Subtype-DCC: decoupled contrastive clustering method for cancer subtype identification based on multi-omics data.

Due to the high heterogeneity and complexity of cancers, patients with different cancer subtypes often have distinct groups of genomic and clinical characteristics. Therefore, the discovery and identification of cancer subtypes are crucial to cancer diagnosis, prognosis and treatment. Recent technological advances have accelerated the increasing availability of multi-omics data for cancer subtyping. To take advantage of the complementary information from multi-omics data, it is necessary to develop computational models that can represent and integrate different layers of data into a single framework. Here, we propose a decoupled contrastive clustering method (Subtype-DCC) based on multi-omics data integration for clustering to identify cancer subtypes. The idea of contrastive learning is introduced into deep clustering based on deep neural networks to learn clustering-friendly representations. Experimental results demonstrate the superior performance of the proposed Subtype-DCC model in identifying cancer subtypes over the currently available state-of-the-art clustering methods. The strength of Subtype-DCC is also supported by the survival and clinical analysis.

Soluble expression of recombinant human interleukin-2 in Escherichia coli and its facile production.

Recombinant human interleukin-2 (rhIL-2) represents one of the most difficult-to-produce cytokines in E. coli due to its extreme hydrophobicity and high tendency to formation of inclusion bodies. Refolding of rhIL-2 inclusion bodies always represents cumbersome downstream processes and low production efficiency. Herein, we disclosed a fusion strategy for efficiently soluble expression and facile production of rhIL-2 in E. coli Origami B (DE3) host. A two-tandem SUMO fusion partner (His-2SUMO) with a unique SUMO protease cleavage site at C-terminus was devised to fuse with the N-terminus of rhIL-2 and the fusion protein (His-2SUMO-rhIL-2) was almost completely expressed in a soluble from. The fusion partner could be efficiently removed by Ulp1 cleavage and the rhIL-2 was simply produced by a two-step Ni-NTA affinity chromatography with a considerable purity and whole recovery. The eventually obtained rhIL-2 was well-characterized and the results showed that the purified rhIL-2 exhibits a compact and ordered structure. Although the finally obtained rhIL-2 exists in a soluble aggregates form and the aggregation probably has been occurred during expression stage, the soluble rhIL-2 aggregates remain exhibit comparable bioactivity with the commercially available rhIL-2 drug formulation.

Labile Carbon from Artificial Roots Alters the Patterns of N2O and N2 Production in Agricultural Soils.

Labile carbon (C) continuously delivered from the rhizosphere profoundly affects terrestrial nitrogen (N) cycling. However, nitrous oxide (N2O) and dinitrogen (N2) production in agricultural soils in the presence of continuous root C exudation with applied N remains poorly understood. We conducted an incubation experiment using artificial roots to continuously deliver small-dose labile C combined with 15N tracers to investigate N2O and N2 emissions in agricultural soils with pH and organic C (SOC) gradients. A significantly negative exponential relationship existed between N2O and N2 emissions under continuous C exudation. Increasing soil pH significantly promoted N2 emissions while reducing N2O emissions. Higher SOC further promoted N2 emissions in alkaline soils. Native soil-N (versus fertilizer-N) was the main source of N2O (average 67%) and N2 (average 80%) emissions across all tested soils. Our study revealed the overlooked high N2 emissions, mainly derived from native soil-N and strengthened by increasing soil pH, under relatively real-world conditions with continuous root C exudation. This highlights the important role of N2O and N2 production from native soil-N in terrestrial N cycling when there is a continuous C supply (e.g., plant-root exudate) and helps mitigate emissions and constrain global budgets of the two concerned nitrogenous gases.

Digestate induces significantly higher N2O emission compared to urea under different soil properties and moisture.

Digestate is considered as an option for recycling resources and a part of the substitution for chemical fertilizers to reduce environmental impacts. However, its application may lead to significant nitrous oxide (N2O) emissions because of its high concentration of ammonium and degradable carbon. The research objectives are to evaluate how N2O emissions respond to digestate as compared to urea application and whether this depends on soil properties and moisture. Either digestate or urea (100 mg N kg-1) was applied with and without a nitrification inhibitor of 3,4-dimethylpyrazole phosphate (DMPP) to three soil types (fluvo-aquic soil, black soil, and latosol) under three different soil moisture conditions (45, 65, and 85% water-filled pore space (WFPS)) through microcosm incubations. Results showed that digestate- and urea-induced N2O emissions increased exponentially with soil moisture in the three studied soils, and the magnitude of the increase was much greater in the alkaline fluvo-aquic soil, coinciding with high net nitrification rate and transient nitrite accumulation. Compared with urea-amended soils, digestate led to significantly higher peaks in N2O and carbon dioxide (CO2) emissions, which might be due to stimulated rapid oxygen consumption and mineralized N supply. Digestate-induced N2O emissions were all more than one time higher than those induced by urea at the three moisture levels in the three studied soils, except at 85% WFPS in the fluvo-aquic soil. DMPP was more effective at mitigating N2O emissions (inhibitory efficacy: 73%-99%) in wetter digestate-fertilized soils. Overall, our study shows the contrasting effect of digestate to urea on N2O emissions under different soil properties and moisture levels. This is of particular value for determining the optimum of applying digestate under varying soil moisture conditions to minimize stimulated N2O emissions in specific soil properties.

RTCB deficiency triggers colitis in mice by influencing the NF-κB and Wnt/ß-catenin signaling pathways.

RNA terminal phosphorylase B (RTCB) has been shown to play a significant role in multiple physiological processes. However, the specific role of RTCB in the mouse colon remains unclear. In this study, we employ a conditional knockout mouse model to investigate the effects of RTCB depletion on the colon and the potential molecular mechanisms. We assess the efficiency and phenotype of Rtcb knockout using PCR, western blot analysis, histological staining, and immunohistochemistry. Compared with the control mice, the Rtcb-knockout mice exhibit compromised colonic barrier integrity and prominent inflammatory cell infiltration. In the colonic tissues of Rtcb-knockout mice, the protein levels of TNF-α, IL-8, and p-p65 are increased, whereas the levels of IKKß and IκBα are decreased. Moreover, the level of GSK3ß is increased, whereas the levels of Wnt3a, ß-catenin, and LGR5 are decreased. Collectively, our findings unveil a close association between RTCB and colonic tissue homeostasis and demonstrate that RTCB deficiency can lead to dysregulation of both the NF-κB and Wnt/ß-catenin signaling pathways in colonic cells.

Partial substitution of manure increases N2O emissions in the alkaline soil but not acidic soils.

The fertilization regimes of combining manure with synthetic fertilizer are benefits for crop yields and soil fertility in cropping systems as compared to sole synthetic fertilization, but the responses of nitrous oxide (N2O) emissions to these practices are inconsistent in the literatures. We hypothesized that it is caused by different proportions of nitrogen (N) applied as manure and various soil properties. Here, we conducted a microcosm experiment, and measured the N2O emissions from control (no N) and five manure substitution treatments (supplied 100 mg N kg-1 using the combination of urea with manure) with a range of proportions of N applied as manure (0, 25%, 50%, 75%, and 100%) in three different soil types (fluvo-aquic soil, black soil, and latosol) under aerobic condition. The stimulated effect on N2O emissions was more pronounced after manure application in an alkaline soil with high nitrification rate, due to relatively rapid soil DOC depletion and N mineralization of manure. N2O emissions from partial substitution of urea with manure were significantly higher than manure-only addition under high soil pH due to abundant labile C from manure. However, there was no difference between manure substitution treatments under acid soils. Nitrification inhibitor substantially decreased N2O emissions with increasing soil pH, but it was less effective in mitigating N2O emissions with larger proportion of manure. This is likely due to the slow nitrification under low soil pH, and denitrification derived N2O increased with increasing manure application rate. Collectively, our study shows that the application of manure substitution to alkaline soils requires careful consideration, which might have rapid nitrification potential and hence trigger significant N2O emissions. The knowledge gained in this work will help the decision-makers in optimizing a sound N fertilization regime interacted with soil properties for sustainable crop production and N2O mitigation.

AOB Nitrosospira cluster 3a.2 (D11) dominates N2O emissions in fertilised agricultural soils.

Ammonia-oxidation process directly contribute to soil nitrous oxide (N2O) emissions in agricultural soils. However, taxonomy of the key nitrifiers (within ammonia oxidising bacteria (AOB), archaea (AOA) and complete ammonia oxidisers (comammox Nitrospira)) responsible for substantial N2O emissions in agricultural soils is unknown, as is their regulation by soil biotic and abiotic factors. In this study, cumulative N2O emissions, nitrification rates, abundance and community structure of nitrifiers were investigated in 16 agricultural soils from major crop production regions of China using microcosm experiments with amended nitrogen (N) supplemented or not with a nitrification inhibitor (nitrapyrin). Key nitrifier groups involved in N2O emissions were identified by comparative analyses of the different treatments, combining sequencing and random forest analyses. Soil cumulative N2O emissions significantly increased with soil pH in all agricultural soils. However, they decreased with soil organic carbon (SOC) in alkaline soils. Nitrapyrin significantly inhibited soil cumulative N2O emissions and AOB growth, with a significant inhibition of the AOB Nitrosospira cluster 3a.2 (D11) abundance. One Nitrosospira multiformis-like OTU phylotype (OTU34), which was classified within the AOB Nitrosospira cluster 3a.2 (D11), had the greatest importance on cumulative N2O emissions and its growth significantly depended on soil pH and SOC contents, with higher growth at high pH and low SOC conditions. Collectively, our results demonstrate that alkaline soils with low SOC contents have high N2O emissions, which were mainly driven by AOB Nitrosospira cluster 3a.2 (D11). Nitrapyrin can efficiently reduce nitrification-related N2O emissions by inhibiting the activity of AOB Nitrosospira cluster 3a.2 (D11). This study advances our understanding of key nitrifiers responsible for high N2O emissions in agricultural soils and their controlling factors, and provides vital knowledge for N2O emission mitigation in agricultural ecosystems.

Metagenomic and network analyses reveal key players in nitrification in upland agricultural soils.

Nitrification, a key step in soil nitrogen cycling, is a biologically mediated process crucial to the ecological environment. However, how nitrifiers drive nitrification under different soil properties and climatic factors at large spatial scales is poorly understood. Here, using metagenomic sequencing and network-based approaches, we identified key nitrifying species of upland agricultural soils in northern China, which spans a wide range of climates and geographic distances. We found that potential nitrification rates (PNRs) varied in different soils and were positively correlated with soil pH (5.42-8.46) and mean annual temperature (MAT) and negatively correlated with the C/N ratio. Network analysis revealed that one module (module 3) was significantly correlated with PNR. In this module, 16 dominant nodes were associated with AOB Nitrosomonas and most nodes were significantly correlated with environmental factors, suggesting that abiotic conditions are important for determining the assembly of these key nitrifiers. Our study advanced the understanding of the key nitrifying populations and their environmental drivers in upland agricultural soil across different soil and climate types.

In situ 15 N-N2 O site preference and O2 concentration dynamics disclose the complexity of N2 O production processes in agricultural soil.

Arable soil continues to be the dominant anthropogenic source of nitrous oxide (N2 O) emissions owing to application of nitrogen (N) fertilizers and manures across the world. Using laboratory and in situ studies to elucidate the key factors controlling soil N2 O emissions remains challenging due to the potential importance of multiple complex processes. We examined soil surface N2 O fluxes in an arable soil, combined with in situ high-frequency measurements of soil matrix oxygen (O2 ) and N2 O concentrations, in situ 15 N labeling, and N2 O 15 N site preference (SP). The in situ O2 concentration and further microcosm visualized spatiotemporal distribution of O2 both suggested that O2 dynamics were the proximal determining factor to matrix N2 O concentration and fluxes due to quick O2 depletion after N fertilization. Further SP analysis and in situ 15 N labeling experiment revealed that the main source for N2 O emissions was bacterial denitrification during the hot-wet summer with lower soil O2 concentration, while nitrification or fungal denitrification contributed about 50.0% to total emissions during the cold-dry winter with higher soil O2 concentration. The robust positive correlation between O2 concentration and SP values underpinned that the O2 dynamics were the key factor to differentiate the composite processes of N2 O production in in situ structured soil. Our findings deciphered the complexity of N2 O production processes in real field conditions, and suggest that O2 dynamics rather than stimulation of functional gene abundances play a key role in controlling soil N2 O production processes in undisturbed structure soils. Our results help to develop targeted N2 O mitigation measures and to improve process models for constraining global N2 O budget.

Effect of two-stage heating treatment on physicochemical properties of shrimp (Penaeus vannamei) meat.

BACKGROUND: Changes in the physicochemical properties of shrimp meat treated with two-stage heating were investigated. Currently, shrimp products in the processing process are susceptible to uneven dehydration, shrimp meat shrinkage, which results in rough and hard texture, poor chewiness, and seriously affects the edible quality as well as economic benefits. Improving the utilization value of shrimp resources, expanding its market shares, optimizing the tenderness of shrimp is the key to developing new types of fresh and ready-to-eat shrimp products. RESULTS: The results indicated that preheating at 30 °C could not affect the quality of shrimp meat significantly (P > 0.05). As the preheating temperature increased from 40 °C to 50 °C, the hardness and shear force of shrimp meat decreased due to the exposure of protein hydrophobic groups, protein aggregation and degradation, muscle fraction broken, and weight loss increase. Further increase in preheating temperature would lead to further aggregation and gelation of proteins, causing hardness and shear force increase. Besides, the results of microstructure showed that preheating at 40 °C and 50 °C could cause the shrimp muscles to become loose. CONCLUSION: This study showed that the preheating temperature ranging from 40 °C to 50 °C could effectively improve the tenderness of shrimp meat. This study might be useful for developing tenderized shrimp products in the future. © 2022 Society of Chemical Industry.

Using nitrification inhibitors and deep placement to tackle the trade-offs between NH3 and N2 O emissions in global croplands.

Ammonia (NH3 ) and nitrous oxide (N2 O) are two important air pollutants that have major impacts on climate change and biodiversity losses. Agriculture represents their largest source and effective mitigation measures of individual gases have been well studied. However, the interactions and trade-offs between NH3 and N2 O emissions remain uncertain. Here, we report the results of a two-year field experiment in a wheat-maize rotation in the North China Plain (NCP), a global hotspot of reactive N emissions. Our analysis is supported by a literature synthesis of global croplands, to understand the interactions between NH3 and N2 O emissions and to develop the most effective approaches to jointly mitigate NH3 and N2 O emissions. Field results indicated that deep placement of urea with nitrification inhibitors (NIs) reduced both emissions of NH3 by 67% to 90% and N2 O by 73% to 100%, respectively, in comparison with surface broadcast urea which is the common farmers' practice. But, deep placement of urea, surface broadcast urea with NIs, and application of urea with urease inhibitors probably led to trade-offs between the two gases, with a mitigation potential of -201% to 101% for NH3 and -112% to 89% for N2 O. The literature synthesis showed that deep placement of urea with NIs had an emission factor of 1.53%-4.02% for NH3 and 0.22%-0.36% for N2 O, which were much lower than other fertilization regimes and the default values recommended by IPCC guidelines. This would translate to a reduction of 3.86-5.47 Tg N yr-1 of NH3 and 0.41-0.50 Tg N yr-1 of N2 O emissions, respectively, when adopting deep placement of urea with NIs (relative to current practice) in global croplands. We conclude that the combination of NIs and deep placement of urea can successfully tackle the trade-offs between NH3 and N2 O emissions, therefore avoiding N pollution swapping in global croplands.

Integrated analysis of lncRNA-miRNA-mRNA ceRNA network in human aortic dissection.

BACKGROUND: Many studies on long chain non-coding RNAs (lncRNAs) are published in recent years. But the roles of lncRNAs in aortic dissection (AD) are still unclear and should be further examined. The present work focused on determining the molecular mechanisms underlying lncRNAs regulation in aortic dissection on the basis of the lncRNA-miRNA-mRNA competing endogenous RNA (ceRNA) network. METHODS: This study collected the lncRNAs (GSE52093), mRNAs (GSE52093) and miRNAs (GSE92427) expression data within human tissue samples with aortic dissection group and normal group based on Gene Expression Omnibus (GEO) database. RESULTS: This study identified three differentially expressed lncRNAs (DELs), 19 differentially expressed miRNAs (DEmiRs) and 1046 differentially expressed mRNAs (DEGs) identified regarding aortic dissection. Furthermore, we constructed a lncRNA-miRNA-mRNA network through three lncRNAs (including two with up-regulation and one with down-regulation), five miRNAs (five with up-regulation), as well as 211 mRNAs (including 103 with up-regulation and 108 with down-regulation). Simultaneously, we conducted functional enrichment and pathway analyses on genes within the as-constructed ceRNA network. According to our PPI/ceRNA network and functional enrichment analysis results, four critical genes were found (E2F2, IGF1R, BDNF and PPP2R1B). In addition, E2F2 level was possibly modulated via lncRNA FAM87A-hsa-miR-31-5p/hsa-miR-7-5p or lncRNA C9orf106-hsa-miR-7-5p. The expression of IGF1R may be regulated by lncRNA FAM87A-hsa-miR-16-5p/hsa-miR-7-5p or lncRNA C9orf106-hsa-miR-7-5p. CONCLUSION: In conclusion, the ceRNA interaction axis we identified is a potentially critical target for treating AD. Our results shed more lights on the possible pathogenic mechanism in AD using a lncRNA-associated ceRNA network.

YY1 and RTCB in mouse uterine decidualization and embryo implantation.

As a multifunctional transcription factor, YY1 regulates the expression of many genes essential for early embryonic development. RTCB is an RNA ligase that plays a role in tRNA maturation and Xbp1 mRNA splicing. YY1 can bind in vitro to the response element in the proximal promoter of Rtcb and regulate Rtcb promoter activity. However, the in vivo regulation and whether these two genes are involved in the mother-fetal dialogue during early pregnancy remain unclear. In this study, we validated that YY1 bound in vivo to the proximal promoter of Rtcb in mouse uterus of early pregnancy. Moreover, via building a variety of animal models, our study suggested that both YY1 and RTCB might play a role in mouse uterus decidualization and embryo implantation during early pregnancy.

Systemic Inflammatory Markers for Predicting Overall Survival in Patients with Osteosarcoma: A Systematic Review and Meta-Analysis.

BACKGROUND: Inflammatory markers are associated with tumor genesis and progression, but their prognostic significance in osteosarcoma remains unclear. Therefore, we discussed the prognostic value of related inflammatory markers in osteosarcoma through a meta-analysis and systematic review. These inflammatory markers include C-reactive protein (CRP), neutrophil to lymphocyte ratio (NLR), lymphocyte to monocyte ratio (LMR), platelet to lymphocyte ratio (PLR), and Glasgow prognostic score (GPS). METHODS: The Chinese National Knowledge Infrastructure (CNKI), Wanfang, Chinese Scientific Journals (VIP), PubMed, Embase, and Cochrane libraries were searched. The design of meta-analysis was made based on the PICOS (population, intervention/exposure, control, outcomes, and study design) principles, and STATA 15.1 was used to analyze the data. The Newcastle-Ottawa scale (NOS) was used to assess the quality of included studies. Hazard ratios (HRs) for overall survival (OS) and disease-specific survival (DPS) were extracted for the investigation of the prognostic value of inflammatory markers. RESULTS: Twelve researches with 2162 osteosarcoma patients were included in total. The pooled results showed that elevated NLR, CRP, and GPS are all greatly related to shortening of OS among patients with osteosarcoma (HR = 1.68, P = 0.007, 95% CI: 1.15-2.45; HR = 1.96, P = 0.002, 95% CI: 1.28-3.00; HR = 2.54, P < 0.0001, 95% CI: 1.95-3.31, respectively), and CRP level is significantly associated with shortening of DPS among patients with osteosarcoma (HR = 2.76, 95% CI:2.01-3.80, P < 0.0001), additionally. However, the correlation between LMR or PLR and the prognosis of osteosarcoma is not statistically significant (HR = 0.60, 95% CI: 0.30-1.18, P = 0.138; HR = 1.13, 95% CI: 0.85-1.49, P = 0.405, respectively). The outcomes of subgroup analysis to NLR and CRP suggested that histology, ethnicity, metastasis, and sample size all have an impact on its prognosis of patients with osteosarcoma. CONCLUSION: Worsened prognosis may be related to high levels of NLR, CRP, and GPS before treatment rather than LMR or PLR, which can provide the basis for clinicians to judge the outcomes of prognosis. Trial Registration. PROSPERO (CRD42021249954), https://www.crd.york.ac.uk/prospero/display_record.php?RecordID=249954.

Oxygen Regulates Nitrous Oxide Production Directly in Agricultural Soils.

Oxygen (O2) plays a critical and yet poorly understood role in regulating nitrous oxide (N2O) production in well-structured agricultural soils. We investigated the effects of in situ O2 dynamics on N2O production in a typical intensively managed Chinese cropping system under a range of environmental conditions (temperature, moisture, ammonium, nitrate, dissolved organic carbon, and so forth). Climate and management (fertilization, irrigation, precipitation, and temperature), and their interactions significantly affected soil O2 and N2O concentrations (P < 0.05). Soil O2 concentration was the most significant factor correlating with soil N2O concentration (r = -0.71) when compared with temperature, water-filled pore space, and ammonium concentration (r = 0.30, 0.25, and 0.26, respectively). Soil N2O concentration increased exponentially with decreasing soil O2 concentrations. The exponential model of N treatments and fertilization with irrigation/precipitation events predicted 74-90% and 58% of the variance in soil N2O concentrations, respectively. Our results highlight that the soil O2 status is the proximal, direct, and the most decisive environmental trigger for N2O production, outweighing the effects of other factors and could be a key variable integrating the aggregated effects of various complex interacting variables. This study offers new opportunities for developing more sensitive approaches to predicting and through appropriate management interventions mitigating N2O emissions from agricultural soils.

Nitrous Oxide Emissions Increase Exponentially When Optimum Nitrogen Fertilizer Rates Are Exceeded in the North China Plain.

The IPCC assume a linear relationship between nitrogen (N) application rate and nitrous oxide (N2O) emissions in inventory reporting, however, a growing number of studies show a nonlinear relationship under specific soil-climatic conditions. In the North China plain, a global hotspot of N2O emissions, covering a land as large as Germany, the correlation between N rate and N2O emissions remains unclear. We have therefore specifically investigated the N2O response to N applications by conducting field experiments with five N rates, and high-frequency measurements of N2O emissions across contrasting climatic years. Our results showed that cumulative and yield-scaled N2O emissions both increased exponentially as N applications were raised above the optimum rate in maize ( Zea mays L.). In wheat ( Triticum aestivum L.) there was a corresponding quadratic increase in N2O emissions with the magnitude of the response in 2012-2013 distinctly larger than that in 2013-2014 owing to the effects of extreme snowfall. Existing empirical models (including the IPCC approach) of the N2O response to N rate have overestimated N2O emissions in the North China plain, even at high N rates. Our study therefore provides a new and robust analysis of the effects of fertilizer rate and climatic conditions on N2O emissions.

Synthesis of 6-hydroxyaurone analogues and evaluation of their α-glucosidase inhibitory and glucose consumption-promoting activity: Development of highly active 5,6-disubstituted derivatives.

A series of 6-hydroxyaurones and their analogues have been synthesized and evaluated for their in vitro α-glucosidase inhibitory and glucose consumption-promoting activity. These compounds exhibited varying degrees of α-glucosidase inhibitory activity, 11 of them showing higher potency than that of the control standard acarbose (IC50=50.30µM). Surprisingly, analogues devoid of a substituent at C-2 but having an aryl group at C-5 were found to be highly active (e.g., 7f, IC50=9.88µM). Docking analysis substantiated these findings. The kinetic analysis of compound 7f, the most potent α-glucosidase inhibitor of this study, revealed that it inhibited α-glucosidase in an irreversible and mixed competitive mode. In addition, compounds 7f and 10c exhibited significant glucose consumption promoting activity at 1µM.

CD123-Engager T Cells as a Novel Immunotherapeutic for Acute Myeloid Leukemia.

Immunotherapy with CD123-specific T-cell engager proteins or with T cells expressing CD123-specific chimeric antigen receptors is actively being pursued for acute myeloid leukemia. T cells secreting bispecific engager molecules (ENG-T cells) may present a promising alternative to these approaches. To evaluate therapeutic potential, we generated T cells to secrete CD123/CD3-bispecific engager molecules. CD123-ENG T cells recognized primary acute myeloid leukemia (AML) cells and cell lines in an antigen-dependent manner as judged by cytokine production and/or tumor killing, and redirected bystander T cells to AML cells. Infusion of CD123-ENG T cells resulted in regression of AML in xenograft models conferring a significant survival advantage of treated mice in comparison to mice that received control T cells. At high effector to target ratios, CD123-ENG T cells recognized normal hematopoietic stem and progenitor cells (HSPCs) with preferential recognition of HSPCs from cord blood compared to bone marrow. We therefore introduced the CD20 suicide gene that can be targeted in vivo with rituximab into CD123-ENG T cells. The expression of CD20 did not diminish the anti-AML activity of CD123-ENG T cells, but allowed for rituximab-mediated ENG-T cell elimination. Thus, ENG-T cells coexpressing CD20 suicide and CD123 engager molecules may present a promising immunotherapeutic approach for AML.

Regulation of Insulin Resistance by Multiple MiRNAs via Targeting the GLUT4 Signalling Pathway.

BACKGROUND/AIMS: Type 2 Diabetes Mellitus (T2DM) is characterized by insulin resistance (IR), but the underlying molecular mechanisms are incompletely understood. MicroRNAs (miRNAs) have been demonstrated to participate in the signalling pathways relevant to glucose metabolism in IR. The purpose of this study was to test whether the multiple-target anti-miRNA antisense oligonucleotides (MTg-AMO) technology, an innovative miRNA knockdown strategy, can be used to interfere with multiple miRNAs that play critical roles in regulating IR. METHODS: An MTg-AMO carrying the antisense sequences targeting miR-106b, miR-27a and miR-30d was constructed (MTg-AMO106b/27a/30d). Protein levels were determined by Western blot analysis, and transcript levels were detected by real-time RT-PCR (qRT-PCR). Insulin resistance was analysed with glucose consumption and glucose uptake assays. RESULTS: We found that the protein level of glucose transporter 4 (GLUT4), Mitogen-activated protein kinase 14 (MAPK 14), Phosphatidylinositol 3-kinase regulatory subunit beta (PI3K regulatory subunit beta) and mRNA level of Slc2a4 (encode GLUT4), Mapk14 (encode MAPK 14) and Pik3r2 (encode PI3K regulatory subunit beta) were all significantly down-regulated in the skeletal muscle of diabetic rats and in insulin-resistant L6 cells. Overexpression of miR-106b, miR-27a and miR-30d in L6 cells decreased glucose consumption and glucose uptake, and reduced the expression of GLUT4, MAPK 14 and PI3K regulatory subunit beta. Conversely, silencing of endogenous miR-106b, miR-27a and miR-30d in insulin-resistant L6 cells enhanced glucose consumption and glucose uptake, and increased the expression of GLUT4, MAPK 14 and PI3K regulatory subunit beta. MTg-AMO106b/27a/30d up-regulated the protein levels of GLUT4, MAPK 14 and PI3K regulatory subunit beta, enhanced glucose consumption and glucose uptake. CONCLUSION: Our data suggested that miR-106b, miR-27a and miR-30d play crucial roles in the regulation of glucose metabolism by targeting the GLUT4 signalling pathway in L6 cells. Moreover, MTg-AMO106b/27a/30d offers more potent effects than regular singular AMOs.

Engager T cells: a new class of antigen-specific T cells that redirect bystander T cells.

Adoptive immunotherapy with antigen-specific T cells has shown promise for the treatment of malignancies. However, infused T cells are unable to redirect resident T cells, limiting potential benefit. While the infusion of bispecific T-cell engagers can redirect resident T cells to tumors, these molecules have a short half-life, and do not self amplify. To overcome these limitations, we generated T cells expressing a secretable T-cell engager specific for CD3 and EphA2, an antigen expressed on a broad range of human tumors (EphA2-ENG T cells). EphA2-ENG T cells were activated and recognized tumor cells in an antigen-dependent manner, redirected bystander T cells to tumor cells, and had potent antitumor activity in glioma and lung cancer severe combined immunodeficiency (SCID) xenograft models associated with a significant survival benefit. This new class of tumor-specific T cells, with the unique ability to redirect bystander T cells, may be a promising alternative to current immunotherapies for cancer.