Identification and Analysis of Immune Microenvironment-Related Genes for Keloid Risk Prediction and Their Effects on Keloid Proliferation and Migration.

Keloid is a kind of proliferative scar with continuous growth, no restriction and easy recurrence, which cannot be cured and bring serious physical injury and psychological burden to patients. The main reason is that the pathological mechanism is not clear. Therefore, this project is expected to reveal the immune microenvironment-related genes and their functions in keloid progression, and provide effective targets for the treatment of keloid. Firstly, 8 kinds of immune infiltrating cells and 19 potential characteristic genes were identified by immune infiltration analysis, ssGSEA, LASSO regression (glmnet algorithm and lars algorithm) and WGCNA, indicating that keloid was closely related to the changes of immune microenvironment. Then, 4 pathological biomarkers of keloid (MAPK1, PTPRC, STAT3 and IL1R1) were identified by differentially analysis, univariate analysis, LASSO regression (lars algorithm), support vector machine recursive feature elimination (SVM-REF) algorithm, multivariate logical regression analysis and six machine learning algorithms. Based on the 4 feature genes, the risk prediction model and nomogram were constructed. Calibration curve and ROC analysis (AUC = 0.930) showed that the model had reliable clinical value. Subsequently, consistent cluster analysis was used to find that there were 2 immune microenvironment subsets in keloid patients, of which subgroup II was immune subgroup. Multiple independent datasets and RT-qPCR showed that the expression trend of the 4 genes was consistent with the analysis. Cell gain-loss experiment confirmed that 4 genes regulated the proliferation and migration of keloid cells. The above data shows that MAPK1, PTPRC, STAT3 and IL1R1 may be personalized therapeutic targets for keloid patients.

Controllable Site-Selective Construction of 4- and 5-Hydroxyalkyl-Substituted Imidazoles from Amidines, Ynals, and Water.

The first example of controllable site-selective pathways to construct 4- and 5-hydroxyalkyl-substituted imidazoles through a three-component reaction of amidines, ynals, and water has been documented. Particularly, the high regioselectivity of the reaction was simply switched by changing the additives. In addition, further 18O-labeled experiments to probe a plausible mechanism and the gram-scale synthesis were studied.

Cu(I)-Catalyzed Three-Component Cyclization for the Construction of Functionalized Thiazoles.

A novel and straightforward strategy for the synthesis of functionalized thiazoles from thioamides, ynals, and alcohols via a copper(I)-catalyzed three-component reaction has been described. Through the formation of new C-S, C-N, and C-O bonds in one pot, it is easy to produce various valuable thiazoles fixed with aryl or heteroaryl groups. In addition, the reaction also exhibits other unique advantages, such as high step economics, good functional group tolerance, and good regioselectivity.

Synthesis of Pyrrolo[2,1,5-cd]indolizine Rings via Visible-Light-Induced Intermolecular [3+2] Cycloaddition of Indolizines and Alkynes.

A range of indolizine smoothly underwent visible-light-induced intermolecular [3+2] annulations with internal alkynes to afford pyrrolo[2,1,5-cd]indolizine in good to excellent yields with high regioselectivity. Through this cascade reaction, a series of fluoroactive fused indolizines with a large π-system were conveniently synthesized. The usage of visible light as energy source with air as a stoichiometric oxidant under simple conditions makes this process attractive and practical.

Differences in responses of grass carp to different types of grass carp reovirus (GCRV) and the mechanism of hemorrhage revealed by transcriptome sequencing.

BACKGROUND: Grass carp is an important farmed fish in China that is affected by serious disease, especially hemorrhagic disease caused by grass carp reovirus (GCRV). The mechanism underlying the hemorrhagic symptoms in infected fish remains to be elucidated. Although GCRV can be divided into three distinct subtypes, differences in the pathogenesis and host immune responses to the different subtypes are still unclear. The aim of this study was to provide a comprehensive insight into the grass carp response to different GCRV subtypes and to elucidate the mechanism underlying the hemorrhagic symptoms. RESULTS: Following infection of grass carp, GCRV-I was associated with a long latent period and low mortality (42.5%), while GCRV-II was associated with a short latent period and high mortality (81.4%). The relative copy number of GCRV-I remained consistent or decreased slightly throughout the first 7 days post-infection, whereas a marked increase in GCRV-II high copy number was detected at 5 days post-infection. Transcriptome sequencing revealed 211 differentially expressed genes (DEGs) in Group I (66 up-regulated, 145 down-regulated) and 670 (386 up-regulated, 284 down-regulated) in Group II. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed significant enrichment in the terms or pathways involved in immune responses and correlating with blood or platelets. Most of the DEGs in Group I were also present in Group II, although the expression profiles differed, with most DEGs showing mild changes in Group I, while marked changes were observed in Group II, especially the interferon-related genes. Many of the genes involved in the complement pathway and coagulation cascades were significantly up-regulated at 7 days post-infection in Group II, suggesting activation of these pathways. CONCLUSION: GCRV-I is associated with low virulence and a long latent period prior to the induction of a mild host immune response, whereas GCRV-II is associated with high virulence, a short latent period and stimulates a strong and extensive host immune response. The complement and coagulation pathways are significantly activated at 7 days post-infection, leading to the endothelial cell and blood cell damage that result in hemorrhagic symptoms.

Cloning and characterization of Bax1 and Bax2 genes of Ctenopharyngodon idellus and evaluation of transcript expression in response to grass carp reovirus infection.

Multidomain proapoptotic Bcl-2-associated X (Bax) protein is an essential effector responsible for mitochondrial outer membrane permeabilization, resulting in cell death via apoptosis. In this study, two Bax genes of grass carp (Ctenopharyngodon idellus), designated as CiBax1 and CiBax2, were isolated and analyzed. The obtained CiBax1 cDNA is 2058 bp long, with a 579 bp open reading frame (ORF) coding a protein of 192 amino acid residues. The full-length cDNA of CiBax2 is 1161 bp, with a 618 bp ORF coding 205 amino acids. Both CiBax1 and CiBax2 are typical members of Bcl-2 family containing conserved Bcl and C-terminal domains, and they share conserved synteny with zebrafish Bax genes despite the grass carp Bax mapping to different linkage groups. Phylogenetic analysis showed that CiBax1 was clustered with Bax from most teleost fish, and CiBax2 was close to Bax2 from teleost fish but far separated from that of Salmo salar. Quantitative real-time PCR analysis revealed broad expression of CiBax1 and CiBax2 in tissues from healthy grass carp, but the relative expression level differed. The mRNA expression of CiBax1 and CiBax2 was both upregulated significantly and peaked in all examined tissues at days 5 or 6 post-infection with grass carp reovirus. Subcellular localization indicated that CiBax1 protein was localized in both nucleus and cytosol, while CiBax2 protein only in cytosol. Moreover, CiBax2, but not CiBax1 was colocalized with mitochondrion under normal condition. Taken together, the findings would be helpful for further understanding of the function of Bax in teleost fish.

Global gene expression patterns of grass carp following compensatory growth.

BACKGROUND: Compensatory growth is accelerated compared with normal growth and occurs when growth-limiting conditions are overcome. Most animals, especially fish, are capable of compensatory growth, but the mechanisms remain unclear. Further investigation of the mechanism of compensatory growth in fish is needed to improve feeding efficiency, reduce cost, and explore growth-related genes. RESULTS: In the study, grass carp, an important farmed fish in China, were subjected to a compensatory growth experiment followed by transcriptome analysis by RNA-sequencing. Samples of fish from starved and re-feeding conditions were compared with the control. Under starved conditions, 4061 and 1988 differentially expressed genes (DEGs) were detected in muscle and liver tissue when compared the experimental group with control group, respectively. After re-feeding, 349 and 247 DEGs were identified in muscle and liver when the two groups were compared. Moreover, when samples from experimental group in starved and re-feeding conditions were compared, 4903 and 2444 DEGs were found in muscle and liver. Most of these DEGs were involved in metabolic processes, or encoded enzymes or proteins with catalytic activity or binding functions, or involved in metabolic and biosynthetic pathways. A number of the more significant DEGs were subjected to further analysis. Under fasting conditions, many up-regulated genes were associated with protein ubiquitination or degradation, whereas many down-regulated genes were involved in the metabolism of glucose and fatty acids. Under re-feeding conditions, genes participating in muscle synthesis and fatty acid metabolism were up-regulated significantly, and genes related to protein ubiquitination or degradation were down-regulated. Moreover, Several DEGs were random selected for confirmation by real-time quantitative PCR. CONCLUSIONS: Global gene expression patterns of grass carp during compensatory growth were determined. To our knowledge, this is a first reported for a teleost fish. The results will enhance our understanding of the mechanism of compensatory growth in teleost fish.

Expression pattern and transcriptional regulatory mechanism of noxa gene in grass carp (Ctenopharyngodon idella).

Noxa, a pro-apoptotic protein, plays an important role in cell apoptosis. The researches about noxa gene were concentrated in mammalians, whereas the role and transcriptional regulatory mechanism of noxa in fish were still unclear. In this study, the expression pattern and transcriptional regulatory mechanism of noxa gene in grass carp were analyzed. Noxa was constitutively expressed in all the examined tissues but the relative expression level differed. After exposure to grass carp reovirus (GCRV), mRNA expression level of noxa was down-regulated at the early phase whereas up-regulated at the late phase of infection. Luciferase assays showed that the promoter region -867 ∼ +107 of noxa had high activity and the region -678 ∼ -603 was important in the response to GCRV infection. By deleting the predicted transcription factor binding sites, transcription factors FOXO1 and CEBPß were found important for noxa in response to GCRV infection. Moreover, the noxa promoter was biotin-labeled and incubated with nuclear extracts from GCRV infected cells. Mass spectrometry analysis showed that transcription factors FOXO1 and CEBPß were also enriched in the combined proteins. Therefore, the results suggested that transcription factors FOXO1 and CEBPß may play an important role in the regulation of noxa. Our study would provide new insight into the transcriptional regulatory mechanism of noxa in teleost fish.

Isolation and expression of grass carp toll-like receptor 5a (CiTLR5a) and 5b (CiTLR5b) gene involved in the response to flagellin stimulation and grass carp reovirus infection.

Toll-like receptor 5 (TLR5), a member of Toll-like receptors (TLRs) family and is responsible for the bacterial flagellin recognition in vertebrates, play an important role in innate immunity. In the study, two TLR5 genes of grass carp (Ctenopharyngodon idellus), named CiTLR5a and CiTLR5b, were cloned and analyzed. Both CiTLR5a and CiTLR5b are typical TLR proteins, including LRR motif, transmembrane region and TIR domain. The full-length cDNA of CiTLR5a is 3054 bp long, with a 2646 bp open reading frame (ORF), 78 bp 5' untranslated regions (UTR), and 330 bp 3' UTR. The full-length cDNA of CiTLR5b is 3326 bp, with a 2627 bp ORF, 95 bp 5' UTR, and 594 bp 3' UTR. Phylogenetic analysis showed that CiTLR5a and CiTLR5b were closed to the TLR5 of cirrhinus mrigala, cyprinus_carpio, and danio rerio. Subcellular localization indicated that CiTLR5a and CiTLR5b shared similar localization pattern and may locate in the plasma membrane of transfected cells. Real-time quantitative PCR revealed CiTLR5a and CiTLR5b were constitutively expressed in all examined tissues, whereas the highest expressed tissue differed. Following exposure to flagellin and GCRV, CiTLR5a and CiTLR5b were up-regulated significantly. Moreover, the downstream genes of TLR5 signal pathway such as MyD88, NF-κB, IRF7, IL-1ß, and TNF-α also up-regulated significantly, whereas the IκB gene was down-regulated, suggesting that CiTLR5a and CiTLR5b involved in response to flagellin stimulation and GCRV infection. The results obtained in the study would provide a new insight for further understand the function of TLR5 in teleost fish.

Analysis of Immune and Prognostic-Related lncRNA PRKCQ-AS1 for Predicting Prognosis and Regulating Effect in Sepsis.

Background: Sepsis was a high mortality and great harm systemic inflammatory response syndrome caused by infection. lncRNAs were potential prognostic marker and therapeutic target. Therefore, we expect to screen and analyze lncRNAs with potential prognostic markers in sepsis. Methods: Transcriptome sequencing and limma was used to screen dysregulated RNAs. Key RNAs were screened by correlation analysis, lncRNA-mRNA co-expression and weighted gene co-expression network analysis. Immune infiltration, gene set enrichment analysis and gene set variation analysis were used to analyze the immune correlation. Kaplan-Meier curve, receiver operator characteristic curve, Cox regression analysis and nomogram were used to analyze the correlation between key RNAs and prognosis. Sepsis model was established by lipopolysaccharide-induced HUVECs injury, and then cell viability and migration ability were detected by cell counting kit-8 and wound healing assay. The levels of apoptosis-related proteins and inflammatory cytokines were detected by RT-qPCR and Western blot. Reactive Oxygen Species and superoxide dismutase were detected by commercial kit. Results: Fourteen key differentially expressed lncRNAs and 663 key differentially expressed genes were obtained. And these lncRNAs were closely related to immune cells, especially T cell activation, immune response and inflammation. Subsequently, Subsequently, lncRNA PRKCQ-AS1 was identified as the regulator for further investigation in sepsis. RT-qPCR results showed that PRKCQ-AS1 expression was up-regulated in clinical samples and sepsis model cells, which was an independent prognostic factor in sepsis patients. Immune correlation analysis showed that PRKCQ-AS1 was involved in the immune response and inflammatory process of sepsis. Cell function tests confirmed that PRKCQ-AS1 could inhibit sepsis model cells viability and promote cell apoptosis, inflammatory damage and oxidative stress. Conclusion: We constructed immune-related lncRNA-mRNA regulatory networks in the progression of sepsis and confirmed that PRKCQ-AS1 is an important prognostic factor affecting the progression of sepsis and is involved in immune response.

Construction and evaluation of Alzheimer's disease diagnostic prediction model based on genes involved in mitophagy.

Introduction: Alzheimer's disease (AD) is a common neurodegenerative disease. The concealment of the disease is the difficulty of its prevention and treatment. Previous studies have shown that mitophagy is crucial to the development of AD. However, there is a lack of research on the identification and clinical significance of mitophagy-related genes in AD. Therefore, the purpose of this study was to identify the mitophagy-related genes with the diagnostic potential for AD and establish a diagnostic model for AD. Methods: Firstly, we download the AD gene expression profile from Gene Expression Omnibus (GEO). Limma, PPI, functional enrichment analysis and WGCNA were used to screen the differential expression of mitophagy-related AD gene. Then, machine learning methods (random forest, univariate analysis, support vector machine, LASSO regression and support vector machine classification) were used to identify diagnostic markers. Finally, the diagnostic model was established and evaluated by ROC, multiple regression analysis, nomogram, calibration curve and other methods. Moreover, multiple independent datasets, AD cell models and AD clinical samples were used to verify the expression level of characteristic genes in the diagnostic model. Results: In total, 72 differentially expressed mitophagy-related related genes were identified, which were mainly involved in biological functions such as autophagy, apoptosis and neurological diseases. Four mitophagy-related genes (OPTN, PTGS2, TOMM20, and VDAC1) were identified as biomarkers. A diagnostic prediction model was constructed, and the reliability of the model was verified by receiver operating characteristic (ROC) curve analysis of GSE122063 and GSE63061. Then, we combine four mitophagy-related genes with age to establish a nomogram model. The ROC, C index and calibration curve show that the model has good prediction performance. Finally, multiple independent datasets, AD cell model samples and clinical peripheral blood samples confirmed that the expression levels of four mitophagy-related genes were consistent with the results of bioinformatics analysis. Discussion: The analysis results and diagnostic model of this study are helpful for the follow-up clinical work and mechanism research of AD.

High-throughput sequencing approach for the identification of lncRNA biomarkers in hepatocellular carcinoma and revealing the effect of ZFAS1/miR-150-5p on hepatocellular carcinoma progression.

Aims: To screen abnormal lncRNAs and diagnostic biomarkers in the progression of hepatocellular carcinoma through high-throughput sequencing and explore the underlying mechanisms of abnormal lncRNAs in the progression of hepatocellular carcinoma. Methods: The transcriptome sequencing was used to analyze the RNA expression profile and identify differentially expressed RNAs. Hub lncRNAs were screened by combining (WGCNA, ceRNA regulatory network, PPI, GO and KEGG analyses, Kaplan-Meier curve analysis, Cox analysis, risk model construction and qPCR). Thereafter, the correlation between the expression of hub lncRNAs and tumor clinicopathological parameters was analyzed, and the hub lncRNAs were analyzed by GSEA. Finally, the effects of hub RNAs on the proliferation, migration and invasion of HepG2 cells were investigated in vitro. Results: Compared with the control group, a total of 610 lncRNAs, 2,593 mRNAs and 26 miRNAs were screened in patients with hepatocellular carcinoma. Through miRNA target prediction and WGCNA, a ceRNA was constructed, comprising 324 nodes and 621 edges. Enrichment analysis showed that mRNAs in ceRNA were involved mainly in cancer development progression. Then, the ZFAS1/miR-150-5p interaction pair was screened out by Kaplan Meier curve analysis, Cox analysis and qPCR analysis. Its expression was related to tumor stage, TNM stage and patient age. ROC curve analysis showed that it has a good predictive value for the risk of hepatocellular carcinoma. GSEA showed that ZFAS1 was also enriched in the regulation of immune response, cell differentiation and proliferation. Loss-of-function experiments revealed that ZFAS1 inhibition could remarkably suppress HepG2 cell proliferation, migration and invasion in vitro. Bioinformatic analysis and luciferase reporter assays revealed that ZFAS1 directly interacted with miR-150-5p. Rescue experiments showed that a miR-150-5p inhibitor reversed the cell proliferation, migration and invasion functions of ZFAS1 knockdown in vitro. Conclusion: ZFAS1 is associated with the malignant status and prognosis of patients with hepatocellular carcinoma, and the ZFAS1/miR-150-5p axis is involved in hepatocellular carcinoma progression.

Identification of miRNA-mRNA Pairs in the Alzheimer's Disease Expression Profile and Explore the Effect of miR-26a-5p/PTGS2 on Amyloid-ß Induced Neurotoxicity in Alzheimer's Disease Cell Model.

Alzheimer's disease (AD) is a progressive neurodegenerative disease and the most common type of dementia. MicroRNAs (miRNAs) have been extensively studied in many diseases, including AD. To identify the AD-specific differentially expressed miRNAs and mRNAs, we used bioinformatics analysis to study candidate miRNA-mRNA pairs involved in the pathogenesis of AD. These miRNA-mRNAs may serve as promising biomarkers for early diagnosis or targeted therapy of AD patients. In this study, based on the AD mRNA and miRNA expression profile data in Gene Expression Omnibus (GEO), through differential expression analysis, functional annotation and enrichment analysis, weighted gene co-expression network analysis, miRNA-mRNA regulatory network, protein-protein interaction network, receiver operator characteristic and Least absolute shrinkage and selection operator (LASSO) regression and other analysis, we screened the key miRNA-mRNA in the progress of AD: miR-26a-5p/PTGS2. Dual-luciferase and qPCR experiments confirmed that PTGS2 is a direct target gene of miR-26a-5p. The expression of miR-26a-5p in the peripheral blood of AD patients and AD model cells (SH-SY5Y cells treated with Aß25-35) was up-regulated, and the expression of PTGS2 was down-regulated. Functional gain -loss experiments confirmed that PTGS2 protects AD model cells from damage by inhibiting proliferation and migration. However, the expression of miR-26a-5p promotes the proliferation of AD model cells. It is further found that PTGS2 is involved in the regulation of miR-26a-5p and can reverse the effect of miR-26a-5p on the proliferation of AD model cells. In addition, through network pharmacology, qPCR and CCK-8, we found that baicalein may affect the progression of AD by regulating the expression of PTGS2. Therefore, PTGS2 can be used as a target for AD research, and miR-26a-5p/PTGS2 can be used as an axis of action to study the pathogenesis of AD.

Use of miRNA Sequencing to Reveal Hub miRNAs and the Effect of miR-582-3p/SMAD2 in the Progression of Hepatocellular Carcinoma.

Hepatocellular carcinoma is a common tumor with a high fatality rate worldwide, and exploring its pathogenesis and deterioration mechanism is a focus for many researchers. Increasing evidence has shown that miRNAs are involved in the occurrence and progression of a variety of cancers, including hepatocellular carcinoma. Therefore, this study mainly aimed identify key miRNAs related to hepatocellular carcinoma and explore their potential functions and clinical significance. In this study, we performed miRNA sequencing on three pairs of hepatocellular carcinoma tissue samples and screened 26 differentially expressed miRNAs. Then 2 key miRNAs (miR-139-5p and miR-582-3p) were screened by Kaplan-Meier curve analysis, Cox multivariate analysis and qPCR methods. The expression of miR-582-3p was positively correlated with clinicopathological parameters in patients with hepatocellular carcinoma. Subsequently, miRwalk and starbase were used to predict the target genes of key miRNAs, and then the key pairs miR-582-3p/SMAD2 identified by WGCNA, PPI, qPCR and Pearson correlation analysis. Finally, a dual luciferase experiment, the rescue-of-function experiment and qPCR confirmed that miR-582-3p directly targets SMAD2 and regulates the proliferation, migration and invasion of HepG2 cells by targeting SMAD2. At the same time, interference with SMAD2 can influence the effect of miR-582-3p on HepG2 cells. In conclusion, our findings confirm that miR-582-3p is an independent factor for the prognosis of hepatocellular carcinoma patients, and can regulate the progression of hepatocellular carcinoma cells by targeting SMAD2.

Integrated Dissection of lncRNA-miRNA-mRNA Pairs and Potential Regulatory Role of lncRNA PCAT19 in Lung Adenocarcinoma.

Lung adenocarcinoma (LUAD) is the main cause of cancer-related deaths worldwide. Long noncoding RNAs have been reported to play an important role in various cancers due to their special functions. Therefore, identifying the lncRNAs involved in LUAD tumorigenesis and development can help improve therapeutic strategies. The TCGA-LUAD RNA expression profile was downloaded from The Cancer Genome Atlas, and a total of 49 differential lncRNAs, 112 differential miRNAs, and 2,953 differential mRNAs were screened. Through Kaplan-Meier curves, interaction networks, hub RNAs (lncRNAs, miRNAs, and mRNAs) were obtained. These hub genes are mainly involved in cell proliferation, cell cycle, lung development, and tumor-related signaling pathways. Two lncRNAs (SMIM25 and PCAT19) more significantly related to the prognosis of LUAD were screened by univariate Cox analysis, multivariate Cox analysis, and risk model analysis. The qPCR results showed that the expression levels of SMIM25 and PCAT19 were downregulated in clinical tissues, A549 and SPC-A1 cells, which were consistent with the bioinformatics analysis results. Subsequently, the PCAT19/miR-143-3p pairs were screened through the weighted gene co-expression network analysis and miRNA-lncRNA regulatory network. Dual luciferase detection confirmed that miR-143-3p directly targets PCAT19, and qPCR results indicated that the expression of the two is positively correlated. Cell function tests showed that overexpression of PCAT19 could significantly inhibit the proliferation, migration, and invasion of A549 and SPC-A1 cells. In contrast, knockout of PCAT19 can better promote the proliferation and migration of A549 and SPC-A1 cells. The expression of PCAT19 was negatively correlated with tumor grade, histological grade, and tumor mutation load in LUAD. In addition, co-transfection experiments confirmed that the miR-143-3p mimic could partially reverse the effect of PCAT19 knockout on the proliferation of A549 and SPC-A1 cells. In summary, PCAT19 is an independent prognostic factor in patients with LUAD that can regulate the proliferation, migration, and invasion of LUAD cells and may be a potential biomarker for the diagnosis of LUAD. PCAT19/miR-143-3p plays a very important regulatory role in the occurrence and development of LUAD.

Identification of Potential Hub Genes and miRNA-mRNA Pairs Related to the Progression and Prognosis of Cervical Cancer Through Integrated Bioinformatics Analysis.

In recent years, the incidence and mortality of cervical cancer have increased worldwide. At the same time, increasing data have confirmed that miRNA-mRNA plays a positive or negative regulatory role in many cancers. This study attempted to screen effective miRNA-mRNA in the progression of cervical cancer, and to study the mechanism of miRNA-mRNA in the progression of cervical cancer. The expression profile data of GSE7410, GSE 63514, GSE 86100 and TCGA-CESC were downloaded, and 34 overlapping differentially expressed genes (22 up-regulated and 12 down-regulated) and 166 miRNAs (74 down-regulated and 92 up-regulated) were screened through limma package. Then, miR-197-3p/TYMS pairs were obtained by PPI, functional enrichment, Kaplan-Meier plotter analysis, Cox univariate and multivariate analysis, risk modeling, WGCNA, qPCR and dual-luciferase experiments. The results showed that TYMS was an independent prognostic factor of cervical cancer, and its expression level was negatively correlated with cervical cancer tissue grade (TMN), tumor grade, age, microsatellite stability and tumor mutation load, and positively correlated with methyl expression in DNMT1, DNMT2, DNMT3A and DNMT3B. Functional experiments showed that TYMS knockout could promote the proliferation, migration and invasion of HeLa cells and reduce apoptosis. Overexpression of TYMS showed the opposite trend, miR-197-3p was negatively correlated with the expression of TYMS. MiR-197-3p inhibitor reversed the effect of si-TYMS on the proliferation of HeLa cells. In conclusion, these results reveal that TYMS plays a very important role in the prognosis and progression of cervical cancer, and has the potential to be thought of as cervical cancer biomarkers. At the same time, miR-197-3p/TYMS axis can regulate the deterioration of cervical cancer cells, which lays a foundation for the molecular diagnosis and treatment of cervical cancer.

Visible Light-Induced Cascade Cyclization of 3-Aminoindazoles, Ynals, and Chalcogens: Access to Chalcogen-Containing Pyrimido[1,2-b]-indazoles.

A direct cascade cyclization of 3-aminoindazoles, ynals, and accessible chalcogens facilitated by visible light has been developed. A series of fluoroactive selenium/tellurium-substituted pyrimido[1,2-b]-indazoles were easily accessed in moderate to good yields with a broad scope. Furthermore, we surveyed the spectral properties of selenide pyrimido[1,2-b]-indazoles prepared by this method.

A rapid "on-off-on" mitochondria-targeted phosphorescent probe for selective and consecutive detection of Cu2+ and cysteine in live cells and zebrafish.

The detection of mitochondrial Cu2+ and cysteine is very important for investigating cellular functions or dysfunctions. In this study, we designed a novel cyclometalated iridium(iii) luminescence chemosensor Ir bearing a bidentate chelating pyrazolyl-pyridine ligand as a copper-specific receptor. The biocompatible and photostable Ir complex exhibited not only mitochondria-targeting properties but also an "on-off-on" type phosphorescence change for the reversible dual detection of Cu2+ and cysteine. Ir had a highly sensitive (detection limit = 20 nM) and selective sensor performance for Cu2+ in aqueous solution due to the formation of a non-phosphorescent Ir-Cu(ii) ensemble through 1 : 1 binding. According to the displacement approach, Ir was released from the Ir-Cu(ii) ensemble accompanied with "turn-on" phosphorescence in the presence of 0-10 µM cysteine, with a low detection limit of 54 nM. This "on-off-on" process could be accomplished within 30 s and repeated at least five times without significant loss of signal strength. Moreover, benefiting from its good permeability, low cytotoxicity, high efficiency, and anti-interference properties, Ir was found to be suitable for imaging and detecting mitochondrial Cu2+ and cysteine in living cells and zebrafish.

RNA-seq profiles from grass carp tissues after reovirus (GCRV) infection based on singular and modular enrichment analyses.

Hemorrhagic disease of the grass carp, Ctenopharyngodon idella, is a fatal disease in fingerlings and yearlings caused by a reovirus, GCRV. RNA-seq data from four diseased grass carp tissues (gill, intestine, liver and spleen) were obtained at 2h before and six times after (2h, 24h, 48h, 72h, 96h and 120h) GCRV challenge. A total of 7.25±0.18 million (M) clean reads and 3.53±0.37M unique reads were obtained per RNA-seq analysis. Compared with controls, there were 9060 unique differentially expressed genes (DEGs) in the four tissues at the six time points post-GCRV challenge. Hierarchical clustering analysis of the DEGs showed that the data from the six time points fell into three branches: 2h, 24h/48h, and 72h/96h/120h. Singular (SEA) and modular enrichment analyses of DEGs per RNA-seq dataset were performed based on gene ontology. The results showed that immune responses occurred in all four tissues, indicating that GCRV probably does not target any tissue specifically. Moreover, during the course of disease, disturbances were observed in lipid and carbohydrate metabolism in each of the organs. SEA of DEGs based on the Kyoto Encyclopedia of Genes and Genomes database was also performed, and this indicated that the complement system and cellular immunity played an important role during the course of hemorrhagic disease. The qPCR of pooled samples of duplicate challenge experiment were used to confirm our RNA-seq approach.