LINC00330/CCL2 axis-mediated ESCC TAM reprogramming affects tumor progression.

BACKGROUND: Tumor-associated macrophages (TAMs) significantly influence the progression, metastasis, and recurrence of esophageal squamous cell carcinoma (ESCC). The aberrant expression of long noncoding RNAs (lncRNAs) in ESCC has been established, yet the role of lncRNAs in TAM reprogramming during ESCC progression remains largely unexplored. METHODS: ESCC TAM-related lncRNAs were identified by intersecting differentially expressed lncRNAs with immune-related lncRNAs and performing immune cell infiltration analysis. The expression profile and clinical relevance of LINC00330 were examined using the TCGA database and clinical samples. The LINC00330 overexpression and interference sequences were constructed to evaluate the effect of LINC00330 on ESCC progression. Single-cell sequencing data, CIBERSORTx, and GEPIA were utilized to analyze immune cell infiltration within the ESCC tumor microenvironment and to assess the correlation between LINC00330 and TAM infiltration. ESCC-macrophage coculture experiments were conducted to investigate the influence of LINC00330 on TAM reprogramming and its subsequent effect on ESCC progression. The interaction between LINC00330 and C-C motif ligand 2 (CCL2) was confirmed through transcriptomic sequencing, subcellular localization analysis, RNA pulldown, silver staining, RNA immunoprecipitation, and other experiments. RESULTS: LINC00330 is significantly downregulated in ESCC tissues and strongly associated with poor patient outcomes. Overexpression of LINC00330 inhibits ESCC progression, including proliferation, invasion, epithelial-mesenchymal transition, and tumorigenicity in vivo. LINC00330 promotes TAM reprogramming, and LINC00330-mediated TAM reprogramming inhibits ESCC progression. LINC00330 binds to the CCL2 protein and inhibits the expression of CCL2 and downstream signaling pathways. CCL2 is critical for LINC00330-mediated TAM reprogramming and ESCC progression. CONCLUSIONS: LINC00330 inhibited ESCC progression by disrupting the CCL2/CCR2 axis and its downstream signaling pathways in an autocrine fashion; and by impeding CCL2-mediated TAM reprogramming in a paracrine manner. The new mechanism of TAM reprogramming mediated by the LINC00330/CCL2 axis may provide potential strategies for targeted and immunocombination therapies for patients with ESCC.

An active tunable terahertz functional metamaterial based on hybrid-graphene vanadium dioxide.

In this paper, we propose a switchable and tunable functional metamaterial device based on hybrid graphene-vanadium dioxide (VO2). Using the properties of the metal-insulator transition in VO2, the proposed metamaterials can enable switching between tunable circular dichroism (CD) and dual-band perfect absorption in the terahertz region. When VO2 is in the insulator state, a polarization-selective single-band perfect absorption can be achieved for circularly polarized waves, thus resulting in a strong CD response with a maximum value of 0.84. When VO2 acts as a metal, there is a tunable dual-band perfect absorption for the designed metamaterial device under the illumination of x-polarization waves. The operation mechanism behind the phenomena can be explained by utilizing the electric field distribution and the coupled mode theory. Moreover, the influences of the Fermi energy of graphene and geometrical parameters on the CD and absorption spectra are discussed in detail. Our proposed switchable and tunable metamaterial can provide a platform for designing versatile functional devices in the terahertz region.

Morphological and anatomical characteristics of exserted stigma sterility and the location and function of SlLst (Solanum lycopersicum Long styles) gene in tomato.

KEY MESSAGE: Anatomical changes in and hormone roles of the exserted stigma were investigated, and localization and functional analysis of SlLst for the exserted stigma were performed using SLAF-BSA-seq, parental resequencing and overexpression of SlLst in tomato. Tomato accession T431 produces stigmas under relatively high temperatures (> 27 °C, the average temperature in Harbin, China, in June-August), so pollen can rarely reach the stigma properly. This allows the percentage of male sterility exceed 95%, making the use of this accession practical for hybrid seed production. To investigate the mechanism underlying the exserted stigma male sterility, the morphological changes of, anatomical changes of, and comparative endogenous hormone (IAA, ABA, GA3, ZT, SA) changes in flowers during flower development of tomato accessions DL5 and T431 were measured. The location and function of genes controlling exserted stigma sterility were analyzed using super SLAF-BSA-seq, parental resequencing, comparative genomics and the overexpression of SlLst in tomato. The results showed that an increase in cell number mainly caused stigma exsertion. IAA played a major role, while ABA had an opposite effect on stigma exertion. Moreover, 26 candidate genes related to the exserted stigma were found, located on chromosome 12. The Solyc12g027610.1 (SlLst) gene was identified as the key candidate gene by functional analysis. A subcellular localization assay revealed that SlLst is targeted to the nucleus and cell membrane. Phenotypic analysis of SlLst-overexpressing tomato showed that SlLst plays a crucial role during stigma exsertion.

The α-Subunit of the Chloroplast ATP Synthase of Tomato Reinforces Resistance to Gray Mold and Broad-Spectrum Resistance in Transgenic Tobacco.

Chloroplast ATP synthase (cpATPase) is responsible for ATP production during photosynthesis. Our previous studies showed that the cpATPase CF1 α subunit (AtpA) is a key protein involved in Clonostachys rosea-induced resistance to the fungus Botrytis cinerea in tomato. Here, we show that expression of the tomato atpA gene was upregulated by B. cinerea and Clonostachys rosea. The tomato atpA gene was then isolated, and transgenic tobacco lines were obtained. Compared with untransformed plants, atpA-overexpressing tobacco showed increased resistance to B. cinerea, characterized by reduced disease incidence, defense-associated hypersensitive response-like reactions, balanced reactive oxygen species, alleviated damage to the chloroplast ultrastructure of leaf cells, elevated levels of ATP content and cpATPase activity, and enhanced expression of genes related to carbon metabolism, photosynthesis, and defense. Incremental Ca2+ efflux and steady H+ efflux were observed in transgenic tobacco after inoculation with B. cinerea. In addition, overexpression of atpA conferred enhanced tolerance to salinity and resistance to the fungus Cladosporium fulvum. Thus, AtpA is a key regulator that links signaling to cellular redox homeostasis, ATP biosynthesis, and gene expression of resistance traits to modulate immunity to pathogen infection and provides broad-spectrum resistance in plants in the process.

Carbon storage regulator CsrA plays important roles in multiple virulence-associated processes of Clostridium difficile.

The carbon storage regulator CsrA is a global regulator that controls multiple virulence-associated processes including host cell invasion, virulence secretion, quorum sensing, biofilm formation, and motility in many pathogenic bacteria. However, the roles of CsrA in Clostridium difficile still remain unclear. In this study, a C. difficile strain overexpressing csrA was constructed to investigate its effects on multiple virulence associated processes. Overexpression of csrA resulted in flagella defect and poor motility in C. difficile 630Δerm, suggesting that CsrA involves in the regulation of flagellum synthesis. The levels of toxin production were increased in the C. difficile 630Δerm overexpressing of csrA. Moreover, csrA overexpression enhanced the adherence ability to Caco-2 cells and solvent production of C. difficile 630Δerm. Altogether, CsrA of C. difficile participates in multiple virulence processes including toxin production, motility, and adherence, and in the regulation of carbon metabolism. These results enhance our understanding of the regulatory functions of CsrA and reveal that CsrA is an important regulator in C. difficile contributing to virulence regulation.

High-mobility group box 1 protein contributes to the immunogenicity of rTcdB-treated CT26 cells.

Clostridium difficile TcdB is a key virulence factor that causes C. difficile-associated diseases. Our previous studies have shown that recombinant full-length TcdB (rTcdB) induces cell death in CT26 cells, and rTcdB-treated CT26 cells with high immunogenicity could stimulate dendritic cell (DC) activation and T cell activation in vitro. The rTcdB-treated CT26 cells also induce antitumor immunity in mice and protect mice from CT26 cells. High-mobility group box 1 protein (HMGB1) is a non-histone nuclear protein, which has various biological functions within the nucleus and also acts as an extracellular signal molecule involving in inflammatory diseases, cancers or autoimmune diseases. In this study, HMGB1 was found to be released from the rTcdB-treated CT26 cells. HMGB1 knockdown by using specific siRNA weakened the capacity of the BMDCs loaded with the rTcdB-treated CT26 cells to prime T cells in vitro and in vivo. The released HMGB1 from CT26 cells could interact with the receptor TLR4, which is closely related to DC activation and immune responses. The knockdown of HMGB1 also affected the phagocytosis of the rTcdB-treated CT26 cells by DCs in vitro. Furthermore, HMGB1 weakened the antitumor immunity of the rTcdB-treated CT26 cells, which protects mice from rechallenge of the live CT26 cells. Taken together, these results suggest that HMGB1 plays an important role on the immunogenicity of the rTcdB-treated dying CT26 cells.

Anemonin ameliorates human diploid fibroblasts 2BS and IMR90 cell senescence by PARP1-NAD+-SIRT1 signaling pathway.

OBJECTIVE: Aging becomes the most predominant risk factor for all age-associated pathological conditions with the increase of life expectancy and the aggravation of social aging. Slowing down the speed of aging is considered an effective way to improve health, but so far, effective anti-aging methods are relatively lacking. METHODS: Anemonin (ANE) was screened from eight existing small-molecule compounds by cell viability assay. The function of ANE was determined by the analysis of cell proliferation, ß -galactosidase (SA-ß -Gal) activity, cell cycle, SASP secretion, NAD+/NADH ratio, and other aging-related indicators. The targets of ANE were predicted by Drug Target Prediction System (DTPS) and Swiss Targe Prediction System. The effect of ANE on PARP-1-NAD+-SIRT1 signaling pathway was assessed by quantitative reverse-transcription polymerase chain reaction (RT-PCR), Western blot, PARP1, NAD+ and SIRT1 activity detection. RESULTS: ANE can delay cell senescence; PARP1 is one of the targets of ANE and plays a crucial role in ANE anti-aging; ANE release more NAD+ by inhibiting PARP1 activity, thereby conversely promoting the function of SIRT1 and delay cell senescence. CONCLUSIONS: Our study indicates that ANE can delay cellular senescence through the PARP1-NAD+-SIRT1 signaling pathway, which may be considered as an effective anti-aging strategy.

Aspirin suppresses hepatocellular carcinoma progression by inhibiting platelet activity.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most common malignant liver disease in the world. Platelets (PLTs) are known to play a key role in the maintenance of liver homeostasis and the pathophysiological processes of a variety of liver diseases. Aspirin is the most classic antiplatelet agent. However, the molecular mechanism of platelet action and whether aspirin can affect HCC progression by inhibiting platelet activity need further study. AIM: To explore the impact of the antiplatelet effect of aspirin on the development of HCC. METHODS: Platelet-rich plasma, platelet plasma, pure platelet, and platelet lysate were prepared, and a coculture model of PLTs and HCC cells was established. CCK-8 analysis, apoptosis analysis, Transwell analysis, and real-time polymerase chain reaction (RT-PCR) were used to analyze the effects of PLTs on the growth, metastasis, and inflammatory microenvironment of HCC. RT-PCR and Western blot were used to detect the effects of platelet activation on tumor-related signaling pathways. Aspirin was used to block the activation and aggregation of PLTs both in vitro and in vivo, and the effect of PLTs on the progression of HCC was detected. RESULTS: PLTs significantly promoted the growth, invasion, epithelial-mesenchymal transition, and formation of an inflammatory microenvironment in HCC cells. Activated PLTs promoted HCC progression by activating the mitogen-activated protein kinase/protein kinase B/signal transducer and activator of transcription three (MAPK/ AKT/STAT3) signaling axis. Additionally, aspirin inhibited HCC progression in vitro and in vivo by inhibiting platelet activation. CONCLUSION: PLTs play an important role in the pathogenesis of HCC, and aspirin can affect HCC progression by inhibiting platelet activity. These results suggest that antiplatelet therapy has promising application prospects in the treatment and combined treatment of HCC.

Escaping drought: The pectin methylesterase inhibitor gene Slpmei27 can significantly change drought resistance in tomato.

Drought stress will lead to a decrease in tomato yield and poor flavour, yield and quality, resulting in economic losses in agricultural production. Mining the key genes regulating tomato drought resistance is of great significance to improve the drought resistance of tomato plants. The cell wall can directly participate in the plant drought stress response as one of the main components of the cell wall, and the regulation of pectin content in plant drought resistance is still unclear. Here, the candidate gene Solyc08g006690 (Slpmei27) was obtained by fine mapping based on genome sequencing technology (BSA-seq) of late-maturing stress-resistant tomato mutants found in the field. Slpmei27 is expressed in the cell wall. The transient silencing of Slpmei27 by VIGS significantly improved the drought resistance of tomato. Meanwhile, Slpmei27 silencing could significantly change the cell wall structure of plants, change the stomatal pass rate, reduce the water loss rate of plants, improve the scavenging ability of reactive oxygen species, change the redox balance in plants, and thus improve the drought resistance of tomato. The promoter region of this gene contains a large number of hormone-response and stress-response binding sites. The promoter region of the Slpmei27 gene in the mutant could lower the expression of downstream genes. Through this study, the mechanism by which Slpmei27 improves tomato drought resistance was revealed, and the relationship between pectin methyl ester metabolism and plant drought resistance was established, providing a theoretical basis for the production of high-quality tomato materials with high drought resistance.

Slym1 control the color etiolation of leaves by facilitating the decomposition of chlorophyll in tomato.

Photosynthesis, as an important biological process of plants, produces organic substances for plant growth and development. Although the molecular mechanisms of photosynthesis had been well investigated, the relationship between chlorophyll synthesis and photosynthesis remains largely unknown. The leaf-color mutant was an ideal material for studying photosynthesis and chlorophyll synthesis, which had been seldom investigated in tomato. Here, we obtained a yellow leaf tomato mutant ym (The mutant plants from the line of zs4) in field. Transmission electron microscopy (TEM) and photosynthetic parameters results demonstrated that chloroplast's structure was obviously destroyed and photosynthetic capacity gets weak. The mutant was hybridized with the control to construct the F2 segregation population for sequencing. Slym1 gene, controlling yellow mutant trait, was identified using Bulked Segregation Analysis. Slym1 was up-regulated in the mutant and Slym1 was located in the nucleus. The genes associated with photosynthesis and chlorophyll synthesis were down-regulated in Slym1-OE transgenic tomato plants. The results suggested that Slym1 negatively regulate photosynthesis. Photosynthetic pigment synthesis related genes HEMA, HEMB1, CHLG and CAO were up-regulated in Slym1 silencing plants. The redundant Slym1 binding the intermediate proteins MP resulting in hindering the interaction between MP and HY5 due to the Slym1 with a high expression level in ym mutant, lead to lots of the HY5 with unbound state accumulates in cells, that could accelerate the decomposition of chlorophyll. Therefore, the yellow leaf-color mutant ym could be used as an ideal material for further exploring the relationship between leaf color mutant and photosynthesis and the specific mechanism.

Dynamics of vaginal microbiome in female beagles at different ages.

Age-related changes in human vaginal microbiota composition have been reported, and such changes might be influenced by humidity, external stimuli, hormone levels, drug to use, and other factors. However, there is no report about the vaginal microbiota composition of female beagles at different ages. To investigate the effects of aging on the vaginal microbiota independent of other effects, we analyzed the vaginal microbiomes of 23 beagles at a wide range of ages from 1 year to 7 years old (except the 3rd year), 1-2 y were categorized into youth stage (YS), 4-5 y were categorized into middle stage (MS), and 6-7 y were categorized into elderly stage (ES) based on age. Samples were collected by scraping the vaginal mucosa of YS (n = 8), MS (n = 5) and ES (n = 10), and analyzed by 16S-rRNA gene high-throughput-sequencing. The diversity of the vaginal microbiome in female beagles was found to continuously change with age. We also found associations between age and specific members and functions of the vaginal microbiome. The metabolism of terpenoids and polyketide and the cell motility are significantly enhanced with age. Our results suggest that the proportion of Tenericutes might be a biomarker which could distinguish between YS and others.

Advantages of Lateral Flow Assays Based on Fluorescent Submicrospheres and Quantum Dots for Clostridium difficile Toxin B Detection.

Clostridium difficile colitis is caused by a cytotoxin produced by the anaerobic bacteria C. difficile in the epithelial cells of the large intestine, particularly C. difficile toxin B (TcdB). However, the sensitivity of currently utilized C. difficile endotoxin determination methods has been called into question, and, therefore, more accurate and convenient detection methods are needed. Our study is the first to systematically compare fluorescent submicrosphere-based and quantum-dot nanobead-based lateral fluidity measurement methods (FMs-LFA and QDNBs-LFA) with toxin B quantification in fecal samples via sandwich analysis. The limits of detection (LOD) of FMs-LFA and QDNBs-LFA in the fecal samples were 0.483 and 0.297 ng/mL, respectively. TcdB analyses of the fecal samples indicated that the results of QDNBs-LFA and FMs-LFA were consistent with those of a commercial enzyme-linked immunosorbent assay (ELISA) test kit. The sensitivity of QDNBs-LFA was highly correlated with clinical diagnoses. Therefore, quantum dot nanobeads (QDNBs) are deemed highly suitable for lateral fluidity analyses, which would facilitate the implementation of portable and rapid on-the-spot applications, such as food hygiene and safety tests and onsite medical testing.

Rapid and fully-automated detection of Clostridium difficile Toxin B via magnetic-particle-based chemiluminescent immunoassay.

Clostridium difficile colitis is caused by a cytotoxin produced by the anaerobic bacteria C. difficile on the epithelial cells of the large intestine, particularly C. difficile toxin B (Tcd B). Current C. difficile toxin assays have proven to be insensitive and have thus been ruled out from diagnostic purposes. Therefore, Tcd B detection via sandwich-type chemiluminescent immunoassay was proposed as a straightforward approach with potential diagnostic applicability. Here, two high-affinity anti-Tcd B monoclonal antibodies were successfully identified and implemented in a fully-automated magnetic-particle-based chemiluminescent immunoassay (CLEIA). In this test, toxin B was sandwiched between the anti-toxin B antibody-coated magnetic particles and alkaline phosphate-labeled anti-toxin B antibodies. Compared with traditional techniques, the proposed immunoassay demonstrated high sensitivity for toxin B identification and was further optimized to achieve a linear response ranging from 0.12 to 150 ng/mL with a limit of detection (LOD) of 0.47 ng/mL. Importantly, the entire process could be completed in less than 30 minutes. The proposed assay was used to detect toxin B in 104 randomly-selected human stool samples and delivered similar results to those of a commercial ELISA kit, highlighting its great potential for rapid and efficient toxin B determination in human stool specimens.

Impact of biochar amendment on the uptake, fate and bioavailability of pharmaceuticals in soil-radish systems.

Crops grown in soils receiving wastewaters, biosolids, or manures can accumulate pharmaceuticals in edible parts, raising concerns over potential human exposure to multiple pharmaceuticals. Nonetheless, viable mitigation options for minimizing plant uptake of pharmaceuticals are limited. This study evaluated how biochar amendment could influence the uptake of 15 pharmaceuticals by radish (Raphanus sativus) grown in a sandy loam at two amendment rates (0.1 and 1% w/w). Comparing with that in the unamended soil, the accumulation of acetaminophen, carbamazepine, sulfadiazine, sulfamethoxazole, lamotrigine, carbadox, trimethoprim, oxytetracycline, tylosin, estrone, and triclosan in radish grown in the soil amended with 1.0% of biochar was significantly decreased by 33.3-83.0%. However, the concentration of lincomycin in radish was increased by 36.7-48.2% in the soil amended with 1% biochar. While the soil amended with 1.0% of biochar had increased sorption of all 15 pharmaceuticals, the persistence of 7 pharmaceuticals in the soil were prolonged, including caffeine, sulfadiazine, sulfamethoxazole, lincomycin, estrone, 17 ß-estradiol and triclosan. The reduced plant uptake of pharmaceuticals was mainly due to their lowered concentrations in pore water by the presence of biochar. Overall, the estimated daily intake data suggest that biochar amendment could potentially decrease total human exposure to a mixture of pharmaceuticals.

The micropeptide LEMP plays an evolutionarily conserved role in myogenesis.

In recent years, micropeptides have been increasingly identified as important regulators in various biological processes. However, whether micropeptides are functionally conserved remains largely unknown. Here, we uncovered a micropeptide with evolutionarily conserved roles in myogenesis. RNA-seq data analysis of proliferating mouse satellite cells (SCs) and differentiated myotubes identified a previously annotated lncRNA, MyolncR4 (1500011K16RIK), which is upregulated during muscle differentiation. Significantly, MyolncR4 is highly conserved across vertebrate species. Multiple lines of evidence demonstrate that MyolncR4 encodes a 56-aa micropeptide, which was named as LEMP (lncRNA encoded micropeptide). LEMP promotes muscle formation and regeneration in mouse. In zebrafish, MyolncR4 is enriched in developing somites and elimination of LEMP results in impaired muscle development, which could be efficiently rescued by expression of the mouse LEMP. Interestingly, LEMP is localized at both the plasma membrane and mitochondria, and associated with multiple mitochondrial proteins, suggestive of its involvement in mitochondrial functions. Together, our work uncovers a micropeptide that plays an evolutionarily conserved role in skeletal muscle differentiation, pinpointing the functional importance of this growing family of small peptides.

Endothelial CDS2 deficiency causes VEGFA-mediated vascular regression and tumor inhibition.

The response of endothelial cells to signaling stimulation is critical for vascular morphogenesis, homeostasis and function. Vascular endothelial growth factor-a (VEGFA) has been commonly recognized as a pro-angiogenic factor in vertebrate developmental, physiological and pathological conditions for decades. Here we report a novel finding that genetic ablation of CDP-diacylglycerol synthetase-2 (CDS2), a metabolic enzyme that controls phosphoinositide recycling, switches the output of VEGFA signaling from promoting angiogenesis to unexpectedly inducing vessel regression. Live imaging analysis uncovered the presence of reverse migration of the angiogenic endothelium in cds2 mutant zebrafish upon VEGFA stimulation, and endothelium regression also occurred in postnatal retina and implanted tumor models in mice. In tumor models, CDS2 deficiency enhanced the level of tumor-secreted VEGFA, which in-turn trapped tumors into a VEGFA-induced vessel regression situation, leading to suppression of tumor growth. Mechanistically, VEGFA stimulation reduced phosphatidylinositol (4,5)-bisphosphate (PIP2) availability in the absence of CDS2-controlled-phosphoinositide metabolism, subsequently causing phosphatidylinositol (3,4,5)-triphosphate (PIP3) deficiency and FOXO1 activation to trigger regression of CDS2-null endothelium. Thus, our data indicate that the effect of VEGFA on vasculature is context-dependent and can be converted from angiogenesis to vascular regression.

Time-resolved transcriptome analysis of Clostridium difficile R20291 response to cysteine.

The incidence of Clostridium difficile infection has been steadily rising over the past decade. The increase in the rate of incidence is associated with the specific NAP1/BI/027 strains which are "hypervirulent" and have led to several large outbreaks since their emergence. However, the relation between these outbreaks and virulence regulation mechanisms remains unclear. It has been reported that the major virulence factor TcdA and TcdB in C. difficile could be repressed by cysteine. Here, we investigated the functional and virulence-associated regulation of C. difficile R20291 response to cysteine by using a time-resolved genome-wide transcriptome analysis. Dramatic changes of gene expression in C. difficile revealed functional processes related to transport, metabolism, and regulators in the presence of cysteine during different phases of growth. Flagellar and ribosomal genes were significantly down-regulated in long-term response to cysteine. Many NAP1/BI/027- specific genes were also modulated by cysteine. In addition, cdsB inactivation in C. difficile R20291 could remove the repression of toxin synthesis but could not remove the repression of butyrate production in the presence of cysteine. This suggests that toxin synthesis and butyrate production might have different regulatory controls in response to cysteine. Altogether, our research provides important insights into the regulatory mechanisms of C. difficile response to cysteine.