry) measurements are recommended to accurately assess the patient's subsequent metabolic requirements.

A common method for calculating energy needs (PEE) in acute trauma patients is multiplying the Harris-Benedict equation (BEE) by activity factors (AF) and variable stress factors (SF) depending on the injury severity. Selection of the SF can be an arbitrary and potentially inaccurate decision. The purposes of this study were: (1) to investigate the relationship between injury severity score (ISS) to postinjury energy expenditure (MEE), and (2) to compare the MEE to PEE when using the SF of 1.75. Thirty-five severely injured patients (mean ISS = 27.5 +/- 10.7 SD) admitted to our Level I Trauma Center of the University of New Mexico were prospectively assessed for energy needs [PEE = BEE x 1.2(AF) x 1.75(SF)] and ISS. Total nutritional support delivered estimated needs. Indirect calorimetry measurements were obtained on all of the patients within the first 7-10 days following injury. No correlation (r = -0.042) existed between the MEE and ISS. There was a significant correlation (r = 0.772, P < 0.05) between PEE and MEE when using the SF of 1.75 for all of the patients. These results suggest that there is not a correlation between ISS and subsequent MEE in major trauma patients. In addition, using the SF of 1.75 will closely estimate energy needs in acute trauma patients.

Enteral nutrition has become the preferred route of nutrient administration. Because of vigorous attempts to deliver nutrient enterally in expanded patient groups, many different locations for enteral access have been advocated along with a variety of methods related to rate and pattern of delivery. Because all modes of delivery are not compatible with all sites of access and both need to be tailored to specific subsets of patients, confusion can develop regarding where and how enteral nutrients are best delivered and why. In an era when such a high priority has been placed on feeding through the enteral route, a review of the methods and rationale behind the ever-expanding choices of enteral access is timely.

Conventional total parenteral nutrition regimens (TPN-C) involve concentrations of dextrose/protein which necessitate administration of 2.5-4 liters/day to meet target nutritional needs. Although this is frequently acceptable, certain clinical settings mandate a volume-restricted (VR) approach. This study compares a VR TPN regimen (TPN-VR) involving the use of 25% dextrose and 9.5% amino acid with D17.5 AA 5.0 (TPN-C). The two groups were compared for adequacy of nutritional delivery, balance, and tolerance. Twenty patients received the TPN-VR (Group 1) and 20 patients received TPN-C (Group 2). The groups were comparable in age, sex, injury severity, and APACHE 2 scores. Harris-Benedict (BEE) x 2 and 2 g/protein/kg of ideal body weight were delivered by the second day of TPN. A 27% reduction in administered fluid was achieved in Group 1 (P < 0.001). Metabolic cart data in both groups demonstrated that delivered calories exceeded REE. The average RQ in Group 1 was 0.84 and in Group 2 was 0.90 (P > 0.1). There was no significant difference between the two groups in nitrogen balance, mean serum bilirubin levels, PT and PTT, serum albumin levels, and triglycerides (P > 0.20). SGPT and alkaline phosphatase levels were significantly higher in Group 2 (P < 0.001). Group 2 received an average of 22% more carbohydrate than Group 1 and 45% required insulin compared to 25% in Group 1 (P < 0.01). In summary, TPN-VR is comparable to TPN-C in terms of effectiveness of delivery, nutritional balance, and tolerance.

Nutritional support of stressed geriatric patients remains empiric and has classically been limited by tolerance. Although the hypermetabolic response is known to increase protein and calorie demands, tolerance to increased loads of delivered nutrients in older patients has been questioned. We compared tolerance to nutrient delivery and nitrogen metabolism in 38 stressed surgical patients over age 65 to 38 Injury Severity Score or disease matched younger controls. Twenty-seven of the 31 geriatric patients (87%) who maintained normal renal function (serum creatinine less than 2.0 mg/dl) became azotemic (BUN greater than 30) while receiving 1.5 to 2.0 g of protein per kilogram of ideal body weight compared to only 21% of controls. This phenomenon led to inaccuracies in 17% of geriatric nitrogen balance studies because of unaccounted for serum accumulation of urea nitrogen (compared to only 6% in the control group). When calculated protein requirements were administered to the geriatric group, the mean nitrogen balance was -1.6. Resting energy expenditure as measured by indirect calorimetry demonstrated a strong correlation between actual calorie expenditures and calculated needs based on the Harris-Benedict basal energy expenditure (BEE) multiplied by an activity factor of 1.2 and a stress factor of 1.75 for trauma (r = 0.86, P less than 0.05) or 1.5 for general surgery patients (r = 0.72, P less than 0.05). In summary, energy requirements by stressed geriatric patients can be closely defined by calculation of the Harris-Benedict BEE in conjunction with appropriate activity and stress factors. However, attempts to deliver traditional levels of protein lead to azotemia and are frequently unsuccessful in achieving positive nitrogen balance.(ABSTRACT TRUNCATED AT 250 WORDS)

Obligatory nitrogen losses due to paralysis in the spinal cord-injured (SCI) patient prevent positive nitrogen balance (NB) regardless of the calorie and protein intakes. Ten patients with SCI and 20 controls with nonspinal cord injury (NSCI) matched for time, sex, age, and injury severity score (ISS) were admitted to our Level I trauma center. In both groups, total nutritional support was delivered within 72 hours of admission based on predicted energy expenditures (PEE = Harris-Benedict equation x 1.2 x 1.6) and 2 g of protein/kg of ideal body weight (IBW). Subsequent changes in nutrient delivery were based on NB. No SCI patient established positive NB during the 7-week period following injury despite an average delivery of 2.4 g of protein/kg IBW and 120% of the PEE at the time of peak negative NB (-10.5). In six SCI patients, an average increase of 25% in delivered protein and 12% in delivered calories over a 1-week period effected no change in average NB (-7.4 vs -6.8). Indirect calorimetry in five SCI patients showed that calorie intakes were 110% more than average measured energy expenditures. In contrast, 17 of 20 NCSI patients achieved positive NB within 3 weeks of admission. They required an average delivery of 2.3 g of protein/kg IBW and 110% of PEE to reach positive NB. These data demonstrate the phenomenon of obligatory negative NB acutely following SCI. Aggressive attempts to achieve positive NB in these patients will fail and result in overfeeding.

Extracts from Ulex europaeus L. (Fabaceae) were tested for antiviral activity against HSV-1 and poliovirus. It was found that the n-butanolic extract significantly inhibited the replication of these viruses, as shown by inhibition of cytophathic effects and reduction of plaques; the extract has neither a prophylactic nor a virucidal activity. From this extract has been isolated and identified by chromatographic, chemical, and spectroscopic methods, the 5-O-methylgenistein 7-O-beta D-glucopyranoside. This isoflavonic glycoside has not been previously described and it is responsible at least partially for the antiviral activity.

Discrete-time models of growth of populations with nonoverlapping generations and density regulation in two life stages are studied. It is assumed that there is no delay in the effects of density. Assigning exponential, linear, or hyperbolic functions to describe the dependence of preadult survival and fecundity on density, nine models are obtained. The dynamics of the model resulting from using the exponential function to describe the density dependence of both preadult survival and fecundity is analyzed: for large values of the intrinsic rate of increase there may exist up to three equilibrium population sizes, two stable. This indicates that a life history with two episodes of density regulation can give origin to alternative stable states. The models are fitted to recruitment data from growth experiments of Drosophila laboratory populations obtained with the Serial Transfer System Type 2 (Ayala et al., 1973. Theor. Pop. Biol. 4, 331-356) and collected by other authors. The results of the fittings suggest that this recruitment data can be adequately described with the models.

Previous studies demonstrated that prostaglandins are local or tissue hormones which can be released from blood vessel walls. In the present study, we investigated the capacity of bovine ductus arteriosus to synthetize prostaglandins in vitro. After incubation of slices of ductus arteriosus in Krebs' solution with (1-14C) arachidonic acid for 3 hours, more than 40% of the radiolabeled material recovered from the incubating medium were metabolites of arachidonic acid. The major product was indistinguishable from 6 keto-PGF1alpha as determined by its chromatographic motility and resistance to alkaline conversion to PGB. The PGI2 synthetic capacity of the ductus arteriosus, as revealed by the predominance of its major metabolite 6 keto-PGF1alpha, suggests that this metabolic pathway of arachidonic acid may contribute to the hemodynamic changes occurring during fetal life and at birth.