Effect of Growth Stage on Nutrition, Fermentation Quality, and Microbial Community of Semidry Silage from Forage Soybean.

Soybean (Glycine max (Linn.) Merr.) is highly suitable as animal feed. The silage quality and microbial characteristics of soybean silage are still unclear. Forage soybean (HN389), at six different growth stages (R2-R7), were used as experimental materials to investigate the changes in fermentation, nutritional quality, and microbial characteristics of semidry silage after 0, 7, 14, 30, and 45 d. As the growth period extended, the content of crude protein (CP) and crude fat (EE) gradually increased, while the neutral detergent fiber (NDF) and the acid detergent fiber (ADF) content decreased. The pH value also decreased gradually with fermentation time, accompanied by increases in the proportion of ammonia-N and the content of lactic acid (LA) and acetic acid (AA). In addition, competitive inhibition was observed in the microbial fermentation. With the process of ensiling, Lactobacillus became the dominant bacterial species. The results indicate that the most active stage of fermentation during ensiling occurred within the first 7 days, the fermentation and nutritional quality of the soybean forage were improved, and the optimal mowing stage was the grain stage. Comparison of the microbial abundance showed that all microorganisms entered a stable stage at 30 days of silage. After storage, the dominant bacteria were Lactobacillus, Enterobacter, and Pantoea.

Metagenomics study of soil microorganisms involved in the carbon cycle in a saline-alkaline meadow steppe in the Songnen Plain in Northeast China.

Soil microorganisms play an important role in regulating and contributing to carbon cycling processes in grassland ecosystems. Soil salinization is one of the major problems causing soil degradation, and its effects on carbon cycle immobilization-related functional genes in soil microorganisms remain unknown. Therefore, we took Songnen salinization grassland as the research object, selected grasslands with different salinization levels, and explored the diversity of soil microorganisms and functional genes related to carbon cycling in Songnen grassland with different salinization levels through metagenomic technology. The results showed that with the increase of salinity, the relative abundance of Ascomycetes increased, while the relative abundance of Proteus and Firmicutes decreased. In addition, the relative abundance of functional genes related to carbon cycling fixation has also decreased. As the degree of soil salinization increases, the relative abundance of glycoside hydrolases (GH)130 family significantly increases, while the relative abundance of soil carbohydrate enzymes belonging to GH3 and GH55 families significantly decreases. Using structural equation modeling (SEM), it was found that soil pH and conductivity (EC) have a significant impact on soil microbial diversity and functional genes related to carbon cycling fixation. The increase in soil pH directly reduces the Shannon diversity of soil microbial diversity and functional genes related to carbon cycling fixation. Therefore, it can be concluded that the intensification of grassland salinization reduces the diversity of bacteria and fungi, and affects the diversity of functional genes related to carbon cycling fixation by reducing the total diversity of bacteria. The increase in salinity has a negative feedback effect on grassland soil carbon cycling. This study provides a theoretical framework for grassland soil carbon sequestration and degradation restoration.

CDK6 inhibits de novo lipogenesis in white adipose tissues but not in the liver.

Increased de novo lipogenesis (DNL) in white adipose tissue is associated with insulin sensitivity. Under both Normal-Chow-Diet and High-Fat-Diet, mice expressing a kinase inactive Cyclin-dependent kinase 6 (Cdk6) allele (K43M) display an increase in DNL in visceral white adipose tissues (VAT) as compared to wild type mice (WT), accompanied by markedly increased lipogenic transcriptional factor Carbohydrate-responsive element-binding proteins (CHREBP) and lipogenic enzymes in VAT but not in the liver. Treatment of WT mice under HFD with a CDK6 inhibitor recapitulates the phenotypes observed in K43M mice. Mechanistically, CDK6 phosphorylates AMP-activated protein kinase, leading to phosphorylation and inactivation of acetyl-CoA carboxylase, a key enzyme in DNL. CDK6 also phosphorylates CHREBP thus preventing its entry into the nucleus. Ablation of runt related transcription factor 1 in K43M mature adipocytes reverses most of the phenotypes observed in K43M mice. These results demonstrate a role of CDK6 in DNL and a strategy to alleviate metabolic syndromes.

CDK6 is essential for mesenchymal stem cell proliferation and adipocyte differentiation.

Background: Overweight or obesity poses a significant risk of many obesity-related metabolic diseases. Among all the potential new therapies, stem cell-based treatments hold great promise for treating many obesity-related metabolic diseases. However, the mechanisms regulating adipocyte stem cells/progenitors (precursors) are unknown. The aim of this study is to investigate if CDK6 is required for mesenchymal stem cell proliferation and adipocyte differentiation. Methods: Cyclin-dependent kinase 6 (Cdk6) mouse models together with stem cells derived from stromal vascular fraction (SVF) or mouse embryonic fibroblasts (MEFs) of Cdk6 mutant mice were used to determine if CDK6 is required for mesenchymal stem cell proliferation and adipocyte differentiation. Results: We found that mice with a kinase inactive CDK6 mutants (K43M) had fewer precursor residents in the SVF of adult white adipose tissue (WAT). Stem cells from the SVF or MEFs of K43M mice had defects in proliferation and differentiation into the functional fat cells. In contrast, mice with a constitutively active kinase CDK6 mutant (R31C) had the opposite traits. Ablation of RUNX1 in both mature and precursor K43M cells, reversed the phenotypes. Conclusion: These results represent a novel role of CDK6 in regulating precursor numbers, proliferation, and differentiation, suggesting a potential pharmacological intervention for using CDK6 inhibitors in the treatment of obesity-related metabolic diseases.

Transcriptome and Metabolome Analysis of Selenium Treated Alfalfa Reveals Influence on Phenylpropanoid Biosynthesis to Enhance Growth.

Selenium (Se) plays an important role in the growth of plants. Alfalfa (Medicago sativa L.) is a perennial legume forage crop with high nutritional value and Se-rich functions. Many studies have shown that selenium can promote alfalfa growth, but few have explored the molecular biology mechanisms behind this effect. In this study, alfalfa was divided into two groups. One group was sprayed with sodium selenite (Na2SeO3) and the other group was sprayed with distilled water as a control. This study determined the growth, reproductive traits, physiological changes, transcriptome and metabolome of both groups of alfalfa. We found that foliar spraying of 100 mg/L Na2SeO3 could significantly increase the growth rate, dry weight, total Se content, amount of pollen per flower, pollen viability, pod spirals, and seed number per pod of alfalfa plants. The level of chlorophyll, soluble protein, proline, and glutathione also increased dramatically in Na2SeO3-sprayed alfalfa seedlings. After transcriptome and metabolome analysis, a total of 614 differentially expressed genes (DEGs) and 1500 differentially expressed metabolites (DEMs), including 26 secondary differentially metabolites were identified. The DEGs were mainly enriched in MAPK signaling pathway, phenylpropanoid biosynthesis, isoflavonoid biosynthesis, cutin, suberine, and wax biosynthesis, and glycerolipid metabolism. The DEMs were mainly enriched in flavone and flavonol biosynthesis, carbon metabolism, glyoxylate and dicarboxylate metabolism, nitrogen metabolism, and phenylpropanoid biosynthesis. Integrative analysis of transcriptome and metabolome showed that the foliar spraying of Na2SeO3 mainly affects phenylpropanoid biosynthesis to promote alfalfa growth.

Autophagy inhibition and ferroptosis activation during atherosclerosis: Hypoxia-inducible factor 1α inhibitor PX-478 alleviates atherosclerosis by inducing autophagy and suppressing ferroptosis in macrophages.

OBJECTIVE: To elucidate the key regulator responsible for autophagy and ferroptosis, and if specific pharmacological inhibitor of upregulated gene exerted the pro-autophagic and anti-ferroptotic effect on macrophage to alleviate the atherosclerosis. METHODS: Autophagy and ferroptosis were evaluated in atherosclerotic lesions and THP-1 macrophages exposed to ox-LDL. Autophagy/ferroptosis-related differentially expressed genes (DEGs) in atherosclerosis were identified by bioinformatic analysis of GSE97210 dataset, and were validated in atherosclerotic cells and tissues. The efficacy and mechanism of pharmacological inhibition of the validated DEGs on alleviating atherosclerosis were explored in vivo and in vitro. RESULTS: Atherosclerotic lesions were characterized by autophagy inhibition and ferroptosis activation in macrophages. The crosslink between autophagy and ferroptosis were demonstrated. Ox-LDL induced THP-1 macrophage foam cell formation, autophagy dysfunction, and ferroptosis occurrence. Rapamycin ameliorated and, conversely, erastin deteriorated the effect of ox-LDL on THP-1 macrophages. Eleven autophagy/ferroptosis-related DEGs were identified in atherosclerosis vs. normal. The up-regulated expression of HIF-1α was verified in atherosclerotic lesions and THP-1 macrophages induced by ox-LDL. HIF-1α inhibitor PX-478 restored autophagy function, depressed ferroptosis, and reduced lipid accumulation in ox-LDL induced THP-1 macrophage. Autophagy inhibitor 3-MA obviously abrogated the pro-autophagic, anti-ferroptotic, and anti-atherosclerotic effects of PX-478. PX-478 treatment down-regulated HIF-1α expression and reduced atherosclerotic plaques in the mice model. CONCLUSIONS: Autophagy is inhibited, ferroptosis is activated, and crosslink occurs between autophagy and ferroptosis during atherosclerosis. HIF-1α, an upregulated DEG between atherosclerosis and normal, co-regulates autophagy and ferroptosis. HIF-1α inhibitor PX-478 attenuates foam cell formation and lessens atherosclerosis by enhancing autophagy and depressing ferroptosis in macrophages.

Glial receptor PLXNB2 regulates schizophrenia-related stress perception via the amygdala.

Stress is a trigger for the development of psychiatric disorders. However, how stress trait differs in schizophrenia patients is still unclear. Stress also induces and exacerbates immune activation in psychiatric disorders. Plexins (Plxn) and its ligands semaphorins (Sema) are important cellular receptors with plural functions in both the brain and the immune system. Recently, the role of Plxn/Sema in regulation of neuroinflammation was also noticed. Here, when investigating immune mechanisms underlying stress susceptibility in schizophrenia, we discovered the role of Plxnb2 in stress response. Patients of first-episode schizophrenia (FES) with high stress (FES-hs, n=51) and low stress (FES-ls, n=50) perception and healthy controls (HCs) (n=49) were first recruited for neuroimaging and blood bulk RNA sequencing (RNA-seq). A mouse model of chronic unpredictable stress (CUS) and intra-amygdaloid functional blocking of Plxnb2 were further explored to depict target gene functions. Compared to HCs, FES-hs patients had bigger caudate and thalamus (FDR=0.02&0.001, respectively) whereas FES-ls patients had smaller amygdala (FDR=0.002). Blood RNA-seq showed differentially expressed PLXNB2 and its ligands among patient groups and HCs (FDR<0.05~0.01). Amygdaloid size and PLXNB2 level were both negatively correlated with stress perception (p<0.01&0.05, respectively), which fully mediated the amygdaloid positive association with PLXNB2 expression (ß=0.9318, 95% CI: 0.058~1.886) in FES-hs patients. In mice, Plxnb2 was enriched in astrocytes and microglia and CUS reduced its expression in astrocytes (p<0.05). Inhibition of amygdaloid Plxnb2 by its functional blocking monoclonal antibody (mAb)-102 induced mice anxiety (p<0.05), amygdaloid enlargement (p<0.05), and microglial ramification (p<0.001) compared to saline. These data suggest that PLXNB2 regulates amygdala-dependent stress responses.

Management Strategy and Radiologic Outcomes of Symptomatic Spontaneous Isolated Superior Mesenteric Artery Dissection Based on Angiographic Classification: The Follow-Up Experience in a Single Center.

PURPOSE: To investigate the demographics, clinical features, radiologic measurement, treatment, and outcomes of symptomatic spontaneous isolated superior mesenteric artery dissection (SISMAD) according to computed tomography (CT) classification. METHODS: This retrospective study included 201 patients diagnosed with symptomatic SISMAD from November 2014 to December 2020. Symptomatic spontaneous isolated superior mesenteric artery dissection was categorized into four types based on CT images by Yun's angiographic classification. Their clinical characteristics, images features, treatment methods, and radiological outcomes were comparatively analyzed by CT angiographic types. RESULTS: SISMADs were categorized into type I (13.9%) patent false lumen (FL) with both entry and re-entry; type IIa (37.3%), blind pouch of FL; type IIb (43.3%), thrombosed FL; and type III (5.5%), and the occlusion of superior mesenteric artery (SMA). Type IIb, the most common SISMAD, showed the largest true lumen (TL) residual diameter and the lowest percentage of TL stenosis. Type III positioned most proximally to SMA origin and had the maximum dissection length. Symptomatic spontaneous isolated superior mesenteric artery dissections underwent conservative (75.1%), endovascular (22.4%), and surgical (2.5%) treatment. Conservative treatment was more frequent in type I (85.7%) and type IIb (83.9%) than in type IIa (65.3%) and type III (45.5%). Endovascular intervention was more commonly utilized in type IIa (32.0%) and type III (36.4%) than in type I (14.3%) and type IIb (14.9%). Conservative patients achieved FL vanishment/shrinkage (57.8%), stabilization (26.6%), and enlargement (15.6%). After conservative treatment, type I showed angiographic FL stabilization; type IIa achieved FL shrinkage (48.1%), stabilization (22.2%), and enlargement (29.6%); type IIb exhibited FL vanishment/shrinkage (92.0%) and enlargement (8.0%). Cumulative rate of stent patency was 92.3% during 6-year follow-up. CONCLUSIONS: Conservative management with close follow-up is initially provided especially for types I and IIb. Morphological stabilization is more frequent in type I of patent FL with entry and re-entry. False lumen vanishment or shrinkage was more likely to occur in type IIb due to the thrombus absorption. Endovascular intervention has excellent long-term in-stent patency and is predominantly utilized in types IIa and III. Blood flow sustained into a blind-ending FL causes the TL compression and stenosis in type IIa. Type III with the occlusion of SMA has the high risk of bowel ischemia. CLINICAL IMPACT: According to Yun's angiographic classification of spontaneous isolated superior mesenteric artery dissection (SISMAD), type I (13.9%) has patent true and false lumen and the morphological pattern is maintained stable; type IIa (37.3%) possesses a patent blind-ending false lumen which might shrink, remain unchanged, or enlarge; and endovascular intervention is suggested when conservative treatment failed; type IIb (43.3%) recovers spontaneously due to the absorption of false lumen thrombus and conservative treatment is preferentially considered; type III (5.5%) with the occlusion of main trunk carries a high risk of bowel necrosis, early endovascular intervention is proposed, and open surgery might be necessary.

Percutaneous endovenous intervention versus anticoagulation in the treatment of lower extremity deep vein thrombosis: a systematic review and meta-analysis.

Background: Deep vein thrombosis (DVT) of the lower extremity (LE) might lead to pulmonary embolism (PE) and post-thrombolytic syndrome (PTS). Recently, percutaneous endovenous intervention (PEVI) has been advocated for early removal of thrombus clot and restoration of venous patency. This study aims to review the safety and efficacy outcomes of PEVI versus anticoagulation in the treatment of acute LE-DVT. Methods: We searched the databases of PubMed, Embase, and the Cochrane Library for randomized controlled trials (RCTs) comparing catheter-directed thrombolysis (CDT) and/or pharmacomechanical thrombectomy (PMT) versus anticoagulation for acute proximal LE-DVT, published before August 2022. Efficacy outcomes were PTS and venous patency. Safety outcomes included recurrent thromboembolism, bleeding complications, and PE. Results: Overall, 1,266 patients were included from 6 RCTs. The overall risk of bias was small due to enrolled high-quality RCTs. Compared to anticoagulation, PEVI moderately reduced PTS incidence [odds ratio (OR) 0.47, 95% confidence interval (CI) 0.23-0.99], obviously inhibited moderate-to-severe PTS (OR 0.60, 95% CI: 0.40-0.88), significantly decreased PE (OR 0.16, 95% CI: 0.05-0.48), and substantially increased venous patency (OR 7.95, 95% CI: 1.00-63.16). There was no significant difference in recurrent thromboembolism between PEVI and anticoagulation (OR 0.76, 95% CI: 0.34-1.73). Bleeding events did not differ statistically between PEVI and anticoagulation (OR 1.36, 95% CI: 0.87-2.11). We conducted single-arm meta-analysis of the PEVI or anticoagulation. Pooled proportion of PTS was less after PEVI (0.295, 95% CI: 0.123-0.505) than after anticoagulation (0.459, 95% CI: 0.306-0.616). Pooled proportion of moderate-to-severe PTS was lower after PEVI (0.098, 95% CI: 0.033-0.191) than after anticoagulation (0.183, 95% CI: 0.126-0.247). Pooled proportion of PE was smaller after PEVI (0.006, 95% CI: 0.00-0.020) as compared to anticoagulation (0.075, 95% CI: 0.038-0.122). Pooled proportion of recurrent thromboembolism was similar between PEVI (0.095, 95% CI: 0.054-0.146) and anticoagulation (0.124, 95% CI: 0.061-0.206). Pooled proportion of bleeding was not different statistically between PEVI (0.026, 95% CI: 0.00-0.131) and anticoagulation (0.008, 95% CI: 0.00-0.094). Conclusions: PEVI, consisting of PMT and/or CDT, is an extremely effective and feasible approach for patients with acute LE-DVT. In comparison to therapeutic anticoagulation, PEVI restores venous patency, inhibits the PTS development, reduces the PE occurrence, does not markedly increase the bleeding risk, but does not reduce recurrent thromboembolism.

Ribonuclease 4 is associated with aggressiveness and progression of prostate cancer.

Prostate specific antigen screening has resulted in a decrease in prostate cancer-related deaths. However, it also has led to over-treatment affecting the quality of life of many patients. New biomarkers are needed to distinguish prostate cancer from benign prostate hyperplasia (BPH) and to predict aggressiveness of the disease. Here, we report that ribonuclease 4 (RNASE4) serves as such a biomarker as well as a therapeutic target. RNASE4 protein level in the plasma is elevated in prostate cancer patients and is positively correlated with disease stage, grade, and Gleason score. Plasma RNASE4 level can be used to predict biopsy outcome and to enhance diagnosis accuracy. RNASE4 protein in prostate cancer tissues is enhanced and can differentiate prostate cancer and BPH. RNASE4 stimulates prostate cancer cell proliferation, induces tumor angiogenesis, and activates receptor tyrosine kinase AXL as well as AKT and S6K. An RNASE4-specific monoclonal antibody inhibits the growth of xenograft human prostate cancer cell tumors in athymic mice.

Angiogenin and plexin-B2 axis promotes glioblastoma progression by enhancing invasion, vascular association, proliferation and survival.

BACKGROUND: Angiogenin is a multifunctional secreted ribonuclease that is upregulated in human cancers and downregulated or mutationally inactivated in neurodegenerative diseases. A role for angiogenin in glioblastoma was inferred from the inverse correlation of angiogenin expression with patient survival but had not been experimentally investigated. METHODS: Angiogenin knockout mice were generated and the effect of angiogenin deficiency on glioblastoma progression was examined. Angiogenin and plexin-B2 genes were knocked down in glioblastoma cells and the changes in cell proliferation, invasion and vascular association were examined. Monoclonal antibodies of angiogenin and small molecules were used to assess the therapeutic activity of the angiogenin-plexin-B2 pathway in both genetic and xenograft animal models. RESULTS: Deletion of Ang1 gene prolonged survival of PDGF-induced glioblastoma in mice in the Ink4a/Arf-/-:Pten-/- background, accompanied by decreased invasion, vascular association and proliferation. Angiogenin upregulated MMP9 and CD24 leading to enhanced invasion and vascular association. Inhibition of angiogenin or plexin-B2, either by shRNA, monoclonal antibody or small molecule inhibitor, decreases sphere formation of patient-derived glioma stem cells, reduces glioblastoma proliferation and invasion and inhibits glioblastoma growth in both genetic and xenograft animal models. CONCLUSIONS: Angiogenin and its receptor, plexin-B2, are a pair of novel regulators that mediate invasion, vascular association and proliferation of glioblastoma cells. Inhibitors of the angiogenin-plexin-B2 axis have therapeutic potential against glioblastoma.

Angiogenin mediates paternal inflammation-induced metabolic disorders in offspring through sperm tsRNAs.

Paternal environmental inputs can influence various phenotypes in offspring, presenting tremendous implications for basic biology and public health and policy. However, which signals function as a nexus to transmit paternal environmental inputs to offspring remains unclear. Here we show that offspring of fathers with inflammation exhibit metabolic disorders including glucose intolerance and obesity. Deletion of a mouse tRNA RNase, Angiogenin (Ang), abolished paternal inflammation-induced metabolic disorders in offspring. Additionally, Ang deletion prevented the inflammation-induced alteration of 5'-tRNA-derived small RNAs (5'-tsRNAs) expression profile in sperm, which might be essential in composing a sperm RNA 'coding signature' that is needed for paternal epigenetic memory. Microinjection of sperm 30-40 nt RNA fractions (predominantly 5'-tsRNAs) from inflammatory Ang+/+ males but not Ang-/- males resulted in metabolic disorders in the resultant offspring. Moreover, zygotic injection with synthetic 5'-tsRNAs which increased in inflammatory mouse sperm and decreased by Ang deletion partially resembled paternal inflammation-induced metabolic disorders in offspring. Together, our findings demonstrate that Ang-mediated biogenesis of 5'-tsRNAs in sperm contributes to paternal inflammation-induced metabolic disorders in offspring.

Integrated Analysis of the Transcriptome and Metabolome Revealed Candidate Genes Involved in GA3-Induced Dormancy Release in Leymus chinensis Seeds.

Leymus chinensis is a perennial forage grass that has good palatability, high yield and high feed value, but seed dormancy is a major problem limiting the widespread cultivation of L. chinensis. Here, we performed transcriptomic and metabolomic analysis of hulled and de-hulled seeds of L. chinensis treated with or without GA3 to investigate the changes in gene and metabolites associated with dormancy release induced by GA3. The germination test revealed that the optimum concentration of GA3 for disruption of L. chinensis seed dormancy was 577 µM. A total of 4327 and 11,919 differentially expressed genes (DEGs) and 871 and 650 differentially abundant metabolites were identified in de-hulled and hulled seeds treated with GA3, respectively, compared with seeds soaked in sterile water. Most of the DEGs were associated with starch and sucrose metabolism, protein processing in the endoplasmic reticulum, endocytosis and ribosomes. Furthermore, isoquinoline alkaloid biosynthesis, tyrosine metabolism, starch and sucrose metabolism, arginine and proline metabolism, and amino sugar and nucleotide sugar metabolism were significantly enriched pathways. Integrative analysis of the transcriptomic and metabolomic data revealed that starch and sucrose metabolism is one of the most important pathways that may play a key role in providing carbon skeletons and energy supply for the transition of L. chinensis seeds from a dormant state to germination by suppressing the expression of Cel61a, egID, cel1, tpsA, SPAC2E11.16c and TPP2, enhancing the expression of AMY1.1, AMY1.2, AMY1.6 and GLIP5, and inhibiting the synthesis of cellobiose, cellodextrin, and trehalose while promoting the hydrolysis of sucrose, starch, cellobiose, cellodextrin, and trehalose to glucose. This study identified several key genes and provided new insights into the molecular mechanism of seed dormancy release induced by GA3 in L. chinensis. These putative genes will be valuable resources for improving the seed germination rate in future breeding studies.

De novo transcriptome in roots of switchgrass (Panicum virgatum L.) reveals gene expression dynamic and act network under alkaline salt stress.

BACKGROUND: Soil salinization is a major limiting factor for crop cultivation. Switchgrass is a perennial rhizomatous bunchgrass that is considered an ideal plant for marginal lands, including sites with saline soil. Here we investigated the physiological responses and transcriptome changes in the roots of Alamo (alkaline-tolerant genotype) and AM-314/MS-155 (alkaline-sensitive genotype) under alkaline salt stress. RESULTS: Alkaline salt stress significantly affected the membrane, osmotic adjustment and antioxidant systems in switchgrass roots, and the ASTTI values between Alamo and AM-314/MS-155 were divergent at different time points. A total of 108,319 unigenes were obtained after reassembly, including 73,636 unigenes in AM-314/MS-155 and 65,492 unigenes in Alamo. A total of 10,219 DEGs were identified, and the number of upregulated genes in Alamo was much greater than that in AM-314/MS-155 in both the early and late stages of alkaline salt stress. The DEGs in AM-314/MS-155 were mainly concentrated in the early stage, while Alamo showed greater advantages in the late stage. These DEGs were mainly enriched in plant-pathogen interactions, ubiquitin-mediated proteolysis and glycolysis/gluconeogenesis pathways. We characterized 1480 TF genes into 64 TF families, and the most abundant TF family was the C2H2 family, followed by the bZIP and bHLH families. A total of 1718 PKs were predicted, including CaMK, CDPK, MAPK and RLK. WGCNA revealed that the DEGs in the blue, brown, dark magenta and light steel blue 1 modules were associated with the physiological changes in roots of switchgrass under alkaline salt stress. The consistency between the qRT-PCR and RNA-Seq results confirmed the reliability of the RNA-seq sequencing data. A molecular regulatory network of the switchgrass response to alkaline salt stress was preliminarily constructed on the basis of transcriptional regulation and functional genes. CONCLUSIONS: Alkaline salt tolerance of switchgrass may be achieved by the regulation of ion homeostasis, transport proteins, detoxification, heat shock proteins, dehydration and sugar metabolism. These findings provide a comprehensive analysis of gene expression dynamic and act network induced by alkaline salt stress in two switchgrass genotypes and contribute to the understanding of the alkaline salt tolerance mechanism of switchgrass and the improvement of switchgrass germplasm.

H2S attenuates oxidative stress via Nrf2/NF-κB signaling to regulate restenosis after percutaneous transluminal angioplasty.

Restenosis after angioplasty of peripheral arteries is a clinical problem involving oxidative stress. Hydrogen sulfide (H2S) participates in oxidative stress regulation and activates nuclear factor erythroid 2-related factor 2 (Nrf2). This study investigated the effect of H2S and Nrf2 on restenosis-induced arterial injury. Using an in vivo rat model of restenosis, we investigated whether H2S inhibits restenosis after percutaneous transluminal angioplasty (PTA) and the oxidative stress-related mechanisms implicated therein. The involvement of Nrf2 was explored using Nrf2-shRNA. Neointimal formation and the deposition of elastic fibers were assessed histologically. Inflammatory cytokine secretion and the expression of proteins associated with oxidative stress and inflammation were evaluated. The artery of rats subjected to restenosis showed increased arterial intimal thickness, with prominent elastic fiber deposition. Sodium hydrosulfide (NaHS), an H2S donor, counteracted these changes in vivo. Restenosis caused a decrease in anti-oxidative stress signaling. This phenomenon was inhibited by NaHS, but Nrf2-shRNA counteracted the effects of NaHS. In terms of inflammation, inflammatory cytokines were upregulated, whereas NaHS suppressed the induced inflammatory reaction. Similarly, Nrf2 downregulation blocked the effect of NaHS. In vitro studies using aortic endothelial and vascular smooth muscle cells isolated from experimental animals showed consistent results as those of in vivo studies, and the participation of the nuclear factor-kappa B signaling pathway was demonstrated. Collectively, H2S played a role in regulating post-PTA restenosis by alleviating oxidative stress, modulating anti-oxidant defense, and targeting Nrf2-related pathways via nuclear factor-kappa B signaling.

Angiogenin maintains gut microbe homeostasis by balancing α-Proteobacteria and Lachnospiraceae.

OBJECTIVE: Antimicrobial peptides (AMPs) play essential roles in maintaining gut health and are associated with IBD. This study is to elucidate the effect of angiogenin (ANG), an intestine-secreted AMP, on gut microbiota and its relevance with IBD. DESIGN: The effect of ANG on microbiota and its contribution to colitis were evaluated in different colitis models with co-housing and faecal microbiota transplantation. ANG-regulated bacteria were determined by 16S rDNA sequencing and their functions in colitis were analysed by bacterial colonisation. The species-specific antimicrobial activity of ANG and its underlying mechanism were further investigated with microbiological and biochemical methods. ANG level and the key bacteria were characterised in IBD faecal samples. RESULTS: ANG regulated microbiota composition and inhibited intestinal inflammation. Specifically, Ang1 deficiency in mice led to a decrease in the protective gut commensal strains of Lachnospiraceae but an increase in the colitogenic strains of α-Proteobacteria. Direct binding of ANG to α-Proteobacteria resulted in lethal disruption of bacterial membrane integrity, and consequently promoted the growth of Lachnospiraceae, which otherwise was antagonised by α-Proteobacteria. Oral administration of ANG1 reversed the dysbiosis and attenuated the severity of colitis in Ang1-deficient mice. The correlation among ANG, the identified bacteria and IBD status was established in patients. CONCLUSION: These findings demonstrate a novel role of ANG in shaping gut microbe composition and thus maintaining gut health, suggesting that the ANG-microbiota axis could be developed as a potential preventive and/or therapeutic approach for dysbiosis-related gut diseases.

Myeloid cells protect intestinal epithelial barrier integrity through the angiogenin/plexin-B2 axis.

Communication between myeloid cells and epithelium plays critical role in maintaining intestinal epithelial barrier integrity. Myeloid cells interact with intestinal epithelial cells (IECs) by producing various mediators; however, the molecules mediating their crosstalk remain incompletely understood. Here, we report that deficiency of angiogenin (Ang) in mouse myeloid cells caused impairment of epithelial barrier integrity, leading to high susceptibility to DSS-induced colitis. Mechanistically, myeloid cell-derived angiogenin promoted IEC survival and proliferation through plexin-B2-mediated production of tRNA-derived stress-induced small RNA (tiRNA) and transcription of ribosomal RNA (rRNA), respectively. Moreover, treatment with recombinant angiogenin significantly attenuated the severity of experimental colitis. In human samples, the expression of angiogenin was significantly down-regulated in patients with inflammatory bowel disease (IBD). Collectively, we identified, for the first time to our knowledge, a novel mediator of myeloid cell-IEC crosstalk in maintaining epithelial barrier integrity, suggesting that angiogenin may serve as a new preventive agent and therapeutic target for IBD.

Angiogenin regulates PKD activation and COX-2 expression induced by TNF-α and bradykinin in the colonic myofibroblast.

INTRODUCTION: The myofibroblast is a gastrointestinal stromal cell that is a target of tumor necrosis factor-alpha (TNF-α), a pro-inflammatory cytokine strongly implicated in colitis-associated cancer. Crosstalk between TNF-α and other pro-inflammatory mediators amplify inflammatory signaling but the mechanism is unknown. Angiogenin (ANG) is a 14-kDa angiogenesis protein that is regulated in patients with inflammatory bowel disease. However, the role of ANG on inflammatory mediator crosstalk in the myofibroblast is unknown. METHODS: The human colonic myofibroblast cell line 18Co, as well as primary mouse and human colonic myofibroblasts, were exposed to TNF-α (10 ng/ml) and bradykinin (BK, 100 nM). ANG was quantified by ELISA. The expression of cyclo-oxygenase-2 (COX-2) and phosphorylation of PKD was assessed by Western Blot. RESULTS: Primary mouse and human colonic myofibroblasts exposed to TNF-α/BK led to enhanced PKD phosphorylation and synergistic COX-2 expression. 18Co cells secrete high levels of ANG (24h, 265 ± 5 pg/ml). The monoclonal antibody 26-2F, which neutralizes ANG, inhibited TNF-α/BK-mediated PKD phosphorylation and synergistic COX-2 expression in primary human myofibroblasts. Likewise, in primary mouse myofibroblasts that do not express ANG (ANG-KO), TNF-α/BK failed to enhance PKD phosphorylation and COX-2 expression. CONCLUSIONS: TNF-α/BK enhance PKD phosphorylation and COX-2 expression in primary mouse and human colonic myofibroblasts. Angiogenin is produced by the myofibroblast, and inhibition of ANG signaling, either by its absence (ANG-KO) or by pharmacologic inhibition, blocks enhanced PKD phosphorylation and synergistic COX-2 expression induced by TNF-α/BK. ANG mediates crosstalk signaling between TNF-α/BK in the regulation of stroma-derived COX-2 and may be a novel therapeutic target for the management of colitis-associated cancer.

Chemosensitization of prostate cancer stem cells in mice by angiogenin and plexin-B2 inhibitors.

Cancer stem cells (CSCs) are an obstacle in cancer therapy and are a major cause of drug resistance, cancer recurrence, and metastasis. Available treatments, targeting proliferating cancer cells, are not effective in eliminating quiescent CSCs. Identification of CSC regulators will help design therapeutic strategies to sensitize drug-resistant CSCs for chemo-eradication. Here, we show that angiogenin and plexin-B2 regulate the stemness of prostate CSCs, and that inhibitors of angiogenin/plexin-B2 sensitize prostate CSCs to chemotherapy. Prostate CSCs capable of self-renewal, differentiation, and tumor initiation with a single cell inoculation were identified and shown to be regulated by angiogenin/plexin-B2 that promotes quiescence and self-renewal through 5S ribosomal RNA processing and generation of the bioactive 3'-end fragments of 5S ribosomal RNA, which suppress protein translation and restrict cell cycling. Monoclonal antibodies of angiogenin and plexin-B2 decrease the stemness of prostate CSCs and sensitize them to chemotherapeutic agents in vitro and in vivo.

Full-length transcriptome sequencing reveals the low-temperature-tolerance mechanism of Medicago falcata roots.

BACKGROUND: Low temperature is one of the main environmental factors that limits crop growth, development, and production. Medicago falcata is an important leguminous herb that is widely distributed worldwide. M. falcata is related to alfalfa but is more tolerant to low temperature than alfalfa. Understanding the low temperature tolerance mechanism of M. falcata is important for the genetic improvement of alfalfa. RESULTS: In this study, we explored the transcriptomic changes in the roots of low-temperature-treated M. falcata plants by combining SMRT sequencing and NGS technologies. A total of 115,153 nonredundant sequences were obtained, and 8849 AS events, 73,149 SSRs, and 4189 lncRNAs were predicted. A total of 111,587 genes from SMRT sequencing were annotated, and 11,369 DEGs involved in plant hormone signal transduction, protein processing in endoplasmic reticulum, carbon metabolism, glycolysis/gluconeogenesis, starch and sucrose metabolism, and endocytosis pathways were identified. We characterized 1538 TF genes into 45 TF gene families, and the most abundant TF family was the WRKY family, followed by the ERF, MYB, bHLH and NAC families. A total of 134 genes, including 101 whose expression was upregulated and 33 whose expression was downregulated, were differentially coexpressed at all five temperature points. PB40804, PB75011, PB110405 and PB108808 were found to play crucial roles in the tolerance of M. falcata to low temperature. WGCNA revealed that the MEbrown module was significantly correlated with low-temperature stress in M. falcata. Electrolyte leakage was correlated with most genetic modules and verified that electrolyte leakage can be used as a direct stress marker in physiological assays to indicate cell membrane damage from low-temperature stress. The consistency between the qRT-PCR results and RNA-seq analyses confirmed the validity of the RNA-seq data and the analysis of the regulatory mechanism of low-temperature stress on the basis of the transcriptome. CONCLUSIONS: The full-length transcripts generated in this study provide a full characterization of the transcriptome of M. falcata and may be useful for mining new low-temperature stress-related genes specific to M. falcata. These new findings could facilitate the understanding of the low-temperature-tolerance mechanism of M. falcata.