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Find video protocols related to scientific articles indexed in Pubmed.
Analysis of Candida albicans mutants defective in the Cdk8 module of mediator reveal links between metabolism and biofilm formation.
PLoS Genet.
PUBLISHED: 10-01-2014
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Candida albicans biofilm formation is a key virulence trait that involves hyphal growth and adhesin expression. Pyocyanin (PYO), a phenazine secreted by Pseudomonas aeruginosa, inhibits both C. albicans biofilm formation and development of wrinkled colonies. Using a genetic screen, we identified two mutants, ssn3?/? and ssn8?/?, which continued to wrinkle in the presence of PYO. Ssn8 is a cyclin-like protein and Ssn3 is similar to cyclin-dependent kinases; both proteins are part of the heterotetrameric Cdk8 module that forms a complex with the transcriptional co-regulator, Mediator. Ssn3 kinase activity was also required for PYO sensitivity as a kinase dead mutant maintained a wrinkled colony morphology in the presence of PYO. Furthermore, similar phenotypes were observed in mutants lacking the other two components of the Cdk8 module-Srb8 and Srb9. Through metabolomics analyses and biochemical assays, we showed that a compromised Cdk8 module led to increases in glucose consumption, glycolysis-related transcripts, oxidative metabolism and ATP levels even in the presence of PYO. In the mutant, inhibition of respiration to levels comparable to the PYO-treated wild type inhibited wrinkled colony development. Several lines of evidence suggest that PYO does not act through Cdk8. Lastly, the ssn3 mutant was a hyperbiofilm former, and maintained higher biofilm formation in the presence of PYO than the wild type. Together these data provide novel insights into the role of the Cdk8 module of Mediator in regulation of C. albicans physiology and the links between respiratory activity and both wrinkled colony and biofilm development.
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Pregnancy-associated miRNA-clusters.
J. Reprod. Immunol.
PUBLISHED: 02-26-2013
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MicroRNAs (miRNAs) are expressed in the placenta and can be detected in maternal plasma. An increasing number of studies have been published on the cellular origin, distribution and function of miRNAs in pregnancy. Specific miRNA profiles have been described for the placenta, maternal plasma and several pregnancy disorders. It has been observed that numerous miRNAs, which are predominantly or exclusively expressed during pregnancy, are clustered in chromosomal regions, may be controlled by the same promoters, may have similar seed regions and targets, and work synergistically. The three most eminent clusters are the chromosome 19 miRNA cluster (C19MC), C14MC and miR-371-3 cluster, which is also localized on chromosome 19. MiRNA members of these clusters are not only detected in the placenta, but also in other compartments, e.g. in serum where they have the potential to become novel biomarkers of pregnancy disorders. Additionally, some members are also expressed in a variety of tumors. Antagonism of selected miRNAs or their targets may lead to novel strategies for the development of new drug classes in pregnancy disorders or other diseases. This review summarizes current knowledge on the pregnancy-related miRNA clusters - the C19MC, C14MC and miR-371-3 cluster - in regard to pregnancy and also other, mostly pathological circumstances.
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YWHAE/14-3-3?: a potential novel genetic risk factor and CSF biomarker for HIV neurocognitive impairment.
J. Neurovirol.
PUBLISHED: 02-11-2013
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YWHAE (14-3-3?) protein levels are considered to be a reliable biomarker for neurodegeneration. The YWHAE protein interacts both directly and indirectly with human immunodeficiency virus (HIV) accessory proteins, leading to cell death. The purpose of this study was to examine the relationship between YWHAE polymorphisms and HIV-associated neurocognitive disorder (HAND) and the relationship between YWHAE protein levels and HAND. A cross-sectional study using random samples of HIV-seropositive (n?=?20) and HIV-seronegative (controls) (n?=?16) women from the Hispanic-Latino Longitudinal Cohort of Women was conducted. Individuals who are HIV-seropositive and heterozygous at the rs4790084/rs1204828 loci in the YWHAE gene were 3× more likely to display reduced cognitive functioning, to have received a HAND diagnosis, and to have less YHWAE protein expressed than homozygotes. Western blots from cerebral spinal fluid indicate that the HIV-seropositive women with HAND expressed 4.5× less YWHAE compared to HIV-seropositive cognitively normal women (94 % sensitivity, 84 % specificity; HIV-seropositive vs. controls). Therefore, polymorphism in YWHAE may be a genetic risk factor for HAND and levels of YWHAE protein are a likely biomarker for neurocognitive status in HIV-seropositive women.
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Control of Candida albicans metabolism and biofilm formation by Pseudomonas aeruginosa phenazines.
MBio
PUBLISHED: 01-31-2013
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Candida albicans has developmental programs that govern transitions between yeast and filamentous morphologies and between unattached and biofilm lifestyles. Here, we report that filamentation, intercellular adherence, and biofilm development were inhibited during interactions between Candida albicans and Pseudomonas aeruginosa through the action of P. aeruginosa-produced phenazines. While phenazines are toxic to C. albicans at millimolar concentrations, we found that lower concentrations of any of three different phenazines (pyocyanin, phenazine methosulfate, and phenazine-1-carboxylate) allowed growth but affected the development of C. albicans wrinkled colony biofilms and inhibited the fungal yeast-to-filament transition. Phenazines impaired C. albicans growth on nonfermentable carbon sources and led to increased production of fermentation products (ethanol, glycerol, and acetate) in glucose-containing medium, leading us to propose that phenazines specifically inhibited respiration. Methylene blue, another inhibitor of respiration, also prevented the formation of structured colony biofilms. The inhibition of filamentation and colony wrinkling was not solely due to lowered extracellular pH induced by fermentation. Compared to smooth, unstructured colonies, wrinkled colony biofilms had higher oxygen concentrations within the colony, and wrinkled regions of these colonies had higher levels of respiration. Together, our data suggest that the structure of the fungal biofilm promotes access to oxygen and enhances respiratory metabolism and that the perturbation of respiration by bacterial molecules such as phenazines or compounds with similar activities disrupts these pathways. These findings may suggest new ways to limit fungal biofilms in the context of disease. IMPORTANCE Many of the infections caused by Candida albicans, a major human opportunistic fungal pathogen, involve both morphological transitions and the formation of surface-associated biofilms. Through the study of C. albicans interactions with the bacterium Pseudomonas aeruginosa, which often coinfects with C. albicans, we have found that P. aeruginosa-produced phenazines modulate C. albicans metabolism and, through these metabolic effects, impact cellular morphology, cell-cell interactions, and biofilm formation. We suggest that the structure of C. albicans biofilms promotes access to oxygen and enhances respiratory metabolism and that the perturbation of respiration by phenazines inhibits biofilm development. Our findings not only provide insight into interactions between these species but also provide valuable insights into novel pathways that could lead to the development of new therapies to treat C. albicans infections.
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Intranuclear Crosstalk between Extracellular Regulated Kinase1/2 and Signal Transducer and Activator of Transcription 3 Regulates JEG-3 Choriocarcinoma Cell Invasion and Proliferation.
ScientificWorldJournal
PUBLISHED: 01-01-2013
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Invasiveness of trophoblast and choriocarcinoma cells is in part mediated via leukemia inhibitory factor- (LIF-) induced activation of signal transducer and activator of transcription 3 (STAT3). The regulation of STAT3 phosphorylation at its ser727 binding site, possible crosstalk with intracellular MAPK signaling, and their functional implications are the object of the present investigation. JEG-3 choriocarcinoma cells were cultured in presence/absence of LIF and the specific ERK1/2 inhibitor (U0126). Phosphorylation of signaling molecules (p-STAT3 (ser727 and tyr705) and p-ERK1/2 (thr 202/tyr 204)) was assessed per Western blot. Immunocytochemistry confirmed results, but also pinpointed the location of phosphorylated signaling molecules. STAT3 DNA-binding capacity was studied with a colorimetric ELISA-based assay. Cell viability and invasion capability were assessed by MTS and Matrigel assays. Our results demonstrate that LIF-induced phosphorylation of STAT3 (tyr705 and ser727) is significantly increased after blocking ERK1/2. STAT3 DNA-binding capacity and cell invasiveness are enhanced after LIF stimulation and ERK1/2 blockage. In contrast, proliferation is enhanced by LIF but reduced after ERK1/2 inhibition. The findings herein show that blocking ERK1/2 increases LIF-induced STAT3 phosphorylation and STAT3 DNA-binding capacity by an intranuclear crosstalk, which leads to enhanced invasiveness and reduced proliferation.
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Understanding the link between the IL-6 cytokine family and pregnancy: implications for future therapeutics.
Expert Rev Clin Immunol
PUBLISHED: 09-08-2011
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Cytokines are involved in almost all processes during the menstrual cycle, the fertilization period and pregnancy. They are expressed in numerous reproduction-related body fluids and tissues. Disorders of cytokine expression patterns may cause pregnancy pathologies. Therefore, cytokines have the potential as new biomarkers in different body compartments for a variety of such pathologies. Furthermore, cytokines may also serve to treat fertility and pregnancy disorders. The IL-6-like family of cytokines is an intensively investigated group of cytokines with well-accepted functions in fertility and pregnancy. This article summarizes current knowledge on IL-6-like cytokines in regard of their role in reproduction and their potential for new strategies in the treatment of reproductive pathologies.
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Reduction in miR-141 is induced by leukemia inhibitory factor and inhibits proliferation in choriocarcinoma cell line JEG-3.
Am. J. Reprod. Immunol.
PUBLISHED: 07-06-2011
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Starting from the peri-implantation period, leukemia inhibitory factor (LIF) is a major regulator of trophoblast functions. Micro-RNAs (miRNA) are short non-coding RNA sequences, which regulate expression of genes at post-transcriptional level. The influence of LIF on miRNA expression in trophoblastic cells has not yet been analyzed and was focus of this investigation.
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Leukaemia inhibitory factor mediated proliferation of HTR-8/SVneo trophoblast cells is dependent on activation of extracellular signal-regulated kinase 1/2.
Reprod. Fertil. Dev.
PUBLISHED: 06-04-2011
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Leukaemia inhibitory factor (LIF) is one of the cytokines that is indispensable for embryo implantation. The aim of the present study was to investigate the role of activation of extracellular signal-regulated kinase (ERK) 1/2 in LIF-mediated proliferation of HTR-8/SVneo cells. Stimulation of HTR-8/SVneo cells with LIF (50 ng mL(-1)) resulted in an increase in cell proliferation (P < 0.05) via increased transition of cells to the G(2)/M phase of cell cycle. Stimulation with LIF resulted in the activation of both signal transducer and activator of transcription (STAT) 3 Tyr(705) and ERK1/2, but inhibition of ERK1/2 signalling by pretreatment of cells with U0126 (10 µM) for 2h resulted in abrogation of LIF-mediated increases in G(2)/M transition, with a significant decrease (P < 0.05) in absolute cell numbers compared with control. Although STAT3 silencing had no effect on LIF-dependent proliferation of HTR-8/SVneo cells, it did result in an increase in cell apoptosis, which increased further upon inhibition of ERK1/2 activation irrespective of LIF stimulation. Stimulation of cells with LIF increased the Bcl-2/Bax ratio, whereas ERK1/2 inhibition decreased the Bcl-2/Bax ratio, even after LIF stimulation. Hence, it can be inferred that ERK1/2 activation is essential for LIF-mediated increases in proliferation and that both STAT3 and ERK1/2 activation are important for the survival of HTR-8/SVneo cells.
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Detection of circulating tumor cells in metastatic and clinically localized urothelial carcinoma.
Urology
PUBLISHED: 03-24-2011
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To examine the incidence and prognostic value of circulating tumor cells (CTCs) in urothelial cancer (UC). The detection of CTCs is prognostic in several cancer types.
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MicroRNAs in pregnancy.
J. Reprod. Immunol.
PUBLISHED: 01-12-2011
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Since the discovery of non-coding RNAs, several families of small regulatory molecules have been described including small nucleolar RNAs, piwi-interacting RNAs and microRNAs (miRNAs). MiRNAs are small single-stranded RNA molecules which play an important role in the regulation of gene expression at the transcriptional level. Recent studies demonstrated that about 30% of human genes are regulated by miRNAs and their deregulation has been associated with malignancies and poor outcome. Therefore, it is not surprising that profiling of miRNAs expression and studies on their regulation became a great field of interest in the last decade. However, miRNA-mediated regulation in pregnancy remains poorly investigated although several independent processes associated with placenta development have been shown to be miRNA-regulated. This review provides a general overview of the current data on profiles and functions of microRNAs in the peri-implantation period, embryonic stem cells, placentation and pregnancy, as well as in several pregnancy-related pathologies. We conclude that miRNAs present in the maternal circulation may provide a new promising diagnostic tool for pregnancy disorders.
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Antifungal mechanisms by which a novel Pseudomonas aeruginosa phenazine toxin kills Candida albicans in biofilms.
Mol. Microbiol.
PUBLISHED: 10-18-2010
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Pseudomonas aeruginosa produces several phenazines including the recently described 5-methyl-phenazine-1-carboxylic acid (5MPCA), which exhibits a novel antibiotic activity towards pathogenic fungi such as Candida albicans. Here we characterize the unique antifungal mechanisms of 5MPCA using its analogue phenazine methosulphate (PMS). Like 5MPCA, PMS induced fungal red pigmentation and killing. Mass spectrometry analyses demonstrated that PMS can be covalently modified by amino acids, a process that yields red derivatives. Furthermore, soluble proteins from C. albicans grown with either PMS or P. aeruginosa were also red and demonstrated absorbance and fluorescence spectra similar to that of PMS covalently linked to either amino acids or proteins in vitro, suggesting that 5MPCA modification by protein amine groups occurs in vivo. The red-pigmented C. albicans soluble proteins were reduced by NADH and spontaneously oxidized by oxygen, a reaction that likely generates reactive oxygen species (ROS). Additional evidence indicated that ROS generation precedes 5MPCA-induced fungal death. Reducing conditions greatly enhanced PMS uptake by C. albicans and killing. Since 5MPCA was more toxic than other phenazines that are not modified, such as pyocyanin, we propose that the covalent binding of 5MPCA promotes its accumulation in target cells and contributes to its antifungal activity in mixed-species biofilms.
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Candida albicans-produced farnesol stimulates Pseudomonas quinolone signal production in LasR-defective Pseudomonas aeruginosa strains.
Microbiology (Reading, Engl.)
PUBLISHED: 07-23-2010
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Candida albicans has been previously shown to stimulate the production of Pseudomonas aeruginosa phenazine toxins in dual-species colony biofilms. Here, we report that P. aeruginosa lasR mutants, which lack the master quorum sensing system regulator, regain the ability to produce quorum-sensing-regulated phenazines when cultured with C. albicans. Farnesol, a signalling molecule produced by C. albicans, was sufficient to stimulate phenazine production in LasR(-) laboratory strains and clinical isolates. P. aeruginosa ?lasR mutants are defective in production of the Pseudomonas quinolone signal (PQS) due to their inability to properly induce pqsH, which encodes the enzyme necessary for the last step in PQS biosynthesis. We show that expression of pqsH in a ?lasR strain was sufficient to restore PQS production, and that farnesol restored pqsH expression in ?lasR mutants. The farnesol-mediated increase in pqsH required RhlR, a transcriptional regulator downstream of LasR, and farnesol led to higher levels of N-butyryl-homoserine lactone, the small molecule activator of RhlR. Farnesol promotes the production of reactive oxygen species (ROS) in a variety of species. Because the antioxidant N-acetylcysteine suppressed farnesol-induced RhlR activity in LasR(-) strains, and hydrogen peroxide was sufficient to restore PQS production in las mutants, we propose that ROS are responsible for the activation of downstream portions of this quorum sensing pathway. LasR mutants frequently arise in the lungs of patients chronically infected with P. aeruginosa. The finding that C. albicans, farnesol or ROS stimulate virulence factor production in lasR strains provides new insight into the virulence potential of these strains.
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Specific control of Pseudomonas aeruginosa surface-associated behaviors by two c-di-GMP diguanylate cyclases.
MBio
PUBLISHED: 07-07-2010
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The signaling nucleotide cyclic diguanylate (c-di-GMP) regulates the transition between motile and sessile growth in a wide range of bacteria. Understanding how microbes control c-di-GMP metabolism to activate specific pathways is complicated by the apparent multifold redundancy of enzymes that synthesize and degrade this dinucleotide, and several models have been proposed to explain how bacteria coordinate the actions of these many enzymes. Here we report the identification of a diguanylate cyclase (DGC), RoeA, of Pseudomonas aeruginosa that promotes the production of extracellular polysaccharide (EPS) and contributes to biofilm formation, that is, the transition from planktonic to surface-dwelling cells. Our studies reveal that RoeA and the previously described DGC SadC make distinct contributions to biofilm formation, controlling polysaccharide production and flagellar motility, respectively. Measurement of total cellular levels of c-di-GMP in ?roeA and ?sadC mutants in two different genetic backgrounds revealed no correlation between levels of c-di-GMP and the observed phenotypic output with regard to swarming motility and EPS production. Our data strongly argue against a model wherein changes in total levels of c-di-GMP can account for the specific surface-related phenotypes of P. aeruginosa.
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[Acute bilateral lesion of the patellar tendon associated to diabetes mellitus. Case report].
Acta Ortop Mex
PUBLISHED: 04-10-2010
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The bilateral patellar tendon tear is very infrequent. Various pathological conditions may affect the extensor mechanism including kidney disease, diabetes mellitus, hyperparathyroidism, rheumatoid arthritis, systemic lupus erythematosus, osteomalacia, infection, obesity, corticosteroid therapy and even osteogenesis imperfect. We present the case of a 43-year-old male with a history of type-2 diabetes mellitus, systemic hypertension and exogenous obesity, who sustained a bilateral patellar tendon tear. He received surgical and medical treatment and was followed-up for one year, with satisfactory functional outcomes.
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[Development of a laminar bioadsorbent with hypertolerant Phanerochaete chrysosporium to Cd, Ni and Pb for wastewater treatment].
Rev Iberoam Micol
PUBLISHED: 01-15-2010
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The use of basidiomycetes for metal removal is an alternative to traditional methods. In this, the biomass acts as a natural ionic exchanger removing metals from solution.
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Translational spatial task and its relationship to HIV-associated neurocognitive disorders and apolipoprotein E in HIV-seropositive women.
J. Neurovirol.
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HIV-associated neurocognitive disorders (HAND) continue to be a neurological complication of HIV infection in the era of combined antiretroviral therapy. Hippocampal neurodegeneration and dysfunction occurs as a result of HIV infection, but few studies to date have assesses spatial learning and memory function in patients with HAND. We used the Memory Island (MI) test to study the effects of HIV infection, apolipoprotein E (ApoE) allele status, and cerebral spinal fluid (CSF) ApoE protein levels on spatial learning and memory in our cohort of Hispanic women. The MI test is a virtual reality-based computer program that tests spatial learning and memory and was designed to resemble the Morris Water Maze test of hippocampal function widely used in rodent studies. In the current study, HIV-seropositive women (n?=?20) and controls (n?=?16) were evaluated with neuropsychological (NP) tests, the MI test, ApoE, and CSF ApoE assays. On the MI, the HIV-seropositive group showed significant reduced learning and delayed memory performance compared with HIV-seronegative controls. When stratified by cognitive performance on NP tests, the HIV-seropositive, cognitively impaired group performed worse than HIV-seronegative controls in ability to learn and in the delayed memory trial. Interestingly, differences were observed in the results obtained by the NP tests and the MI test for ?4 carriers and noncarriers: NP tests showed effects of the ?4 allele in HIV-seronegative women but not HIV-seropositive ones, whereas the converse was true for the MI test. Our findings suggest that the MI test is sensitive in detecting spatial deficits in HIV-seropositive women and that these deficits may arise relatively early in the course of HAND.
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14-3-3s are potential biomarkers for HIV-related neurodegeneration.
J. Neurovirol.
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Over the last decade, it has become evident that 14-3-3 proteins are essential for primary cell functions. These proteins are abundant throughout the body, including the central nervous system and interact with other proteins in both cell cycle and apoptotic pathways. Examination of cerebral spinal fluid in humans suggests that 14-3-3s including 14-3-3? (YWHAE) are up-regulated in several neurological diseases, and loss or duplication of the YWHAE gene leads to Miller-Dieker syndrome. The goal of this review is to examine the utility of 14-3-3s as a marker of human immune deficiency virus (HIV)-dependent neurodegeneration and also as a tool to track disease progression. To that end, we describe mechanisms implicating 14-3-3s in neurological diseases and summarize evidence of its interactions with HIV accessory and co-receptor proteins.
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AP-1 transcription factors, mucin-type molecules and MMPs regulate the IL-11 mediated invasiveness of JEG-3 and HTR-8/SVneo trophoblastic cells.
PLoS ONE
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This study examines the IL-11 mediated activation of downstream signaling and expression of effector molecules to resolve the controversies associated with the IL-11 mediated regulation of the invasiveness of two commonly used trophoblastic cell models viz. JEG-3 and HTR-8/SVneo cells. It has been reported that IL-11 increases the invasiveness of JEG-3 cells while, reduces the invasiveness of HTR-8/SVneo cells. Invasion assay performed simultaneously for both the cell lines confirmed the above findings. In addition, HTR-8/SVneo cells showed a higher basal invasiveness than JEG-3 cells. Western blot showed the IL-11 mediated activation of STAT3(tyr705) and STAT1(tyr701) in both the cell lines. However, IL-11 activated the ERK1/2 phosphorylation in JEG-3 cells but, inhibited it in HTR-8/SVneo cells. Within 10 min of IL-11 treatment, p-STAT3(tyr705) was localized inside the nucleus of both the cell lines but, there was enhanced co-localization of protein inhibitor of activated STAT1/3 (PIAS1/3) and p-STAT3(tyr705) in HTR-8/SVneo cells and not in JEG-3 cells. This could be reason for the poor responsiveness of STAT3 responsive genes like mucin 1 (MUC1) in HTR-8/SVneo cells and not in JEG-3 cells. Further, microarray analysis of the IL-11 treated cells revealed differential responsiveness of JEG-3 as compared to HTR-8/SVneo cells. Several family of genes like activator protein-1 (AP-1) transcription factors (Jun and Fos), mucin-type molecules, MMP23B etc showed enhanced expression in IL-11 treated JEG-3 cells while, there was no response or decrease in their expression in IL-11 treated HTR-8/SVneo cells. Expression of these molecules was confirmed by quantitative RT-PCR. In addition, HTR-8/SVneo cells also showed a significant decrease in the expression of MMP2, MMP3 and MMP9 upon IL-11 treatment. Hence, IL-11 mediated differential activation of signaling and expression of effector molecules is responsible for the differential invasive response of JEG-3 and HTR-8/SVneo cells.
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JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.