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Genetic variants can affect gene expression by altering the level of N6-methyladenosine (m6A) modifications. A better understanding of the association of these genetic variants with susceptibility to cervical cancer (CC) can promote advances in disease screening and treatment. Genome-wide identification of m6A-associated functional SNPs for CC was performed using the TCGA and JENGER databases, incorporating the data from RNA-seq and MeRIP-seq. The screened risk-associated SNP rs1059288 (A>G), which is located in the 3′ UTR of TAPBP, was further validated in a case–control study involving 921 cases and 1077 controls. The results revealed a significant association between rs1059288 and the risk of CC (OR 1.48, 95% CI 1.13–1.92). Mechanistically, the presence of the risk G allele of rs1059288 was associated with increased m6A modification of TAPBP compared with the A allele. This modification was facilitated by the m6A methyltransferase METTL14 and the reading protein YTHDF2. Immunohistochemical staining of tissue microarrays containing 61 CC and 45 normal tissues showed an overexpression of TAPBP in CC. Furthermore, the upregulation of TAPBP promoted the growth and migration of CC cells as well as tumor-forming ability, inhibited apoptosis, and conferred increased resistance to commonly used chemotherapeutic drugs such as bleomycin, cisplatin, and doxorubicin. Knockdown of TAPBP inhibited the JAK/STAT/MICB signaling pathway in CC cells and upregulated certain immune genes including ISG15, IRF3, PTPN6, and HLA-A. These findings offer insights into the involvement of genetic variations in TAPBP in the development and progression of CC. [ABSTRACT FROM AUTHOR]

The post-genomic era has ushered in the extensive application of epigenetic editing tools, allowing for precise alterations of gene expression. The use of reprogrammable editors that carry transcriptional corepressors has significant potential for long-term epigenetic silencing for the treatment of human diseases. The ideal scenario involves precise targeting of a specific genomic location by a DNA-binding domain, ensuring there are no off-target effects and that the process yields no genetic remnants aside from specific epigenetic modifications (i.e., DNA methylation). A notable example is a recent study on the mouse Pcsk9 gene, crucial for cholesterol regulation and expressed in hepatocytes, which identified synthetic zinc-finger (ZF) proteins as the most effective DNA-binding editors for silencing Pcsk9 efficiently, specifically, and persistently. This discussion focuses on enhancing the specificity of ZF-array DNA binding by optimizing interactions between specific amino acids and DNA bases across three promoters containing CpG islands. [ABSTRACT FROM AUTHOR]

Introduction: Electroconvulsive therapy (ECT) is widely used for treatment‐resistant depression. However, it is unclear whether/how ECT can be targeted to affect brain regions and circuits in the brain to dynamically regulate mood and cognition. Methods: This study used brain entropy (BEN) to measure the irregular levels of brain systems in 46 major depressive disorder (MDD) patients before and after ECT treatment. Functional connectivity (FC) was further adopted to reveal changes of functional couplings. Moreover, transcriptomic and neurotransmitter receptor data were used to reveal genetic and molecular basis of the changes of BEN and functional connectivities. Results: Compared to pretreatment, the BEN in the posterior cerebellar lobe (PCL) significantly decreased and FC between the PCL and the right temporal pole (TP) significantly increased in MDD patients after treatment. Moreover, we found that these changes of BEN and FC were closely associated with genes' expression profiles involved in MAPK signaling pathway, GABAergic synapse, and dopaminergic synapse and were significantly correlated with the receptor/transporter density of 5‐HT, norepinephrine, glutamate, etc. Conclusion: These findings suggest that loops in the cerebellum and TP are crucial for ECT regulation of mood and cognition, which provides new evidence for the antidepressant effects of ECT and the potential molecular mechanism leading to cognitive impairment. [ABSTRACT FROM AUTHOR]

Objective: Existing research suggests that bone marrow-derived mesenchymal stem cells (BMSCs) may promote endogenous bone repair. This may be through the secretion of factors that stimulate repair processes or directly through differentiation into osteoblast-progenitor cells. However, the osteogenic potential of BMSCs varies among different tissue sources (e.g., mandibular versus long BMSCs). The main aim of this study was to investigate the difference in osteogenic differentiation capacity between mandibular BMSCs (mBMSCs) and tibial BMSCs (tBMSCs). Materials and Methods: Bioinformatics analysis of the GSE81430 dataset taken from the Gene Expression Omnibus (GEO) database was performed using GEO2R. BMSCs were isolated from mandibular and tibial bone marrow tissue samples. Healthy pigs (n = 3) (registered at the State Office for Nature, Environment, and Consumer Protection, North Rhine-Westphalia (LANUV) 81-02.04.2020.A215) were used for this purpose. Cell morphology and osteogenic differentiation were evaluated in mBMSCs and tBMSCs. The expression levels of toll-like receptor 4 (TLR4) and nuclear transcription factor κB (NF-κB) were analyzed using quantitative polymerase chain reaction (qPCR) and Western blot (WB), respectively. In addition, mBMSC-derived extracellular vesicles (mBMSC-EVs) were gained and used as osteogenic stimuli for tBMSCs. Cell morphology and osteogenic differentiation capacity were assessed after mBMSC-EV stimulation. Results: Bioinformatic analysis indicated that the difference in the activation of the TLR4/NF-κB pathway was more pronounced compared to all other examined genes. Specifically, this demonstrated significant downregulation, whereas only 5–7 upregulated genes displayed significant variances. The mBMSC group showed stronger osteogenic differentiation capacity compared to the tBMSC group, confirmed via ALP, ARS, and von Kossa staining. Furthermore, qPCR and WB analysis revealed a significant decrease in the expression of the TLR4/NF-κB pathway in the mBMSC group compared to the tBMSC group (TLR4 fold changes: mBMSCs vs. tBMSCs p < 0.05; NF-κB fold changes: mBMSCs vs. tBMSCs p < 0.05). The osteogenic differentiation capacity was enhanced, and qPCR and WB analysis revealed a significant decrease in the expression of TLR4 and NF-κB in the tBMSC group with mBMSC-EVs added compared to tBMSCs alone (TLR4 fold changes: p < 0.05; NF-κB fold changes: p < 0.05). Conclusion: Our results indicate that mBMSC-EVs can promote the osteogenic differentiation of tBMSCs in vitro. The results also provide insights into the osteogenic mechanism of mBMSCs via TLR4/NF-κB signaling pathway activation. This discovery promises a fresh perspective on the treatment of bone fractures or malunions, potentially offering a novel therapeutic method. [ABSTRACT FROM AUTHOR]

Endometritis is an inflammation of the surface of the endometrium that does not penetrate the submucosa and can cause infertility and increase the elimination rate in cows. Endometrial epithelial cells are the first barrier of the endometrium against foreign stimuli and bacterial infection. Understanding the genetic changes in stimulated endometrial epithelial cells will help in the efforts to prevent and treat endometritis. This study investigated changes in bovine endometrial epithelial (BEEC) gene expression induced by lipopolysaccharide (LPS)-induced inflammation and compared transcriptome-wide gene changes between LPS- and phosphate-buffered saline (PBS)- treated BEECs by RNA sequencing. Compared with the PBS group, the LPS group showed 60 differentially expressed genes (DEGs) (36 upregulated, 24 downregulated). Gene Ontology enrichment analysis revealed that most enrichment occurred during CXCR chemokine receptor binding, inflammatory response, and neutrophil migration. Kyoto Encyclopedia of Genes and Genomes pathway analysis showed DEGs mainly concentrated in cytokine–cytokine receptor interactions; IL-17, tumor necrosis factor, NOD-like receptor, chemokine, Toll-like receptor, and nuclear factor-κB signaling pathways; and the cytoplasmic DNA sensing pathway. Moreover, results revealed that cytokines SAA3 and HP increased significantly after LPS treatment. These effects of LPS on BEECs transcriptome and the molecular mechanism of endometritis provide a basis for improved clinical treatment and novel drug development. [ABSTRACT FROM AUTHOR]

Mastitis is usually caused by a variety of pathogenic bacteria that seriously impact the health and milk-production ability of dairy cows, with consequent, economically detrimental effects on the dairy industry. Forsythoside A (FTA), isolated from the fruit and leaves of Forsythia suspensa (Thunb.) Vahl (Oleaceae), has been reported to have significant antioxidant, anti-inflammatory, and antibacterial effects. However, it is not clear whether FTA exerts a protective effect against lipopolysaccharide (LPS)-induced bovine mastitis and its potential gene signature. In this study, high-throughput sequencing technology was performed to analyze the differences between the mRNA and enrichment pathway of bovine mammary epithelial cells of the control, LPS, and LPS + FTA groups. The results showed that there were 139 differentially expressed genes (DEGs) (p-value < 0.05, |log2FoldChange| > 1, FPKM > 1) in the LPS group compared with the control group, including 121 up-regulated genes and 18 down-regulated genes, which were mainly enriched in the cellular response to lipopolysaccharide, cytokine activity, protein binding, and IL-17 signaling pathway based on Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, respectively. Compared with the control group and LPS + FTA group, there were 349 DEGs, including 322 up-regulated genes and 27 down-regulated genes. They were mainly enriched in protein localization to organelles, centrosomes, binding, and the IL-17 signaling pathway, based on GO and KEGG analysis. Compared to the LPS group, the LPS + FTA group had 272 DEGs, including 259 up-regulated genes and 13 down-regulated genes, which were mainly enriched in RNA processing, IL-6 receptor binding, and the lysosome pathway, based on GO and KEGG analyses. It can be seen that LPS stimulation induced the expression of inflammation-related genes, IL-17 and IL-6, whereas FTA treatment promoted the expression of the spliceosome-, lysosome-, and oxidative stress-related genes HSP70, HSPA8, and PARP2. The study utilized RNA-sequencing analysis of FTA against LPS-challenged bovine mammary epithelial cells to explore key mRNA findings that may be strongly associated with inflammation and oxidative stress, and provides a theoretical reference for further elucidation of molecular mechanisms of bovine mastitis and therapeutic effects of FTA against bovine mastitis. [ABSTRACT FROM AUTHOR]

Purpose: The potential role of epithelium-specific genes through the adenoma-carcinoma sequence in the development of colorectal cancer (CRC) remains unknown. Therefore, we integrated single-cell RNA sequencing and bulk RNA sequencing data to select diagnosis and prognosis biomarkers for CRC. Methods: The CRC scRNA-seq dataset was used to describe the cellular landscape of normal intestinal mucosa, adenoma and CRC and to further select epithelium-specific clusters. Differentially expressed genes (DEGs) of epithelium-specific clusters were identified between intestinal lesion and normal mucosa in the scRNA-seq data throughout the adenoma-carcinoma sequence. Diagnostic biomarkers and prognostic biomarker (the risk score) for CRC were selected in the bulk RNA-seq dataset based on DEGs shared by the adenoma epithelium-specific cluster and the CRC epithelium-specific cluster (shared-DEGs). Results: Among the 1063 shared-DEGs, we selected 38 gene expression biomarkers and 3 methylation biomarkers that had promising diagnostic power in plasma. Multivariate Cox regression identified 174 shared-DEGs as prognostic genes for CRC. We combined 1000 times LASSO-Cox regression and two-way stepwise regression to select 10 prognostic shared-DEGs to construct the risk score in the CRC meta-dataset. In the external validation dataset, the 1- and 5-year AUCs of the risk score were higher than those of stage, the pyroptosis-related genes (PRG) score and the cuproptosis-related genes (CRG) score. In addition, the risk score was closely associated with the immune infiltration of CRC. Conclusion: The combined analysis of the scRNA-seq dataset and the bulk RNA-seq dataset in this study provides reliable biomarkers for the diagnosis and prognosis of CRC. [ABSTRACT FROM AUTHOR]

Purpose: The abnormal regulation of lncRNA CARMN has been proved to be a tumor suppressor gene of cervical cancer (CC). However, its role in CC is still elusive. The regulation of CARMN post-transcriptional level by m6A modification and miRNA has not been studied. This study aims to analyze the molecular mechanism of m6A modification and miRNA on the abnormal expression of CARMN in CC cells, so as to provide a new theoretical basis for the diagnosis and treatment of CC. Methods: MeRIP-seq was used to identify the differential m6A-modified genes between tumor and normal cervical tissues. RT-qPCR assay was used to detect gene expression levels in tissues or cells. The m6A modification sites of CARMN was predicted by bioinformatics, and the modification of m6A and its regulatory effect on CARMN were analyzed by MeRIP-qPCR, Actinomycin D assay and RIP assay. RIP-microarray combined with bioinformatics methods to screen miRNAs that may target CARMN. The regulation mechanism between miRNA and CARMN was verified by RT-qPCR, nucleo-plasmic separation assay, mRNA stability assay, dual-luciferase reporter assay, and in vivo experiments. Results: MeRIP-seq found that CARMN is a significant different gene in the abundance of m6A in CC, and the modification level of m6A in CC tissues was higher than that in normal cervical tissues. Further, this study verified that m6A reader YTHDF2 could recognize m6A-modified CARMN and promote its degradation in CC cells. miR-21-5p was proved to be the downstream target gene of CARMN, and miR-21-5p could negatively regulate the expression of CARMN. Further experiments showed that miR-21-5p could directly bind to CARMN and lead to the degradation of CARMN. The in vivo experimental results indicated that the level of miR-21-5p in the overexpressed CARMN group was significantly lower than that in the control group. Conclusion: m6A modification and miR-21-5p play important roles in promoting the occurrence and development of tumors by regulating CARMN, provide new potential targets for the treatment of CC. [ABSTRACT FROM AUTHOR]

The wounding‐responsive KED gene, named for its coding for a lysine (K), glutamic acid (E), and aspartic acid (D)‐rich protein, is widely present among land plants. However, little is known about its regulation or function. In this study, we found that transcription of the tomato (Solanum lycopersicum) KED gene, SlKED, was rapidly and transiently elevated by wounding or ethephon treatment. Compared to the wild‐type plants, the CRISPR/Cas9‐mediated SlKED knockout plants did not exhibit altered expression patterns for genes involved in hormone biosynthesis or stress signaling, suggesting a lack of pleiotropic effect on other stress‐responsive genes. Conversely, jasmonic acid did not appear to directly regulate SlKED expression. Wounded leaves of the KED‐lacking plants exhibited higher binding of Evans blue dye than the wild‐type, indicating a possible role for KED in healing damaged tissues. The SlKED knockout plants showed a similar dietary effect as the wild‐type on the larval growth of tobacco hornworm. But a higher frequency of larval mandible (mouth) movement was recorded during the first 2 minutes of feeding on the wounded KED‐lacking SlKED knockout plants than on the wounded KED‐producing wild‐type plants, probably reflecting an initial differential response by the feeding larvae to the SlKED knockout plants. Our findings suggest that SlKED may be an ethylene‐mediated early responder to mechanical stress in tomato, acting downstream of the wound stress response pathways. Although its possible involvement in response to other biotic and abiotic stresses is still unclear, we propose that SlKED may play a role in plant's rapid, short‐term, early wounding responses, such as in cellular damage healing. [ABSTRACT FROM AUTHOR]

Intrahepatic cholangiocarcinoma (ICC) is the second common primary hepatic malignancy tumor. In this study, an integrative analysis of differentially expressed genes (DEGs) and miRNAs from the ICC onset and adjacent normal tissues were performed to explore the regulatory roles of miRNA–mRNA interaction. A total of 1018 DEGs and 39 miRNAs were likely involved in ICC pathogenesis, suggesting the changes in cell metabolism in ICC development. The built network indicated that 30 DEGs were regulated by 16 differentially expressed miRNA. The screened DEGs and miRNA together were probably considered the biomarkers of ICC, and their important roles in ICC pathogenesis remain to be elucidated. This study could provide a good basis to uncover the regulatory mechanism of miRNA and mRNAs in ICC pathogenesis. [ABSTRACT FROM AUTHOR]

Background. Kawasaki disease (KD) is characterized by a disordered inflammation response of unknown etiology. Immune cells are closely associated with its onset, although the immune-related genes' expression and possibly involved immune regulatory mechanisms are little known. This study aims to identify KD-implicated significant immune- and inflammation-related biomarkers and pathways and their association with immune cell infiltration. Patients and Methods. Gene microarray data were collected from the Gene Expression Omnibus database. Differential expression analysis, weighted gene coexpression network analysis (WGCNA), least absolute shrinkage and selection operator (LASSO) regression, Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and gene set enrichment analysis (GSEA) were used to find KD hub markers. GSEA was used to assess the infiltration by 28 immune cell types and their connections to essential gene markers. Receiver operating characteristic (ROC) curves were used to examine hub markers' diagnostic effectiveness. Finally, hub genes' expressions were validated in Chinese KD patients by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Results. One hundred and fifty-one unique genes were found. Among 10 coexpression modules at WGCNA, one hub module exhibited the strongest association with KD. Thirty-six overlapping genes were identified. Six hub genes were potential biomarkers according to LASSO analysis. Immune infiltration revealed connections among activated and effector memory CD4+ T cells, neutrophils, activated dendritic cells, and macrophages. The six hub genes' diagnostic value was shown by ROC curve analysis. Hub genes were enriched in immunological and inflammatory pathways. RT-qPCR verification results of FCGR1B (P < 0.001), GPR84 (P < 0.001), KREMEN1 (P < 0.001), LRG1 (P < 0.001), and TDRD9 (P < 0.001) upregulated expression in Chinese KD patients are consistent with our database analysis. Conclusion. Neutrophils, macrophages, and activated dendritic cells are strongly linked to KD pathophysiology. Through immune-related signaling pathways, hub genes such as FCGR1B, GPR84, KREMEN1, LRG1, and TDRD9 may be implicated in KD advancement. [ABSTRACT FROM AUTHOR]

Low temperature is a limiting factor of seed germination and plant growth. Although there is a lot information on the response of maize to low temperatures, there is still poorly description of how histone methylation affects maize germination and growth development at low temperatures. In this study, the germination rate and physiological indexes of wild-type maize inbred lines B73 (WT), SDG102 silencing lines (AS), SDG102 overexpressed lines (OE) at germination stage and seedling stage were measured under low temperature stress (4 ℃), and transcriptome sequencing was applied to analyze the differences of gene expression in panicle leaves among different materials. The results showed that the germination rate of WT and OE maize seeds at 4 ℃ was significantly lower than 25 ℃. The content of MDA, SOD and POD of 4 ℃ seeding leaves higher than contrast. Transcriptome sequencing results showed that there were 409 different expression genes (DEGs) between WT and AS, and the DEGs were mainly up-regulated expression in starch and sucrose metabolism and phenylpropanoid biosynthesis. There were 887 DEGs between WT and OE, which were mainly up-regulated in the pathways of plant hormone signal transduction, porphyrin and chlorophyll metabolism. This result could provide a theoretical basis for analyzing the growth and development of maize from the perspective of histone methylation modification. [ABSTRACT FROM AUTHOR]

The enzyme 1-acylglycerol-3-phosphate O-acyltransferase 2 (AGPAT2) is an intermediate enzyme in triglyceride synthesis. The aim was to study the regulatory mechanism of AGPAT2 on Nile tilapia, Oreochromis niloticus. In this study, antisense RNA technology was used to knock-down AGPAT2 in Nile tilapia. Compared with the control groups (transfected with ultrapure water or the blank expression vector), the AGPAT2 knock-down group showed a significantly higher weight gain rate, special growth rate, visceral somatic index, and hepatopancreas somatic index; and significantly increased the total cholesterol, triglycerides, glucose, low-density lipoprotein cholesterol, and insulin levels in serum. In addition, the contents of total cholesterol and triglycerides and the abundance of superoxide dismutase, catalase, and glutathione peroxidase in the liver significantly increased, while the malondialdehyde content significantly decreased. The liver cells became severely vacuolated and accumulated lipids in the AGPAT2 knock-down group. Comparative transcriptome analyses (AGPAT2 knock-down vs. control group) revealed 1789 differentially expressed genes (DEGs), including 472 upregulated genes and 1313 downregulated genes in the AGPAT2 knock-down group. Functional analysis showed that the main pathway of differentially expressed genes enrichment was lipid metabolism and oxidative stress, such as steroid biosynthesis, unsaturated fatty acid biosynthesis, the PPAR signaling pathway, and the P53 pathway. We used qRT-PCR to verify the mRNA expression changes of 13 downstream differential genes in related signaling pathways. These findings demonstrate that knock-down of AGPAT2 in tilapia leads to abnormal lipid metabolism and oxidative stress. [ABSTRACT FROM AUTHOR]

Background: A rigorous search for alternatives to autogenous bone grafts to avoid invasiveness at the donor site in the treatment of maxillomandibular bone defects. Researchers have used alloplastic, allogeneic, and xenogeneic bone graft substitutes in clinical studies with varying degrees of success, although their in vitro effects on stem cells remain unclear. Dental pulp stem cells (DPSCs) can potentially enhance the bone regeneration of bone graft substitutes. The present in vitro study investigates the osteogenic capability of DPSCs on alloplastic (biphasic calcium phosphate [BCP]), allogeneic (freeze-dried bone allografts [FDBAs]), and xenogeneic (deproteinized bovine bone mineral [DBBM]) bone grafts. Methods: Human DPSCs were seeded on 0.5 mg/ml, 1 mg/ml, and 2 mg/ml of BCP, FDBA, and DBBM to evaluate the optimal cell growth and cytotoxicity. Scaffolds and cell morphologies were analyzed by scanning electron microscopy (SEM). Calcein AM and cytoskeleton staining were performed to determine cell attachment and proliferation. Alkaline phosphatase (ALP) and osteogenesis-related genes expressions was used to investigate initial osteogenic differentiation. Results: Cytotoxicity assays showed that most viable DPSCs were present at a scaffold concentration of 0.5 mg/ml. The DPSCs on the DBBM scaffold demonstrated a significantly higher proliferation rate of 214.25 ± 16.17 (p < 0.001) cells, enhancing ALP activity level and upregulating of osteogenesis-related genes compared with other two scaffolds. Conclusion: DBBP scaffold led to extremely high cell viability, but also promoted proliferation, attachment, and enhanced the osteogenic differentiation capacity of DPSCs, which hold great potential for bone regeneration treatment; however, further studies are necessary. [ABSTRACT FROM AUTHOR]

[Display omitted] • In erythroid cells, ZNF410 regulates a single target gene, CHD4 , encoding a NuRD complex component. • Selective glue degraders have been developed for members of IKAROS family of ZF proteins. • A highly selective and non-genotoxic histone H3 lysine 9 methyltransferase G9a inhibitor induces fetal globin expression in both human erythroid cells and mice. • Non-nucleoside, dicyanopyridine-containing DNMT1-selective inhibitors/degraders have shown promising therapeutic potential in a transgenic mouse model of sickle cell disease. In humans, specific aberrations in β-globin results in sickle cell disease and β-thalassemia, symptoms of which can be ameliorated by increased expression of fetal globin (HbF). Two recent CRISPR-Cas9 screens, centered on ∼1500 annotated sequence-specific DNA binding proteins and performed in a human erythroid cell line that expresses adult hemoglobin, uncovered four groups of candidate regulators of HbF gene expression. They are (1) members of the nucleosome remodeling and deacetylase (NuRD) complex proteins that are already known for HbF control; (2) seven C2H2 zinc finger (ZF) proteins, including some (ZBTB7A and BCL11A) already known for directly silencing the fetal γ-globin genes in adult human erythroid cells; (3) a few other transcription factors of different structural classes that might indirectly influence HbF gene expression; and (4) DNA methyltransferase 1 (DNMT1) that maintains the DNA methylation marks that attract the MBD2-associated NuRD complex to DNA as well as associated histone H3 lysine 9 methylation. Here we briefly discuss the effects of these regulators, particularly C2H2 ZFs, in inducing HbF expression for treating β-hemoglobin disorders, together with recent advances in developing safe and effective small-molecule therapeutics for the regulation of this well-conserved hemoglobin switch. [ABSTRACT FROM AUTHOR]

Objective: To study the sedative and hypnotic effects and underlying mechanisms of Polygala tenuifolia (PT) on treating aged insomnia rats. Methods: Sixty Sprague-Dawley male rats were divided into 6 groups by a random number table, including control group, model group, diazepam group (0.92 mg/kg), as well as PT low-, medium- and high-dose groups (0.0875, 0.175, 0.35 g/kg, respectively), 10 rats in each group. Aged insomnia rat model was established with subcutaneous injection of D-galactose for 42 days and then intraperitoneal injection of para-chlorophenylalanine for 3 days. PT and diazepam were respectively given to aged insomnia rats by intragastric administration for 7 days after model establishment. Then the rats were investigated by body weight, Morris water maze test, pentobarbital test, enzyme-linked immunosorbent assay, and transcriptome sequencing. Results: Compared with the model group, PT increased the body weight, improved memory ability, and prolonged pentobarbital-induced sleep time of aged insomnia rats (P<0.01 or P<0.05). The medium dose of PT also increased the neurotransmitter levels of 5-hydroxytryptamine (5-HT) and gamma-aminobutyric acid (GABA), and decreased the level of Glu in the hippocampus of aged insomnia rats (P<0.05 or P<0.01). Twenty-four differentially expressed genes (DEGs) were overlapped among model group, medium-dose PT group, and diazepam group in transcriptome analysis. Fuom and Pcp2 were down-regulated by the treatment of medium-dose PT (P<0.01 or P<0.05). The metabolic pathways of PT were relatively less than diazepam (91 vs. 104). Conclusions: The sedative and hypnotic effects of PT in aged insomnia rats might be related to neuro, metabolism pathways, especially through GABAergic signaling pathway. It provided more effective herb choice for the treatment of senile insomnia. [ABSTRACT FROM AUTHOR]

Background: SINEs are a type of nonautonomous retrotransposon that can transpose from one site to be integrated elsewhere in an organism genome. SINE insertion can give rise to genetic variants and regulate gene expression, allowing organisms to acquire new adaptive capacity. Studies on this subject have focused on the impacts of SINEs on genes. However, ecological disparities in fish have not yet been explained by SINEs. Results: New SINEs were isolated from Coilia nasus, which has two ecotypes—migratory and resident—that differ in their spawning and migration behaviors. The SINEs possess two structures that resemble a tRNA gene and a LINE retrotransposon tail. Comparison of olfactory tissue transcriptomes, intact SINE transcript copies were detected in only the migratory fish at the initial retrotransposition stage. The SINE DNA copy numbers were higher in the resident type than in the migratory type, while the frequency of SINE insertion was higher in the migratory type than in the resident type. Furthermore, SINE insertions can lead to new repeats of short DNA fragments in the genome, along with target site duplications. SINEs in the resident type have undergone excision via a mechanism in which predicted cleavage sites are formed by mutations, resulting in gaps that are then filled by microsatellites via microhomology-induced replication. Conclusions: Notably, SINEs in the resident type have undergone strong natural selection, causing genomic heteroplasmy and driving ecological diversity of C. nasus. Our results reveal possible evolutionary mechanisms underlying the ecological diversity at the interface between SINE mobilization and organism defense. [ABSTRACT FROM AUTHOR]

Abstract Ethnopharmacological relevance Jianpi Yangzheng Xiaozheng Decoction (JPYZXZ) is an empirical compound prescription based on the theory of traditional Chinese medicine. JPYZXZ, which is "Qi-invigorating, spleen-strengthening and stasis-removing," can improve the quality of life of gastric cancer patients and prolong their survival; however, the exact mechanism underlying the antitumor effects of this compound is still not clear. Aim of the study The aim of this study is to clearly define the effect of JPYZXZ and its components, Jianpi Yangzheng Decoction (JPYZ) and Xiao Zheng San Jie Decoction (XZSJ), on inhibiting the progression of gastric cancer. Materials and methods The effect of JPYZXZ and its components on the motility of gastric cancer MGC-803 cells was measured by MTT, adhesion, transwell assays and wound-healing assays. JPYZXZ, JPYZ and XZSJ were administered to 615 mice with gastric cancer xenografts, and their effect on the inhibition of subcutaneous transplantation was analyzed. THP-1 monocyte cells were used to establish tumor-associated macrophage (TAM) models. The polarized state of the TAMs was detected by Flow Cytometry, ELISA and Immunohistochemistry. The mRNA and protein expression of tumor epithelial-mesenchymal transition (EMT) and TAM-related genes was determined by Real-time PCR and Western Blot, respectively. Results We determined that both JPYZXZ and its components inhibited the progress of gastric cancer in vitro , and JPYZXZ was clearly more effective than JPYZ or XZSJ. The in vivo results demonstrated that the JPYZXZ and XZSJ group exhibited a significant decrease in the tumor weight compared to the control group. Further analysis indicated that JPYZXZ was more active than JPYZ or XZSJ in inhibiting the gastric cancer EMT transformation both in vivo and in vitro. However, JPYZ was more effective compared with JPYZXZ for inducing the phenotypic change in macrophages from M2 to M1. Conclusions Our results demonstrate that both JPYZXZ and its components prevent the progress of gastric cancer. JPYZXZ inhibits the gastric cancer EMT more effectively than JPYZ and XZSJ, but JPYZ primarily works to regulate the phenotypic change in macrophages from M2 to M1. Graphical abstract fx1 [ABSTRACT FROM AUTHOR]

The ecological importance of the duplication and diversification of gene clusters that synthesize secondary metabolites in fungi remains poorly understood. Here, we demonstrated that the duplication and subsequent diversification of a gene cluster produced two polyketide synthase gene clusters in the cosmopolitan fungal genus Metarhizium. Diversification occurred in the promoter regions and the exon-intron structures of the two Pks paralogs (Pks1 and Pks2). These two Pks genes have distinct expression patterns, with Pks1 highly expressed during conidiation and Pks2 highly expressed during infection. Different upstream signaling pathways were found to regulate the two Pks genes. Pks1 is positively regulated by Hog1-MAPK, Slt2-MAPK and Mr-OPY2, while Pks2 is positively regulated by Fus3-MAPK and negatively regulated by Mr-OPY2. Pks1 and Pks2 have been subjected to positive selection and synthesize different secondary metabolites. PKS1 is involved in synthesis of an anthraquinone derivative, and contributes to conidial pigmentation, which plays an important role in fungal tolerance to UV radiation and extreme temperatures. Disruption of the Pks2 gene delayed formation of infectious structures and increased the time taken to kill insects, indicating that Pks2 contributes to pathogenesis. Thus, the duplication of a Pks gene cluster and its subsequent functional diversification has increased the adaptive flexibility of Metarhizium species. [ABSTRACT FROM AUTHOR]

The rigidity of bacterial cell walls synthesized by a complicated pathway limit the cell shapes as coccus, bar or ellipse or even fibers. A less rigid bacterium could be beneficial for intracellular accumulation of poly-3-hydroxybutyrate (PHB) as granular inclusion bodies. To understand how cell rigidity affects PHB accumulation, E. coli cell wall synthesis pathway was reinforced and weakened, respectively. Cell rigidity was achieved by thickening the cell walls via insertion of a constitutive gltA (encoding citrate synthase) promoter in front of a series of cell wall synthesis genes on the chromosome of several E. coli derivatives, resulting in 1.32–1.60 folds increase of Young's modulus in mechanical strength for longer E. coli cells over-expressing fission ring FtsZ protein inhibiting gene sulA . Cell rigidity was weakened by down regulating expressions of ten genes in the cell wall synthesis pathway using CRISPRi, leading to elastic cells with more spaces for PHB accumulation. The regulation on cell wall synthesis changes the cell rigidity: E. coli with thickened cell walls accumulated only 25% PHB while cell wall weakened E. coli produced 93% PHB. Manipulation on cell wall synthesis mechanism adds another possibility to morphology engineering of microorganisms. [ABSTRACT FROM AUTHOR]

Metarhizium robertsii occupies a wide array of ecological niches and has diverse lifestyle options (saprophyte, insect pathogen and plant symbiont), that renders it an unusually effective model for studying genetic mechanisms for fungal adaptation. Here over 20,000 M. robertsii T-DNA mutants were screened in order to elucidate genetic mechanism by which M. robertsii replicates and persists in diverse niches. About 287 conidiation, colony sectorization or pathogenicity loci, many of which have not been reported in other fungi were identified. By analysing a series of conidial pigmentation mutants, a new fungal pigmentation gene cluster, which contains Mr-Pks1, Mr-EthD and Mlac1 was identified. A conserved conidiation regulatory pathway containing Mr-BrlA, Mr-AbaA and Mr-WetA regulates expression of these pigmentation genes. During conidiation Mr-BlrA up-regulates Mr-AbaA, which in turn controls Mr-WetA. It was found that Hog1-MAPK regulates fungal conidiation by controlling the conidiation regulatory pathway, and that all three pigmentation genes exercise feedback regulation of conidiation. This work provided the foundation for deeper understanding of the genetic processes behind M. robertsii adaptive phenotypes, and advances our insights into conidiation and pigmentation in this fungus. [ABSTRACT FROM AUTHOR]

Tripterygium wilfordiiHook. f. is the traditional medicinal plants in China. Triptolide, wilforgine, and wilforine are the bioactive compounds inT. wilfordii.In this study, the contents of three metabolites and transcription levels of 21 genes involved in three metabolites biosynthesis inT. wilfordiiwere examined using high-performance liquid chromatography and reverse transcription PCR after application of methyl jasmonate (MeJA) on hairy roots in time course experiment (3–24 h). The results indicated that application of MeJA inhibited triptolide accumulation and promoted wilforgine and wilforine metabolites biosynthesis. In hairy roots, wilforgine content reached 693.36 μg/g at 6 h after adding MeJA, which was 2.23-fold higher than control. The accumulation of triptolide and wilforine in hairy roots increased the maximum at 9 h, which was 1.3- and 1.6-folds more than the control. Most of the triptolide secretes into the medium, but wilforgine and wilforine cannot secrete into the medium. The expression levels of unigenes which involved terpenoid backbone biosynthesis exist the correlation with marker metabolites (triptolide, wilforgine and wilforine) after induction by MeJA, and can be then used to infer flux bottlenecks inT. wilfordiisecondary metabolites accumulation. These results showed that these genes may have potential applications in the metabolic engineering ofT. wilfordiimetabolites production. [ABSTRACT FROM PUBLISHER]

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This study aimed to investigate the expression and clinical significance of enoyl coenzyme A hydratase, short chain, 1 (ECHS1), in patients with colorectal cancer (CRC). The ECHS1 protein expression as detected by immunohisto-chemistry in 148 CRC specimens was evaluated and compared by clinical pathology and prognosis; 38 specimens from proximal non-cancerous colorectal tissues were included as controls. The ECHS1 protein expression was also measured by western blot analysis in 46 fresh CRC tissue specimens and 22 normal colorectal tissue specimens. The rate of positive ECHS1 expression differed significantly between the CRC tissues (56.76%, 84/148) and the proximal non-cancerous colorectal tissues (5.26%, 2/38) (P<0.001). The ECHS1 protein expression was confirmed not to be associated with gender or age. However, the positive expression of ECHS1 tended to be positively associated with clinical TNM stage (P=0.015), lymph node metastasis (P=0.011) and histological differentiation (P=0.028). The expression of the ECHS1 protein on western blot analysis was significantly increased in CRC vs. normal tissues. In addition, the overall survival curves estimated with the Kaplan-Meier method demonstrated that CRC patients exhibiting low ECHS1 expression survived significantly longer compared to patients with high ECHS1 levels (P=0.039). Our data suggested that ECHS1 protein expression may contribute to the occurrence, progression and metastasis of CRC, is closely associated with prognosis and may provide useful information for CRC molecular-targeted therapy. [ABSTRACT FROM AUTHOR]

AZD6244, an ATP-uncompetitive inhibitor of mitogen-activated protein kinase 1/2 (MEK1/2), has shown activity in several malignant tumours. However, whether AZD6244 has a function in lymphoma cells is not known. We report that AZD6244 treatment represses the growth of Raji and MOLT4 cells by inducing apoptosis and G1-phase arrest. Using miRNAs array and quantitative RT-PCR, miR-92a was downregulated byAZD6244 treatment through the ERK1/2-AP1 signalling pathway. Overexpression of miR-92a abrogated AZD6244-induced apoptosis and G1-phase arrest, indicating that it is involved in the cytotoxicity of AZD6244 in lymphoma cells. A luciferase reporter assay showed that miR-92a directly targetsthe 3′-UTRs of Bim. Overexpression of miR-92a mimics downregulated Bim mRNA and protein expression level, indicating that miR-92a negatively regulates its expression at both levels. Silencing Bim decreases AZD6244-induced apoptosis and G1-phase arrest, suggesting that Bim contributes to the growth arrest. Thus, miR-92a mediates AZD6244-induced cytotoxicity of lymphoma cells by targeting Bim. Downregulation of miR-92a by AZD6244 is mediated by the ERK1/2-AP1 signalling pathway. [ABSTRACT FROM AUTHOR]

The apical hook is an essential structure that enables epigeal plants to protrude through the soil. Arabidopsis thaliana HOOKLESS1 (HLS1) is reported to be a key regulator of hook development and a direct target gene of the ethylene (ET)-activated transcription factors ETHYLENE INSENSITIVE3 (EIN3) and its close homolog EIN3-Like1. Previous research has shown that the phytohormones jasmonate (JA) and ET antagonistically regulate apical hook development, although the underlying molecular mechanism is largely unknown. Here, we report that JA represses hook formation by reducing HLS1 expression. Our results further reveal that the JA -activated transcription factor MYC2 represses EIN3 function to reduce HLS1 expression through at least the following two layers of regulation: (1) MYC2 binds to the promoter of an F-box gene, EIN3 BINDING F-BOX PROTEIN1 , to induce its expression and thus promote EIN3 degradation; and (2) MYC2 physically interacts with EIN3 and inhibits its DNA binding activity. Collectively, our findings shed light on the molecular mechanism underlying the antagonism between JA and ET during apical hook development and provide insight into the coaction of multiple phytohormones in the regulation of plant growth and development. [ABSTRACT FROM AUTHOR]

Aim:Aptamers are oligonucleic acid or peptide molecules that bind to a specific target molecule in cells, thus may act as effective vehicles for drug or siRNA delivery. In this study we investigated the DNA aptamers that target human glioblastoma multiforme (GBM) cells overexpressing epidermal growth factor receptor variant III (EGFRvIII), which was linked to radiation and chemotherapeutic resistance of this most aggressive brain tumor.Methods:A 73-mer ssDNA library containing molecules with 30 nt of random sequence flanked by two primer hybridization sites was chosen as the initial library. Cell systematic evolution of ligands by exponential enrichment (Cell-SELEX) method was used to select the DNA aptamers that target EGFRvIII. The binding affinity of the aptamers was measured using a cell-based biotin-avidin ELISA.Results:After 14 rounds of selection, four DNA aptamers (32, 41, 43, and 47) that specifically bound to the EGFRvIII-overexpressing human glioma U87Δ cells with Kd values of less than 100 nmol/L were discovered. These aptamers were able to distinguish the U87Δ cells from the negative control human glioma U87MG cells and HEK293 cells. Aptamer 32 specifically bound to the EGFRvIII protein with an affinity similar to the EGFR antibody (Kd values of aptamer 32 and the EGFR antibody were 0.62±0.04 and 0.32±0.01 nmol/L, respectively), and this aptamer was localized in the cell nucleus.Conclusion:The DNA aptamers are promising molecular probes for the diagnosis and treatment of GBM. [ABSTRACT FROM AUTHOR]

Hexavalent chromium [Cr(IV)], a well-known industrial waste product and an environmental pollutant, is recognized as a human carcinogen. But its mechanisms of carcinogenicity remain unclear, and recent studies suggest that DNA methylation may play an important role in the carcinogenesis of Cr(IV). The aim of our study was to investigate the effects of Cr(IV) on cell cycle progress, global DNA methylation, and DNA methylation of p16 gene. A human B lymphoblastoid cell line and a human lung cell line A549 were exposed to 5–15 µM potassium dichromate or 1.25–5 µg/cm2 lead chromate for 2–24 hours. Cell cycle was arrested at G1 phase by both compounds in 24 hours exposure group, but global hypomethylation occurred earlier than cell cycle arrest, and the hypomethylation status maintained for more than 20 hours. The mRNA expression of p16 was significantly up-regulated by Cr(IV), especially by potassium dichromate, and the mRNA expression of cyclin-dependent kinases (CDK4 and CDK6) was significantly down-regulated. But protein expression analysis showed very little change of p16 gene. Both qualitative and quantitative results showed that DNA methylation status of p16 remained unchanged. Collectively, our data suggested that global hypomethylation was possibly responsible for Cr(IV) - induced G1 phase arrest,but DNA methylation might not be related to up-regulation of p16 gene by Cr(IV). [ABSTRACT FROM AUTHOR]

Reactive oxygen species (ROS) from nicotinamide adenine dinucleotide phosphate (NADPH) oxidases and their related dual oxidases are known to have significant roles in innate immunity and cell proliferation. In this study, the 5,545 bp cDNA of the silkworm Bombyx mori dual oxidase (BmDuox) gene containing a full-length open reading frame was cloned. It was shown to include an N-terminal signal peptide consisting of 28 amino acid residues, a 240 bp 5′-terminal untranslated region (5′-UTR), an 802 bp 3′-terminal region (3′-UTR), which contains nine ATTTA motifs, and a 4,503 bp open reading frame encoding a polypeptide of 1,500 amino acid residues. Structural analysis indicated that BmDuox contains a typical peroxidase domain at the N-terminus followed by a calcium-binding domain, a ferric-reducing domain, six transmembrane regions and binding domains for flavin adenine dinucleotide (FAD) and nicotinamide adenine dinucleotide (NAD). Transcriptional analysis revealed that BmDuox mRNA was expressed more highly in the head, testis and trachea compared to the midgut, hemocyte, Malpighian tube, ovary, fat bodies and silk glands. BmDuox mRNA was expressed during all the developmental stages of the silkworm. Subcellular localization revealed that BmDoux was present mainly in the periphery of the cells. Some cytoplasmic staining was detected, with rare signals in the nucleus. Expression of BmDuox was induced significantly in the larval midgut upon challenge by Escherichia coli and Bombyx mori nucleopolyhedrovirus (BmNPV). BmDuox-deleted larvae showed a marked increase in microbial proliferation in the midgut after ingestion of fluorescence-labeled bacteria compared to the control. We conclude that reducing BmDuox expression greatly increased the bacterial load, suggesting BmDuox has an important role in inhibiting microbial proliferation and the maintenance of homeostasis in the silkworm midgut. [ABSTRACT FROM AUTHOR]

Abstract: To maintain the normal length of female reproductive life, the majority of primordial follicles must be maintained in a quiescent state for later use. In this study, we aimed to study the effects of rapamycin on primordial follicle development and investigate the role of mTOR and sirtuin signaling. Rats were treated every other day with an intraperitoneal injection of rapamycin (5mg/kg) or vehicle. After 10weeks of treatment, ovaries were harvested for hematoxylin and eosin (HE) staining, and analysis by immunohistochemistry and Western blotting. HE staining showed that the number and percentage of primordial follicles in the rapamycin-treated group were twice the control group (P<0.001). Immunohistochemical analysis showed that mTOR and phosphorylated-p70S6K were extensively expressed in surviving follicles with strong staining observed in the cytoplasm of the oocyte. Western blotting showed decreased expression of phosphorylated mTOR and phosphorylated p70S6K in the rapamycin-treated group, and increased the expression of both SIRT1 and SIRT6 compared to the control group (P<0.05). Taken together, these results suggest that rapamycin may inhibit the transition from primordial to developing follicles and preserve the follicle pool reserve, thus extending the ovarian lifespan of female rats via the modulation of mTOR and sirtuin signalings. [Copyright &y& Elsevier]

Premise of research. Formerly sparsely distributed southern cattail (Typha domingensis) is now rapidly replacing the historically predominant sawgrass (Cladium jamaicense) in Florida Everglades wetlands. Previous studies have attributed anthropogenic phosphorus (P) enrichment as a causal factor . We investigated various traits of known importance to P acquisition and use in cattail and sawgrass, in order to determine the underlying mechanisms driving the sawgrass-cattail habitat shift. Methodology. An integration of morphological, physiological, biochemical, and molecular approaches were used to examine growth, root structure, photosynthesis, enzyme activities, and gene expression in plants grown under different P conditions. Pivotal results. Cattail and sawgrass exhibited distinct patterns of P responses between roots and shoots. While cattail displayed plasticity in shoots and inflexibility in roots, sawgrass demonstrated the opposite pattern, exhibiting a remarkable capacity for modulating its root system architecture, acid phosphatase activity, and phosphate transporter abundance. Likely advantageous under P impoverishment for sawgrass in the historic Everglades undisturbed by man, these adaptations are nullified under current anthropogenic P en- richment, thus opening the door for competition from cattail. In response to high P availability, cattail exhibited enhanced protein synthesis, photosynthesis, and growth in its plastic shoots, empowering its spatial expansion at the expense of sawgrass. Conclusions. The species-specific patterns of root/shoot-disparate plasticity may be an important mechanism underlying the spatial redistribution of cattail and sawgrass in the anthropogenic resource-altered environment. High plasticity in shoots in response to environmental nutrient enrichment is likely a common trait shared by invasive-prone plants. [ABSTRACT FROM AUTHOR]

We have previously reported that the mitochondria inhibitor 3-nitropropionic acid (3-NP), induces the expression of DNA damage-regulated autophagy modulator1 (DRAM1) and activation of autophagy in rat striatum. Although the role of DRAM1 in autophagy has been previously characterized, the detailed mechanism by which DRAM1 regulates autophagy activity has not been fully understood. The present study investigated the role of DRAM1 in regulating autophagy flux. In A549 cells expressing wilt-type TP53, 3-NP increased the protein levels of DRAM1 and LC3-II, whereas decreased the levels of SQSTM1 (sequestosome 1). The increase in LC3-II and decrease in SQSTM1 were blocked by the autophagy inhibitor 3-methyl-adenine. Lack of TP53 or knock-down of TP53 in cells impaired the induction of DRAM1. Knock-down of DRAM1 with siRNA significantly reduced 3-NP-induced upregulation of LC3-II and downregulation of SQSTM1, indicating DRAM1 contributes to autophagy activation. Knock-down of DRAM1 robustly decreased rate of disappearance of induced autophagosomes, increased RFP-LC3 fluorescence dots and decreased the decline of LC3-II after withdraw of rapamycin, indicating DRAM1 promotes autophagy flux. DRAM1 siRNA inhibited lysosomal V-ATPase and acidification of lysosomes. As a result, DRAM1 siRNA reduced activation of lysosomal cathepsin D. Similar to DRAM1 siRNA, lysosomal inhibitors E64d and chloroquine also inhibited clearance of autophagosomes and activation of lysosomal cathapsin D after 3-NP treatment. These data suggest that DRAM1 plays important roles in autophagy activation induced by mitochondria dysfunction. DRAM1 affects autophagy through argument of lysosomal acidification, fusion of lysosomes with autophagosomes and clearance of autophagosomes. [ABSTRACT FROM AUTHOR]

To effectively treat articular cartilage defect with tissue engineering there is an urgent need to develop safe and cheap drugs that can substitute or cooperate with growth factors for chondrogenesis promotion. Here, we demonstrate the chondrogenic effect of icariin, the major pharmacological active constituent of Herb Epimedium (HEP). Rabbit chondrocytes were isolated from articular cartilage and cultured in vitro with different concentrations of icariin. Icariin at concentrations under 1 × 10−5 m showed low cytotoxicity toward chondrocytes, but icariin at 5 × 10−5 m inhibited the proliferation of chondrocytes. Icariin hardly affected the cell morphology with concentrations ranging from 1 × 10−7 m to 5 × 10−5 m. However, the higher concentration of icariin produced more extracellular matrix (ECM) synthesis and expression of chondrogenesis genes of chondrocytes. Indeed, the promotion of icariin on the synthesis of glycosaminoglycans (GAGs) and collagen of chondrocytes, and finally exerting a potent chondrogenic effect, might be due to its ability to up-regulate the expression of aggrecan, collagen II and Sox9 genes and to down-regulate the expression of the collagen I gene of chondrocytes. These preliminary results imply that icariin might be an effective accelerant for chondrogenesis and that icariin-loaded biomaterials might have the potential for cartilage tissue engineering. 1 × 10−5 m may be a suitable concentration of icariin with chondrogenic effect for tissue engineering. Copyright © 2012 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Abstract: 10–23 DNAzyme (DrzBC) could block the expression of HBV e-gene with application limit of dependence on exogenous delivery. Stearic acid grafted chitosan oligosaccharide (CSSA) could self-aggregate to form micelles in aqueous medium and bind with DrzBC by electrostatic interaction. Compared with Lipofectamine™ 2000, the CSSA micelles showed much lower cytotoxicity (412.5μg/mL) and advantaged target in subcellular organelles-cytoplasm. Both including DrzBC of 1μmol/L, Lipofectamine™ 2000/DrzBC complex showed maximum inhibition rate (IR) on HBeAg expression of 46.53±2.00% at 48h, and then decreased rapidly, while CSSA/DrzBC complex showed maximum IR of 82.51±1.28% at 72h, and hold on IR above 70% until 96h. Moreover, within a concentration range of 0.1–2.0μmol/L, and equally incubating for 48h, the IR of Lipofectamine™ 2000/DrzBC complex was from 23.20±1.61% to 66.27±1.96%, while the IR of CSSA/DrzBC complex increased from 40.23±3.28% to 80.95±1.69%. CSSA/DrzBC complex is a promising effective system for inhibiting HBeAg expression. [Copyright &y& Elsevier]

Abstract: Apolipoprotein E (apoE) has an intricate biological function in modulating immune responses and apoE isoforms exhibit diverse effects on neurodegenerative and neuroinflammatory disorders. In the present study, we investigated the individual roles of apoE isoforms in the kainic acid (KA)-induced hippocampal neurodegeneration with focus on immune response and microglia functions. ApoE2, 3 and 4 transgenic mice as well as wild-type (WT) mice were treated with KA by intranasal route. ApoE4 overexpressing mice revealed several peculiarities as compared with other transgenic mice and WT mice, i.e. (1) they had more severe KA-induced seizures than apoE2 and 3 mice, (2) they exhibited neuron loss in hippocampus that was higher than in apoE2, 3 and WT mice, (3) KA administration resulted in higher counts of their head drops in the cross-area of elevated plus-maze, (4) they showed lower KA-induced rearing activity than apoE2 mice in the open-field test, (5) their KA-induced microglial expression of MHC-II and CD86 was elevated compared to apoE3 mice, (6) the KA-induced increase of microglial iNOS was higher than that in the other groups of mice, and (7) the TNF-α and IL-6 expression was decreased 7days after KA application compared to untreated mice and mice treated 1day with KA. However, the signaling pathway of NFκB or Akt seemed not to be involved in apoE-isoform dependent susceptibility to KA-induced neurotoxicity. In conclusion, over-expression of apoE4 deteriorated KA-induced hippocampal neurodegeneration in C57BL/6 mice, which might result from a higher up-regulation of microglia activation compared to apoE2 and 3 transgenic mice and WT mice. [Copyright &y& Elsevier]

Abstract: Infection with duck circovirus (DuCV) is associated with growth retardation and developmental problems in farmed ducks. To detect DuCV-specific antibody in duck serum, an indirect enzyme-linked immunosorbent assay (iELISA) method was developed using the recombinant capsid protein antigen prepared by cloning the cap (Cap) gene of DuCV FJ0601 strain into pET-32a (+) vector and expressed in Escherichia coli. Using the optimized iELISA method, DuCV-specific antibodies were detected in 157 (12.96%) of 1211 samples obtained from 17 (89.47%) of 19 meat duck flocks aged from 25 to 40 days and in 89 (22.08%) of 403 samples obtained from 9 (75%) of 12 breeder flocks aged from 14 to 61 weeks. These results indicated that the iELISA method is useful for serological diagnosis of DuCV infection and epidemiological investigation. [ABSTRACT FROM AUTHOR]

Histone lysine methylation is an important epigenetic mark that regulates gene expression and chromatin organization. G9a and G9a-like protein (GLP) are euchromatin-associated methyltransferases that repress transcription by methylating histone H3 Lys9. BIX-01294 was originally identified as a G9a inhibitor during a chemical library screen of small molecules and has previously been used in the generation of induced pluripotent stem cells. Here we present the crystal structure of the catalytic SET domain of GLP in complex with BIX-01294 and S-adenosyl-L-homocysteine. The inhibitor is bound in the substrate peptide groove at the location where the histone H3 residues N-terminal to the target lysine lie in the previously solved structure of the complex with histone peptide. The inhibitor resembles the bound conformation of histone H3 Lys4 to Arg8, and is positioned in place by residues specific for G9a and GLP through specific interactions. [ABSTRACT FROM AUTHOR]

Ascomycete fungi usually produce small hydrophobic asexual conidia that are easily dispersed and essential for long-term survival under a variety of environmental conditions. Several upstream signaling regulators have been documented to control conidiation via regulation of the central regulatory pathway that contains the transcription factors BrlA, AbaA and WetA. Here, we showed that the Slt2-MAPK signaling pathway and the transcription factor RNS1 constitute a novel upstream signaling cascade that activates the central regulatory pathway for conidiation in the Ascomycetes fungus M. robertsii. The BrlA gene has two overlapping transcripts BrlAα and BrlAβ; they have the same major ORF, but the 5' UTR of BrlAβ is 835 bp longer than the one of BrlAα. During conidiation, Slt2 phosphorylates the serine residue at the position 306 in RNS1, which self-induces. RNS1 binds to the BM2 motif in the promoter of the BrlA gene and induces the expression of the transcript BlrAα, which in turn activates the expression of the genes AbaA and WetA. In conclusion, the Slt2/RNS1 cascade represents a novel upstream signaling pathway that initiates conidiation via direct activation of the central regulatory pathway. This work provides significant mechanistic insights into the production of asexual conidia in an Ascomycete fungus. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the growth pattern of the large uterine leiomyomata (ULM), we examined the spatial gene distributions, vessel density, proliferative activity, and hyaline degeneration. Design: Tissue sections from three-dimensional large ULM, matched myometrium, and small ULM were collected and microarrayed. The spatial difference of the tumor activity was mapped in large ULM. Setting: University clinical research laboratory. Patient(s): Hysterectomy specimens from 7 patients with large (>10 cm) ULM and 3 patients with large (>10 cm) uterine leiomyosarcomas. Intervention(s): Tissue microarray analysis by the immunohistochemistry. Main Outcome Measure(s): Selected gene products, vessel density, and the percentage of hyaline degeneration were all scored in tissue cores/sections of large and small ULM against matched myometrium. Result(s): We found that there was a spherical spatial difference of the tumor activities in large ULM. The tumor region next to the periphery, the most biologically active zone, demonstrated higher levels of gene expression, a higher density of vessels, a higher proliferative rate and a lower level of hyaline degeneration. The large ULM have higher levels of gene products (except for estrogen and progesterone receptors) than small ULM. Conclusion(s): In comparison of the spatial patterns of the gene activity between the large ULM and the large uterine leiomyosarcoma, the large ULM illustrate a growth pattern of nutritional dependence. [Copyright &y& Elsevier]

Abstract: To examine whether the selected antisense oligodeoxynucleotides (AS-ODN) targeting against human brain acetylcholinesterase (AChE) mRNA could improve the cognitive deficit in the Alzheimer''s disease (AD) model mice induced by amyloid-β peptide (Aβ), we determined the time–effect relationship of AChE activity and the learning and memory after AS-ODN delivery. The results showed that the AChE activity decreased gradually along with time, initiating at 8 h and lasting 42 h. The time–effect curves of acetylcholine (ACh) behaved consistency with that of AChE activity. The animal cognition studies showed that in step-through test, the error number of the AS-ODN-treated AD model mice was significantly decreased, and the memory retention was increased. In the water maze performance, the swimming time obviously shortened. Our results indicated that antisense therapy is of potential use in the treatment of cognitive deficit in the Aβ model mice. [Copyright &y& Elsevier]

Huntington Disease (HD) is caused by an abnormal expansion of polyQ tract in the protein named huntingtin (Htt). HD pathology is featured by accumulation and aggregation of mutant Htt in striatal and cortical neurons. Aberrant Htt degradation is implicated in HD pathogenesis. The aim of this study was to investigate the regulatory role of chaperone-mediated autophagy (CMA) components, heat shock protein cognate 70 (Hsc70) and lysosome-associated protein 2A (LAMP-2A) in degradation of Htt fragment 1-552aa (Htt-552). A cell model of HD was produced by overexpression of Htt-552 with adenovirus. The involvement of CMA components in degradation of Htt-552 was determined with over-expression or silencing of Hsc70 and LAMP-2A. The results confirmed previous reports that both macroautophagy and CMA were involved in degradation of Htt-552. Changing the levels of CMA-related proteins affected the accumulation of Htt-552. The lysosomal binding and luminal transport of Htt-552 was demonstrated by incubation of Htt-552 with isolated lysosomes. Expansion of the polyQ tract in Htt-552 impaired its uptake and degradation by lysosomes. Mutation of putative KFERQ motif in wild-type Htt-552 interfered with interactions between Htt-552 and Hsc70. Endogenous Hsc70 and LAMP-2A interacted with exogenously expressed Htt-552. Modulating the levels of CMA related proteins degraded endogenous full-length Htt. These studies suggest that Hsc70 and LAMP-2A through CMA play a role in the clearance of Htt and suggest a novel strategy to target the degradation of mutant Htt.

Ginseng is an ancient medicinal and edible plant with many health benefits, and can serve as a drug and dietary supplement, but there are few relevant studies on its use to ease ultraviolet (UV) irradiation damage. After 0.8 mg/mL ginseng extract (GE) was added to the medium of female Drosophila melanogaster subjected to UV irradiation, the lifespan, climbing ability, sex ratio, developmental cycle, and antioxidant capacity of flies were examined to evaluate the GE function. In addition, the underlying mechanism by which GE enhances the irradiation tolerance of D. melanogaster was explored. With GE supplementation, female flies subjected to UV irradiation exhibited an extension in their lifespan, enhancement in their climbing ability, improvement in their offspring sex ratio, and restoration of the normal development cycle by increasing their antioxidant activity. Finally, further experiments indicated that GE could enhance the irradiation tolerance of female D. melanogaster by upregulating the gene expressions of SOD, GCL, and components of the autophagy signaling pathway. Finally, the performance of r4-Gal4;UAS-AMPKRNAi flies confirmed the regulatory role of the autophagy signaling pathway in mitigating UV irradiation injury. [ABSTRACT FROM AUTHOR]

The transcription start sites (TSSs) of porcine mitochondrial genome were firstly identified in this study, including heavy-strand promoter 1 and 2 (HSP1 and HSP2) harbored at nt 903 and nt 1369 in H strand, respectively, and light-strand promoter (LSP) located at nt 166 in L strand. HSP1 structure and expression features were investigated by analyzing mtDNA copy number, expression of 11 nucleoplasmic genes, mtDNA methylation levels, and gene expression levels of methyl-modifying enzymes, DNMT1 and TETs. The mtDNA copy number presented large differences among 15 organs/tissues, and the largest disparity, nearly 17 times, was found between pancreas (~1890 relative copy numbers) and spleen (~110 relative copy numbers, P <.01). The expression levels of HSP1 strand in these organs/tissues presented similar trends with mtDNA copy number (P <.05), and all of 11 nucleoplasmic genes (POLG , POLRMT , TERT , TFAM , TFB1M , TFB2M , NRF-1 , PPARα , ESRRA , SP1 and TUFM) detected in this study displayed significantly higher expression values in pancreas than those in spleen (P <.05). Besides, bisulfite sequencing showed that all cytosine residues in the detected region (D-loop) existed methylation with different levels, and the methylation level in spleen was significantly higher than that in pancreas (P <.05). Unlike nuclear DNA, the tested region contained four types of methylation mode (CA, CC, CT, and CG). In addition, the expression of TET1 in pancreas was significantly higher than that in spleen (P <.05). Collectively, our findings indicated that mtDNA TSSs had correlation to mtDNA copy number, expression of nucleoplasmic gene, and mtDNA methylation level. • The TSSs of porcine mitochondrial genome were firstly identified. • Porcine mtDNA TSSs were correlated to mtDNA copy number, expressions of nucleoplasmic genes, and mtDNA methylation levels. • Pancreas and brain possessed the highest mtDNA copies compared with other organs/tissues detected in this study. • Mitochondrial genome methylation was negatively correlated with mtDNA transcription activity. [ABSTRACT FROM AUTHOR]

NLRC5, a newly found member of the NLR family and the largest member of nucleotide-binding, has been reported to regulate immune responses and is associated with hepatocellular carcinoma (HCC). We investigated the mechanisms and signaling pathways of NLRC5 in HCC progression. Increased expression of NLRC5, vascular endothelial growth factor-A (VEGF-A) were found in human HCC tissue. There was a positive correlation between NLRC5 and VEGF-A expression and cell proliferation were enhanced in NLRC5-overexpressing HepG2 cells, but inhibited in cells with NLRC5 silencing treatment. Interestingly, we found that up-regulation of NLRC5 also coordinated the activation of PI3K/AKT signaling pathway. An AKT inhibitor LY294002 blocked VEGF-A expression and AKT phosphorylation in HepG2 cells and NLRC5-overexpressing HepG2 cells. These results demonstrate that NLRC5 promotes HCC progression via the AKT/VEGF-A signaling pathway. [ABSTRACT FROM AUTHOR]