Negative Fraction Research Articles

Cadmium uptake and transport in vegetables near a zinc–lead mine: Novel insights from Cd isotope fractionation

In this study, Cd isotope analysis was conducted on drought-tolerant (cowpea and sesame) and less drought-tolerant vegetables (water spinach, green pepper, and mung bean) to elucidate the mechanisms underlying Cd uptake and transport. Cd isotopes in plants were identical to or lighter than those in the available pool and exhibited negative fractionation from roots to straws (Δ114/110Cd = −0.22 ‰ to −0.17 ‰) in drought-tolerant vegetables, whereas contrasting results were obtained for less drought-tolerant vegetables (Δ114/110Cd = −0.050 ‰ to 0.39 ‰). Positive Cd isotope fractionation from straws to fruits in drought-tolerant vegetables (Δ114/110Cd = 0.33 ‰ ± 0.03 ‰ and 0.10 ‰ ± 0.03 ‰, respectively) was observed, whereas negligible or negative fractionation was found in less drought-tolerant vegetables (Δ114/110Cd = 0.01 ‰ ± 0.04 ‰ and −0.34 ‰ ± 0.02 ‰, respectively). The vast secretion of organic acids might have led to positive available pool-to-roots and negative roots-to-straws isotope fractionation in drought-tolerant vegetables. In contrast, preferential xylem transport resulted in negative straws-to-fruits isotope fractionation in less drought-tolerant vegetables. This study demonstrated that Cd isotope fractionation in the soil-plant system is associated with plant drought tolerance, and drought-tolerant and less-tolerant plants developed a distinct Cd detoxification mechanism, corresponding to a reversed fractionation of Cd isotopes.

Refined selection of individuals for preventive cardiovascular disease treatment with a transformer-based risk model

Abstract Background Although statistical models have been commonly used to identify patients at risk of cardiovascular disease (CVD) for preventive therapy, these models tend to over-recommend therapy [1]. Moreover, in populations with pre-existing diseases, the current approach is to indiscriminately treat all individuals, as modelling in this context is currently inadequate. Aims To develop and validate a novel deep learning model, the Transformer-based Risk assessment survival model (TRisk) model, for prediction of 10-year risk of CVD in both the general population and individuals with diabetes. Methods An open cohort of 3 million adults aged 25 to 84 years free of CVD and statin use was identified using linked primary and secondary electronic health records from 291 and 98 general practices in England, which were used for TRisk model development and external validation, respectively. Comparison against two commonly recommended statistical models (QRISK3 [2] and SCORE2 [3]) and a deep learning derivation of QRISK3 was conducted. Additional analyses compared discriminatory performance in other age groups, by sex, and across categories of socioeconomic status. Similar analyses were conducted in a subgroup of 59 thousand patients with diabetes. Results TRisk demonstrated superior discrimination (C index in the general population: 0.910; 95% confidence interval [CI]: 0.906 to 0.913). TRisk’s performance was found to be less sensitive to population age range than the benchmark models and outperformed other models also in all stratified analyses. All models were overall well-calibrated. In decision curve analyses, TRisk demonstrated greater net benefit than benchmark models across the range of relevant thresholds. In impact analyses, at the widely recommended 10% risk threshold and the higher 15% threshold for the general population, TRisk reduced both the total number of patients classified at high risk (by 27% and 39% respectively) and the number of false negatives as compared with currently recommended strategies. TRisk similarly outperformed other models in patients with diabetes. Compared with the widely recommended treat-all policy approach for patients with diabetes, TRisk at a 10% risk threshold would lead to deselection of 24% of individuals with a small fraction of false negatives (0.2% of cohort). Conclusion TRisk enabled a more targeted selection of individuals at risk of CVD compared to benchmark statistical and deep learning models, in both the general population and patients with diabetes. Incorporation of TRisk into routine clinical care would allow a reduction in the number of treatment-eligible patients in general population and subgroup with diabetes by approximately one-third and one-quarter respectively while preventing at least as many events as with currently adopted approaches.Models' discrimination on both cohortsImpact analyses for both cohorts

Isotope-Based Characterization of Soil Elemental Mercury Emissions from Historical Mercury Mining Areas: Driving Pathways and Relative Contributions.

Photo-, microbial, and abiotic dark reduction of soil mercury (Hg) may all lead to elemental mercury (Hg(0)) emissions. Utilizing lab incubations, isotope signatures of Hg(0) emitted from mining soils were characterized to quantify the interplay and contributions of various Hg reduction pathways, which have been scarcely studied. At 15 °C, microbial reduced Hg(0) showed a negative mass-dependent fractionation (MDF) (δ202Hg = -0.30 ± 0.08‰, 1SD) and near-zero mass-independent fractionation (MIF) (Δ199Hg = 0.01 ± 0.04‰, 1SD), closely resembling dark reduced Hg(0) (δ202Hg = -0.18 ± 0.05‰, Δ199Hg = -0.01 ± 0.03‰, 1SD). In comparison, photoreduced Hg(0) exhibited significant MDF and MIF (δ202Hg = -0.55 ± 0.05‰, Δ199Hg = -0.20 ± 0.07‰, 1SD). In the dark, Hg isotopic signatures remained constant over the temperature range of 15-35 °C. Nonetheless, light exposure and temperature changes together altered Hg(0) MIF signatures significantly. Isotope mixing models along with Hg(0) emission flux data highlighted photo- and microbial reduction contributing 79-88 and 12-21%, respectively, of the total Hg(0) emissions from mining soils, with negligible abiotic dark reduction. Microorganisms are the key driver of soil Hg(0) emissions by first dissolving HgS and then promoting ionic Hg formation, followed by facilitating the photo- and microbial reduction of organically bound Hg. These insights deepen our understanding of the biogeochemical processes that influence Hg(0) releases from surface soils.

Experimental investigation of an electronegative cylindrical capacitively coupled geometrically asymmetric plasma discharge with an axisymmetric magnetic field

In this study, we have investigated the production of negative ions by mixing electronegative oxygen gas with electropositive argon gas in a geometrically asymmetric cylindrical capacitively coupled radio frequency plasma discharge. The plasma parameters such as density (electron, positive, and negative ion), negative ion fraction, and electron temperature are investigated for fixed gas pressure and increasing axial magnetic field strength. The axisymmetric magnetic field creates an E × B drift in the azimuthal direction, leading to the confinement of high-energy electrons at the radial edge of the chamber, resulting in decreased species density and negative ion fraction in the plasma bulk. However, the electron temperature increases with the magnetic field. It is concluded that low magnetic fields are better suited for negative ion production in such devices. Furthermore, in addition to the percentage ratio of the two gases, the applied axial magnetic field also plays a vital role in controlling negative ion fraction.

Cadmium Isotope Fractionation during Adsorption onto Edge Sites and Vacancies in Phyllomanganate.

Cadmium (Cd) geochemical behavior is strongly influenced by its adsorption onto natural phyllomanganates, which contain both layer edge sites and vacancies; however, Cd isotope fractionation mechanisms at these sites have not yet been addressed. In the present work, Cd isotope fractionation during adsorption onto hexagonal (containing both types of sites) and triclinic birnessite (almost only edge sites) was investigated using a combination of batch adsorption experiments, extended X-ray absorption fine structure (EXAFS) spectroscopy, surface complexation modeling, and density functional theory (DFT) calculations. Light Cd isotopes are preferentially enriched on solid surfaces, and the isotope fractionation induced by Cd2+ adsorption on edge sites (Δ114/110Cdedge-solution = -1.54 ± 0.11‰) is smaller than that on vacancies (Δ114/110Cdvacancy-solution = -0.71 ± 0.21‰), independent of surface coverage or pH. Both Cd K-edge EXAFS and DFT results indicate the formation of double corner-sharing complexes on layer edge sites and mainly triple cornering-sharing complexes on vacancies. The distortion of both complexes results in the negative isotope fractionation onto the solids, and the slightly longer first Cd-O distances and a smaller number of nearest Mn atoms around Cd at edge sites probably account for the larger fractionation magnitude compared to that of vacancies. These results provide deep insights into Cd isotope fractionation mechanisms during interactions with phyllomanganates.

Zinc isotope fractionation during coprecipitation with amorphous iron (hydr)oxides

Coprecipitation facilitates the incorporation of Zn into the lattice of Fe (hydr)oxide, which is a crucial mechanism causing low Zn bioavailability in widespread Zn-deficient soils. Zn isotopes are a potential indicator of the coprecipitation of Zn and Fe (hydr)oxides in soils. However, the Zn isotope fractionation caused by coprecipitation with amorphous Fe (hydr)oxides (e.g., ferrihydrite) and the impact of environmentally relevant conditions remain unknown. Here, the molecular mechanism and impact factor of Zn isotope fractionation during Zn2+-Fe3+ coprecipitation under natural soil conditions (including different pH values and initial Zn/Fe ratios) were investigated by combining isotope ratio measurements, extended X-ray absorption fine structure (EXAFS) analyses, and density functional theory (DFT) geometry optimization. The results show that aqueous Zn (Znaq) can be complexed on the ferrihydrite surface (Znads) with positive isotope fractionation (Δ66Znads–aq = 0.42 ± 0.09 ‰) or directly incorporated into the ferrihydrite lattice with slightly negative isotope fractionation (Δ66Znstru-aq = −0.08 ± 0.03 ‰). In addition, over time, the surface-complexed Zn can further transform to a layered structure on the surface but exhibits limited Zn isotope fractionation. According to Zn K-edge EXAFS, the Zn isotope fractionation of surface complexation and incorporation of Znaq are related to the decrease in the Zn–O bond strength in the order surface complexed Zn (RZn–O = 2.00 Å) > aqueous Zn (RZn–O = 2.08 Å) > directly incorporated Zn (RZn–O = 2.06–2.07 Å with slight distortion). The limited Zn isotope fractionation associated with the transformation of Znads is related to the reconfiguration of Znads during the nucleation of octahedral Zn layer structures. Furthermore, the primary process controlling Zn isotope fractionation is different under varying experimental conditions, leading to distinct Zn isotope fractionation between the bulk solid and solution during Zn–Fe coprecipitation. Low pH values (5.0–6.0) and initial Zn/Fe ratios (0.01–0.02) favor the direct precipitation of Znaq, resulting in slightly negative Zn isotope fractionation between the bulk solid and solution. In contrast, high pH values (7.0–7.5) and initial Zn/Fe ratios (0.1–0.2) favor surface complexation and the transformation of Znads, leading to heavy Zn isotope enrichment in the bulk solid. These findings offer novel insights into the primary mechanism through which Fe (hydr)oxides regulate Zn bioavailability in Zn-deficient soils and provide experimental evidence supporting the potential application of Zn isotopes as an indicator for comprehending the fate of Zn controlled by Fe (hydr)oxides in soils.

Cross-section without factors: a string model for expected returns

Many asset pricing models assume that expected returns are driven by common factors. We formulate a model where returns are driven by a string, and no-arbitrage restricts each expected return to capture the asset's granular exposure to all other asset returns: a correlation premium. The model predicts fresh properties for big stocks, which display higher connectivity in bad times, but also work as correlation hedges: they contribute to a negative fraction of the correlation premium, and portfolios that are more exposed to them command a lower premium. The string model performs at least as well as many existing linear factor models.

Combretum aculeatum Vent (Combretaceae) hydroethanolic root bark extract attenuates D-galactoseinduced cognitive impairment, oxidative, and hippocampi dysfunction in mice

Introduction: Combretum aculeatum is a plant of Combretaceae family. In traditional medicine, it is used to treat schizophrenia. The aim of this study was to assess the possible impacts of hydroethanolic extract of C. aculeatum root bark on D-galactose (D-Gal)-induced amnesia in mice. Methods: Memory was tested using Y-maze test, radial arms maze (RAM), and new item appreciation. Mice brains were collected for histological and biochemical testing. Results: Combretum aculeatum substantially (P < 0.001) improved the ratio of spontaneous alternation versus negative control. In addition, discrimination index, and the time required to appreciate the new item were reversed considerably (P < 0.001) in mice receiving the extract opposed to the negative control fraction in the novel object appreciation test. The frequency of working memory mistakes was reversed in receiving extract categories versus negative control animals in RAM essay. Various doses of the extract substantially (P < 0.001) diminished the level of malondialdehyde (MDA), and crucially enlarged superoxide dismutase (SOD) and catalase activity as opposed to the negative control. Furthermore, all doses of the extract had a restructuring effect on the organization of hippocampal cells. Conclusion: Combretum aculeatum improved cognitive impairment possibly thought its antioxidant activity.

Trophic role and predatory interactions between the blue crab, Callinectes sapidus, and native species in open waters of the Ebro Delta

The Ebro Delta has become a major blue crab (Callinectes sapidus) fishery area in the NW Mediterranean, but there is limited information on factors controlling the abundance of populations in open waters, a crucial habitat for ovigerous females. Here, we use a stable isotope approach (δ15N and δ13C), to assess blue crab trophic position, the potential consumption of food items using mixing models, and the isotopic niche overlap with local commercial species. For Octopus vulgaris, a potential blue crab key predator, a trophic enrichment experiment was also conducted to further assess predation control in wild populations. The blue crab showed 1.6 times higher trophic position than in other habitats of the Ebro Delta, and similar to that found in the harbor crab, Liocarcinus depurator, and several predatory fish. Additionally, the isotopic niche of blue crabs showed overlaps from 46.2 to 14.9% with native predators, and mixing models also suggest even dietary contributions throughout the food web. For O. vulgaris, field results showed a trophic position of 3.93, lower than that of blue crab, and lower δ15N signatures were also obtained in a captivity experiment drawing negative fractionation (−1.1‰). We conclude that high dietary contribution of animal prey might provide a high protein diet that could be crucial for allowing the maintenance of a large local population, but the overall functional trophic similarity could also disfavor local native species. The similarity between experimental fractionation and field differences between predator and prey (−1.6‰) suggests that predation of blue crab is possible, but further research is needed to clarify the metabolic routes involved in octopus δ15N fractionation.

Advancing CAR-T Therapy in Acute Lymphoblastic Leukemia: Multi-Omic Analyses of CD19-Directed CAR-T Cells Enabled By an Ex Vivo Co-Culture Platform

Background: Acute Lymphoblastic Leukemia (ALL) has witnessed remarkable progress in the treatment of patients with Chimeric Antigen Receptor T-cell (CAR-T) therapies, leading to high response rates and durable remissions. However, challenges persist for patients who do not respond to CAR-T treatment, experience relapse, or encounter adverse events such as cytokine release syndrome. Overcoming these challenges necessitates a deeper understanding of mechanisms and the development of advanced technologies and assays. This study focuses on harnessing the potential of an innovative microfluidic platform for high-throughput cell-based assays, enabling multi-omics and functional analysis using a minimal amount of primary patient cells. By utilizing this platform, we aim to gain insight into the dynamics of primary patient material by evaluating responses to aid in selection of promising CAR-T enhancements or combination therapies designed to overcome mechanisms of CAR-T failure or relapse to improve clinical outcomes. Methods: For the cytotoxicity assay, we utilized autologous or allogeneic pairs of CD19-directed CAR-T cells and T-cell negative fractions from ALL patients' blood apheresis samples, containing CD19-expressing cancer cells. A negative control was established using untransduced T-cells. To ensure high cell viability and purity, the cells underwent careful processing and were labeled with long-lived dyes for tracking. Multiple cell suspensions with precise Effector to Target to Negative Fraction ratios were prepared. Only a few thousand cells from each coculture suspension were seeded in microfluidic culture chambers, which required no fluidic control system. Experiments were conducted for up to 96 hours with media replacement every 24 hours. At each endpoint, media was collected for cytokine quantification, and cells were stained for live/dead counting as well as quantification of intracellular or surface marker expression by imaging. After image acquisition, cells were collected for RNA extraction and gene expression analysis. Results: Results demonstrated that the developed co-culture platform allows precise control over culture conditions, ensuring culture of accurate ratios of highly complex cell populations. The viability of primary cells in the device guaranteed functional relevance of the cytotoxicity assays for up to a 96-hour timepoint. Strong CAR-T-mediated cell killing was observed, with a dose-dependent effect depending on the co-culture's relative CAR-T content. Less than 50% cell survival was observed in cocultures with Effector to Target ratio 1:1, as compared to the untransduced T-cell control. Furthermore, strong CAR-T activation was observed with over 60% of CART expressing CD25 and CD69 surface markers, as measured by imaging. A multi-color staining panel allowed for precise monitoring of changes over time within the assay of immune cell populations, including cancer cells, monocytes/macrophages, and NK cells, among others. Cytokine analysis revealed an increased secretion of IL-2, TNFa, IFNg, and Granzyme B in coculture with CAR-T cells compared to untransduced T-cells, with a dose-dependent effect based on the CAR-T cell ratio. Finally, RNA isolated from remaining cells at the end of the assay was of high quality with yields sufficient to enable transcriptome analysis for each treatment condition. Discussion: This study contributes to the development of innovative multiplexed high throughput strategies for in vitro functionality assessments in cell therapy, including combination studies and deeper mechanistic understandings. The platform's ability to generate fully integrated, multi-omics readings from complex allogeneic and autologous cocultures of CAR-T and patient cells offers a new frontier in early discovery and translational cancer research. This opens new possibilities for drug discovery, drug screening, and disease modeling, ultimately advancing our understanding of complex biological processes and accelerating therapeutic development. Future directions include exploring mechanisms of resistance and sensitivity, including donor-specific effector cell functionality, combination studies, impacts of immunosuppressive cell types, patient heterogeneity of target antigen expression or other co-stimulatory molecules, and evaluation of product potency or fitness of incoming apheresis material.

Analysis of Metabolic Mechanism of CD19CAR-T with CD79A/CD40 Co-Stimulatory Domain Based on Comparison with Established CAR-Ts

Introduction: Anti-CD19 chimeric antigen receptor T cell (CD19CAR-T) therapy has achieved significant clinical outcomes and changed treatment strategy in B-cell malignancies. However, recurrence after CAR-T therapy remains a major obstacle due to poor peak expansion and short persistence of CAR-T cells. We have reported that CD19CAR-T incorporating a co-stimulatory domain consisting of CD79A and CD40 (79A40CAR) provided superior anti-tumor efficacy and survival compared to CD19CAR-T with CD28 or 4-1BB (CD28CAR/4-1BBCAR) in mice xenograft models. 79A40CAR exhibited sustained NF-κB phosphorylation and superior in vivo efficacy compared to the CD28CAR or 4-1BBCAR. (Molecular Therapy 2021, Julamanee J et al.) In this study, we aimed to clarify the molecular mechanism of 79A40CAR by comparing with CD28/4-1BBCAR in terms of T-cell metabolism. Methods: CD8+T cells were isolated from PBMCs of healthy donors and retrovirally transduced with either CD28IC-, 4-1BBIC-, or 79A40IC-CD19CARs. CD28CAR, 4-1BBCAR, or 79A40CAR were cocultured with 100 Gy irradiated NALM6 at E:T ratio = 1:5, respectively. Repeated antigen stimulations (Ag-stim) were performed, and day 7 after each stimulation was subjected to analysis. RNA-seq, expansion capacity, T-cell phenotype, and metabolic evaluation using a seahorse flux analyzer (Agilent Technologies) were performed. After Ag-stim, CD45RA negative CD62L positive and CD45RA negative CD62L positive CAR-T cell fractions were sorted as central memory (T CM) and effector memory (T EM), respectively. T CM and T EM received Ag-stim. After Ag-stim, we performed evaluation of mitochondrial functionality with MitoTracker based on mitochondrial mass and membrane potential. Results: All CD19CAR genes were successfully transduced into primary CD8+ T-cells. To understand the differences between co-stimulatory domains, we first confirmed bulk RNA-seq, expansion capacity, and T-cell phenotype. After Ag-stim, 79A40CAR showed enhancement of the NF-κB pathway. We detected few DEGs when 79A40CAR compared to 4-1BBCAR, however, we detected 162 upregulated DEGs when 79A40CAR compared to CD28CAR. Principal component analysis of gene expression demonstrated similarity between 4-1BBCAR and 79A40CAR. We confirmed that 79A40CAR had superior expansion capacity than CD28, or 4-1BBCAR after repeated Ag-stim. PD-1 and Tim-3 expression in 79A40CAR was lowest after repeated Ag-stim. In the evaluation of T-cell differentiation, 4-1BB and 79A40CAR showed an earlier transition to the effector phenotype compared to CD28CAR after Ag-stim. To evaluate differences in terms of T-cell metabolism, we performed metabolic analysis using a seahorse flux analyzer. After repeated Ag-stim, 4-1BB and 79A40CAR showed a predominant oxidative phosphorylation metabolic pathway (Fig1A) and higher spare respiratory capacity (SRC), which has been correlated with mitochondrial functionality (Fig1B), than CD28CAR. After repeated Ag-stim, 4-1BB and 79A40CAR demonstrated higher ECAR than CD28CAR. Because we observed differences in the expression of T-cell differentiation phenotype, we performed cell sorting to evaluate T-cell metabolism of each differentiation phenotype. After applying repeated Ag-stim to T CM and T EM-derived CD19CAR-T, mitochondrial functionality was evaluated using Mitotracker. T CM- (Fig1C) and T EM- (Fig1D) derived 4-1BB and 79A40CAR had more functional mitochondria and fewer dysfunctional mitochondria compared to CD28CAR. Conclusion: 79A40CAR seems to preserve the mitochondrial oxidative phosphorylation-dominant metabolic pathway and supply energy demands for a longer period, especially when compared to CD28CAR. 79A40CAR showed functionally very similar to 4-1BBCAR. We showed that CD19CAR-Ts had different metabolic pathway according to the different co-stimulatory domains. To reveal additional metabolic pathway differences between 4-1BBCAR and 79A40CAR, we are now conducting proteomics analysis.

Characterization of Multiple Myeloma Subpopulations Associated with Treatment Resistance By Single-Cell RNA Sequencing

Multiple myeloma (MM) has extensive genomic and transcriptomic heterogeneity with identical variable-diversity-joining (VDJ) region recombination of immunoglobulin loci (called repertoire). Even though new treatment approaches have been developed, most cases of MM remain incurable. Recent evidence suggests that epigenetic cell plasticity and changes in cell state underlie the therapy resistance, but the mechanisms remain largely unknown. We attempted to understand the comprehensive architecture of the entire tumor cell populations using repertoire clonality as a fingerprint. First, we subdivided bone marrow samples of primary MM at different stages (n=8) into 11 fractions and performed bulk RNA sequencing (bulk RNA-seq). Analysis of the repertoire revealed that tumor cells were present at very low frequencies in some minor fractions other than the main fraction (Lineage marker - (Lin -)/CD19 -/CD38 +/CD138 +) as the disease stage progressed. Importantly, MM cells were consistently present in CD138 negative fraction (Lin -/CD19 -/CD38 +/CD138 -) regardless of disease stage. Flow cytometry (FCM) analysis confirmed that CD138negative MM cells were present at a rate of less than 10% across the disease stages. Compared with CD138 positive cells, they were slowly cycling and showed unique morphology with large cytoplasm (n=59). To achieve a selective and comprehensive evaluation of MM population, we performed single-cell RNA sequencing (scRNA-seq) coupled with VDJ targeted sequencing (scVDJ-seq) on Lin -/CD19 -/CD38 +(without CD138 gating) samples from 11 MM patients, including those at diagnosis and in remission with minimal residual disease (MRD). This method allowed us to discriminate MM cells (20,084 cells) clearly from others by clonal repertoire sequence even in the post-treatment samples with extremely small populations of tumor cells. Because mRNA levels of CD138 did not correspond to the differential intensity in FCM, we defined gene signatures of CD138 positive and negative cells respectively by bulk RNA-seq (n=6). As a result of integration of these gene signatures using Gene Set Enrichment Analysis (GSEA), the heterogeneity on the CD138 axis was also unraveled in scRNA-seq data. Interestingly, gene ontology (GO) analysis of bulk RNA-seq data revealed that several terms related to H3K4 methylation were positively enriched in CD138 negative fraction, which included KMT2A, KMT2B, KMT2C and KMT2D. Western blotting confirmed significant reductions in H3K4me1 and H3K4me3 levels in CD138 negative cells. Furthermore, primary MM cell culture indicated the plasticity between CD138 positive and negative cells, which was also suggested by trajectory/velocity analysis of scRNA-seq data. Collectively, it was suggested that epigenetic changes, including those in histone modifications, contribute to the heterogeneity along the CD138 axis. We next compared the size of each cluster before and after treatment in scRNA-seq data to identify the MM populations associated with treatment resistance. Remarkably, we identified unique clusters which tended to remain at high proportions against treatment in the CD138 negative population. They were characterized by low expression of MHC class-I genes and TNFRSF17/BCMA, as well as 198 significantly upregulated genes including MCL1, MALAT1 and NEAT1, which have been previously implicated in treatment resistance of MM. To systematically interrogate their contribution in MM cell survival, we performed CRISPR/Cas9 knockout screening of selected 100 genes, which were differentially up-regulated in the resistant clusters and associated with poor prognosis in a MMRF CoMMpass Study database. GO analysis of indispensable genes reproducibly identified among three MM cell lines revealed enrichment of several pathways related to mRNA splicing. These results suggested that the RNA splicing pathway is a promising therapeutic target to eliminate resistant populations. Indeed, we confirmed that inhibition of a splicing factor by CRISPR/Cas9-mediated knockout and pharmacological protein degradation suppresses MM cell proliferation effectively. Taken together, we achieved a complete view of MM diversity by using repertoire clonality and confirmed heterogeneity along the CD138 axis, which may be characterized by epigenetic plasticity. Our results highlight a novel subpopulation as a candidate therapeutic target for cure.

CD70/CD27 Signaling Promotes Expansion of Clonal Plasma Cells in Multiple Myeloma and Is a Promising Therapeutic Target in Advanced Disease

Introduction: Multiple Myeloma (MM) is a hematological malignancy characterized by the clonal expansion of malignant plasma cells within the bone marrow (BM). In the last decade, substantial advances in the treatment of MM patients have been made resulting in improved progression-free and overall survival rates. However, to date, MM remains incurable. CD70 and CD27 belong to the TNF receptor family. CD70 is the only known ligand for its receptor CD27. Physiologically, the CD70/CD27 interaction is involved in the regulation of immune cell expansion. However, numerous solid and hematological cancer types overexpress CD70. Aside, CD70 and CD27 co-expression has been described on leukemic stem cells (LSC) in acute myeloid leukemia patients where an autonomous signaling activation drives LSC self-renewal and expansion. While CD70/CD27 signaling plays a key role in the differentiation process of B lymphocytes into plasma cells, only little is known about its impact on myelomagenesis and MM progression. In this study, we analyzed the role of CD70/CD27 signaling in the pathogenesis of MM and studied the possibility to target CD70 by monoclonal antibodies to treat the disease. Materials and Methods: We evaluated CD70 and CD27 gene expression levels and its correlations with overall survival (OS), MM progression and high-risk molecular alterations using different databases (CoMMpass IA15, GSE6477). CD70 and CD27 protein expression patterns were analyzed in a newly established MM tissue microarray (TMA) and a collection of matched BM samples from initial and advanced disease stages (in collaboration with the Institutes of Pathology in Bern, Basel and Tübingen). To assess the functional role of CD70/CD27 signaling on MM initiation and expansion, we generated CD70 knock-out MM cell lines and newly established patient-derived xenografts (PDX) using CRISPR/Cas9 genome editing. CD70-expressing MM cell lines or patient-derived myeloma cells were grown in NOD/SCID (NSG) mice. Moreover, different monoclonal antibodies (Abs) and Ab-derivates were tested including a CD70 blocking antibody (αCD70) and an antibody-dependent cell-mediated cytotoxicity (ADCC) optimized antibody (cusatuzumab, ArgenX) that activates FcγR-expressing effector cells to assess the therapeutic potential of targeting CD70 in MM. Results: High CD70 gene expression levels correlate with increased MKI67 gene expression and elevated lactate dehydrogenase serum levels indicating a more rapid proliferation of CD70-expressing myeloma cells. In addition, high CD70 expression in MM patients correlated with shorter OS. Longitudinal analyses of CD70 gene expression in the precursor stages monoclonal gammopathy of undetermined significance and smoldering myeloma as well as MM and relapsed MM stages showed highest CD70 gene expression levels in relapsed MM patients. Assessment of the molecular background further revealed increased frequencies of the prognostically unfavorable genomic translocations t(4;14) and t(14;16), as well as gain/amplification of the chromosomal region 1q21 (1q+) in CD70 high expressing MM patients. In line with our previous findings, a TMA analysis of over 200 MM core biopsies as well as a longitudinal analysis of matched biopsies from patients at initial diagnosis and MM relapse indicated that CD70 was overexpressed in advanced disease. Functionally, we found that CD70/CD27 signaling promotes plasma cell survival inducing the expansion of MM cell lines and primary malignant plasma cells. CD70 blockade (αCD70) or CD70 knock-out (CD70 KO) resulted in a significant inhibition or complete abrogation of growth of a CD70-expressing MM patient-derived xenograft (PDX) (EMM01 CD70+) compared to controls (Figure 1). Interestingly, in vivo limiting dilution assays revealed that CD70+CD27+ double-positive myeloma subpopulations showed significantly higher cancer stem cell frequencies compared to CD70 negative fractions suggesting that CD70 expression might be related to an immature or stem-cell like phenotype. The ADCC-optimized CD70 specific antibody custatuzumab efficiently eliminated CD70-expressing MM cells in the presence of PBMC-derived natural killer (NK) cells in vitro and in a PDX model in vivo. Conclusion: Our results indicate that CD70-expressing MM cells have a stem-cell like phenotype and propagate the disease. Targeting CD70 is a promising new therapy especially in advanced disease.

Insights into the reduction of methylmercury accumulation in rice grains through biochar application: Hg transformation, isotope fractionation, and transcriptomic analysis

Methylmercury (MeHg), a potent neurotoxin, easily moves from the soil into rice plants and subsequently accumulates within the grains. Although biochar can reduce MeHg accumulation in rice grains, the precise mechanism underlying biochar-mediated responses to mercury (Hg) stress, specifically regarding MeHg accumulation in rice, remains poorly understood. In the current study, we employed a 4% biochar amendment to remediate Hg-contaminated paddy soil, elucidate the impacts of biochar on MeHg accumulation through a comprehensive analysis involving Hg isotopic fractionation and transcriptomic analyses. The results demonstrated that biochar effectively lowered the levels of MeHg in paddy soils by decreasing bioavailable Hg and microbial Hg methylation. Furthermore, biochar reduced the uptake and translocation of MeHg in rice plants, ultimately leading to a reduction MeHg accumulation in rice grains. During the process of total mercury (THg) uptake, biochar induced a more pronounced negative isotope fractionation magnitude, whereas the effect was less pronounced during the upward transport of THg. Conversely, biochar caused a more pronounced positive isotope fractionation magnitude during the upward transport of MeHg. Transcriptomics analyses revealed that biochar altered the expression levels of genes associated with the metabolism of cysteine, glutathione, and metallothionein, cell wall biogenesis, and transport, which possibly enhance the sequestration of MeHg in rice roots. These findings provide novel insights into the effects of biochar application on Hg transformation and transport, highlighting its role in mitigating MeHg accumulation in rice.

Mercury isotope constraints on the timing and pattern of magmatism during the end-Triassic mass extinction

The End-Triassic Mass Extinction (ETME) was broadly coincident with the emplacement of the Central Atlantic Magmatic Province (CAMP). However, the relationship between evolutionary phases of CAMP (i.e. prolonged shallow intrusive pulses and later extrusives) and the ETME is not clear. Here, a high-resolution record of changing magmatic activity is established using mercury (Hg) isotopes through a Triassic-Jurassic sedimentary succession deposited on the northern flank of CAMP. Successive negative and positive mass independent fractionation (MIF) of odd Hg isotopes through the end-Triassic interval in this succession suggest dominantly thermogenic Hg release from intrusive heating of organic-rich sedimentary rocks, followed by mainly volcanogenic input of mantle-sourced Hg. This inferred pattern is also supported by correlations to available CAMP ages. Minimum calculated volumes of thermogenic and volcanogenic Hg needed to drive the observed shifts at the basin-scale are plausible based on likely CAMP Hg release volumes, and biotic turnover in the studied region was unlikely to have occurred before the onset of volcanogenic Hg release.

High expression of DNMT3A and DNMT3B regulatory factors of TGFB in non-neoplastic liver tissues of HCC.

Increased expression of TGFB regulatory factors DNMT3A and DNMT3B in non-neoplastic liver tissues of HCC patients is the goal of this study. Furthermore, we demonstrate that TGF- is capable of elevating the percentage of CD133+ cells present in liver cancer cell lines in a manner that is both consistent and long-lasting over several cell divisions. This process is linked to stable alterations in DNA methylation that occur over the whole of the genome and continue even after cell division. In addition, the silencing of de novo DNA methyl-transferases with siRNA is able to inhibit the phenotypic changes that are induced by TGF-. According to the findings of our research, there is a self-sustaining interaction between the DNA methylation machinery and the TGF- signaling pathway, which may be significant in the development of cellular phenotypes. CD133 positive and negative fractions expand within liver cancer cell lines in proportions that remain stable throughout time. In contrast to their CD133- counterparts, MACS-sorted CD133+ Huh7cells demonstrated the ability to shape themselves into spheres when grown under non-attachment circumstances. This study also found that the TGF- is responsible for the de novo induction of CD133, which is linked to an increase in the expression of DNMT3 genes and there is a correlation between the TGF-induced transition in the cell subpopulation and a distinct DNA methylome. TGF- has the potential to generate genome-wide alterations in DNA methylation, which ultimately leads to a persistent shift in the fraction of liver cancer cell subpopulations.

Cd isotope fractionation in a soil-rice system: Roles of pH and mineral transformation during Cd immobilization and migration processes

Cd speciation in soil and its transport to rice roots are influenced by the soil pH, oxidation-reduction potential, and mineral transformation; however, the immobilization and migration of Cd in soil-rice systems with different pH values under distinct water regimes remain unclear. This study used Cd isotope fractionation, soil physical analysis, and root gene quantification to elucidate the immobilization and transport of Cd in different soil-rice systems. In drainage soils, the high soil pH enhanced the transformation and magnitude of negative fractionation of Cd from MgCl2 extract to FeMn oxide-bound pool; however, it favored Cd uptake and root-to-grain transport. Compared with drainage regimes, the flooding regimes shifted fractionation toward heavy isotopes from MgCl2-extracted Cd to FeMn oxide-bound Cd in acidic soils (∆114/110CdMgCl2 extract – FeMn oxide-bound Cd = −0.09 ± 0.03 ‰) and to light isotopes from MgCl2-extracted Cd to carbonate-bound Cd in neutral and alkaline soils (∆114/110CdMgCl2 extract – carbonate-bound Cd = 0.29–0.40 ‰). The submerged soils facilitated the forming of carbonate and poorly crystalline minerals (such as ferrihydrite), which were transformed into highly crystalline forms (such as goethite). These results demonstrated that the dissolution-precipitation process of iron oxides was essential for controlling soil Cd availability under flooding regimes, and the relative contribution of carbonate minerals to Cd immobilization was promoted by a high soil pH. Flooding regimes induced lower expressions of OsNRAMP1 and OsNRAMP5 to limit the uptake of light Cd isotopes from MgCl2-extract pool, whereas a teeter-totter effect on gene expression patterns in roots (including those of OsHMA3 and OsHMA2) limited the transport of heavy Cd isotopes from root to grain. These findings demonstrate that flooding regimes could exert multiple effects on soil Cd immobilization and Cd transport to grain. Moreover, alkaline soil was conducive to forming carbonate minerals to sequester Cd.

Solving non-linear fixed-charge transportation problems using nature inspired non-linear particle swarm optimization algorithm

The main objective of this study is to formulate and solve the model of non-linear fixed-charge transportation problem (NFCTP), which is one of the NP-hard problems. This type of problem is challenging to solve using conventional methods because these methods are usually inefficient, costly to compute, and may stuck at the local optimum solution. To overcome this issue, a non-linear particle swarm optimization (NPSO) algorithm with new non-linear acceleration parameters is proposed in this study. Also, to preserve the feasibility condition (non-negative integer solution) of the transportation problem, two novel negative repair and fraction repair strategies have been incorporated into the proposed NPSO. The efficiency of the NPSO algorithm is tested on both the small-scale and large-scale NFCTPs. The dataset are considered from the existing studies. The obtained results are compared with those obtained by the spanning tree-based genetic algorithm (st-GA), priority-based genetic algorithm (pb-GA), and minimum cost flow-based genetic algorithm. The comparative study reveals that for all the considered problems, the proposed NPSO provides better feasible solutions in lesser computational time. Furthermore, the effectiveness of the NPSO algorithm is shown by comparing it to other seven existing variants of PSO and is found to outperformed these variants for the small scale as well as large scale NFCTPs.

Numerical simulation of KVLCC2 self-propulsion with ducted propeller in head waves

In order to meet the Energy Efficiency Design Index (EEDI) requirements, consideration of reducing added power in waves is important for large tankers operating in ocean environments. In this study, the Reynolds-averaged Navier-Stokes (RANS) solver is employed in Computational Fluid Dynamics (CFD) method to simulate the self-propulsion of KVLCC2 model with a ducted KP458 propeller in head waves, especially for comparison with the non-ducted cases. Two methods are adopted for simulating the ducted propeller: the discretized propeller method and the improved body force ducted propeller method. The latter method incorporates an improved principle of momentum conservation to solve the advance speed of the ducted virtual disk precisely in real time, resulting in similar accuracy and higher efficiency compared to the former method. For some wavelengths, the ducted cases display higher propulsive efficiency and lower power than the non-ducted cases, especially under very short wavelength condition, where negative thrust deduction fraction is observed in the ducted case. With the increase of wave height, the effect of the duct becomes more pronounced, resulting in an increase in propulsive efficiency and a reduction in power, both of which can exceed 10%. Moreover, this research carefully analyzes the added values of self-propulsion factors in head waves, as well as flow fields, providing useful insights to inform the application of ducted propellers in ultra-large ocean-going tankers.

The Simulated Source Apportionment of Light Absorbing Aerosols: Effects of Microphysical Properties of Partially‐Coated Black Carbon

AbstractBiomass burning (BB) fractions estimated with the aethalometer model differ from 0 even for pure aerosols from fossil fuel (FF) combustion. We used black carbon (BC) aerosols partially‐coated with non‐absorbing materials to represent typical FF aerosols, and the BB fractions were determined with the aethalometer model. Thus, the estimated BB fractions are the fractions that FF aerosols are incorrectly attributed to BB aerosols. The BC morphology and mixing state have significant effects on the estimation of BB fractions. For freshly emitted BC, the BB fractions do not deviate significantly from 0, and the BB fraction is generally in the range of −1%–10% based on an FF absorption Ångström exponent (AAE) of 1 and a BB AAE of 2. The BB fraction deviates substantially from 0 when BC becomes compact and is coated. The absolute values of the deviations sometimes can be close to 100% for heavily coated BC. The BB fraction is generally greater than 0 for fluffy BC while compact BC generally exhibits a negative BB fraction. The BB fractions of partially‐coated BC are very sensitive to the size distribution and coating ratio, which is consistent with the results of the core‐shell sphere model. We have also performed a series of studies with different configurations for BC morphologies, coating ratios, and mixing states, which show the variations of the estimated BB fractions with atmospheric aging. We also explain the reasons for the uncertainties in the BB fraction estimates and provide suggestions for using different AAE pairs.