Recent advances in ruthenium-based olefin metathesis.

Ruthenium-based olefin metathesis catalysts, known for their functional group tolerance and broad applicability in organic synthesis and polymer science, continue to evolve as an enabling technology in these areas. A discussion of recent mechanistic investigations is followed by an overview of selected applications.

Environmental and Human Health Impacts of Spreading Oil and Gas Wastewater on Roads.

Thirteen states in the United States allow the spreading of O&G wastewaters on roads for deicing or dust suppression. In this study, the potential environmental and human health impacts of this practice are evaluated. Analyses of O&G wastewaters spread on roads in the northeastern, U.S. show that these wastewaters have salt, radioactivity, and organic contaminant concentrations often many times above drinking water standards. Bioassays also indicated that these wastewaters contain organic micropollutants that affected signaling pathways consistent with xenobiotic metabolism and caused toxicity to aquatic organisms like Daphnia magna. The potential toxicity of these wastewaters is a concern as lab experiments demonstrated that nearly all of the metals from these wastewaters leach from roads after rain events, likely reaching ground and surface water. Release of a known carcinogen (e.g., radium) from roads treated with O&G wastewaters has been largely ignored. In Pennsylvania from 2008 to 2014, spreading O&G wastewater on roads released over 4 times more radium to the environment (320 millicuries) than O&G wastewater treatment facilities and 200 times more radium than spill events. Currently, state-by-state regulations do not require radium analyses prior to treating roads with O&G wastewaters. Methods for reducing the potential impacts of spreading O&G wastewaters on roads are discussed.

Single-Molecule Sequencing Reveals Complex Genome Variation of Hepatitis B Virus during 15 Years of Chronic Infection following Liver Transplantation.

UNLABELLED: Chronic hepatitis B (CHB) is prevalent worldwide. The infectious agent, hepatitis B virus (HBV), replicates via an RNA intermediate and is error prone, leading to the rapid generation of closely related but not identical viral variants, including those that can escape host immune responses and antiviral treatments. The complexity of CHB can be further enhanced by the presence of HBV variants with large deletions in the genome generated via splicing (spHBV variants). Although spHBV variants are incapable of autonomous replication, their replication is rescued by wild-type HBV. spHBV variants have been shown to enhance wild-type virus replication, and their prevalence increases with liver disease progression. Single-molecule deep sequencing was performed on whole HBV genomes extracted from samples, including the liver explant, longitudinally collected from a subject with CHB over a 15-year period after liver transplantation. By employing novel bioinformatics methods, this analysis showed that the dynamics of the viral population across a period of changing treatment regimens was complex. The spHBV variants detected in the liver explant remained present posttransplantation, and a highly diverse novel spHBV population as well as variants with multiple deletions in the pre-S genes emerged. The identification of novel mutations outside the HBV reverse transcriptase gene that co-occurred with known drug resistance-associated mutations highlights the relevance of using full-genome deep sequencing and supports the hypothesis that drug resistance involves interactions across the full length of the HBV genome. IMPORTANCE: Single-molecule sequencing allowed the characterization, in unprecedented detail, of the evolution of HBV populations and offered unique insights into the dynamics of defective and spHBV variants following liver transplantation and complex treatment regimens. This analysis also showed the rapid adaptation of HBV populations to treatment regimens with evolving drug resistance phenotypes and evidence of purifying selection across the whole genome. Finally, the new open-source bioinformatics tools with the capacity to easily identify potential spliced variants from deep sequencing data are freely available.

Magnetic trapping of SH radicals.

Magnetic trapping of SH radicals, produced via the photostop technique, has been demonstrated. H2S in a skimmed, supersonic molecular beam was photodissociated at 212.8 nm to produce SH inside a 330 mK deep static magnetic trap. The molecular-beam speed was controlled by the mixing ratio of H2S in Kr to match the recoil velocity of the SH photofragments such that some SH radicals were produced with near-zero laboratory-frame velocity. The density of SH radicals in the 2Π3/2, v = 0, J = 3/2 state was followed by (2 + 1) REMPI over seven orders of magnitude of signal intensity. 5 ms after photodissociation, SH radicals moving faster than the capture velocity of 13 m s-1 had left the trap. The 1/e trap lifetime of the remaining SH radicals was 40 ± 10 ms at an estimated density of 5 × 104 molecules per cm3. Photostop offers a simple and direct way to accumulate absolute ground state molecules in a variety of traps.

Spinal cord injury rehabilitation and mental health, SCReaM.

AIMS: The aim of the study was to investigate whether people with a pre-existing mental health disorder (MHD) benefit from rehabilitation following a spinal cord injury (SCI) and how their outcomes differ from those without a pre-existing MHD. METHODS: Rehabilitation outcomes of a cohort of patients with pre-existing MHD discharged from the London SCI Centre over a 6-year period were investigated. A retrospective matched case-control study design was used to compare the Spinal Cord Independence Measure III between those with an SCI and pre-existing MHD and those without and both compared with published expected outcomes. RESULTS: The study found that, overall, those with MHD do benefit from SCI rehabilitation and that their outcomes do not significantly differ from those without MHD. Furthermore, the outcomes were favourable when compared with published expected outcomes. CONCLUSION: Having a pre-existing MHD does not preclude patients with an SCI from benefiting from rehabilitation. These findings are an important basis on which to ensure equal access to rehabilitation for patients with a pre-existing MHD.

Open-label study of faldaprevir plus peginterferon and ribavirin in hepatitis C virus genotype 1-infected patients who failed placebo plus peginterferon and ribavirin.

Faldaprevir, a hepatitis C virus (HCV) NS3/4A protease inhibitor, was evaluated in HCV genotype 1-infected patients who failed peginterferon and ribavirin (PegIFN/RBV) treatment during one of three prior faldaprevir trials. Patients who received placebo plus PegIFN/RBV and had virological failure during a prior trial were enrolled and treated in two cohorts: prior relapsers (n = 43) and prior nonresponders (null responders, partial responders and patients with breakthrough; n = 75). Both cohorts received faldaprevir 240 mg once daily plus PegIFN/RBV for 24 weeks. Prior relapsers with early treatment success (ETS; HCV RNA <25 IU/mL detectable or undetectable at week 4 and <25 IU/mL undetectable at week 8) stopped treatment at week 24. Others received PegIFN/RBV through week 48. The primary efficacy endpoint was sustained virological response (HCV RNA <25 IU/mL undetectable) 12 weeks post treatment (SVR12). More prior nonresponders than prior relapsers had baseline HCV RNA ≥ 800,000 IU/mL (80% vs 58%) and a non-CC IL28B genotype (91% vs 70%). Rates of SVR12 (95% CI) were 95.3% (89.1, 100.0) among prior relapsers and 54.7% (43.4, 65.9) among prior nonresponders; corresponding ETS rates were 97.7% and 65.3%. Adverse events led to faldaprevir discontinuations in 3% of patients. The most common Division of AIDS Grade ≥ 2 adverse events were anaemia (13%), nausea (10%) and hyperbilirubinaemia (9%). In conclusion, faldaprevir plus PegIFN/RBV achieved clinically meaningful SVR12 rates in patients who failed PegIFN/RBV in a prior trial, with response rates higher among prior relapsers than among prior nonresponders. The adverse event profile was consistent with the known safety profile of faldaprevir.

New tracers identify hydraulic fracturing fluids and accidental releases from oil and gas operations.

Identifying the geochemical fingerprints of fluids that return to the surface after high volume hydraulic fracturing of unconventional oil and gas reservoirs has important applications for assessing hydrocarbon resource recovery, environmental impacts, and wastewater treatment and disposal. Here, we report for the first time, novel diagnostic elemental and isotopic signatures (B/Cl, Li/Cl, δ11B, and δ7Li) useful for characterizing hydraulic fracturing flowback fluids (HFFF) and distinguishing sources of HFFF in the environment. Data from 39 HFFFs and produced water samples show that B/Cl (>0.001), Li/Cl (>0.002), δ11B (25-31‰) and δ7Li (6-10‰) compositions of HFFF from the Marcellus and Fayetteville black shale formations were distinct in most cases from produced waters sampled from conventional oil and gas wells. We posit that boron isotope geochemistry can be used to quantify small fractions (∼0.1%) of HFFF in contaminated fresh water and likely be applied universally to trace HFFF in other basins. The novel environmental application of this diagnostic isotopic tool is validated by examining the composition of effluent discharge from an oil and gas brine treatment facility in Pennsylvania and an accidental spill site in West Virginia. We hypothesize that the boron and lithium are mobilized from exchangeable sites on clay minerals in the shale formations during the hydraulic fracturing process, resulting in the relative enrichment of boron and lithium in HFFF.

Extended haplotype association study in Crohn's disease identifies a novel, Ashkenazi Jewish-specific missense mutation in the NF-κB pathway gene, HEATR3.

The Ashkenazi Jewish population has a several-fold higher prevalence of Crohn's disease (CD) compared with non-Jewish European ancestry populations and has a unique genetic history. Haplotype association is critical to CD etiology in this population, most notably at NOD2, in which three causal, uncommon and conditionally independent NOD2 variants reside on a shared background haplotype. We present an analysis of extended haplotypes that showed significantly greater association to CD in the Ashkenazi Jewish population compared with a non-Jewish population (145 haplotypes and no haplotypes with P-value <10(-3), respectively). Two haplotype regions, one each on chromosomes 16 and 21, conferred increased disease risk within established CD loci. We performed exome sequencing of 55 Ashkenazi Jewish individuals and follow-up genotyping focused on variants in these two regions. We observed Ashkenazi Jewish-specific nominal association at R755C in TRPM2 on chromosome 21. Within the chromosome 16 region, R642S of HEATR3 and rs9922362 of BRD7 showed genome-wide significance. Expression studies of HEATR3 demonstrated a positive role in NOD2-mediated NF-κB signaling. The BRD7 signal showed conditional dependence with only the downstream rare CD-causal variants in NOD2, but not with the background haplotype; this elaborates NOD2 as a key illustration of synthetic association.

Metabolic response at repeat PET/CT predicts pathological response to neoadjuvant chemotherapy in oesophageal cancer.

OBJECTIVES: Reports have suggested that a reduction in tumour 18F-fluorodeoxyglucose (FDG) uptake on positron emission tomography (PET) examination during or after neoadjuvant chemotherapy may predict pathological response in oesophageal cancer. Our aim was to determine whether metabolic response predicts pathological response to a standardised neoadjuvant chemotherapy regimen within a prospective clinical trial. METHODS: Consecutive patients staged with potentially curable oesophageal cancer who underwent treatment within a non-randomised clinical trial were included. A standardised chemotherapy regimen (two cycles of oxaliplatin and 5-fluorouracil) was used. PET/CT was performed before chemotherapy and repeated 24-28 days after the start of cycle 2. RESULTS: Forty-eight subjects were included: mean age 65 years; 37 male. Using the median percentage reduction in SUV(max) (42%) to define metabolic response, pathological response was seen in 71% of metabolic responders (17/24) compared with 33% of non-responders (8/24; P = 0.009, sensitivity 68%, specificity 70%). Pathological response was seen in 81% of subjects with a complete metabolic response (13/16) compared with 38% of those with a less than complete response (12/32; P = 0.0042, sensitivity 52%, specificity 87%). There was no significant histology-based effect. CONCLUSIONS: There was a significant association between metabolic response and pathological response; however, accuracy in predicting pathological response was relatively low.

Learning in the COVID-19 pandemic and beyond: development and rapid uptake of an online learning platform for vascular surgery.

INTRODUCTION: With the onset of the COVID-19 pandemic, alternative methods of delivering medical education were rapidly required. An online learning platform was developed with the aim of providing high-quality, accessible learning to vascular specialty trainees. We describe the design, delivery and analysis of the first 15 months of the platform. Although originally a regional initiative, we discuss how popularity and feedback led to a rapid expansion of the training programme internationally. METHODS: A fully online educational platform for vascular surgery specialist trainees was developed. The primary aims and ethos of the programme were that it should be easily accessible from any location, convenient, flexible, cooperative and collaborative, social and free financially to access. All learning resources were researched carefully and based on the UK vascular surgery curriculum and 20 seminal papers targeted in the Vascular Specialist Fellow of the Royal College of Surgeons (FRCSVasc) examination. RESULTS: The project demonstrated that it is feasible to design, build and deliver a postgraduate clinical teaching platform with minimal time requirement, resources and cost while creating and maintaining high-quality content. Rapid national and international uptake has proven there is demand - in addition to overwhelmingly positive feedback from educators and learners, this demonstrates that previously perceived barriers to online education can be overcome. At present, 53 educational sessions have been delivered and are available in the online library, and in the past year (8 December 2020 to 8 December 2021) the website has been accessed 3,877 times. CONCLUSIONS: Although the programme has grown and evolved, a strong focus is being kept on its original ethos and aims - easily accessible, collaborative, free learning resources for all vascular professionals, based on the UK vascular surgery curriculum. Making learning convenient is key. The COVID-19 pandemic may be a watershed moment for a new era of learning. It is an opportunity for people from different backgrounds to share experiences and to develop cohesion within a hospital and network, nationally and worldwide.

Investigating the Role of Amazonian Mesoscale Wind Patterns and Strength on the Spatial Distribution of Martian Bedrock Exposures.

The Martian highlands contain Noachian-aged areally-extensive (>225 km2) bedrock exposures that have been mapped using thermal and visible imaging datasets. Given their age, crater density and impact gardening should have led to the formation of decameter scale layers of regolith that would overlie and bury these outcrops if composed of competent materials like basaltic lavas. However, many of these regions lack thick regolith layers and show clear exposures of bedrock materials with elevated thermal inertia values compared to the global average. Hypothesized reasons for the lack of regolith include: (a) relatively weaker material properties than lavas, where friable materials are comminuted and deflated during wind erosion, (b) long-term protection from regolith development through burial and later exhumation through one or more surface processes, and (c) spatially concentrated aeolian erosion and wind energetics on well-lithified basaltic substrates. To test the third hypothesis, we used the Mars Regional Atmospheric Modeling System to calculate wind erosive strength at 10 regions throughout the Martian highlands and compared it to their thermophysical properties by using thermal infrared data derived from the Thermal Emission Spectrometer to understand the effect that Amazonian mesoscale wind patterns may have on the exposure of bedrock. We also investigated the effect of planet obliquity, Ls of perihelion, and atmospheric mean pressure on wind erosion potential. We found no evidence for increased aeolian activity over bedrock-containing regions relative to surrounding terrains, including at the mafic floor unit at Jezero crater (Máaz formation), supporting the first or second hypotheses for these regions.

The hepatitis B e antigen suppresses IL-1ß-mediated NF-κB activation in hepatocytes.

Previous clinical studies have demonstrated an association between the hepatitis B e antigen and Toll-like receptor (TLR) expression and signalling. Therefore, the aim of this study was to develop an in vitro assay to measure the effect of hepatitis B virus proteins, including the precore protein, on signalling mediated by members of the Toll-like/interleukin 1 (TIR) superfamily, by measuring NF-κB promoter activity. The basal level of NF-κB reporter activity was measured in three hepatocyte cell lines (Huh7, HepG2 and PH5CH8) and one kidney cell line (HEK293) using a luciferase assay. All cell lines were virtually refractory to stimulation with lipopolysaccharide; however, PH5CH8 cells had a robust activation of NF-κB in response to IL-1ß stimulation, with ∼ 40-fold higher activation than the unstimulated control, a higher degree of activation than that observed in either Huh7 and HepG2, or HEK293 and HEK293-TLR2 cells. In PH5CH8 cells transfected with pCI expression constructs and stimulated with IL-1ß, we showed that the precursor form of the precore protein, p25, inhibits NF-κB activation by up to 30% and the cytosolic form, p22, inhibits NF-κB activation by 70%. The core protein, p21, which shares significant homology with the precore protein except for a 10-amino acid extension at the N-terminus, had no effect on NF-κB activation. We hypothesize that the inhibition of IL-1ß-mediated NF-κB activation by the precore protein may be a mechanism that allows the virus to persist, suggesting a role for the pool of precore protein that remains intracellular.

Immunoglobulins in NZB-BL mice. I. Serum immunoglobulin levels and immunoglobulin class of erythrocyte autoantibody.

Determination of serum immunoglobulin levels in NZB and normal mice has indicated that an elevation of the gammaM-globulin level occurs in NZB mice. This can be demonstrated in young NZB mice well before other autoimmune manifestations are observed. A slight elevation of gammaM-globulin was also observed in NZB hybrid mice. Erythrocyte autoantibodies were analyzed by the direct Coombs' test with specific antiimmunoglobulin reagents. Autoantibodies in NZB mice can be of all immunoglobulin classes, although predominantly of gammaG(1)-class. In (NZB x NZC)F(1) hybrid mice, although Coombs' positivity develops around the same age as in NZB mice, there is a greater restriction of the autoantibodies into gammaM- and gammaG(1)-classes. Preliminary attempts to quantitate immunoglobulins coating NZB red cells have shown that there are approximately 400 molecules of gammaM and 5000 molecules of gammaG-globulin per NZB mouse red cell.

Immunoglobulin isoantigens (allotypes) in the mouse. II. Allotypic analysis of three gammaG2-myeloma proteins from (NZB x BALB/c)F1 hybrids and of normal gammaG2-globulins.

Further analysis of the isoantigens (allotypes) of 2 classes of normal mouse immunoglobulins, gammaG(2a) and gammaG(2b), has shown a minimum of 10 specificities for the Ig-1 locus (controlling gammaG(2a)-antigens) and 3 specificities for the Ig-3 locus (controlling gammaG(2b)-antigens). Three gammaG(2)-myeloma proteins of plasma cell tumors induced in (NZB x BALB/c)F(1) mice have been analyzed for the isoantigens they carry. NZB mice are genotypically Ig-1(e) Ig-3(e), while BALB/c are Ig-1(a) Ig-3(a). Two of the myeloma proteins are gammaG(2a)-globulins. One of these, GPC-7, carries all the isoantigenic specificities of the Ig-1(e) allele while the other, GPC-8, carries all the isoantigenic specificities of the Ig-1(a) allele. Thus only one of the parental alleles of the mouse in which the tumor arose is expressed in each of these myeloma proteins. The third myeloma protein GPC-5, also carries the antigens of only one parental strain (NZB). However GPC-5, a gammaG(2b)-globulin, carries only one of the Ig-3 specificities normally associated with gammaG(2b)-globulins of NZB. Most remarkably it also carries one Ig-1 specificity normally associated with gammaG(2a)-globulins of NZB. This is the first analyzed mouse myeloma shown (a) to express some but not all the antigenic specificities normally associated with an allele and (b) to carry antigenic specificities controlled by two distinct immunoglobulin loci. The implications of these findings are discussed in relation to the genetic control of immunoglobulins.

Transplantation behavior of A.TH and A.TL T-cell lymphomas in congenic resistant and hybrid strains.

Seven spontaneously arising T-cell lymphomas originating in A.TH or A.TL mice, which are congenic for the immune response gene (I) chromosomal segment were described. When transplanted into partner strains which were incompatible at the I region, the tumors were rapidly rejected. Rejection was proposed to be due to the presence of antigens controlled by I-region genes.

Defects in hematopoietic differentiation in NZB and NZC mice.

Hematopoietic stem cell activity in inbred NZB and NZC mice has been determined by transplantation and endogenous spleen colony assays. Whereas NZB mice show normal colony-forming unit (CFU) activity in the transplantation assay, they show markedly elevated endogenous CFU. NZC mice also show this markedly elevated endogenous CFU activity, but in the transplantation assay show only about 5-10% of normal CFU counts. When NZC stem cells are tested for CFU activity in irradiated recipients of the H-2(d) type, almost normal colony numbers occur. NZB stem cells however also cannot form colonies in NZC mice. These results suggest that NZC mice have a defect in the micro-environment of the spleen which renders them incapable of allowing transplanted CFU to form colonies. Genetic analysis of both the NZC defect as a CFU recipient, and the elevated endogenous count in NZB and NZC, shows that both are controlled by single recessive genes which are not linked to either coat color, agouti, H-2 or Ig loci. Of even more relevance is the finding that these hematopoietic abnormalities are not linked to the genes involved in controlling autoantibody formation to red cells in the NZB mice. These mice therefore appear to show two distinct hematopoietic abnormalities, the analysis of which may be of considerable value in understanding the detailed events of hematopoietic stem cell differentiation.

Tolerance and immunity to maternally derived incompatible IgG 2a -globulin in mice.

Progeny mice were confronted with maternal gamma-globulin of a different allotype by either back-cross mating, intercross mating, or by foster nursing. In all cases, many mice subsequently produced alloantibodies directed against the incompatible maternal type of IgG(2a)-globulin. In one series of experiments, immunologic tolerance to the maternally derived gamma-globulin was demonstrated to exist in the period before formation of spontaneous antibody. The state of tolerance was then lost, unless maintenance injections of foreign gamma-globulin were given. These studies demonstrate in a natural situation that maternally derived foreign proteins can first induce a state of immunological tolerance which is followed, after disappearance of the antigen, by a state of immunity. As such, this parallels the experimental induction of tolerance to foreign proteins by neonatal injections.

Surface immunoglobulins on thymus and thymus-derived lymphoid cells.

Lymphoid cells from thymus, thoracic duct lymph (TDL), and thoracic duct lymph in irradiated animals reconstituted with allogeneic thymus cells (TTDL) were labeled with radioiodinated anti-immunoglobulins using radioautographic techniques. Thymus and TTDL were labeled (14.4 and 37.0%, respectively) with anti-light chain protein after prolonged exposures (30-60 days). No labeling was observed on thymus and TTDL with anti-polyheavy chain globulin. In contrast 18.5-19.0% of TDL labeled on short exposure (6 days) with anti-polyheavy chain and anti-light chain materials. It is proposed that the difference between the labeling observed on short exposures versus long exposures can be related to the difference in surface density of immunoglobulins between nonthymus-derived (B) and thymus-derived (T) cells. The distribution of labeled cells in the thymus was preferentially among the larger cells (greater than 10 micro diameter). The TTDL population was mostly composed of a larger, blast-like population and the distribution of label was independent of size. As the thymus and TTDL preparations contain almost exclusively T cells, this represents a direct demonstration of surface immunoglobulin light chains on T lymphoid cells.

A receptor for antibody on B lymphocytes. II. Immunochemical and electron microscopy characteristics.

Binding of antibody to the surface of B lymphocytes was shown to involve the Fc piece of the immunoglobulin molecule. This property was not shared equally by all immunoglobulin classes as revealed by direct binding and inhibition studies. Total IgG globulin was found to label cells more heavily than IgM, and IgG1-containing fractions more heavily than IgG2 fractions lacking IgG1. The ability of various purified myeloma proteins to inhibit attachment of antibody to B cells was examined. Pretreatment of B cells with excess IgG(2a), IgA, or light chain proteins failed to inhibit, whereas IgG1 proteins and to a lesser extent Ig(2b) and IgM proteins at the same concentrations did so. At lower protein concentrations, IgG1 myeloma protein alone retained the capacity to inhibit binding. The conclusion was reached that the receptor on B cells for antibody has a marked predilection for the IgG1 class. Although IgM and IgG(2b) antibody may bind, they do so with lower avidity and probably in insignificant amounts if IgG1 antibody is present in excess. No evidence was found to implicate complement in the binding process. For example, heat-inactivated sera at high dilution retained the ability to label B cells, while the use of purified low molecular weight anticomplementary factor, a potent inhibitor of C'3, did not interfere with the formation of the bond between antibody and cell surface. The failure of anti-mouse immunoglobulin F(ab)'(2) fragments to prevent access of antibody to B cells implied that the antibody-binding receptor and antigen-binding (immunoglobulin) receptor are discrete entities on the B cell membrane. Despite this, a marked similarity between their surface distribution was observed on electron microscopy. The antibody-binding receptor was shown to be a marker for mature B cells. It did not appear to be present on hematopoietic precursor stem cells and was lost during differentiation to antibody-forming cells.

Concomitant induction of the cell surface expression of Ia determinants and accessory cell function by a murine macrophage tumor cell line.

This study demonstrates that an uncharacterized soluble factor produced in concanavalin A-induced rat spleen cell suspensions has the capacity to induce the increased expression of cell surface H-2K and H-2D molecules and the expression of I-region gene products on murine monocyte-macrophage lineage tumors that are not Ia positive in the absence of the factor. In parallel with induction of serologically defined Ia specificities, Ia-induced WEHI-3 macrophage tumor cells are capable of providing accessory cell function in stimulating IL-2 production by T-T hybridomas that are activated in a major histocompatibility complex-restricted, antigen-dependent fashion. The uninduced Ia-negative WEHI-3 tumor cells do not trigger a comparable response in this assay system.