Analysis of differentially expressed long non-coding RNAs in LPS-induced human HMC3 microglial cells.

BACKGROUND: Long non-coding RNAs (lncRNAs) are emerging as key modulators of inflammatory gene expression, but their roles in neuroinflammation are poorly understood. Here, we identified the inflammation-related lncRNAs and correlated mRNAs of the lipopolysaccharide (LPS)-treated human microglial cell line HMC3. We explored their potential roles and interactions using bioinformatics tools such as gene ontology (GO), kyoto encyclopedia of genes and genomes (KEGG), and weighted gene co-expression network analysis (WGCNA). RESULTS: We identified 5 differentially expressed (DE) lncRNAs, 4 of which (AC083837.1, IRF1-AS1, LINC02605, and MIR3142HG) are novel for microglia. The DElncRNAs with their correlated DEmRNAs (99 total) fell into two network modules that both were enriched with inflammation-related RNAs. However, treatment with the anti-inflammatory agent JQ1, an inhibitor of the bromodomain and extra-terminal (BET) protein BRD4, neutralized the LPS effect in only one module, showing little or even enhancing effect on the other. CONCLUSIONS: These results provide insight into, and a resource for studying, the regulation of microglia-mediated neuroinflammation and its potential therapy by small-molecule BET inhibitors.

Basal-type lumenogenesis in extraembryonic endoderm stem cells models the early visceral endoderm.

Cultured rat primitive extraembryonic endoderm (pXEN) cells easily form free-floating multicellular vesicles de novo, exemplifying a poorly studied type of morphogenesis. Here, we reveal the underlying mechanism and the identity of the vesicles. We resolve the morphogenesis into vacuolization, vesiculation and maturation, and define the molecular characteristics and requirements of each step. Vacuolization is fueled by macropinocytosis and occurs by default if not blocked by high cell density or matrix proteins. Fine-tuned cell-cell contact then forms nascent three-cell vesicles with vacuole-derived lumina. In maturation, the vesicles complete epithelialization, expand via mitosis and continued fluid uptake, and differentiate further. The mature vesicles consist of a simple squamous epithelium with an apical-outside/basal-inside polarity that we trace back to the single cell stage. The polarity and gene expression pattern of the vesicles are similar to those of the early visceral endoderm. pXEN cells provide a useful in vitro model for study of matrix-independent, basal-type lumenogenesis and the physiology of the visceral endoderm.This article has an associated First Person interview with the first author of the paper.

Forkhead box O1 (FOXO1) controls the migratory response of Toll-like receptor (TLR3)-stimulated human mesenchymal stromal cells.

Mesenchymal stromal cells (MSCs) can potently regulate the functions of immune cells and are being investigated for the management of inflammatory diseases. Toll-like receptor 3 (TLR3)-stimulated human MSCs (hMSCs) exhibit increased migration and chemotaxis within and toward damaged tissues. However, the regulatory mechanisms underlying these migratory activities are unclear. Therefore, we analyzed the migration capability and gene expression profiles of TLR3-stimulated hMSCs using RNA-Seq, wound healing, and transwell cell migration assay. Along with increased cell migration, the TLR3 stimulation also increased the expression of cytokines, chemokines, and cell migration-related genes. The promoter regions of the latter showed an enrichment of putative motifs for binding the transcription factors forkhead box O1 (FOXO1), FOXO3, NF-κB (NF-κB1), and RELA proto-oncogene and NF-κB subunit. Of note, FOXO1 inhibition by the FOXO1-selective inhibitor AS1842856 significantly reduced both migration and the expression of migration-related genes. In summary, our results indicate that TLR3 stimulation induces hMSC migration through the expression of FOXO1-activated genes.

Maternal alcohol consumption and altered miRNAs in the developing fetus: Context and future perspectives.

Alcohol is a teratogenic agent that can cause a wide range of developmental disorders, and sometimes, the effects persist throughout an individual's lifetime. Researchers have shown the involvement of epigenetic mechanisms in alcohol-mediated disorders. Non-coding RNAs are one of the major sources of epigenetic modifications, especially microRNAs. The association of microRNAs with alcohol consumption leads to a new focus on finding the molecular mechanisms of alcohol toxicity. It has been suggested that alcohol alters the relative expression of microRNAs and regulates target mRNA expression in both in vitro and in vivo models. Currently, we lack information regarding the relationship between altered microRNA expression and disease phenotypes in alcohol-mediated disorders. In this review, we tried to gather all of the available information about the alcohol-mediated dysregulation of microRNA expression in utero. We hope that our efforts will help future researchers identify major microRNAs in the field of prenatal alcohol toxicity and related therapeutics.

Deletion of a putative NlpC/P60 endopeptidase BAS1812 affects germination, long-term survival and endospore formation in Bacillus anthracis.

Bacillus anthracis, an aetiologic agent of the zoonotic disease anthrax, encodes a putative NlpC/P60 endopeptidase BAS1812. It harbours a signal peptide, three bacterial SH3 domains and an NlpC/P60 family domain. Previous studies showed that BAS1812 is immunogenic in infected hosts and is a potential biomarker for anthrax treatment. To date, however, little information is known about its function and involvement in anthrax pathogenesis. Here we describe the phenotypic effect of BAS1812 deletion in B. anthracis Sterne strain. Transcriptional analysis showed that BAS1812 expression in a host-like environment was enhanced at the end of log phase, started to diminish after entry to stationary phase and increased again late in stationary phase. The constructed BAS1812 mutant showed impaired long-term survival in the stationary growth phase, less resilience to detergent, lesser endospore formation and delayed germination. The mutant also showed diminished ability to degrade peptidoglycan, but its ability to produce anthrax exotoxins was not affected. We hypothesize that BAS1812 is a cell wall hydrolase involved in biological activities related to maintaining cell wall integrity, sporulation and spore germination.

Gestational Alcohol Exposure Altered DNA Methylation Status in the Developing Fetus.

Ethanol is well known as a teratogenic factor that is capable of inducing a wide range of developmental abnormalities if the developing fetus is exposed to it. Duration and dose are the critical parameters of exposure that affect teratogenic variation to the developing fetus. It is suggested that ethanol interferes with epigenetic processes especially DNA methylation. We aimed to organize all of the available information on the alteration of DNA methylation by ethanol in utero. Thus, we have summarized all published information regarding alcohol-mediated alterations in DNA methylation during gestation. We tried to arrange information in a way that anyone can easily find the alcohol exposure time, doses, sampling time, and major changes in genomic level. Manuscript texts will also represent the correlation between ethanol metabolites and subsequent changes in methylome patterns. We hope that this review will help future researchers to further examine the issues associated with ethanol exposure.

Changes in Bacillus anthracis CodY regulation under host-specific environmental factor deprived conditions.

BACKGROUND: Host-specific environmental factors induce changes in Bacillus anthracis gene transcription during infection. A global transcription regulator, CodY, plays a pivotal role in regulating central metabolism, biosynthesis, and virulence in B. anthracis. In this study, we utilized RNA-sequencing to assess changes in the transcriptional patterns of CodY-regulated B. anthracis genes in response to three conditions of environmental starvation: iron, CO2, or glucose deprivation. In addition, we performed chromatin immunoprecipitation on newly identified CodY-mediated genes. RESULTS: Environmental deprivation induced transcriptional changes in CodY-regulated genes in both wild-type and codY null strains, and both CodY-specific and environment-specific patterns were observed. In the iron-depleted condition, overexpression of iron homeostasis genes was observed independent of codY deletion; however, transcription of siderophore and amino acid biosynthesis genes was CodY dependent. Although CodY has a significant regulatory role in central metabolism and the carbon overflow pathway, metabolism-associated genes exhibited CodY-independent expression patterns under glucose starvation. Genes that were differentially expressed in response to CO2 availability showed CodY-dependent regulation, though their maximal expression did require a supply of CO2/bicarbonate. CONCLUSIONS: We speculate that CodY regulates the expression of environmental-responsive genes in a hierarchical manner and is likely associated with other transcription regulators that are specific for a particular environmental change.

Transcriptome sequencing reveals that LPS-triggered transcriptional responses in established microglia BV2 cell lines are poorly representative of primary microglia.

BACKGROUND: Microglia are resident myeloid cells in the CNS that are activated by infection, neuronal injury, and inflammation. Established BV2 microglial cell lines have been the primary in vitro models used to study neuroinflammation for more than a decade because they reduce the requirement of continuously maintaining cell preparations and animal experimentation models. However, doubt has recently been raised regarding the value of BV2 cell lines as a model system. METHODS: We used triplicate RNA sequencing (RNA-seq) to investigate the molecular signature of primary and BV2 microglial cell lines using two transcriptomic techniques: global transcriptomic biological triplicate RNA-seq and quantitative real-time PCR. We analyzed differentially expressed genes (DEGs) to identify transcription factor (TF) motifs (-950 to +50 bp of the 5' upstream promoters) and epigenetic mechanisms. RESULTS: Sequencing assessment and quality evaluation revealed that primary microglia have a distinct transcriptomic signature and express a unique cluster of transcripts in response to lipopolysaccharide. This microglial signature was not observed in BV2 microglial cell lines. Importantly, we observed that previously unidentified TFs (i.e., IRF2, IRF5, IRF8, STAT1, STAT2, and STAT5A) and the epigenetic regulators KDM1A, NSD3, and SETDB2 were significantly and selectively expressed in primary microglia (PM). Although transcriptomic alterations known to occur in BV2 microglial cell lines were identified in PM, we also observed several novel transcriptomic alterations in PM that are not frequently observed in BV2 microglial cell lines. CONCLUSIONS: Collectively, these unprecedented findings demonstrate that established BV2 microglial cell lines are probably a poor representation of PM, and we establish a resource for future studies of neuroinflammation.

Late-Exponential Gene Expression in codY-Deficient Bacillus anthracis in a Host-Like Environment.

CodY is a pleiotropic regulator commonly found in Gram-positive bacteria and regulates various biological processes during the stringent response in a nutrient-limiting environment. CodY also participates in virulence factor expression in many low G+C Gram-positive pathogens, as observed in Bacillus anthracis. However, the mechanism by which B. anthracis CodY regulates metabolism and virulence factors in response to environmental changes is unclear. Here, we attempted to identify the link between CodY and B. anthracis regulation with codY-deficient and codY-overexpressing mutants using high-throughput transcriptional analysis. Growth pattern analyses of codY mutants in both rich and minimal media showed defects in early cell proliferation, with opposite patterns in the early stationary phase: CodY overexpression prolonged bacterial growth, whereas deletion inhibited growth. RNA sequencing of codY-deficient B. anthracis showed both positive and negative changes in the gene expression of proteases and virulence factors as well as genes related to stringent response-related metabolism and biosynthetic processing. We also found that changes in codY expression could alter virulence gene expression of B. anthracis, suggesting modes of regulation in its virulence in a CodY concentration-dependent manner. Collectively, we conclude from these results that CodY can both positively and negatively regulate its regulon via direct and/or indirect approaches, and that its mode of regulation may be concentration dependent.

Transcriptome sequencing of microglial cells stimulated with TLR3 and TLR4 ligands.

BACKGROUND: Resident macrophages in the CNS microglia become activated and produce proinflammatory molecules upon encountering bacteria or viruses. TLRs are a phylogenetically conserved diverse family of sensors that drive innate immune responses following interactions with PAMPs. TLR3 and TLR4 recognize viral dsRNA Poly (I:C) and bacterial endotoxin LPS, respectively. Importantly, these receptors differ in their downstream adaptor molecules. Thus far, only a few studies have investigated the effects of TLR3 and TLR4 in macrophages. However, a genome-wide search for the effects of these TLRs has not been performed in microglia using RNA-seq. Gene expression patterns were determined for the BV-2 microglial cell line when stimulated with viral dsRNA Poly (I:C) or bacterial endotoxin LPS to identify novel transcribed genes, as well as investigate how differences in downstream signaling could influence gene expression in innate immunity. RESULTS: Sequencing assessment and quality evaluation revealed that common and unique patterns of proinflammatory genes were significantly up-regulated in response to TLR3 and TLR4 stimulation. However, the IFN/viral response gene showed a stronger response to TLR3 stimulation than to TLR4 stimulation. Unexpectedly, TLR3 and TLR4 stimulation did not activate IFN-ß and IRF3 in BV-2 microglia. Most importantly, we observed that previously unidentified transcription factors (TFs) (i.e., IRF1, IRF7, and IRF9) and the epigenetic regulators KDM4A and DNMT3L were significantly up-regulated in both TLR3- and TLR4-stimulated microglia. We also identified 29 previously unidentified genes that are important in immune regulation. In addition, we confirmed the expressions of key inflammatory genes as well as pro-inflammatory mediators in the supernatants were significantly induced in TLR3-and TLR4-stimulated primary microglial cells. Moreover, transcriptional start sites (TSSs) and isoforms, as well as differential promoter usage, revealed a complex pattern of transcriptional and post-transcriptional gene regulation upon infection with TLR3 and TLR4. Furthermore, TF motif analysis (-950 to +50 bp of the 5' upstream promoters) revealed that the DNA sequences for NF-κB, IRF1, and STAT1 were significantly enriched in TLR3- and TLR4-stimulated microglia. CONCLUSIONS: These unprecedented findings not only permit a comparison of TLR3-and TLR4-stimulated genes but also identify new genes that have not been previously implicated in innate immunity.

RNA sequencing reveals distinct mechanisms underlying BET inhibitor JQ1-mediated modulation of the LPS-induced activation of BV-2 microglial cells.

BACKGROUND: Microglial cells become rapidly activated through interaction with pathogens, and their persistent activation is associated with the production and secretion of various pro-inflammatory genes, cytokines, and chemokines, which may initiate or amplify neurodegenerative diseases. Bromodomain and extraterminal domain (BET) proteins are a group of epigenetic regulators that associate with acetylated histones and facilitate the transcription of target genes. A novel synthetic BET inhibitor, JQ1, was proven to exert immunosuppressive activities by inhibiting the expression of IL-6 and Tnf-α in macrophages. However, a genome-wide search for JQ1 molecular targets is largely unexplored in microglia. METHODS: The present study was aimed at evaluating the anti-inflammatory function and underlying genes targeted by JQ1 in lipopolysaccharide (LPS)-stimulated BV-2 microglial cells using two transcriptomic techniques: global transcriptomic biological duplicate RNA sequencing and quantitative real-time PCR. Associated biological pathways and functional gene ontology were also evaluated. RESULTS: With a cutoff value of P ≤ 0.01 and fold change ≥1.5 log2, the expression level of 214 and 301 genes, including pro-inflammatory cytokine, chemokine, and transcription factors, was found to be upregulated in BV-2 cells stimulated with LPS for 2 and 4 h, respectively. Among these annotated genes, we found that JQ1 selectively reduced the expression of 78 and 118 genes (P ≤ 0.01, and fold change ≥ 1.5, respectively). Importantly, these inflammatory genes were not affected by JQ1 treatment alone. Furthermore, we confirmed that JQ1 reduced the expression of key inflammation- and immunity-related genes as well as cytokines/chemokines in the supernatants of LPS-treated primary microglial cells isolated from 3-day-old ICR mice. Utilizing functional group analysis, the genes affected by JQ1 were classified into four categories related to biological regulation, immune system processes, and response to stimuli. Moreover, the biological pathways and functional genomics obtained in this study may facilitate the suppression of different key inflammatory genes through JQ1-treated BV-2 microglial cells. CONCLUSIONS: These unprecedented results suggest the BET inhibitor JQ1 as a candidate for the prevention or therapeutic treatment of inflammation-mediated neurodegenerative diseases.

Transcriptomic study of mouse embryonic neural stem cell differentiation under ethanol treatment.

Neural stem cells (NSCs) can be differentiated into one of three cell lineages: neurons, astrocytes or, oligodendrocytes. Some neurotoxins have the ability to deregulate this dynamic process. NSC cell fate can be altered by ethanol as reported previously. Our aim was to investigate the alteration of genes by ethanol during NSC differentiation and to explore the molecular mechanism underlying this phenomenon. Here, mouse fetal forebrain derived NSCs were differentiated for 2 days with or without of ethanol (50 mM). We performed a comparative microarray analysis at day two using GeneChip(®) Mouse Genome 430A 2.0 arrays. Microarray analysis showed that the expressions of 496 genes were altered by ethanol (56 and 440 were up- and down-regulated, respectively). Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed the association of the following altered genes in the Wnt signaling pathway: Wnt5a, Csnk2a1, Tcf7l2, Ccnd2, Nlk, Tbl1x, Tbl1xr1, Rac2 and Nfatc3. Quantitative real time PCR analysis also demonstrated the relative expression levels of these genes. As Wnt signaling is a player of brain development, ethanol-induced alterations may contribute to improper development of the brain. Our data could be a useful resource for elucidating the mechanism behind the ethanol neurotoxicity in developing brain.

JMJD2A attenuation affects cell cycle and tumourigenic inflammatory gene regulation in lipopolysaccharide stimulated neuroectodermal stem cells.

JMJD2A is a lysine trimethyl-specific histone demethylase that is highly expressed in a variety of tumours. The role of JMJD2A in tumour progression remains unclear. The objectives of this study were to identify JMJD2A-regulated genes and understand the function of JMJD2A in p53-null neuroectodermal stem cells (p53(-/-) NE-4Cs). We determined the effect of LPS as a model of inflammation in p53(-/-) NE-4Cs and investigated whether the epigenetic modifier JMJD2A alter the expression of tumourigenic inflammatory genes. Global gene expression was measured in JMJD2A knockdown (kd) p53(-/-) NE-4Cs and in LPS-stimulated JMJD2A-kd p53(-/-) NE-4C cells. JMJD2A attenuation significantly down-regulated genes were Cdca2, Ccnd2, Ccnd1, Crebbp, IL6rα, and Stat3 related with cell cycle, proliferation, and inflammatory-disease responses. Importantly, some tumour-suppressor genes including Dapk3, Timp2 and TFPI were significantly up-regulated but were not affected by silencing of the JMJD2B. Furthermore, we confirmed the attenuation of JMJD2A also down-regulated Cdca2, Ccnd2, Crebbp, and Rest in primary NSCs isolated from the forebrains of E15 embryos of C57/BL6J mice with effective p53 inhibitor pifithrin-α (PFT-α). Transcription factor (TF) motif analysis revealed known binding patterns for CDC5, MYC, and CREB, as well as three novel motifs in JMJD2A-regulated genes. IPA established molecular networks. The molecular network signatures and functional gene-expression profiling data from this study warrants further investigation as an effective therapeutic target, and studies to elucidate the molecular mechanism of JMJD2A-kd-dependent effects in neuroectodermal stem cells should be performed.

Ethanol-related alterations in gene expression patterns in the developing murine hippocampus.

It is well known that consuming alcohol prior to and during pregnancy can cause harm to the developing fetus. Fetal alcohol spectrum disorder is a term commonly used to describe a range of disabilities that may arise from prenatal alcohol exposure such as fetal alcohol syndrome, partial fetal alcohol syndrome, alcohol-related neurodevelopmental disorders, and alcohol-related birth defects. Here, we report that maternal binge alcohol consumption alters several important genes that are involved in nervous system development in the mouse hippocampus at embryonic day 18. Microarray analysis revealed that Nova1, Ntng1, Gal, Neurog2, Neurod2, and Fezf2 gene expressions are altered in the fetal hippocampus. Pathway analysis also revealed the association of the calcium signaling pathway in addition to other pathways with the differentially expressed genes during early brain development. Alteration of such important genes and dynamics of the signaling pathways may cause neurodevelopmental disorders. Our findings offer insight into the molecular mechanism involved in neurodevelopmental disorders associated with alcohol-related defects.

Proteomic analysis reveals KRIT1 as a modulator for the antioxidant effects of valproic acid in human bone-marrow mesenchymal stromal cells.

Valproic acid (VPA) protects human bone marrow-mesenchymal stromal cells (hBM-MSCs) against oxidative stress and improves their migratory ability through increasing the secretion of trophic factors. This suggests that VPA may be an excellent candidate for improving stem cell function. However, the molecular mechanisms of VPA in BM-MSCs are not known. In this study, we used a proteomic approach to investigate VPA-associated targets under oxidative stress conditions. Krev/Rap1 interaction Trapped-1 (KRIT1), a modulator for the homeostasis of intracellular reactive oxygen species (ROS), was identified as a target protein by two-dimensional gel electrophoresis and matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF-MS) analyses. The up-regulation of KRIT1 and its target proteins (SOD2 and FoxO1) with VPA treatment of hBM-MSCs was revealed by qPCR and immunoblot analysis. Damage from oxidative stress was reduced in VPA-pretreated BM-MSCs, which was also confirmed by qPCR and immunoblot analysis. In addition, increased in intracellular ROS by H2O2 were also reduced by VPA pretreatment in BM-MSCs. This suggests that VPA reduces intracellular ROS level by the modulation of KRIT1 and its correlated proteins, FoxO1, SOD2, and cyclin D1. Thus, this study is the first to provide evidence that VPA modulates KRIT1 and intracellular ROS in BM-MSCs.

Identification of stringent response-related and potential serological proteins released from Bacillus anthracis overexpressing the RelA/SpoT homolog, Rsh Bant.

RelA and SpoT synthesize ppGpp, a key effector molecule that facilitates the adaptation of bacteria to nutrient starvation and other stresses, known as the stringent response. To investigate the role of Rsh Bant , a putative RelA/SpoT homolog (encoded by BAS4302) in Bacillus anthracis, we examined the alteration of the secretome profiles after the overexpression of a functional His-Rsh Bant protein in the B. anthracis strain Sterne at the stationary growth phase. In the ppGpp-deficient E. coli mutant strain CF1693, overexpression of Rsh Bant restored a ppGpp-dependent growth defect on minimal glucose media. The secretome profiles obtained using a two-dimensional electrophoresis (2-DE) analysis were altered by overexpression of Rsh Bant in B. anthracis. Among the 66 protein spots differentially expressed >1.5-fold, the 29 proteins were abundant for further identification using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Functional categorization of those proteins implicated their involvement in various biological activities. Taken together, our results imply that overexpression of a functional His-Rsh Bant can lead to the increased levels of intracellular ppGpp in B. anthracis, resulting in the significant changes in its secretome profiling. The stringent response-controlled proteins identified are likely useful as potential targets for serodiagnostic applications.

Chronic ethanol exposure increases goosecoid (GSC) expression in human embryonic carcinoma cell differentiation.

Fetal alcohol spectrum disorder (FASD) is a set of developmental malformations caused by excess alcohol consumption during pregnancy. Using an in vitro system, we examined the role that chronic ethanol (EtOH) exposure plays in gene expression changes during the early stage of embryonic differentiation. We demonstrated that EtOH affected the cell morphology, cell cycle progression and also delayed the down-regulation of OCT4 and NANOG during differentiation. Gene expression profiling and pathway analysis demonstrated that EtOH deregulates many genes and pathways that are involved in early embryogenesis. Follow-up analyzes revealed that EtOH exposure to embryoid bodies (EBs) induced the expression of an organizer-specific gene, goosecoid (GSC), in comparison to controls. Moreover, EtOH treatment altered several important genes that are involved in embryonic structure formation, nervous system development, and placental and embryonic vascularization, which are all common processes that FASD can disrupt. Specifically, EtOH treatment let to a reduction in ALDOC, ENO2 and CDH1 expression, whereas EtOH treatment induced the expression of PTCH1, EGLN1, VEGFA and DEC2 in treated EBs. We also found that folic acid (FA) treatment was able to correct the expression of the majority of genes deregulated by EtOH exposure during early embryo development. Finally, the present study identified a gene set including GSC, which was deregulated by EtOH exposure that may contribute to the etiology of fetal alcohol syndrome (FAS). We also reported that EtOH-induced GSC expression is mediated by Nodal signaling, which may provide a new avenue for analyzing the molecular mechanisms behind EtOH teratogenicity in FASD individuals.

Identification of differentially expressed mRNA/lncRNA modules in acutely regorafenib-treated sorafenib-resistant Huh7 hepatocellular carcinoma cells.

The multikinase inhibitor sorafenib is the standard first-line treatment for advanced hepatocellular carcinoma (HCC), but many patients become sorafenib-resistant (SR). This study investigated the efficacy of another kinase inhibitor, regorafenib (Rego), as a second-line treatment. We produced SR HCC cells, wherein the PI3K-Akt, TNF, cAMP, and TGF-beta signaling pathways were affected. Acute Rego treatment of these cells reversed the expression of genes involved in TGF-beta signaling but further increased the expression of genes involved in PI3K-Akt signaling. Additionally, Rego reversed the expression of genes involved in nucleosome assembly and epigenetic gene expression. Weighted gene co-expression network analysis (WGCNA) revealed four differentially expressed long non-coding RNA (DElncRNA) modules that were associated with the effectiveness of Rego on SR cells. Eleven putative DElncRNAs with distinct expression patterns were identified. We associated each module with DEmRNAs of the same pattern, thus obtaining DElncRNA/DEmRNA co-expression modules. We discuss the potential significance of each module. These findings provide insights and resources for further investigation into the potential mechanisms underlying the response of SR HCC cells to Rego.

A global proteome study of Mycobacterium gilvum PYR-GCK grown on pyrene and glucose reveals the activation of glyoxylate, shikimate and gluconeogenetic pathways through the central carbon metabolism highway.

Various hydrocarbons have been released into the environment as a result of industrialization. An effective way of removing these materials without further environmental contamination is microbial bioremediation. Mycobacterium gilvum PYR-GCK, a bacteria isolated from a PAH polluted estuary, was studied using comparative shotgun proteomics to gain insight on its molecular activity while using pyrene and glucose as sole carbon and energy sources. Based on annotated genomic information, a confirmation analysis was first performed to confirm its pyrene degradation activity, using gas chromatography-mass spectrometry technology. One dimensional gel electrophoresis and liquid chromatography-mass spectrometry technologies employed in the proteomics analysis revealed the expression of pyrene degrading gene products along with upregulated expression of proteins functioning in the glyoxylate and shikimate pathways, in the pyrene-induced cells. The study also revealed the pathway of pyrene degraded intermediates, via partial gluconeogenesis, into the pentose phosphate pathway to produce precursors for nucleotides and amino acids biosynthesis.

Defined Conditions Control the Morphological Dualism of Rat Primitive Extraembryonic Endoderm Stem Cells.

Rat primitive extraembryonic endoderm (pXEN) stem cell lines indefinitely preserve the characteristic features of the early extraembryonic endoderm (ExEn) in vitro, but require unknown serum factors and exhibit a hybrid (mesenchymal-epithelial) phenotype. We report two chemically defined conditions that differ by the addition of the cytokine leukemia inhibitory factor (Lif) and the ß-catenin-stabilizing drug Chir99021, and enable permanent self-renewal as mesenchymal and epithelial morphotypes, respectively. The morphotypes are interconvertible and equipotent, as shown by the formation of well-differentiated organoids. Surprisingly, the proliferation of both morphotypes requires Lif-type Gp130/Stat3 signaling (autocrine in the absence of added Lif) and noncanonical Wnt signaling (autocrine). In addition, the epithelial version requires ß-catenin for proliferation and morphology. Interestingly, the mesenchymal cells also express key epithelial markers, but those are improperly structured and/or not functional, indicating a primed state. These results provide an improved platform for studying the proliferation and plasticity of the early ExEn, which occurs in mesenchymal and epithelial forms in vivo.