Science.gov

Sample records for activity ectopic expression

  1. Normalisation to blood activity is required for the accurate quantification of Na/I symporter ectopic expression by SPECT/CT in individual subjects.

    PubMed

    Richard-Fiardo, Peggy; Franken, Philippe R; Lamit, Audrey; Marsault, Robert; Guglielmi, Julien; Cambien, Béatrice; Graslin, Fanny; Lindenthal, Sabine; Darcourt, Jacques; Pourcher, Thierry; Vassaux, Georges

    2012-01-01

    The utilisation of the Na/I symporter (NIS) and associated radiotracers as a reporter system for imaging gene expression is now reaching the clinical setting in cancer gene therapy applications. However, a formal assessment of the methodology in terms of normalisation of the data still remains to be performed, particularly in the context of the assessment of activities in individual subjects in longitudinal studies. In this context, we administered to mice a recombinant, replication-incompetent adenovirus encoding rat NIS, or a human colorectal carcinoma cell line (HT29) encoding mouse NIS. We used (99m)Tc pertechnetate as a radiotracer for SPECT/CT imaging to determine the pattern of ectopic NIS expression in longitudinal kinetic studies. Some animals of the cohort were culled and NIS expression was measured by quantitative RT-PCR and immunohistochemistry. The radioactive content of some liver biopsies was also measured ex vivo. Our results show that in longitudinal studies involving datasets taken from individual mice, the presentation of non-normalised data (activity expressed as %ID/g or %ID/cc) leads to 'noisy', and sometimes incoherent, results. This variability is due to the fact that the blood pertechnetate concentration can vary up to three-fold from day to day. Normalisation of these data with blood activities corrects for these inconsistencies. We advocate that, blood pertechnetate activity should be determined and used to normalise the activity measured in the organ/region of interest that expresses NIS ectopically. Considering that NIS imaging has already reached the clinical setting in the context of cancer gene therapy, this normalisation may be essential in order to obtain accurate and predictive information in future longitudinal clinical studies in biotherapy.

  2. Normalisation to Blood Activity Is Required for the Accurate Quantification of Na/I Symporter Ectopic Expression by SPECT/CT in Individual Subjects

    PubMed Central

    Richard-Fiardo, Peggy; Franken, Philippe R.; Lamit, Audrey; Marsault, Robert; Guglielmi, Julien; Cambien, Béatrice; Graslin, Fanny; Lindenthal, Sabine; Darcourt, Jacques; Pourcher, Thierry; Vassaux, Georges

    2012-01-01

    The utilisation of the Na/I symporter (NIS) and associated radiotracers as a reporter system for imaging gene expression is now reaching the clinical setting in cancer gene therapy applications. However, a formal assessment of the methodology in terms of normalisation of the data still remains to be performed, particularly in the context of the assessment of activities in individual subjects in longitudinal studies. In this context, we administered to mice a recombinant, replication-incompetent adenovirus encoding rat NIS, or a human colorectal carcinoma cell line (HT29) encoding mouse NIS. We used 99mTc pertechnetate as a radiotracer for SPECT/CT imaging to determine the pattern of ectopic NIS expression in longitudinal kinetic studies. Some animals of the cohort were culled and NIS expression was measured by quantitative RT-PCR and immunohistochemistry. The radioactive content of some liver biopsies was also measured ex vivo. Our results show that in longitudinal studies involving datasets taken from individual mice, the presentation of non-normalised data (activity expressed as %ID/g or %ID/cc) leads to ‘noisy’, and sometimes incoherent, results. This variability is due to the fact that the blood pertechnetate concentration can vary up to three-fold from day to day. Normalisation of these data with blood activities corrects for these inconsistencies. We advocate that, blood pertechnetate activity should be determined and used to normalise the activity measured in the organ/region of interest that expresses NIS ectopically. Considering that NIS imaging has already reached the clinical setting in the context of cancer gene therapy, this normalisation may be essential in order to obtain accurate and predictive information in future longitudinal clinical studies in biotherapy. PMID:22470517

  3. Drosophila enhancer-Gal4 lines show ectopic expression during development

    PubMed Central

    Arnés, Mercedes; Ferrús, Alberto

    2017-01-01

    In Drosophila melanogaster the most widely used technique to drive gene expression is the binary UAS/Gal4 system. We show here that a set of nervous system specific enhancers (elav, D42/Toll-6, OK6/RapGAP1) display ectopic activity in epithelial tissues during development, which is seldom considered in experimental studies. This ectopic activity is variable, unstable and influenced by the primary sequence of the enhancer and the insertion site in the chromosome. In addition, the ectopic activity is independent of the protein expressed, Gal4, as it is reproduced also with the expression of Gal80. Another enhancer, LN2 from the sex lethal (Sxl) gene, shows sex-dependent features in its ectopic expression. Feminization of LN2 expressing males does not alter the male specific pattern indicating that the sexual dimorphism of LN2 expression is an intrinsic feature of this enhancer. Other X chromosome enhancers corresponding to genes not related to sex determination do not show sexual dimorphism in their ectopic expressions. Although variable and unstable, the ectopic activation of enhancer-Gal4 lines seems to be regulated in terms of tissue and intensity. To characterize the full domain of expression of enhancer-Gal4 constructs is relevant for the design of transgenic animal models and biotechnology tools, as well as for the correct interpretation of developmental and behavioural studies in which Gal4 lines are used.

  4. dBRWD3 Regulates Tissue Overgrowth and Ectopic Gene Expression Caused by Polycomb Group Mutations

    PubMed Central

    Shih, Hsueh-Tzu; Chen, Wei-Yu; Liu, Kwei-Yan; Shih, Zong-Siou; Chen, Yi-Jyun; Hsieh, Paul-Chen; Kuo, Kuan-Lin; Huang, Kuo-How; Hsu, Pang-Hung; Liu, Ya-Wen; Tsai, Yu-Chen; Wu, June-Tai

    2016-01-01

    To maintain a particular cell fate, a unique set of genes should be expressed while another set is repressed. One way to repress gene expression is through Polycomb group (PcG) proteins that compact chromatin into a silent configuration. In addition to cell fate maintenance, PcG proteins also maintain normal cell physiology, for example cell cycle. In the absence of PcG, ectopic activation of the PcG-repressed genes leads to developmental defects and malignant tumors. Little is known about the molecular nature of ectopic gene expression; especially what differentiates expression of a given gene in the orthotopic tissue (orthotopic expression) and the ectopic expression of the same gene due to PcG mutations. Here we present that ectopic gene expression in PcG mutant cells specifically requires dBRWD3, a negative regulator of HIRA/Yemanuclein (YEM)-mediated histone variant H3.3 deposition. dBRWD3 mutations suppress both the ectopic gene expression and aberrant tissue overgrowth in PcG mutants through a YEM-dependent mechanism. Our findings identified dBRWD3 as a critical regulator that is uniquely required for ectopic gene expression and aberrant tissue overgrowth caused by PcG mutations. PMID:27588417

  5. Ectopic KNOX Expression Affects Plant Development by Altering Tissue Cell Polarity and Identity[OPEN

    PubMed Central

    Rebocho, Alexandra B.

    2016-01-01

    Plant development involves two polarity types: tissue cell (asymmetries within cells are coordinated across tissues) and regional (identities vary spatially across tissues) polarity. Both appear altered in the barley (Hordeum vulgare) Hooded mutant, in which ectopic expression of the KNOTTED1-like Homeobox (KNOX) gene, BKn3, causes inverted polarity of differentiated hairs and ectopic flowers, in addition to wing-shaped outgrowths. These lemma-specific effects allow the spatiotemporal analysis of events following ectopic BKn3 expression, determining the relationship between KNOXs, polarity, and shape. We show that tissue cell polarity, based on localization of the auxin transporter SISTER OF PINFORMED1 (SoPIN1), dynamically reorients as ectopic BKn3 expression increases. Concurrently, ectopic expression of the auxin importer LIKE AUX1 and boundary gene NO APICAL MERISTEM is activated. The polarity of hairs reflects SoPIN1 patterns, suggesting that tissue cell polarity underpins oriented cell differentiation. Wing cell files reveal an anisotropic growth pattern, and computational modeling shows how polarity guiding growth can account for this pattern and wing emergence. The inverted ectopic flower orientation does not correlate with SoPIN1, suggesting that this form of regional polarity is not controlled by tissue cell polarity. Overall, the results suggest that KNOXs trigger different morphogenetic effects through interplay between tissue cell polarity, identity, and growth. PMID:27553356

  6. Ectopic KNOX Expression Affects Plant Development by Altering Tissue Cell Polarity and Identity.

    PubMed

    Richardson, Annis Elizabeth; Rebocho, Alexandra B; Coen, Enrico S

    2016-08-23

    Plant development involves two polarity types: tissue cell (asymmetries within cells are coordinated across tissues) and regional (identities vary spatially across tissues) polarity. Both appear altered in the barley (Hordeum vulgare) Hooded mutant, in which ectopic expression of the KNOTTED1-like Homeobox (KNOX) gene, BKn3, causes inverted polarity of differentiated hairs and ectopic flowers, in addition to wing-shaped outgrowths. These lemma-specific effects allow the spatiotemporal analysis of events following ectopic BKn3 expression, determining the relationship between KNOXs, polarity, and shape. We show that tissue cell polarity, based on localization of the auxin transporter SISTER OF PINFORMED1 (SoPIN1), dynamically reorients as ectopic BKn3 expression increases. Concurrently, ectopic expression of the auxin importer LIKE AUX1 and boundary gene NO APICAL MERISTEM is activated. The polarity of hairs reflects SoPIN1 patterns, suggesting that tissue cell polarity underpins oriented cell differentiation. Wing cell files reveal an anisotropic growth pattern, and computational modeling shows how polarity guiding growth can account for this pattern and wing emergence. The inverted ectopic flower orientation does not correlate with SoPIN1, suggesting that this form of regional polarity is not controlled by tissue cell polarity. Overall, the results suggest that KNOXs trigger different morphogenetic effects through interplay between tissue cell polarity, identity, and growth.

  7. Ventricular ectopic activity and its relevance to aircrew licensing.

    PubMed

    Campbell, R W

    1984-03-01

    Ventricular ectopic complex ( VEC ) activity is seen frequently, both in patients with cardiovascular disease and in normal subjects. It has been claimed to be predictive of cardiovascular disease but in the asymptomatic individual it may be benign. Unifocal ventricular ectopic complexes with activity greater than 2 per 100 on the resting electrocardiogram of an applicant for a flying licence should be assessed. Further investigation including exercise electrocardiography and tape monitoring may be indicated. Even following investigation, empirically high counts may be acceptable only for multi-crew operation. Complex ventricular ectopic activity including multiformity, pairs, ventricular tachycardia, R-on-T ectopic beats, bigeminy and trigeminy, and parasystole, are incompatible with unrestricted certification and most are incompatible with any form of certification as individual risk-stratification is not yet possible. Of these, bigeminal arrhythmias are, however, likely to be the most benign, while R-on-T ectopic complexes and ventricular tachycardia are the most malignant.

  8. Ectopic DNMT3B expression delays leukemogenesis.

    PubMed

    Stanley, Robert F; Steidl, Ulrich

    2016-03-24

    In this issue of Blood, Schulze et al use a tetracycline-inducible Dnmt3b knock-in mouse model to investigate how DNMT3B-mediated DNA methylation affects leukemogenesis. Increased DNMT3B expression prolonged survival in retrovirally induced Myc-Bcl2– or MLL-AF9–driven leukemia, and acute myeloid leukemia (AML) patients with high expression of DNMT3B target genes showed inferior overall survival.

  9. Ectopic ERK Expression Induces Phenotypic Conversion of C10 Cells and Alters DNA Methyltransferase Expression

    SciTech Connect

    Sontag, Ryan L.; Weber, Thomas J.

    2012-05-04

    In some model systems constitutive extracellular signal regulated kinase (ERK) activation is sufficient to promote an oncogenic phenotype. Here we investigate whether constitutive ERK expression influences phenotypic conversion in murine C10 type II alveolar epithelial cells. C10 cells were stably transduced with an ERK1-green fluorescent protein (ERK1-GFP) chimera or empty vector and ectopic ERK expression was associated with the acquisition of soft agar focus-forming potential in late passage, but not early passage cells. Late passage ERK1-GFP cells exhibited a significant increase in the expression of DNA methyl transferases (DNMT1 and 3b) and a marked increase in sensitivity to 5-azacytidine (5-azaC)-mediated toxicity, relative to early passage ERK1-GFP cells and vector controls. The expression of xeroderma pigmentosum complementation group A (XPA) and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) were significantly increased in late passage cells, suggesting enhanced DNA damage recognition and repair activity which we interpret as a reflection of genomic instability. Phospho-ERK levels were dramatically decreased in late passage ERK1-GFP cells, relative to early passage and vector controls, and phospho-ERK levels were restored by treatment with sodium orthovanadate, indicating a role for phosphatase activity in this response. Collectively these observations suggest that ectopic ERK expression promotes phenotypic conversion of C10 cells that is associated with latent effects on epigenetic programming and phosphatase activities.

  10. Expression Profile of Ectopic Olfactory Receptors Determined by Deep Sequencing

    PubMed Central

    Flegel, Caroline; Manteniotis, Stavros; Osthold, Sandra; Hatt, Hanns; Gisselmann, Günter

    2013-01-01

    Olfactory receptors (ORs) provide the molecular basis for the detection of volatile odorant molecules by olfactory sensory neurons. The OR supergene family encodes G-protein coupled proteins that belong to the seven-transmembrane-domain receptor family. It was initially postulated that ORs are exclusively expressed in the olfactory epithelium. However, recent studies have demonstrated ectopic expression of some ORs in a variety of other tissues. In the present study, we conducted a comprehensive expression analysis of ORs using an extended panel of human tissues. This analysis made use of recent dramatic technical developments of the so-called Next Generation Sequencing (NGS) technique, which encouraged us to use open access data for the first comprehensive RNA-Seq expression analysis of ectopically expressed ORs in multiple human tissues. We analyzed mRNA-Seq data obtained by Illumina sequencing of 16 human tissues available from Illumina Body Map project 2.0 and from an additional study of OR expression in testis. At least some ORs were expressed in all the tissues analyzed. In several tissues, we could detect broadly expressed ORs such as OR2W3 and OR51E1. We also identified ORs that showed exclusive expression in one investigated tissue, such as OR4N4 in testis. For some ORs, the coding exon was found to be part of a transcript of upstream genes. In total, 111 of 400 OR genes were expressed with an FPKM (fragments per kilobase of exon per million fragments mapped) higher than 0.1 in at least one tissue. For several ORs, mRNA expression was verified by RT-PCR. Our results support the idea that ORs are broadly expressed in a variety of tissues and provide the basis for further functional studies. PMID:23405139

  11. Ectopic Expression of WUS in Hypocotyl Promotes Cell Division via GRP23 in Arabidopsis

    PubMed Central

    Wang, Min; Li, Junhua; Guo, Xiaoyu; Chong, Kang; Xu, Yunyuan

    2013-01-01

    WUSCHEL (WUS) is essential for preventing stem cell differentiation in Arabidopsis. Here we report that in addition to its functions in meristematic stem cell maintenance, WUS is involved in the regulation of cell division. The WUS gain-of-function mutant, stem ectopic flowers (sef), displayed elongated hypocotyls, whereas the loss-of-function wus-1 mutant had shortened hypocotyls. The long hypocotyl in sef was due to the presence of more cells, rather than increased cell elongation. Microscopic observation, flow cytometry assays, quantitative RT-PCR (qRT-PCR), and histochemical staining of CycB1;1::GUS supported the hypothesis that ectopic cell division occurred in the sef hypocotyls after germination. Both immunoblot and qRT-PCR results showed that WUS was ectopically expressed in sef hypocotyls. Luciferase activity, chromatin immunoprecipitation (ChIP) and electrophoretic mobility shift assay (EMSA) showed that GLUTAMINE-RICH PROTEIN 23 (GRP23) expression can be activated by WUS and that GRP23 is a direct target gene of WUS. The phenotypes of 35S::GRP23 plants and GRP23 knockdown lines supported the notion that GRP23 mediates the effects of WUS on hypocotyl length. Together, our data suggest that ectopic expression of WUS in hypocotyl controls cell division through its target gene GRP23. PMID:24086632

  12. Ectopic expression of SUPERMAN suppresses development of petals and stamens.

    PubMed

    Yun, Jae-Young; Weigel, Detlef; Lee, Ilha

    2002-01-01

    The floral regulatory gene SUPERMAN (SUP) encodes a C2H2 type zinc finger protein that is required for maintaining boundaries between floral organs in Arabidopsis. It has been proposed that the main function of SUP is to balance cell proliferation in the third and fourth whorl of developing flowers, thereby maintaining the boundaries between the two whorls. To gain further insight into the function of SUP, we have ectopically expressed SUP using the promoter of APETALA1 (AP1), a gene that is initially expressed throughout floral meristems and later becomes restricted to the first and second whorls. Flowers of AP1::SUP plants have fewer floral organs, consistent with an effect of SUP on cell proliferation. In addition, the AP1::SUP transgene caused the conversion of petals to sepals and suppressed the development of stamens. The expression of the B function homeotic gene APETALA3 (AP3) and its regulator UNUSUAL FLORAL ORGANS (UFO) were delayed and reduced in AP1::SUP flowers. However, SUP does not act merely through UFO, as constitutive expression of UFO did not rescue the defects in petal and stamen development in AP1::SUP flowers. Together, these results suggest that SUP has both indirect and direct effects on the expression of B function homeotic genes.

  13. Ectopic Six3 expression in the dragon eye goldfish.

    PubMed

    Ma, Dong-Mei; Zhu, Hua-Ping; Gui, Jian-Fang

    2008-02-01

    For goldfish (Carassius auratus), there are many varieties with different eye phenotypes due to artificial selection and adaptive evolution. Dragon eye is a variant eye characterized by a large-size eyeball protruding out of the socket similar to the eye of dragon in Chinese legends. In this study, anatomical structure of the goldfish dragon eye was compared with that of the common eye, and a stretching of the retina was observed in the enlarged dragon eye. Moreover, the homeobox-containing transcription factor Six3 cDNAs were cloned from the two types of goldfish, and the expression patterns were analyzed in both normal eye and dragon eye goldfish. No amino acid sequence differences were observed between the two deduced peptides, and the expression pattern of Six3 protein in dragon eye is quite similar to common eye during embryogenesis, but from 2 days after hatching, ectopic Six3 expression began to occur in the dragon eye, especially in the outer nuclear layer cells. With eye development, more predominant Six3 distribution was detected in the outer nuclear layer cells of dragon eye than that of normal eye, and fewer cell-layers in outer nuclear layer were observed in dragon eye retina than in normal eye retina. The highlight of this study is that higher Six3 expression occurs in dragon eye goldfish than in normal eye goldfish during retinal development of larvae.

  14. Ectopic AP4 expression induces cellular senescence via activation of p53 in long-term confluent retinal pigment epithelial cells.

    PubMed

    Wang, Yiping; Wong, Matthew Man-Kin; Zhang, Xiaojian; Chiu, Sung-Kay

    2015-11-15

    When cells are grown to confluence, cell-cell contact inhibition occurs and drives the cells to enter reversible quiescence rather than senescence. Confluent retinal pigment epithelial (RPE) cells exhibiting contact inhibition was used as a model in this study to examine the role of overexpression of transcription factor AP4, a highly expressed transcription factor in many types of cancer, in these cells during long-term culture. We generated stable inducible RPE cell clones expressing AP4 or AP4 without the DNA binding domain (DN-AP4) and observed that, when cultured for 24 days, RPE cells with a high level of AP4 exhibit a large, flattened morphology and even cease proliferating; these changes were not observed in DN-AP4-expressing cells or non-induced cells. In addition, AP4-expressing cells exhibited senescence-associated β-galactosidase activity and the senescence-associated secretory phenotype. We demonstrated that the induced cellular senescence was mediated by enhanced p53 expression and that AP4 regulates the p53 gene by binding directly to two of the three E-boxes present on the promoter of the p53 gene. Moreover, we showed that serum is essential for AP4 in inducing p53-associated cellular senescence. Collectively, we showed that overexpression of AP4 mediates cellular senescence involving in activation of p53 in long-term post-confluent RPE cells.

  15. Engineering secondary metabolism in maize cells by ectopic expression of transcription factors

    PubMed Central

    Grotewold, E; Chamberlin, M; Snook, M; Siame, B; Butler, L; Swenson, J; Maddock, S; Clair, GS; Bowen, B

    1998-01-01

    Manipulation of plant natural product biosynthesis through genetic engineering is an attractive but technically challenging goal. Here, we demonstrate that different secondary metabolites can be produced in cultured maize cells by ectopic expression of the appropriate regulatory genes. Cell lines engineered to express the maize transcriptional activators C1 and R accumulate two cyanidin derivatives, which are similar to the predominant anthocyanin found in differentiated plant tissues. In contrast, cell lines that express P accumulate various 3-deoxy flavonoids. Unexpectedly, P-expressing cells in culture also accumulate phenylpropanoids and green fluorescent compounds that are targeted to different subcellular compartments. Two endogenous biosynthetic genes (c2 and a1, encoding chalcone synthase and flavanone/dihydroflavonol reductase, respectively) are independently activated by ectopic expression of either P or C1/R, and there is a dose-response relationship between the transcript level of P and the degree to which c2 or a1 is expressed. Our results support a simple model showing how the gene encoding P may act as a quantitative trait locus controlling insecticidal C-glycosyl flavone level in maize silks, and they suggest how p1 might confer a selective advantage against insect predation in maize. PMID:9596632

  16. Ectopically Expressed Pro-group X Secretory Phospholipase A2 Is Proteolytically Activated in Mouse Adrenal Cells by Furin-like Proprotein Convertases

    PubMed Central

    Layne, Joseph D.; Shridas, Preetha; Webb, Nancy R.

    2015-01-01

    Group X secretory phospholipase A2 (GX sPLA2) hydrolyzes mammalian cell membranes, liberating free fatty acids and lysophospholipids. GX sPLA2 is produced as a pro-enzyme (pro-GX sPLA2) that contains an N-terminal 11-amino acid propeptide ending in a dibasic motif, suggesting cleavage by a furin-like proprotein convertase (PC). Although propeptide cleavage is clearly required for enzymatic activity, the protease(s) responsible for pro-GX sPLA2 activation have not been identified. We previously reported that GX sPLA2 negatively regulates adrenal glucocorticoid production, likely by suppressing liver X receptor-mediated activation of steroidogenic acute regulatory protein expression. In this study, using a FLAG epitope-tagged pro-GX sPLA2 expression construct (FLAG-pro-GX sPLA2), we determined that adrenocorticotropic hormone (ACTH) enhanced FLAG-pro-GX sPLA2 processing and phospholipase activity secreted by Y1 adrenal cells. ACTH increased the expression of furin and PCSK6, but not other members of the PC family, in Y1 cells. Overexpression of furin and PCSK6 in HEK 293 cells significantly enhanced FLAG-pro-GX sPLA2 processing, whereas siRNA-mediated knockdown of both PCs almost completely abolished FLAG-pro-GX sPLA2 processing in Y1 cells. Expression of either furin or PCSK6 enhanced the ability of GX sPLA2 to suppress liver X receptor reporter activity. The PC inhibitor decanoyl-Arg-Val-Lys-Arg-chloromethyl ketone significantly suppressed FLAG-pro-GX sPLA2 processing and sPLA2 activity in Y1 cells, and it significantly attenuated GX sPLA2-dependent inhibition of steroidogenic acute regulatory protein expression and progesterone production. These findings provide strong evidence that pro-GX sPLA2 is a substrate for furin and PCSK6 proteolytic processing and define a novel mechanism for regulating corticosteroid production in adrenal cells. PMID:25623068

  17. Ectopic Atoh1 expression drives Merkel cell production in embryonic, postnatal and adult mouse epidermis

    PubMed Central

    Ostrowski, Stephen M.; Wright, Margaret C.; Bolock, Alexa M.; Geng, Xuehui; Maricich, Stephen M.

    2015-01-01

    Merkel cells are mechanosensitive skin cells whose production requires the basic helix-loop-helix transcription factor Atoh1. We induced ectopic Atoh1 expression in the skin of transgenic mice to determine whether Atoh1 was sufficient to create additional Merkel cells. In embryos, ectopic Atoh1 expression drove ectopic expression of the Merkel cell marker keratin 8 (K8) throughout the epidermis. Epidermal Atoh1 induction in adolescent mice similarly drove widespread K8 expression in glabrous skin of the paws, but in the whisker pads and body skin ectopic K8+ cells were confined to hair follicles and absent from interfollicular regions. Ectopic K8+ cells acquired several characteristics of mature Merkel cells in a time frame similar to that seen during postnatal development of normal Merkel cells. Although ectopic K8+ cell numbers decreased over time, small numbers of these cells remained in deep regions of body skin hair follicles at 3 months post-induction. In adult mice, greater numbers of ectopic K8+ cells were created by Atoh1 induction during anagen versus telogen and following disruption of Notch signaling by conditional deletion of Rbpj in the epidermis. Our data demonstrate that Atoh1 expression is sufficient to produce new Merkel cells in the epidermis, that epidermal cell competency to respond to Atoh1 varies by skin location, developmental age and hair cycle stage, and that the Notch pathway plays a key role in limiting epidermal cell competency to respond to Atoh1 expression. PMID:26138479

  18. Ectopic Atoh1 expression drives Merkel cell production in embryonic, postnatal and adult mouse epidermis.

    PubMed

    Ostrowski, Stephen M; Wright, Margaret C; Bolock, Alexa M; Geng, Xuehui; Maricich, Stephen M

    2015-07-15

    Merkel cells are mechanosensitive skin cells whose production requires the basic helix-loop-helix transcription factor Atoh1. We induced ectopic Atoh1 expression in the skin of transgenic mice to determine whether Atoh1 was sufficient to create additional Merkel cells. In embryos, ectopic Atoh1 expression drove ectopic expression of the Merkel cell marker keratin 8 (K8) throughout the epidermis. Epidermal Atoh1 induction in adolescent mice similarly drove widespread K8 expression in glabrous skin of the paws, but in the whisker pads and body skin ectopic K8+ cells were confined to hair follicles and absent from interfollicular regions. Ectopic K8+ cells acquired several characteristics of mature Merkel cells in a time frame similar to that seen during postnatal development of normal Merkel cells. Although ectopic K8+ cell numbers decreased over time, small numbers of these cells remained in deep regions of body skin hair follicles at 3 months post-induction. In adult mice, greater numbers of ectopic K8+ cells were created by Atoh1 induction during anagen versus telogen and following disruption of Notch signaling by conditional deletion of Rbpj in the epidermis. Our data demonstrate that Atoh1 expression is sufficient to produce new Merkel cells in the epidermis, that epidermal cell competency to respond to Atoh1 varies by skin location, developmental age and hair cycle stage, and that the Notch pathway plays a key role in limiting epidermal cell competency to respond to Atoh1 expression.

  19. Leptin responsiveness in mice that ectopically express agouti protein.

    PubMed

    Harris, Ruth B S; Mitchell, Tiffany D; Mynatt, Randall L

    Agouti protein is an endogenous antagonist of melanocortin receptors (MCR), including MCR3 and MCR4, which have been implicated as part of the hypothalamic mechanism that mediates leptin-induced hypophagia. In this experiment we examined the effects of peripheral and central leptin administration in male and female beta-actin promoter (BAPa) mice that express agouti protein ectopically and have a phenotype that includes obesity and diabetes which is exaggerated in males compared with females. Intraperitoneal infusion of 10 microg leptin/day for 13 days caused weight loss and a transient inhibition of food intake in wild-type mice, with a greater effect in males than females. Male BAPa mice were resistant to leptin infusion whereas female mice lost weight. All of the mice lost body weight following a single intracerebroventricular injection of leptin but the effect was greater in female BAPa mice than any other group. There also was a delayed suppression of food intake that was the same for wild-type and BAPa female mice, whereas food intake recovered faster in BAPa than wild-type males. The dissociation between food intake and body weight loss implies a significant effect of leptin on energy expenditure in BAPa mice. These results demonstrate that the effect of leptin on energy balance is not entirely dependent upon the melanocortin system.

  20. Genome-wide expressions in autologous eutopic and ectopic endometrium of fertile women with endometriosis

    PubMed Central

    2012-01-01

    Background In order to obtain a lead of the pathophysiology of endometriosis, genome-wide expressional analyses of eutopic and ectopic endometrium have earlier been reported, however, the effects of stages of severity and phases of menstrual cycle on expressional profiles have not been examined. The effect of genetic heterogeneity and fertility history on transcriptional activity was also not considered. In the present study, a genome-wide expression analysis of autologous, paired eutopic and ectopic endometrial samples obtained from fertile women (n = 18) suffering from moderate (stage 3; n = 8) or severe (stage 4; n = 10) ovarian endometriosis during proliferative (n = 13) and secretory (n = 5) phases of menstrual cycle was performed. Methods Individual pure RNA samples were subjected to Agilent’s Whole Human Genome 44K microarray experiments. Microarray data were validated (P < 0.01) by estimating transcript copy numbers by performing real time RT-PCR of seven (7) arbitrarily selected genes in all samples. The data obtained were subjected to differential expression (DE) and differential co-expression (DC) analyses followed by networks and enrichment analysis, and gene set enrichment analysis (GSEA). The reproducibility of prediction based on GSEA implementation of DC results was assessed by examining the relative expressions of twenty eight (28) selected genes in RNA samples obtained from fresh pool of eutopic and ectopic samples from confirmed ovarian endometriosis patients with stages 3 and 4 (n = 4/each) during proliferative and secretory (n = 4/each) phases. Results Higher clustering effect of pairing (cluster distance, cd = 0.1) in samples from same individuals on expressional arrays among eutopic and ectopic samples was observed as compared to that of clinical stages of severity (cd = 0.5) and phases of menstrual cycle (cd = 0.6). Post hoc analysis revealed anomaly in the expressional profiles of several genes

  1. Ectopic Expression of Testis Germ Cell Proteins in Cancer and Its Potential Role in Genomic Instability

    PubMed Central

    Nielsen, Aaraby Yoheswaran; Gjerstorff, Morten Frier

    2016-01-01

    Genomic instability is a hallmark of human cancer and an enabling factor for the genetic alterations that drive cancer development. The processes involved in genomic instability resemble those of meiosis, where genetic material is interchanged between homologous chromosomes. In most types of human cancer, epigenetic changes, including hypomethylation of gene promoters, lead to the ectopic expression of a large number of proteins normally restricted to the germ cells of the testis. Due to the similarities between meiosis and genomic instability, it has been proposed that activation of meiotic programs may drive genomic instability in cancer cells. Some germ cell proteins with ectopic expression in cancer cells indeed seem to promote genomic instability, while others reduce polyploidy and maintain mitotic fidelity. Furthermore, oncogenic germ cell proteins may indirectly contribute to genomic instability through induction of replication stress, similar to classic oncogenes. Thus, current evidence suggests that testis germ cell proteins are implicated in cancer development by regulating genomic instability during tumorigenesis, and these proteins therefore represent promising targets for novel therapeutic strategies. PMID:27275820

  2. Ectopic expression of a soybean phytase in developing seeds of Glycine max to improve phosphorus availability.

    PubMed

    Chiera, Joseph M; Finer, John J; Grabau, Elizabeth A

    2004-12-01

    A transgenic approach was used to alter soybean seed phytate content by expressing a soybean phytase gene (GmPhy) during seed development to degrade accumulating phytic acid (IP6). An expression vector containing the soybean phytase cDNA controlled by the seed-specific beta-conglycinin promoter (alpha'-subunit) was used to transform embryogenic soybean cultures. Plants from four independent transgenic lines were analyzed for transgene integration and seed IP6 levels. The reduction in IP6 levels in transgenic seeds compared to control 'Jack' soybeans ranged from 12.6 to 24.8 as determined by HPLC. A low copy transformant was propagated to the T4 generation and examined in more detail for phytase expression and enzyme activity during seed development. Expression of phytase mRNA and phytase activity increased during seed development, consistent with the use of an embryo-specific promoter. Ectopic phytase expression during seed development offers potential as an effective strategy for reducing phytate content in soybean seed.

  3. Characterization of ectopically-expressed ets1 in human colon-cancer cells - induction of putative ets1-target gene.

    PubMed

    Suzuki, H; Romanospica, V; Georgiou, P; Fisher, R; Papas, T; Bhat, N

    1993-10-01

    We have previously shown that the ETS1 gene is expressed in a tissue-specific manner, and encodes a transcription factor, that may be involved in lymphocyte development, activation and proliferation. To understand the ETS1 function in non-lymphoid cells, we have ectopically expressed ETS1 protein in a human colon cancer cell line, and studied its biochemical properties. The 51 kDa ETS1 protein expressed in transfected cells localized in both the nucleus and the cytoplasm, has similar biochemical properties compared to ETS1 protein expressed in lymphoid cells. The ectopically expressed ETS1 binds to the DNA in sequence-specific manner and the binding activity is affected by the flanking sequences outside the 'GGA' core. Our results also demonstrate that the DNA-binding activity of full-length ETS1 is similar in lymphoid and non-lymphoid cells. The ETS1 expressed in DLD-1 cells is biologically active since it induces a 54.5 kDa polypeptide, whose expression level correlates with the expression of ETS1 in DLD-1 cells.

  4. Ectopic expression of Cripto-1 in transgenic mouse embryos causes hemorrhages, fatal cardiac defects and embryonic lethality

    PubMed Central

    Lin, Xiaolin; Zhao, Wentao; Jia, Junshuang; Lin, Taoyan; Xiao, Gaofang; Wang, Shengchun; Lin, Xia; Liu, Yu; Chen, Li; Qin, Yujuan; Li, Jing; Zhang, Tingting; Hao, Weichao; Chen, Bangzhu; Xie, Raoying; Cheng, Yushuang; Xu, Kang; Yao, Kaitai; Huang, Wenhua; Xiao, Dong; Sun, Yan

    2016-01-01

    Targeted disruption of Cripto-1 in mice caused embryonic lethality at E7.5, whereas we unexpectedly found that ectopic Cripto-1 expression in mouse embryos also led to embryonic lethality, which prompted us to characterize the causes and mechanisms underlying embryonic death due to ectopic Cripto-1 expression. RCLG/EIIa-Cre embryos displayed complex phenotypes between embryonic day 14.5 (E14.5) and E17.5, including fatal hemorrhages (E14.5-E15.5), embryo resorption (E14.5-E17.5), pale body surface (E14.5-E16.5) and no abnormal appearance (E14.5-E16.5). Macroscopic and histological examination revealed that ectopic expression of Cripto-1 transgene in RCLG/EIIa-Cre embryos resulted in lethal cardiac defects, as evidenced by cardiac malformations, myocardial thinning, failed assembly of striated myofibrils and lack of heartbeat. In addition, Cripto-1 transgene activation beginning after E8.5 also caused the aforementioned lethal cardiac defects in mouse embryos. Furthermore, ectopic Cripto-1 expression in embryonic hearts reduced the expression of cardiac transcription factors, which is at least partially responsible for the aforementioned lethal cardiac defects. Our results suggest that hemorrhages and cardiac abnormalities are two important lethal factors in Cripto-1 transgenic mice. Taken together, these findings are the first to demonstrate that sustained Cripto-1 transgene expression after E11.5 causes fatal hemorrhages and lethal cardiac defects, leading to embryonic death at E14.5-17.5. PMID:27687577

  5. Ectopic Activation of Germline and Placental Genes Identifies Aggressive Metastasis-Prone Lung Cancers

    PubMed Central

    Rousseaux, Sophie; Debernardi, Alexandra; Jacquiau, Baptiste; Vitte, Anne-Laure; Vesin, Aurélien; Nagy-Mignotte, Hélène; Moro-Sibilot, Denis; Brichon, Pierre-Yves; Lantuejoul, Sylvie; Hainaut, Pierre; Laffaire, Julien; de Reyniès, Aurélien; Beer, David G.; Timsit, Jean-François; Brambilla, Christian; Brambilla, Elisabeth; Khochbin, Saadi

    2016-01-01

    Activation of normally silent tissue-specific genes and the resulting cell “identity crisis” are the unexplored consequences of malignant epigenetic reprogramming. We designed a strategy for investigating this reprogramming, which consisted of identifying a large number of tissue-restricted genes that are epigenetically silenced in normal somatic cells and then detecting their expression in cancer. This approach led to the demonstration that large-scale “off-context” gene activations systematically occur in a variety of cancer types. In our series of 293 lung tumors, we identified an ectopic gene expression signature associated with a subset of highly aggressive tumors, which predicted poor prognosis independently of the TNM (tumor size, node positivity, and metastasis) stage or histological subtype. The ability to isolate these tumors allowed us to reveal their common molecular features characterized by the acquisition of embryonic stem cell/germ cell gene expression profiles and the down-regulation of immune response genes. The methodical recognition of ectopic gene activations in cancer cells could serve as a basis for gene signature–guided tumor stratification, as well as for the discovery of oncogenic mechanisms, and expand the understanding of the biology of very aggressive tumors. PMID:23698379

  6. Ectopic Expression of a Neospora caninum Kazal Type Inhibitor Triggers Developmental Defects in Toxoplasma and Plasmodium

    PubMed Central

    Goulielmaki, Evi; Kaforou, Sofia; Kim, Kami; Waters, Andrew P.; Carruthers, Vern B.; Siden-Kiamos, Inga; Koussis, Konstantinos

    2015-01-01

    Regulated proteolysis is known to control a variety of vital processes in apicomplexan parasites including invasion and egress of host cells. Serine proteases have been proposed as targets for drug development based upon inhibitor studies that show parasite attenuation and transmission blockage. Genetic studies suggest that serine proteases, such as subtilisin and rhomboid proteases, are essential but functional studies have proved challenging as active proteases are difficult to express. Proteinaceous Protease Inhibitors (PPIs) provide an alternative way to address the role of serine proteases in apicomplexan biology. To validate such an approach, a Neospora caninum Kazal inhibitor (NcPI-S) was expressed ectopically in two apicomplexan species, Toxoplasma gondii tachyzoites and Plasmodium berghei ookinetes, with the aim to disrupt proteolytic processes taking place within the secretory pathway. NcPI-S negatively affected proliferation of Toxoplasma tachyzoites, while it had no effect on invasion and egress. Expression of the inhibitor in P. berghei zygotes blocked their development into mature and invasive ookinetes. Moreover, ultra-structural studies indicated that expression of NcPI-S interfered with normal formation of micronemes, which was also confirmed by the lack of expression of the micronemal protein SOAP in these parasites. Our results suggest that NcPI-S could be a useful tool to investigate the function of proteases in processes fundamental for parasite survival, contributing to the effort to identify targets for parasite attenuation and transmission blockage. PMID:25803874

  7. Ectopic expression of a Neospora caninum Kazal type inhibitor triggers developmental defects in Toxoplasma and Plasmodium.

    PubMed

    Tampaki, Zoi; Mwakubambanya, Ramadhan S; Goulielmaki, Evi; Kaforou, Sofia; Kim, Kami; Waters, Andrew P; Carruthers, Vern B; Siden-Kiamos, Inga; Loukeris, Thanasis G; Koussis, Konstantinos

    2015-01-01

    Regulated proteolysis is known to control a variety of vital processes in apicomplexan parasites including invasion and egress of host cells. Serine proteases have been proposed as targets for drug development based upon inhibitor studies that show parasite attenuation and transmission blockage. Genetic studies suggest that serine proteases, such as subtilisin and rhomboid proteases, are essential but functional studies have proved challenging as active proteases are difficult to express. Proteinaceous Protease Inhibitors (PPIs) provide an alternative way to address the role of serine proteases in apicomplexan biology. To validate such an approach, a Neospora caninum Kazal inhibitor (NcPI-S) was expressed ectopically in two apicomplexan species, Toxoplasma gondii tachyzoites and Plasmodium berghei ookinetes, with the aim to disrupt proteolytic processes taking place within the secretory pathway. NcPI-S negatively affected proliferation of Toxoplasma tachyzoites, while it had no effect on invasion and egress. Expression of the inhibitor in P. berghei zygotes blocked their development into mature and invasive ookinetes. Moreover, ultra-structural studies indicated that expression of NcPI-S interfered with normal formation of micronemes, which was also confirmed by the lack of expression of the micronemal protein SOAP in these parasites. Our results suggest that NcPI-S could be a useful tool to investigate the function of proteases in processes fundamental for parasite survival, contributing to the effort to identify targets for parasite attenuation and transmission blockage.

  8. Ectopic expression of C/EBPalpha in the lung epithelium disrupts late lung development.

    PubMed

    Berg, Tove; Didon, Lukas; Nord, Magnus

    2006-10-01

    The lung develops from the endoderm through a process of branching morphogenesis. This process is highly active during the pseudoglandular stage of lung development and continues into the canalicular stage, resulting in the formation of terminal sacs. CCAAT/enhancer binding proteins (C/EBPs) are transcription factors regulating central aspects of differentiation and proliferation. We report here the developmental expression of C/EBPalpha, -beta, and -delta in the lung. C/EBPalpha exhibits a dynamic expression pattern and is first detected during the late pseudoglandular stage. At this stage, expression is observed in a subset of epithelial cells in the distal parts of the branching tubules. The expression of C/EBPalpha is confined to nonproliferating cells. To examine the role of C/EBPalpha in lung development, we generated transgenic mice ectopically expressing C/EBPalpha in the lung epithelium using the human surfactant protein C promoter. Lungs from these mice were of normal size but exhibited a phenotype characterized by fewer and larger developing epithelial tubules, indicating that the branching process was affected. No effects on overall proliferation or cellular differentiation were observed. When this phenotype was compared with that of mice carrying a targeted mutation of the Cebpa gene, the Cebpa-/- mice exhibited a similar developmental phenotype. In conclusion, our results show a role for C/EBPalpha in lung development and suggest a function in the later stages of lung branching morphogenesis.

  9. Ectopic expression of new alternative splice variant of Smac/DIABLO increases mammospheres formation

    PubMed Central

    Martinez-Ruiz, Gustavo U; Victoria-Acosta, Georgina; Vazquez-Santillan, Karla I; Jimenez-Hernandez, Luis; Muñoz-Galindo, Laura; Ceballos-Cancino, Gisela; Maldonado, Vilma; Melendez-Zajgla, Jorge

    2014-01-01

    Smac-α is a mitochondrial protein that, during apoptosis, is translocated to the cytoplasm, where it negatively regulates members of the inhibitor of apoptosis (IAP) family via the IAP-binding motif (IBM) contained within its amino-terminus. Here, we describe a new alternative splice variant from Smac gene, which we have named Smac-ε. Smac-ε lacks both an IBM and a mitochondrial-targeting signal (MTS) element. Smac-ε mRNA exhibits a tissue-specific expression pattern in healthy human tissues as well as in several cancer cell lines. The steady-state levels of endogenous Smac-ε protein is regulated by the proteasomal pathway. When ectopically expressed, this isoform presents a cytosolic localization and is unable to associate with or to regulate the expression of X-linked Inhibitor of apoptosis protein, the best-studied member of IAP family. Nevertheless, over-expression of Smac-ε increases mammosphere formation. Whole genome expression analyses from these mammospheres show activation of several pro-survival and growth pathways, including Estrogen-Receptor signaling. In conclusion, our results support the functionality of this new Smac isoform. PMID:25337193

  10. Ectopic expression of new alternative splice variant of Smac/DIABLO increases mammospheres formation.

    PubMed

    Martinez-Ruiz, Gustavo U; Victoria-Acosta, Georgina; Vazquez-Santillan, Karla I; Jimenez-Hernandez, Luis; Muñoz-Galindo, Laura; Ceballos-Cancino, Gisela; Maldonado, Vilma; Melendez-Zajgla, Jorge

    2014-01-01

    Smac-α is a mitochondrial protein that, during apoptosis, is translocated to the cytoplasm, where it negatively regulates members of the inhibitor of apoptosis (IAP) family via the IAP-binding motif (IBM) contained within its amino-terminus. Here, we describe a new alternative splice variant from Smac gene, which we have named Smac-ε. Smac-ε lacks both an IBM and a mitochondrial-targeting signal (MTS) element. Smac-ε mRNA exhibits a tissue-specific expression pattern in healthy human tissues as well as in several cancer cell lines. The steady-state levels of endogenous Smac-ε protein is regulated by the proteasomal pathway. When ectopically expressed, this isoform presents a cytosolic localization and is unable to associate with or to regulate the expression of X-linked Inhibitor of apoptosis protein, the best-studied member of IAP family. Nevertheless, over-expression of Smac-ε increases mammosphere formation. Whole genome expression analyses from these mammospheres show activation of several pro-survival and growth pathways, including Estrogen-Receptor signaling. In conclusion, our results support the functionality of this new Smac isoform.

  11. Peritoneal ectopic lesions from women with endometriosis show abnormalities in progesterone-dependent glycan expression.

    PubMed

    Jones, Carolyn J P; Nardo, Luciano G; Litta, Pietro; Fazleabas, Asgerally T

    2009-04-01

    Examination of 12 paired peritoneal ectopic and eutopic endometria for histochemical binding of Dolichos biflorus agglutinin, normally found in the mid-late secretory part of the cycle, showed a failure of lectin binding in 9 of 11 secretory-phase lesions although the eutopic specimens generally stained normally. This failure of glycan expression in the secretory phase may result from various anomalies, including an inability to respond to progesterone, possibly due to a lack of, or to nonfunctional, progesterone receptors, suggesting that an ectopic environment may produce changes in tissue cell biology and hormonal responsiveness compared with that of eutopic endometrium.

  12. Ectopic Expression of WRINKLED1 Affects Fatty Acid Homeostasis in Brachypodium distachyon Vegetative Tissues1[OPEN

    PubMed Central

    Yang, Yang; Munz, Jacob; Cass, Cynthia; Zienkiewicz, Agnieszka; Kong, Que; Ma, Wei; Sedbrook, John; Benning, Christoph

    2015-01-01

    Triacylglycerol (TAG) is a storage lipid used for food purposes and as a renewable feedstock for biodiesel production. WRINKLED1 (WRI1) is a transcription factor that governs fatty acid (FA) synthesis and, indirectly, TAG accumulation in oil-storing plant tissues, and its ectopic expression has led to TAG accumulation in vegetative tissues of different dicotyledonous plants. The ectopic expression of BdWRI1 in the grass Brachypodium distachyon induced the transcription of predicted genes involved in glycolysis and FA biosynthesis, and TAG content was increased up to 32.5-fold in 8-week-old leaf blades. However, the ectopic expression of BdWRI1 also caused cell death in leaves, which has not been observed previously in dicotyledonous plants such as Arabidopsis (Arabidopsis thaliana). Lipid analysis indicated that the free FA content was 2-fold elevated in BdWRI1-expressing leaf blades of B. distachyon. The transcription of predicted genes involved in β-oxidation was induced. In addition, linoleic FA treatment caused cell death in B. distachyon leaf blades, an effect that was reversed by the addition of the FA biosynthesis inhibitor cerulenin. Taken together, ectopic expression of BdWRI1 in B. distachyon enhances FA biosynthesis and TAG accumulation in leaves, as expected, but also leads to increased free FA content, which has cytotoxic effects leading to cell death. Thus, while WRI appears to ubiquitously affect FA biosynthesis and TAG accumulation in diverse plants, its ectopic expression can lead to undesired side effects depending on the context of the specific lipid metabolism of the respective plant species. PMID:26419778

  13. Ectopic Expression of WRINKLED1 Affects Fatty Acid Homeostasis in Brachypodium distachyon Vegetative Tissues.

    PubMed

    Yang, Yang; Munz, Jacob; Cass, Cynthia; Zienkiewicz, Agnieszka; Kong, Que; Ma, Wei; Sedbrook, John; Benning, Christoph

    2015-11-01

    Triacylglycerol (TAG) is a storage lipid used for food purposes and as a renewable feedstock for biodiesel production. WRINKLED1 (WRI1) is a transcription factor that governs fatty acid (FA) synthesis and, indirectly, TAG accumulation in oil-storing plant tissues, and its ectopic expression has led to TAG accumulation in vegetative tissues of different dicotyledonous plants. The ectopic expression of BdWRI1 in the grass Brachypodium distachyon induced the transcription of predicted genes involved in glycolysis and FA biosynthesis, and TAG content was increased up to 32.5-fold in 8-week-old leaf blades. However, the ectopic expression of BdWRI1 also caused cell death in leaves, which has not been observed previously in dicotyledonous plants such as Arabidopsis (Arabidopsis thaliana). Lipid analysis indicated that the free FA content was 2-fold elevated in BdWRI1-expressing leaf blades of B. distachyon. The transcription of predicted genes involved in β-oxidation was induced. In addition, linoleic FA treatment caused cell death in B. distachyon leaf blades, an effect that was reversed by the addition of the FA biosynthesis inhibitor cerulenin. Taken together, ectopic expression of BdWRI1 in B. distachyon enhances FA biosynthesis and TAG accumulation in leaves, as expected, but also leads to increased free FA content, which has cytotoxic effects leading to cell death. Thus, while WRI appears to ubiquitously affect FA biosynthesis and TAG accumulation in diverse plants, its ectopic expression can lead to undesired side effects depending on the context of the specific lipid metabolism of the respective plant species.

  14. Ectopic expression of transcription factor AP-2δ in developing retina: effect on PSA-NCAM and axon routing.

    PubMed

    Li, Xiaodong; Monckton, Elizabeth A; Godbout, Roseline

    2014-04-01

    Retinal ganglion cells transmit the visual signal from the retina to the brain. We have previously shown that the activator protein 2 (AP-2)δ (TFAP2D) transcription factor is expressed in one third of ganglion cells in developing retina suggesting a specialized role for these AP-2δ-expressing cells. Here, we address the role of AP-2δ in retina by in ovo electroporation of RCAS/AP-2δ retroviral constructs into the eyes of chick embryos at day 2 of gestation. Ectopic expression of AP-2δ does not affect lineage differentiation in the developing retina. However, immunostaining of retinal tissue with markers associated with axonal growth such as growth-associated protein 43 and polysialic acid-neural cell adhesion molecule (PSA-NCAM) demonstrates axonal misrouting and abnormal axonal bundling. Treatment of AP-2δ-misexpressing retinal cell cultures with endoneuraminidase, an enzyme that removes PSA from NCAM, decreases AP-2δ-induced axonal bundling. Our data suggest a role for AP-2δ in polysialylation of NCAM, with ectopic expression of AP-2δ resulting in premature bundling of emerging axons and misrouting of axons. We propose that expression of AP-2δ in a subset of ganglion cells contributes to the fine-tuning of axonal growth in the developing retina.

  15. Ectopic expression of Arabidopsis RCI2A gene contributes to cold tolerance in tomato.

    PubMed

    Sivankalyani, Velu; Geetha, Mahalingam; Subramanyam, Kondeti; Girija, Shanmugam

    2015-04-01

    Cold is a major stress that limits the quality and productivity of economically important crops such as tomato (Solanum lycopersicum L.). Generating a cold-stress-tolerant tomato by expressing cold-inducible genes would increase agricultural strategies. Rare cold-inducible 2a (RCI2A) is expressed in Arabidopsis, but its molecular function during cold stress is not fully understood. Here we ectopically expressed Arabidopsis RCI2A in transgenic tomato to evaluate tolerance to cold stress without altering agronomic traits. Biochemical and physiological study demonstrated that expression of RCI2A in transgenic tomato enhanced the activity of peroxidase and ascorbate peroxidase (APX) and reduced the accumulation of H2O2, alleviated lipid peroxidation, increased the accumulation of chlorophyll, reduced chilling-induced membrane damage, retained relative water content and enhanced cold tolerance. A motif search revealed that the motifs of photosystem II (PSII) phosphoproteins PsbJ and PsbH and reaction-center proteins PsbL and PsbK were common to cold-inducible RCI2A and peroxidase proteins RCI3A, tomato peroxidase (TPX1), TPX2, tomato ascorbate peroxidase (APX1), and horseradish peroxidase (HRP-c). In addition to membrane protection, RCI2A may cross talk with PSII-associated proteins or peroxidase family enzymes in response to cold stress. Our findings may strengthen the understanding of the molecular function of RCI2A in cold-stress tolerance. RCI2A could be used to improve abiotic stress tolerance in agronomic crops.

  16. Transcriptional activities of the Pax6 gene eyeless regulate tissue specificity of ectopic eye formation in Drosophila

    PubMed Central

    Weasner, Bonnie M.; Weasner, Brandon; DeYoung, Stephanie M.; Michaels, Scott D.; Kumar, Justin P.

    2009-01-01

    Pax genes encode DNA binding proteins that play pivotal roles in the determination of complex tissues. Members of one subclass, Pax6, function as selector genes and play key roles in the retinal development of all seeing animals. Mutations within the Pax6 homologs including fly eyeless, mouse Small eye and human Pax6 lead to severe retinal defects in their respective systems. In Drosophila eyeless and twin of eyeless, play non-redundant roles in the developing retina. One particularly interesting characteristic of these genes is that, although expression of either gene can induce ectopic eye formation in non-retinal tissues, there are differences in the location and frequencies at which the eyes develop. eyeless induces much larger ectopic eyes, at higher frequencies, and in a broader range of tissues than twin of eyeless. In this report we describe a series of experiments conducted in both yeast and flies that has identified protein modules that are responsible for the differences in tissue transformation. These domains appear to contain transcriptional activator and repressor activity of distinct strengths. We propose a model in which the selective presence of these activities and their relative strengths accounts, in part, for the disparity to which ectopic eyes are induced in response to the forced expression of eyeless and twin of eyeless. The identification of both transcriptional activator and repressor activity within the Pax6 protein furthers our understanding of how this gene family regulates tissue determination. PMID:19406113

  17. Ectopic expression of catalase in Drosophila mitochondria increases stress resistance but not longevity.

    PubMed

    Mockett, Robin J; Bayne, Anne Cécile V; Kwong, Linda K; Orr, William C; Sohal, Rajindar S

    2003-01-15

    The goal of this study was to test the hypothesis that the rate of mitochondrial oxidant production governs the aging process of the fruit fly, Drosophila melanogaster. Catalase, an antioxidative enzyme expressed in the cytosol and peroxisomes of Drosophila, was targetted ectopically to the mitochondrial matrix by fusion of a leader peptide derived from ornithine aminotransferase with its N-terminus. The presence of the transgene encoding this fusion protein was associated with moderate (35 +/- 13%) increases in total catalase activity in most lines, and measurable levels of catalase activity in the mitochondria (30-140 U/mg protein). There was no impact on the life span of the flies at 25 degrees C, even in an exceptional line with a 149% increase in total catalase activity, and there was a small decrease in longevity at 29 degrees C. There were no compensatory changes in the rate of metabolism or physical activity, or in the levels of other major antioxidants, suggesting that the aging process was largely unaffected. Resistance to exogenous hydrogen peroxide, paraquat, and cold stress was enhanced, but there was no appreciable effect on resistance to hyperoxia. The results demonstrate the importance of mitochondrial antioxidant levels in the resistance to oxidative stress at the organismal level, and illustrate that different effects on aging and stress resistance may ensue from a single treatment. The main inferences drawn are that: (i) levels of stress resistance may neither be a cause nor a reliable indicator of the rate of aging, and (ii) bolstering antioxidant levels in Drosophila may not delay or slow down the aging process.

  18. Ectopic Pregnancy

    MedlinePlus

    ... Feeding Your 1- to 2-Year-Old Ectopic Pregnancy KidsHealth > For Parents > Ectopic Pregnancy Print A A ... lower back pain continue What Causes an Ectopic Pregnancy? An ectopic pregnancy usually happens because a fertilized ...

  19. Ectopic Pregnancy

    MedlinePlus

    ... Feeding Your 1- to 2-Year-Old Ectopic Pregnancy KidsHealth > For Parents > Ectopic Pregnancy A A A ... lower back pain continue What Causes an Ectopic Pregnancy? An ectopic pregnancy usually happens because a fertilized ...

  20. Ectopic expression of murine diphosphoinositol polyphosphate phosphohydrolase 1 attenuates signaling through the ERK1/2 pathway.

    PubMed

    Chu, Caryn; Alapat, Daisy; Wen, Xiaping; Timo, Kimberly; Burstein, David; Lisanti, Michael; Shears, Stephen; Kohtz, D Stave

    2004-09-01

    Signals from several receptor tyrosine kinases are transduced by activation of the Ras family of GTP-binding proteins. Activation of Ras initiates a kinase cascade that culminates in activation of the mitogen-activated protein kinases (MAPKs). The MAPKs include the c-jun NH(2)-terminal protein kinases (JNKs) and extracellular signal-regulated kinases (ERKs), both of which phosphorylate Elk-1/TCF, a factor that activates transcription of the c-fos gene. In this report, we identify a novel 19 kDa gene product as a negative regulator of signaling through the ERK1/2 pathway. While these studies were in progress, the human homologue of this gene was characterized as diphosphoinositol polyphosphate phosphohydrolase (DIPP1) [EMBO J. 17 (1998) 6599], a phosphohydrolase that converts diphosphate groups on diphosphoinositol polyphosphates to monophosphates. Ectopic expression of murine DIPP1 (muDIPP1) blocked activation of the c-fos promoter by the ERK1/2 pathway. Inhibition of signal transduction through the ERK1/2 pathway by muDIPP1 occurs at or downstream from activation of MEK. In vitro kinase studies suggest that muDIPP1 is not a direct inhibitor of MEK or ERK activity, although, ectopic expression at near physiological levels results in attenuation of ERK phosphorylation in vivo. Interestingly, a site mutant of muDIPP1 lacking phosphohydrolase activity blocked signaling through the ERK1/2 pathway with greater efficiency than wild-type muDIPP1. This result suggests that inhibition of signaling through the ERK1/2 pathway is a distinct function of muDIPP1 that is not dependent on, but may be regulated by, its activity as a phosphohydrolase.

  1. Receptor-Independent Ectopic Activity of Prolactin Predicts Aggressive Lung Tumors and Indicates HDACi-Based Therapeutic Strategies

    PubMed Central

    Le Bescont, Aurore; Vitte, Anne-Laure; Debernardi, Alexandra; Curtet, Sandrine; Buchou, Thierry; Vayr, Jessica; de Reyniès, Aurélien; Ito, Akihiro; Guardiola, Philippe; Brambilla, Christian; Yoshida, Minoru; Brambilla, Elisabeth

    2015-01-01

    Abstract Aims: Ectopic activation of tissue-specific genes accompanies malignant transformation in many cancers. Prolactin (PRL) aberrant activation in lung cancer was investigated here to highlight its value as a biomarker. Results: PRL is ectopically activated in a subset of very aggressive lung tumors, associated with a rapid fatal outcome, in our cohort of 293 lung tumor patients and in an external independent series of patients. Surprisingly PRL receptor expression was not detected in the vast majority of PRL-expressing lung tumors. Additionally, the analysis of the PRL transcripts in lung tumors and cell lines revealed systematic truncations of their 5′ regions, including the signal peptide-encoding portions. PRL expression was found to sustain cancer-specific gene expression circuits encompassing genes that are normally responsive to hypoxia. Interestingly, this analysis also indicated that histone deacetylase (HDAC) inhibitors could counteract the PRL-associated transcriptional activity. Innovation and Conclusion: Altogether, this work not only unravels a yet unknown oncogenic mechanism but also indicates that the specific category of PRL-expressing aggressive lung cancers could be particularly responsive to an HDAC inhibitor-based treatment. Antioxid. Redox Signal. 23, 1–14. PMID:24512221

  2. Ectopic expression of RESISTANCE TO POWDERY MILDEW8.1 confers resistance to fungal and oomycete pathogens in Arabidopsis.

    PubMed

    Ma, Xian-Feng; Li, Yan; Sun, Jin-Long; Wang, Ting-Ting; Fan, Jing; Lei, Yang; Huang, Yan-Yan; Xu, Yong-Ju; Zhao, Ji-Qun; Xiao, Shunyuan; Wang, Wen-Ming

    2014-08-01

    Broad-spectrum disease resistance is a highly valuable trait in plant breeding and attracts special attention in research. The Arabidopsis gene locus RESISTANCE TO POWDERY MILDEW 8 (RPW8) contains two adjacent homologous genes, RPW8.1 and RPW8.2, and confers broad-spectrum resistance to powdery mildew. Remarkably, the RPW8.2 protein is specifically localized to the extrahaustorial membrane (EHM) encasing the feeding structure of powdery mildew whereby RPW8.2 activates haustorium-targeted defenses. Here, we show that ectopic expression of the yellow fluorescent protein (YFP)-tagged RPW8.1 from the native promoter leads to unique cell death lesions and enhances resistance to virulent fungal and oomycete pathogens that cause powdery mildew and downy mildew diseases, respectively. In powdery mildew-infected plants, RPW8.1-YFP accumulates at higher levels in the mesophyll cells underneath the infected epidermal cells where RPW8.2-YFP is mainly expressed. This cell type-preferential protein accumulation pattern largely correlates with that of H(2)O(2) accumulation, suggesting that RPW8.1 may spatially collaborate with RPW8.2 in activation of resistance to powdery mildew. Interestingly, when ectopically expressed from the RPW8.2 promoter, RPW8.1-YFP is also targeted to the EHM of powdery mildew and the transgenic plants display resistance to both powdery mildew and downy mildew. Using YFP as a reporter, we further reveal that the RPW8.1 promoter is constitutively active but induced to higher levels in cells at the infection site, whereas the RPW8.2 promoter is activated specifically in cells at the infection site. Taken together, our results suggest that RPW8.1 (and its promoter) is functionally distinct from RPW8.2 and may have a higher potential in engineering broad-spectrum resistance in plants.

  3. Ectopic lymphokine gene expression in human peripheral blood lymphocytes in vivo

    SciTech Connect

    Chambers, C.A.; Kang, Joonsoo; Hozumi, Nobumichi Mount Sinai Hospital, Toronto, Ontario )

    1992-02-01

    An animal model to study the effects of ectopic expression of cytokines involved in cell growth and differentiation has been established. Retrovirus vectors containing the human interleukin 6 cDNA were used to produce high titer virus-producing lines. Human peripheral blood lymphocytes (hPBLs) were successfully infected with the retrovirus and engrafted into severe combined immunodeficient mice. The majority of the animals were engrafted with hPBLs, as determined by the presence of human glucose phosphate isomerase. Furthermore, six of seven mice engrafted with hPBLs infected with high titer virus and detectable hPBLs present in the spleen expressed the retroviral human interleukin 6 gene. Importantly, human interleukin 6 protein was expressed at physiologically significant levels in these mice. These results demonstrate that models for human disease and immunotherapy involving retrovirus-mediated gene transfer into human cells can be developed in mice.

  4. Ectopic activity in cutaneous regenerating afferent nerve fibers following nerve lesion in the rat.

    PubMed

    Gorodetskaya, Natalia; Constantin, Cristina; Jänig, Wilfrid

    2003-11-01

    Spontaneous activity, and mechanical and thermal sensitivity were investigated in regenerating afferent nerve fibers within 4-21 days post sural nerve lesion (crush or transection and resuturing) in anaesthetized rats. About 33-40% of the myelinated (A) and 22-27% of the unmyelinated (C) fibers excited by electrical nerve stimulation exhibited at least one of these ectopic discharge properties. In total 177 A- and 169 C-fibers with ectopic activity were analysed. Most A-fibers (161/177) were mechanosensitive. Spontaneous activity (median 1 imp/s) was present in 23/177 and thermosensitivity in 14/177 A-fibers (13 of them being activated by heat stimuli). Almost all A-fibers (159/177) exhibited only one type of ectopic discharge property. Most C-fibers (94/169) were thermosensitive responding either to cold (n = 45) or to heat stimuli (n = 33) or to both (n = 16). Eighty-four of 169 C-fibers were spontaneously active (median 0.3 imp/s) and 75/169 C-fibers were mechanosensitive. Both the proportion and the discharge rate of spontaneously active C-fibers were significantly higher after crush than after section and resuturing of the nerve. About 60% of the C-fibers (101/169) had only one ectopic discharge property and 40% two or three. In conclusion, regenerating cutaneous afferent A- and C-fibers may develop mechano- and/or thermosensitivity as well as spontaneous activity. We suggest that spontaneous and evoked ectopic activity in regenerating cutaneous afferents are a function of the intrinsic functional properties of these neurons and of the interaction between the regenerating nerve fibers and non-neural cells during Wallerian degeneration in the nerve distal to the nerve lesion.

  5. An activated form of UFO alters leaf development and produces ectopic floral and inflorescence meristems.

    PubMed

    Risseeuw, Eddy; Venglat, Prakash; Xiang, Daoquan; Komendant, Kristina; Daskalchuk, Tim; Babic, Vivijan; Crosby, William; Datla, Raju

    2013-01-01

    Plants are unique in their ability to continuously produce new meristems and organ primordia. In Arabidopsis, the transcription factor LEAFY (LFY) functions as a master regulator of a gene network that is important for floral meristem and organ specification. UNUSUAL FLORAL ORGANS (UFO) is a co-activator of LEAFY and is required for proper activation of APETALA3 in the floral meristem during the specification of stamens and petals. The ufo mutants display defects in other parts of the flower and the inflorescence, suggestive of additional roles. Here we show that the normal determinacy of the developing Arabidopsis leaves is affected by the expression of a gain-of-function UFO fusion protein with the VP16 transcriptional activator domain. In these lines, the rosette and cauline leaf primordia exhibit reiterated serration, and upon flowering produce ectopic meristems that develop into flowers, bract leaves and inflorescences. These striking phenotypes reveal that developing leaves maintain the competency to initiate flower and inflorescence programs. Furthermore, the gain-of-function phenotypes are dependent on LFY and the SEPALLATA (SEP) MADS-box transcription factors, indicative of their functional interactions with UFO. The findings of this study also suggest that UFO promotes the establishment of the lateral meristems and primordia in the peripheral zone of the apical and floral meristems by enhancing the activity of LFY. These novel phenotypes along with the mutant phenotypes of UFO orthologs in other plant species suggest a broader function for UFO in plants.

  6. An Activated Form of UFO Alters Leaf Development and Produces Ectopic Floral and Inflorescence Meristems

    PubMed Central

    Risseeuw, Eddy; Venglat, Prakash; Xiang, Daoquan; Komendant, Kristina; Daskalchuk, Tim; Babic, Vivijan; Crosby, William; Datla, Raju

    2013-01-01

    Plants are unique in their ability to continuously produce new meristems and organ primordia. In Arabidopsis, the transcription factor LEAFY (LFY) functions as a master regulator of a gene network that is important for floral meristem and organ specification. UNUSUAL FLORAL ORGANS (UFO) is a co-activator of LEAFY and is required for proper activation of APETALA3 in the floral meristem during the specification of stamens and petals. The ufo mutants display defects in other parts of the flower and the inflorescence, suggestive of additional roles. Here we show that the normal determinacy of the developing Arabidopsis leaves is affected by the expression of a gain-of-function UFO fusion protein with the VP16 transcriptional activator domain. In these lines, the rosette and cauline leaf primordia exhibit reiterated serration, and upon flowering produce ectopic meristems that develop into flowers, bract leaves and inflorescences. These striking phenotypes reveal that developing leaves maintain the competency to initiate flower and inflorescence programs. Furthermore, the gain-of-function phenotypes are dependent on LFY and the SEPALLATA (SEP) MADS-box transcription factors, indicative of their functional interactions with UFO. The findings of this study also suggest that UFO promotes the establishment of the lateral meristems and primordia in the peripheral zone of the apical and floral meristems by enhancing the activity of LFY. These novel phenotypes along with the mutant phenotypes of UFO orthologs in other plant species suggest a broader function for UFO in plants. PMID:24376756

  7. Ectopic expression of Msx-2 in posterior limb bud mesoderm impairs limb morphogenesis while inducing BMP-4 expression, inhibiting cell proliferation, and promoting apoptosis.

    PubMed

    Ferrari, D; Lichtler, A C; Pan, Z Z; Dealy, C N; Upholt, W B; Kosher, R A

    1998-05-01

    During early stages of chick limb development, the homeobox-containing gene Msx-2 is expressed in the mesoderm at the anterior margin of the limb bud and in a discrete group of mesodermal cells at the midproximal posterior margin. These domains of Msx-2 expression roughly demarcate the anterior and posterior boundaries of the progress zone, the highly proliferating posterior mesodermal cells underneath the apical ectodermal ridge (AER) that give rise to the skeletal elements of the limb and associated structures. Later in development as the AER loses its activity, Msx-2 expression expands into the distal mesoderm and subsequently into the interdigital mesenchyme which demarcates the developing digits. The domains of Msx-2 expression exhibit considerably less proliferation than the cells of the progress zone and also encompass several regions of programmed cell death including the anterior and posterior necrotic zones and interdigital mesenchyme. We have thus suggested that Msx-2 may be in a regulatory network that delimits the progress zone by suppressing the morphogenesis of the regions of the limb mesoderm in which it is highly expressed. In the present study we show that ectopic expression of Msx-2 via a retroviral expression vector in the posterior mesoderm of the progress zone from the time of initial formation of the limb bud severely impairs limb morphogenesis. Msx-2-infected limbs are typically very narrow along the anteroposterior axis, are occasionally truncated, and exhibit alterations in the pattern of formation of skeletal elements, indicating that as a consequence of ectopic Msx-2 expression the morphogenesis of large portions of the posterior mesoderm has been suppressed. We further show that Msx-2 impairs limb morphogenesis by reducing cell proliferation and promoting apoptosis in the regions of the posterior mesoderm in which it is ectopically expressed. The domains of ectopic Msx-2 expression in the posterior mesoderm also exhibit ectopic

  8. The morphogen Decapentaplegic employs a two-tier mechanism to activate target retinal determining genes during ectopic eye formation in Drosophila

    PubMed Central

    Aggarwal, Poonam; Gera, Jayati; Mandal, Lolitika; Mandal, Sudip

    2016-01-01

    Understanding the role of morphogen in activating its target genes, otherwise epigenetically repressed, during change in cell fate specification is a very fascinating yet relatively unexplored domain. Our in vivo loss-of-function genetic analyses reveal that specifically during ectopic eye formation, the morphogen Decapentaplegic (Dpp), in conjunction with the canonical signaling responsible for transcriptional activation of retinal determining (RD) genes, triggers another signaling cascade. Involving dTak1 and JNK, this pathway down-regulates the expression of polycomb group of genes to do away with their repressive role on RD genes. Upon genetic inactivation of members of this newly identified pathway, the canonical Dpp signaling fails to trigger RD gene expression beyond a threshold, critical for ectopic photoreceptor differentiation. Moreover, the drop in ectopic RD gene expression and subsequent reduction in ectopic photoreceptor differentiation resulting from inactivation of dTak1 can be rescued by down-regulating the expression of polycomb group of genes. Our results unravel an otherwise unknown role of morphogen in coordinating simultaneous transcriptional activation and de-repression of target genes implicating its importance in cellular plasticity. PMID:27270790

  9. Ectopic expression of the striatal-enriched GTPase Rhes elicits cerebellar degeneration and an ataxia phenotype in Huntington's disease.

    PubMed

    Swarnkar, Supriya; Chen, Youjun; Pryor, William M; Shahani, Neelam; Page, Damon T; Subramaniam, Srinivasa

    2015-10-01

    Huntington's disease (HD) is caused by an expansion of glutamine repeats in the huntingtin protein (mHtt) that invokes early and prominent damage of the striatum, a region that controls motor behaviors. Despite its ubiquitous expression, why certain brain regions, such as the cerebellum, are relatively spared from neuronal loss by mHtt remains unclear. Previously, we implicated the striatal-enriched GTPase, Rhes (Ras homolog enriched in the striatum), which binds and SUMOylates mHtt and increases its solubility and cellular cytotoxicity, as the cause for striatal toxicity in HD. Here, we report that Rhes deletion in HD mice (N171-82Q), which express the N-terminal fragment of human Htt with 82 glutamines (Rhes(-/-)/N171-82Q), display markedly reduced HD-related behavioral deficits, and absence of lateral ventricle dilatation (secondary to striatal atrophy), compared to control HD mice (N171-82Q). To further validate the role of GTPase Rhes in HD, we tested whether ectopic Rhes expression would elicit a pathology in a brain region normally less affected in HD. Remarkably, ectopic expression of Rhes in the cerebellum of N171-82Q mice, during the asymptomatic period led to an exacerbation of motor deficits, including loss of balance and motor incoordination with ataxia-like features, not apparent in control-injected N171-82Q mice or Rhes injected wild-type mice. Pathological and biochemical analysis of Rhes-injected N171-82Q mice revealed a cerebellar lesion with marked loss of Purkinje neuron layer parvalbumin-immunoreactivity, induction of caspase 3 activation, and enhanced soluble forms of mHtt. Similarly reintroducing Rhes into the striatum of Rhes deleted Rhes(-/-)Hdh(150Q/150Q) knock-in mice, elicited a progressive HD-associated rotarod deficit. Overall, these studies establish that Rhes plays a pivotal role in vivo for the selective toxicity of mHtt in HD.

  10. Ectopic expression of the agouti gene in transgenic mice causes obesity, features of type II diabetes, and yellow fur.

    PubMed

    Klebig, M L; Wilkinson, J E; Geisler, J G; Woychik, R P

    1995-05-23

    Mice that carry the lethal yellow (Ay) or viable yellow (Avy) mutation, two dominant mutations of the agouti (a) gene in mouse chromosome 2, exhibit a phenotype that includes yellow fur, marked obesity, a form of type II diabetes associated with insulin resistance, and an increased susceptibility to tumor development. Molecular analyses of these and several other dominant "obese yellow" a-locus mutations suggested that ectopic expression of the normal agouti protein gives rise to this complex pleiotropic phenotype. We have now tested this hypothesis directly by generating transgenic mice that ectopically express an agouti cDNA clone encoding the normal agouti protein in all tissues examined. Transgenic mice of both sexes have yellow fur, become obese, and develop hyperinsulinemia. In addition, male transgenic mice develop hyperglycemia by 12-20 weeks of age. These results demonstrate conclusively that the ectopic agouti expression is responsible for most, if not all, of the phenotypic traits of the dominant, obese yellow mutants.

  11. Modulation of the hormone setting by Rhodococcus fascians results in ectopic KNOX activation in Arabidopsis.

    PubMed

    Depuydt, Stephen; Dolezal, Karel; Van Lijsebettens, Mieke; Moritz, Thomas; Holsters, Marcelle; Vereecke, Danny

    2008-03-01

    The biotrophic actinomycete Rhodococcus fascians has a profound impact on plant development and a common aspect of the symptomatology is the deformation of infected leaves. In Arabidopsis (Arabidopsis thaliana), the serrated leaf margins formed upon infection resemble the leaf phenotype of transgenic plants with ectopic expression of KNOTTED-like homeobox (KNOX) genes. Through transcript profiling, we demonstrate that class-I KNOX genes are transcribed in symptomatic leaves. Functional analysis revealed that BREVIPEDICELLUS/KNOTTED-LIKE1 and mainly SHOOT MERISTEMLESS were essential for the observed leaf dissection. However, these results also positioned the KNOX genes downstream in the signaling cascade triggered by R. fascians infection. The much faster activation of ARABIDOPSIS RESPONSE REGULATOR5 and the establishment of homeostatic and feedback mechanisms to control cytokinin (CK) levels support the overrepresentation of this hormone in infected plants due to the secretion by the pathogen, thereby placing the CK response high up in the cascade. Hormone measurements show a net decrease of tested CKs, indicating either that secretion by the bacterium and degradation by the plant are in balance, or, as suggested by the strong reaction of 35S:CKX plants, that other CKs are at play. At early time points of the interaction, activation of gibberellin 2-oxidase presumably installs a local hormonal setting favorable for meristematic activity that provokes leaf serrations. The results are discussed in the context of symptom development, evasion of plant defense, and the establishment of a specific niche by R. fascians.

  12. Identification of genes in the phenylalanine metabolic pathway by ectopic expression of a MYB transcription factor in tomato fruit

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Altering expression of transcription factors can be an effective means to coordinately modulate entire metabolic pathways in plants. It can also provide useful information concerning the identities of genes that constitute metabolic networks. Here, we used ectopic expression of a MYB transcription f...

  13. Transgenic Expression of Ad4BP/SF-1 in Fetal Adrenal Progenitor Cells Leads to Ectopic Adrenal Formation

    PubMed Central

    Zubair, Mohamad; Oka, Sanae; Parker, Keith L.; Morohashi, Ken-ichirou

    2009-01-01

    Deficiency of adrenal 4 binding protein/steroidogenic factor 1 (Ad4BP/SF-1; NR5A1) impairs adrenal development in a dose-dependent manner, whereas overexpression of Ad4BP/SF-1 is associated with adrenocortical tumorigenesis. Despite its essential roles in adrenal development, the mechanism(s) by which Ad4BP/SF-1 regulates this process remain incompletely understood. We previously identified a fetal adrenal enhancer (FAdE) that stimulates Ad4BP/SF-1 expression in the fetal adrenal gland by a two-step mechanism in which homeobox proteins initiate Ad4BP/SF-1 expression, which then maintains FAdE activity in an autoregulatory loop. In the present study, we examined the effect of transgenic expression of Ad4BP/SF-1 controlled by FAdE on adrenal development. When Ad4BP/SF-1 was overexpressed using a FAdE-Ad4BP/SF-1 transgene, FAdE activity expanded outside of its normal field, resulting in increased adrenal size and the formation of ectopic adrenal tissue in the thorax. The increased size of the adrenal gland did not result from a corresponding increase in cell proliferation, suggesting rather that the increased levels of Ad4BP/SF-1 may divert uncommitted precursors to the steroidogenic lineage. The effects of FAdE-controlled Ad4BP/SF-1 overexpression in mice provide a novel model of ectopic adrenal formation that further supports the critical role of Ad4BP/SF-1 in the determination of steroidogenic cell fate in vivo. PMID:19628584

  14. Attenuated tubal and endometrial urocortin 1 and corticotropin-releasing hormone receptor expression in ectopic pregnancy.

    PubMed

    Borges, L E; Horne, A W; McDonald, S E; Shaw, J L V; Lourenco, P C; Petraglia, F; Critchley, H O D

    2011-03-01

    Fallopian tube (FT) and endometrial urocortin 1 (Ucn1) and corticotropin-releasing hormone (CRH)-receptor (CRH-R1/CRH-R2) expression were examined using quantitative real-time polymerase chain reaction (RT-PCR) and immunohistochemistry in nonpregnant and pregnant women (intrauterine, IUP; ectopic pregnancy, EP). Tubal Ucn1 messenger RNA (mRNA) expression was higher in luteal compared to follicular phase (P < .01) and equivalent to follicular phase in FT from EP. Tubal CRH-R1/CRH-R2 mRNA was lower in luteal phase (P < .05) and in FT from EP compared to follicular phase (P < .01). Ucn1 mRNA was lower in endometrium from EP compared to IUP (P < .05). Corticotropin-releasing hormone-R1 mRNA was higher in endometrium from EP compared to viable IUP (P < .05). No differences were observed in CRH-R2 expression. Corticotropin-releasing hormone-R1 protein was primarily localized to epithelium of FT and endometrium. Quantitative analysis of tubal CRH-R1 protein expression reflected that seen at the mRNA level but endometrial expression was equivocal. The demonstration of attenuated tubal/endometrial Ucn1/CRH-R expression in EP further supports a role of the CRH-family in embryo implantation.

  15. Ectopic AtCBF1 over-expression enhances freezing tolerance and induces cold acclimation-associated physiological modifications in potato.

    PubMed

    Pino, María-Teresa; Skinner, Jeffrey S; Jeknić, Zoran; Hayes, Patrick M; Soeldner, Alfred H; Thomashow, Michael F; Chen, Tony H H

    2008-04-01

    We studied the effect of ectopic AtCBF over-expression on physiological alterations that occur during cold exposure in frost-sensitive Solanum tuberosum and frost-tolerant Solanum commersonii. Relative to wild-type plants, ectopic AtCBF1 over-expression induced expression of COR genes without a cold stimulus in both species, and imparted a significant freezing tolerance gain in both species: 2 degrees C in S. tuberosum and up to 4 degrees C in S. commersonii. Transgenic S. commersonii displayed improved cold acclimation potential, whereas transgenic S. tuberosum was still incapable of cold acclimation. During cold treatment, leaves of wild-type S. commersonii showed significant thickening resulting from palisade cell lengthening and intercellular space enlargement, whereas those of S. tuberosum did not. Ectopic AtCBF1 activity induced these same leaf alterations in the absence of cold in both species. In transgenic S. commersonii, AtCBF1 activity also mimicked cold treatment by increasing proline and total sugar contents in the absence of cold. Relative to wild type, transgenic S. commersonii leaves were darker green, had higher chlorophyll and lower anthocyanin levels, greater stomatal numbers, and displayed greater photosynthetic capacity, suggesting higher productivity potential. These results suggest an endogenous CBFpathway is involved in many of the structural, biochemical and physiological alterations associated with cold acclimation in these Solanum species.

  16. Phytoremediation potential of Arabidopsis thaliana, expressing ectopically a vacuolar proton pump, for the industrial waste phosphogypsum.

    PubMed

    Khoudi, Habib; Maatar, Yafa; Brini, Faïçal; Fourati, Amine; Ammar, Najoua; Masmoudi, Khaled

    2013-01-01

    transgenic A. thaliana lines were more Cd-tolerant than the WT plants. These results suggested that ectopic expression of a vacuolar proton pump in A. thaliana plants can lead to various biotechnological applications including the phytoremediation of industrial wastes.

  17. Ectopic expression of Arabidopsis Target of Rapamycin (AtTOR) improves water-use efficiency and yield potential in rice

    NASA Astrophysics Data System (ADS)

    Bakshi, Achala; Moin, Mazahar; Kumar, M. Udaya; Reddy, Aramati Bindu Madhava; Ren, Maozhi; Datla, Raju; Siddiq, E. A.; Kirti, P. B.

    2017-02-01

    The target of Rapamycin (TOR) present in all eukaryotes is a multifunctional protein, regulating growth, development, protein translation, ribosome biogenesis, nutrient, and energy signaling. In the present study, ectopic expression of TOR gene of Arabidopsis thaliana in a widely cultivated indica rice resulted in enhanced plant growth under water-limiting conditions conferring agronomically important water-use efficiency (WUE) trait. The AtTOR high expression lines of rice exhibited profuse tillering, increased panicle length, increased plant height, high photosynthetic efficiency, chlorophyll content and low ∆13C. Δ13C, which is inversely related to high WUE, was as low as 17‰ in two AtTOR high expression lines. These lines were also insensitive to the ABA-mediated inhibition of seed germination. The significant upregulation of 15 stress-specific genes in high expression lines indicates their contribution to abiotic stress tolerance. The constitutive expression of AtTOR is also associated with significant transcriptional upregulation of putative TOR complex-1 components, OsRaptor and OsLST8. Glucose-mediated transcriptional activation of AtTOR gene enhanced lateral root formation. Taken together, our findings indicate that TOR, in addition to its multiple cellular functions, also plays an important role in response to abiotic stress and potentially enhances WUE and yield related attributes.

  18. Ectopic expression of Arabidopsis Target of Rapamycin (AtTOR) improves water-use efficiency and yield potential in rice

    PubMed Central

    Bakshi, Achala; Moin, Mazahar; Kumar, M. Udaya; Reddy, Aramati Bindu Madhava; Ren, Maozhi; Datla, Raju; Siddiq, E. A.; Kirti, P. B.

    2017-01-01

    The target of Rapamycin (TOR) present in all eukaryotes is a multifunctional protein, regulating growth, development, protein translation, ribosome biogenesis, nutrient, and energy signaling. In the present study, ectopic expression of TOR gene of Arabidopsis thaliana in a widely cultivated indica rice resulted in enhanced plant growth under water-limiting conditions conferring agronomically important water-use efficiency (WUE) trait. The AtTOR high expression lines of rice exhibited profuse tillering, increased panicle length, increased plant height, high photosynthetic efficiency, chlorophyll content and low ∆13C. Δ13C, which is inversely related to high WUE, was as low as 17‰ in two AtTOR high expression lines. These lines were also insensitive to the ABA-mediated inhibition of seed germination. The significant upregulation of 15 stress-specific genes in high expression lines indicates their contribution to abiotic stress tolerance. The constitutive expression of AtTOR is also associated with significant transcriptional upregulation of putative TOR complex-1 components, OsRaptor and OsLST8. Glucose-mediated transcriptional activation of AtTOR gene enhanced lateral root formation. Taken together, our findings indicate that TOR, in addition to its multiple cellular functions, also plays an important role in response to abiotic stress and potentially enhances WUE and yield related attributes. PMID:28230163

  19. Directing Cardiomyogenic Differentiation and Transdifferentiation By Ectopic Gene Expression - Direct Transition Or Reprogramming Detour?

    PubMed

    Andrée, Birgit; Zweigerdt, Robert

    2016-01-01

    Cardiovascular disorders and associated morbidities remain the leading cause of premature death worldwide. Since the regeneration of diseased hearts is very limited and the insufficient supply of donor organs persists, hopes rely on new therapies for heart repair. Reviving the proliferation of endogenous cardiomyocytes (CMs) or the administration of adult stem cells to the heart was of limited curative success to date. Thus, the administration of in vitro generated CMs is under investigation to replenish loss of functional heart muscle tissue. This requires a sustainable source of CMs. Induced pluripotent stem cells (iPSC) have raised hopes for developing autologous cell therapies. To serve for heart repair, efficient and safe iPSC differentiation protocols for CMs production are required. iPSC differentiation into CMs and even functional subtypes was indeed achieved in recent years, either by the ectopic expression of cardiac transcription factors or the supplementation of chemical pathway modulators. An alternative approach aims at the direct transdifferentiation of fibroblasts, which are present in the interstitial tissue of many organs, into functional lineage-specific cell types. As a result the formation of induced cardiomyocyte-like cells (iCMs) by the ectopic expression of specific transcription factors combinations has been demonstrated in vitro and in vivo. This is an important proof-of-concept that the intermediate state of iPSC induction is dispensable. However, most of the early experiments were conducted in mice and translation to more relevant large animal models and subsequently to the clinic are challenging. Progress, drawbacks, and perspectives in this field will be discussed.

  20. Ectopically expressed pro-group X secretory phospholipase A2 is proteolytically activated in mouse adrenal cells by furin-like proprotein convertases: implications for the regulation of adrenal steroidogenesis.

    PubMed

    Layne, Joseph D; Shridas, Preetha; Webb, Nancy R

    2015-03-20

    Group X secretory phospholipase A2 (GX sPLA2) hydrolyzes mammalian cell membranes, liberating free fatty acids and lysophospholipids. GX sPLA2 is produced as a pro-enzyme (pro-GX sPLA2) that contains an N-terminal 11-amino acid propeptide ending in a dibasic motif, suggesting cleavage by a furin-like proprotein convertase (PC). Although propeptide cleavage is clearly required for enzymatic activity, the protease(s) responsible for pro-GX sPLA2 activation have not been identified. We previously reported that GX sPLA2 negatively regulates adrenal glucocorticoid production, likely by suppressing liver X receptor-mediated activation of steroidogenic acute regulatory protein expression. In this study, using a FLAG epitope-tagged pro-GX sPLA2 expression construct (FLAG-pro-GX sPLA2), we determined that adrenocorticotropic hormone (ACTH) enhanced FLAG-pro-GX sPLA2 processing and phospholipase activity secreted by Y1 adrenal cells. ACTH increased the expression of furin and PCSK6, but not other members of the PC family, in Y1 cells. Overexpression of furin and PCSK6 in HEK 293 cells significantly enhanced FLAG-pro-GX sPLA2 processing, whereas siRNA-mediated knockdown of both PCs almost completely abolished FLAG-pro-GX sPLA2 processing in Y1 cells. Expression of either furin or PCSK6 enhanced the ability of GX sPLA2 to suppress liver X receptor reporter activity. The PC inhibitor decanoyl-Arg-Val-Lys-Arg-chloromethyl ketone significantly suppressed FLAG-pro-GX sPLA2 processing and sPLA2 activity in Y1 cells, and it significantly attenuated GX sPLA2-dependent inhibition of steroidogenic acute regulatory protein expression and progesterone production. These findings provide strong evidence that pro-GX sPLA2 is a substrate for furin and PCSK6 proteolytic processing and define a novel mechanism for regulating corticosteroid production in adrenal cells.

  1. Bivalent promoter marks and a latent enhancer may prime the leukaemia oncogene LMO1 for ectopic expression in T-cell leukaemia.

    PubMed

    Oram, S H; Thoms, J; Sive, J I; Calero-Nieto, F J; Kinston, S J; Schütte, J; Knezevic, K; Lock, R B; Pimanda, J E; Göttgens, B

    2013-06-01

    LMO1 is a transcriptional regulator and a T-acute lymphoblastic leukaemia (T-ALL) oncogene. Although first identified in association with a chromosomal translocation in T-ALL, the ectopic expression of LMO1 occurs far more frequently in the absence of any known mutation involving its locus. Given that LMO1 is barely expressed in any haematopoietic lineage, and activation of transcriptional drivers in leukaemic cells is not well described, we investigated the regulation of this gene in normal haematopoietic and leukaemic cells. We show that LMO1 has two promoters that drive reporter gene expression in transgenic mice to neural tissues known to express endogenous LMO1. The LMO1 promoters display bivalent histone marks in multiple blood lineages including T-cells, and a 3' flanking region at LMO1 +57 contains a transcriptional enhancer that is active in developing blood cells in transgenic mouse embryos. The LMO1 promoters become activated in T-ALL together with the 3' enhancer, which is bound in primary T-ALL cells by SCL/TAL1 and GATA3. Taken together, our results show that LMO1 is poised for expression in normal progenitors, where activation of SCL/TAL1 together with a breakdown of epigenetic repression of LMO1 regulatory elements induces ectopic LMO1 expression that contributes to the development and maintenance of T-ALL.

  2. Ectopic expression of Arabidopsis glycosyltransferase UGT85A5 enhances salt stress tolerance in tobacco.

    PubMed

    Sun, Yan-Guo; Wang, Bo; Jin, Shang-Hui; Qu, Xiao-Xia; Li, Yan-Jie; Hou, Bing-Kai

    2013-01-01

    Abiotic stresses greatly influence plant growth and productivity. While glycosyltransferases are widely distributed in plant kingdom, their biological roles in response to abiotic stresses are largely unknown. In this study, a novel Arabidopsis glycosyltransferase gene UGT85A5 was identified as significantly induced by salt stress. Ectopic expression of UGT85A5 in tobacco enhanced the salt stress tolerance in the transgenic plants. There were higher seed germination rates, better plant growth and less chlorophyll loss in transgenic lines compared to wild type plants under salt stress. This enhanced tolerance of salt stress was correlated with increased accumulations of proline and soluble sugars, but with decreases in malondialdehyde accumulation and Na(+)/K(+) ratio in UGT85A5-expressing tobacco. Furthermore, during salt stress, expression of several carbohydrate metabolism-related genes including those for sucrose synthase, sucrose-phosphate synthase, hexose transporter and a group2 LEA protein were obviously upregulated in UGT85A5-expressing transgenic plants compared with wild type controls. Thus, these findings suggest a specific protective role of this glycosyltransferase against salt stress and provide a genetic engineering strategy to improve salt tolerance of crops.

  3. Transient ectopic expression of the histone demethylase JMJD3 accelerates the differentiation of human pluripotent stem cells

    PubMed Central

    Wakabayashi, Shunichi; Soma, Atsumi; Sato, Saeko; Nakatake, Yuhki; Oda, Mayumi; Murakami, Miyako; Sakota, Miki; Chikazawa-Nohtomi, Nana

    2016-01-01

    Harnessing epigenetic regulation is crucial for the efficient and proper differentiation of pluripotent stem cells (PSCs) into desired cell types. Histone H3 lysine 27 trimethylation (H3K27me3) functions as a barrier against cell differentiation through the suppression of developmental gene expression in PSCs. Here, we have generated human PSC (hPSC) lines in which genome-wide reduction of H3K27me3 can be induced by ectopic expression of the catalytic domain of the histone demethylase JMJD3 (called JMJD3c). We found that transient, forced demethylation of H3K27me3 alone triggers the upregulation of mesoendodermal genes, even when the culture conditions for the hPSCs are not changed. Furthermore, transient and forced expression of JMJD3c followed by the forced expression of lineage-defining transcription factors enabled the hPSCs to activate tissue-specific genes directly. We have also shown that the introduction of JMJD3c facilitates the differentiation of hPSCs into functional hepatic cells and skeletal muscle cells. These results suggest the utility of the direct manipulation of epigenomes for generating desired cell types from hPSCs for cell transplantation therapy and platforms for drug screenings. PMID:27802135

  4. Ectopic expression of LEAFY COTYLEDON1-LIKE gene and localized auxin accumulation mark embryogenic competence in epiphyllous plants of Helianthus annuus × H. tuberosus

    PubMed Central

    Chiappetta, A.; Fambrini, M.; Petrarulo, M.; Rapparini, F.; Michelotti, V.; Bruno, L.; Greco, M.; Baraldi, R.; Salvini, M.; Pugliesi, C.; Bitonti, M. B.

    2009-01-01

    Background and Aims The clone EMB-2 of the interspecific hybrid Helianthus annuus × H. tuberosus provides an interesting system to study molecular and physiological aspects of somatic embryogenesis. Namely, in addition to non-epiphyllous (NEP) leaves that expand normally, EMB-2 produces epiphyllous (EP) leaves bearing embryos on the adaxial surface. This clone was used to investigate if the ectopic expression of H. annuus LEAFY COTYLEDON1-LIKE (Ha-L1L) gene and auxin activity are correlated with the establishment of embryogenic competence. Methods Ha-L1L expression was evaluated by semi-quantitative RT-PCR and in situ hybridization. The endogenous level and spatial distribution of free indole-3-acetic acid (IAA) were estimated by a capillary gas chromatography–mass spectrometry–selected ion monitoring method and an immuno-cytochemical approach. Key Results Ectopic expression of Ha-L1L was detected in specific cell domains of the adaxial epidermis of EP leaves prior to the development of ectopic embryos. Ha-L1L was expressed rapidly when NEP leaves were induced to regenerate somatic embryos by in vitro culture. Differences in auxin distribution pattern rather than in absolute level were observed between EP and A-2 leaves. More precisely, a strong IAA immuno-signal was detected in single cells or in small groups of cells along the epidermis of EP leaves and accompanied the early stages of embryo development. Changes in auxin level and distribution were observed in NEP leaves induced to regenerate by in vitro culture. Exogenous auxin treatments lightly influenced Ha-L1L transcript levels in spite of an enhancement of the regeneration frequency. Conclusions In EP leaves, Ha-L1L activity marks the putative founder cells of ectopic embryos. Although the ectopic expression of Ha-L1L seems to be not directly mediated by auxin levels per se, it was demonstrated that localized Ha-L1L expression and IAA accumulation in leaf epidermis domains represent early events of

  5. Cord blood-derived neuronal cells by ectopic expression of Sox2 and c-Myc

    PubMed Central

    Giorgetti, Alessandra; Marchetto, Maria C. N.; Yu, Diana; Fazzina, Raffaella; Mu, Yangling; Adamo, Antonio; Paramonov, Ida; Cardoso, Julio Castaño; Monasterio, Montserrat Barragan; Bardy, Cedric; Cassiani-Ingoni, Riccardo; Liu, Guang-Hui; Gage, Fred H.; Izpisua Belmonte, Juan Carlos

    2012-01-01

    The finding that certain somatic cells can be directly converted into cells of other lineages by the delivery of specific sets of transcription factors paves the way to novel therapeutic applications. Here we show that human cord blood (CB) CD133+ cells lose their hematopoietic signature and are converted into CB-induced neuronal-like cells (CB-iNCs) by the ectopic expression of the transcription factor Sox2, a process that is further augmented by the combination of Sox2 and c-Myc. Gene-expression analysis, immunophenotyping, and electrophysiological analysis show that CB-iNCs acquire a distinct neuronal phenotype characterized by the expression of multiple neuronal markers. CB-iNCs show the ability to fire action potentials after in vitro maturation as well as after in vivo transplantation into the mouse hippocampus. This system highlights the potential of CB cells and offers an alternative means to the study of cellular plasticity, possibly in the context of drug screening research and of future cell-replacement therapies. PMID:22814375

  6. Cord blood-derived neuronal cells by ectopic expression of Sox2 and c-Myc.

    PubMed

    Giorgetti, Alessandra; Marchetto, Maria C N; Li, Mo; Yu, Diana; Fazzina, Raffaella; Mu, Yangling; Adamo, Antonio; Paramonov, Ida; Cardoso, Julio Castaño; Monasterio, Montserrat Barragan; Bardy, Cedric; Cassiani-Ingoni, Riccardo; Liu, Guang-Hui; Gage, Fred H; Izpisua Belmonte, Juan Carlos

    2012-07-31

    The finding that certain somatic cells can be directly converted into cells of other lineages by the delivery of specific sets of transcription factors paves the way to novel therapeutic applications. Here we show that human cord blood (CB) CD133(+) cells lose their hematopoietic signature and are converted into CB-induced neuronal-like cells (CB-iNCs) by the ectopic expression of the transcription factor Sox2, a process that is further augmented by the combination of Sox2 and c-Myc. Gene-expression analysis, immunophenotyping, and electrophysiological analysis show that CB-iNCs acquire a distinct neuronal phenotype characterized by the expression of multiple neuronal markers. CB-iNCs show the ability to fire action potentials after in vitro maturation as well as after in vivo transplantation into the mouse hippocampus. This system highlights the potential of CB cells and offers an alternative means to the study of cellular plasticity, possibly in the context of drug screening research and of future cell-replacement therapies.

  7. Inhibition of SRY-calmodulin complex formation induces ectopic expression of ovarian cell markers in developing XY gonads.

    PubMed

    Sim, Helena; Argentaro, Anthony; Czech, Daniel P; Bagheri-Fam, Stefan; Sinclair, Andrew H; Koopman, Peter; Boizet-Bonhoure, Brigitte; Poulat, Francis; Harley, Vincent R

    2011-07-01

    The transcription factor sex-determining region of the Y chromosome (SRY) plays a key role in human sex determination, because mutations in SRY cause disorders of sex development in XY individuals. During gonadal development, Sry in pre-Sertoli cells activates Sox9 gene transcription, committing the fate of the bipotential gonad to become a testis rather than an ovary. The high-mobility group domain of human SRY contains two independent nuclear localization signals, one bound by calmodulin (CaM) and the other by importin-β. Although XY females carry SRY mutations in these nuclear localization signals that affect SRY nuclear import in transfected cells, it is not known whether these transport mechanisms are essential for gonadal development and sex determination. Here, we show that mouse Sry protein binds CaM and that a CaM antagonist reduces CaM binding, nuclear accumulation, and transcriptional activity of Sry in transfected cells. CaM antagonist treatment of cultured, sexually indifferent XY mouse fetal gonads led to reduced expression of the Sry target gene Sox9, defects in testicular cord formation, and ectopic expression of the ovarian markers Rspondin1 and forkhead box L2. These results indicate the importance of CaM for SRY nuclear import, transcriptional activity, testis differentiation, and sex determination.

  8. Does adenosine deaminase activity play a role in the early diagnosis of ectopic pregnancy?

    PubMed

    Turkmen, G G; Karçaaltıncaba, D; Isık, H; Fidancı, V; Kaayalp, D; Tımur, H; Batıoglu, S

    2016-01-01

    Early diagnosis of ectopic pregnancy (EP) is important due to life-threatening consequences in the first trimester of pregnancy. In this study we aimed to investigate the role of adenosine deaminase (ADA) activity in the prediction of EP. Forty-one patients with unruptured ectopic pregnancy comprised the case group and forty-two first trimester pregnant women with shown foetal heart beating in ultrasound comprised the control group. The mean ADA level in EP (10.9 ± 3.0 IU/L) was higher than that in control group (9.2 ± 3.6 IU/L) (p = 0.018). Receiver operating characteristics or ROC curve identified ADA value of 10.95 IU/L as optimal threshold for the prediction of EP with 56% sensitivity and 67% specificity. High ADA levels are valuable in the early diagnosis of EP. However more comprehensive studies are required.

  9. Ectopic expression of the chemokine CXCL17 in colon cancer cells

    PubMed Central

    Ohlsson, Lina; Hammarström, Marie-Louise; Lindmark, Gudrun; Hammarström, Sten; Sitohy, Basel

    2016-01-01

    Background: The novel chemokine CXCL17 acts as chemoattractant for monocytes, macrophages and dendritic cells. CXCL17 also has a role in angiogenesis of importance for tumour development. Methods: Expression of CXCL17, CXCL10, CXCL9 and CCL2 was assessed in primary colon cancer tumours, colon carcinoma cell lines and normal colon tissue at mRNA and protein levels by real-time qRT–PCR, immunohistochemistry, two-colour immunofluorescence and immunomorphometry. Results: CXCL17 mRNA was expressed at 8000 times higher levels in primary tumours than in normal colon (P<0.0001). CXCL17 protein was seen in 17.2% of cells in tumours as compared with 0.07% in normal colon (P=0.0002). CXCL10, CXCL9 and CCL2 mRNAs were elevated in tumours but did not reach the levels of CXCL17. CXCL17 and CCL2 mRNA levels were significantly correlated in tumours. Concordant with the mRNA results, CXCL10- and CXCL9-positive cells were detected in tumour tissue, but at significantly lower numbers than CXCL17. Two-colour immunofluorescence and single-colour staining of consecutive sections for CXCL17 and the epithelial cell markers carcinoembryonic antigen and BerEP4 demonstrated that colon carcinoma tumour cells indeed expressed CXCL17. Conclusions: CXCL17 is ectopically expressed in primary colon cancer tumours. As CXCL17 enhances angiogenesis and attracts immune cells, its expression could be informative for prognosis in colon cancer patients. PMID:26889977

  10. Enhanced drought and heat stress tolerance of tobacco plants with ectopically enhanced cytokinin oxidase/dehydrogenase gene expression.

    PubMed

    Macková, Hana; Hronková, Marie; Dobrá, Jana; Turečková, Veronika; Novák, Ondřej; Lubovská, Zuzana; Motyka, Václav; Haisel, Daniel; Hájek, Tomáš; Prášil, Ilja Tom; Gaudinová, Alena; Štorchová, Helena; Ge, Eva; Werner, Tomáš; Schmülling, Thomas; Vanková, Radomíra

    2013-07-01

    Responses to drought, heat, and combined stress were compared in tobacco (Nicotiana tabacum L.) plants ectopically expressing the cytokinin oxidase/dehydrogenase CKX1 gene of Arabidopsis thaliana L. under the control of either the predominantly root-expressed WRKY6 promoter or the constitutive 35S promoter, and in the wild type. WRKY6:CKX1 plants exhibited high CKX activity in the roots under control conditions. Under stress, the activity of the WRKY6 promoter was down-regulated and the concomitantly reduced cytokinin degradation coincided with raised bioactive cytokinin levels during the early phase of the stress response, which might contribute to enhanced stress tolerance of this genotype. Constitutive expression of CKX1 resulted in an enlarged root system, a stunted, dwarf shoot phenotype, and a low basal level of expression of the dehydration marker gene ERD10B. The high drought tolerance of this genotype was associated with a relatively moderate drop in leaf water potential and a significant decrease in leaf osmotic potential. Basal expression of the proline biosynthetic gene P5CSA was raised. Both wild-type and WRKY6:CKX1 plants responded to heat stress by transient elevation of stomatal conductance, which correlated with an enhanced abscisic acid catabolism. 35S:CKX1 transgenic plants exhibited a small and delayed stomatal response. Nevertheless, they maintained a lower leaf temperature than the other genotypes. Heat shock applied to drought-stressed plants exaggerated the negative stress effects, probably due to the additional water loss caused by a transient stimulation of transpiration. The results indicate that modulation of cytokinin levels may positively affect plant responses to abiotic stress through a variety of physiological mechanisms.

  11. Ectopic over-expression of the maize beta-glucosidase Zm-p60.1 perturbs cytokinin homeostasis in transgenic tobacco.

    PubMed

    Kiran, Nagavalli S; Polanská, Lenka; Fohlerová, Radka; Mazura, Pavel; Válková, Martina; Smeral, Miloslav; Zouhar, Jan; Malbeck, Jirí; Dobrev, Petre I; Machácková, Ivana; Brzobohaty, Bretislav

    2006-01-01

    The activity of the phytohormone cytokinin depends on a complex interplay of factors such as its metabolism, transport, stability, and cellular/tissue localization. O-glucosides of zeatin-type cytokinins are postulated to be storage and/or transport forms, and are readily deglucosylated. Transgenic tobacco (Nicotiana tabacum L. cv. Petit Havana SR1) plants were constructed over-expressing Zm-p60.1, a maize beta-glucosidase capable of releasing active cytokinins from O- and N3-glucosides, to analyse its potential to perturb zeatin metabolism in planta. Zm-p60.1 in chloroplasts isolated from transgenic leaves has an apparent K(m) more than 10-fold lower than the purified enzyme in vitro. Adult transgenic plants grown in the absence of exogenous zeatin were morphologically indistinguishable from the wild type although differences in phytohormone levels were observed. When grown on medium containing zeatin, inhibition of root elongation was apparent in all seedlings 14 d after sowing (DAS). Between 14 and 21 DAS, the transgenic seedlings accumulated fresh weight leading later (28-32 DAS) to ectopic growths at the base of the hypocotyl. The development of ectopic structures correlated with the presence of the enzyme as demonstrated by histochemical staining. Cytokinin quantification showed that transgenic seedlings grown on medium containing zeatin accumulate active metabolites like zeatin riboside and zeatin riboside phosphate and this might lead to the observed changes. The presence of the enzyme around the base of the hypocotyl and later, in the ectopic structures themselves, suggests that the development of these structures is due to the perturbance in zeatin metabolism caused by the ectopic presence of Zm-p60.1.

  12. Ectopic expression of PtaRHE1, encoding a poplar RING-H2 protein with E3 ligase activity, alters plant development and induces defence-related responses

    PubMed Central

    Mukoko Bopopi, Johnny; Vandeputte, Olivier M.; Himanen, Kristiina; Mol, Adeline; Vaessen, Quentin; El Jaziri, Mondher; Baucher, Marie

    2010-01-01

    RING (really interesting new gene)-H2 domain-containing proteins are widely represented in plants and play important roles in the regulation of many developmental processes as well as in plant–environment interactions. In the present report, experiments were performed to unravel the role of the poplar gene PtaRHE1, coding for a RING-H2 protein. In vitro ubiquitination assays indicate a functional E3 ligase activity for PtaRHE1 with the specific E2 ubiquitin-conjugating enzyme UbcH5a. The overexpression of PtaRHE1 in tobacco resulted in a pleiotropic phenotype characterized by a curling of the leaves, the formation of necrotic lesions on leaf blades, growth retardation, and a delay in floral transition. The plant gene expression response to PtaRHE1 overexpression provided evidence for the up-regulation of defence- and/or programmed cell death-related genes. Moreover, genes coding for WRKY transcription factors as well as for mitogen-activated protein kinases, such as wound-induced protein kinase (WIPK), were also found to be induced in the transgenic lines as compared with the wild type. In addition, histochemical β-glucuronidase staining showed that the PtaRHE1 promoter is induced by plant pathogens and by elicitors such as salicylic acid and cellulase. Taken together, these results suggest that the E3 ligase PtaRHE1 plays a role in the ubiquitination-mediated regulation of defence response, possibly by acting upstream of WIPK and/or in the activation of WRKY factors. PMID:19892745

  13. Ectopic Pregnancy

    MedlinePlus

    ... woman is pregnant. If you have an ectopic pregnancy, the fertilized egg grows in the wrong place, ... tubes. The result is usually a miscarriage. Ectopic pregnancy can be a medical emergency if it ruptures. ...

  14. Senescence-induced ectopic expression of the A. tumefaciens ipt gene in wheat delays leaf senescence, increases cytokinin content, nitrate influx, and nitrate reductase activity, but does not affect grain yield.

    PubMed

    Sykorová, Blanka; Kuresová, Gabriela; Daskalova, Sasha; Trcková, Marie; Hoyerová, Klára; Raimanová, Ivana; Motyka, Václav; Trávnícková, Alena; Elliott, Malcolm C; Kamínek, Miroslav

    2008-01-01

    The manipulation of cytokinin levels by senescence-regulated expression of the Agrobacterium tumefaciens ipt gene through its control by the Arabidopsis SAG12 (senescence-associated gene 12) promoter is an efficient tool for the prolongation of leaf photosynthetic activity which potentially can affect plant productivity. In the present study, the efficiency of this approach was tested on wheat (Triticum aestivum L.)-a monocarpic plant characterized by a fast switch from vegetative to reproductive growth, and rapid translocation of metabolites from leaves to developing grains after anthesis. When compared with the wild-type (WT) control plants, the SAG12::ipt wheat plants exhibited delayed chlorophyll degradation only when grown under limited nitrogen (N) supply. Ten days after anthesis the content of chlorophyll and bioactive cytokinins of the first (flag) leaf of the transgenic plants was 32% and 65% higher, respectively, than that of the control. There was a progressive increase in nitrate influx and nitrate reductase activity. However, the SAG12::ipt and the WT plants did not show differences in yield-related parameters including number of grains and grain weight. These results suggest that the delay of leaf senescence in wheat also delays the translocation of metabolites from leaves to developing grains, as indicated by higher accumulation of ((15)N-labelled) N in spikes of control compared with transgenic plants prior to anthesis. This delay interferes with the wheat reproductive strategy that is based on a fast programmed translocation of metabolites from the senescing leaves to the reproductive sinks shortly after anthesis.

  15. The Ectopic Expression of CaRop1 Modulates the Response of Tobacco Plants to Ralstonia solanacearum and Aphids

    PubMed Central

    Qiu, Ailian; Liu, Zhiqin; Li, Jiazhi; Chen, Yanshen; Guan, Deyi; He, Shuilin

    2016-01-01

    In plants, Rho-related GTPases (Rops) are versatile molecular switches that regulate various biological processes, although their exact roles are not fully understood. Herein, we provide evidence that the ectopic expression of a Rop derived from Capsicum annuum, designated CaRop1, in tobacco plants modulates the response of these plants to Ralstonia solanacearum or aphid attack. The deduced amino acid sequence of CaRop1 harbors a conserved Rho domain and is highly homologous to Rops of other plant species. Transient expression of a CaRop1-GFP fusion protein in Nicotiana benthamiana leaf epidermal cells revealed localization of the GFP signal to the plasma membrane, cytoplasm, and nucleus. Overexpression (OE) of the wild-type CaRop1 or its dominant-negative mutant (DN-CaRop1) conferred substantial resistance to R. solanacearum infection and aphid attack, and this effect was accompanied by enhanced transcriptional expression of the hypersensitive-reaction marker gene HSR201; the jasmonic acid (JA)-responsive PR1b and LOX1; the insect resistance-associated NtPI-I, NtPI-II, and NtTPI; the ethylene (ET) production-associated NtACS1; and NPK1, a mitogen-activated protein kinase kinase kinase (MAPKKK) that interferes with N-, Bs2-, and Rx-mediated disease resistance. In contrast, OE of the constitutively active mutant of CaRop1(CA-CaRop1) enhanced susceptibility of the transgenic tobacco plants to R. solanacearum infection and aphid attack and downregulated or sustained the expression of HSR201, PR1b, NPK1, NtACS1, NtPI-I, NtPI-II, and NtTPI. These results collectively suggest that CaRop1 acts as a signaling switch in the crosstalk between Solanaceaes’s response to R. solanacearum infection and aphid attack possibly via JA/ET-mediated signaling machinery. PMID:27551287

  16. Ectopic expression of GsPPCK3 and SCMRP in Medicago sativa enhances plant alkaline stress tolerance and methionine content.

    PubMed

    Sun, Mingzhe; Sun, Xiaoli; Zhao, Yang; Zhao, Chaoyue; Duanmu, Huizi; Yu, Yang; Ji, Wei; Zhu, Yanming

    2014-01-01

    So far, it has been suggested that phosphoenolpyruvate carboxylases (PEPCs) and PEPC kinases (PPCKs) fulfill several important non-photosynthetic functions. However, the biological functions of soybean PPCKs, especially in alkali stress response, are not yet well known. In previous studies, we constructed a Glycine soja transcriptional profile, and identified three PPCK genes (GsPPCK1, GsPPCK2 and GsPPCK3) as potential alkali stress responsive genes. In this study, we confirmed the induced expression of GsPPCK3 under alkali stress and investigated its tissue expression specificity by using quantitative real-time PCR analysis. Then we ectopically expressed GsPPCK3 in Medicago sativa and found that GsPPCK3 overexpression improved plant alkali tolerance, as evidenced by lower levels of relative ion leakage and MDA content and higher levels of chlorophyll content and root activity. In this respect, we further co-transformed the GsPPCK3 and SCMRP genes into alfalfa, and demonstrated the increased alkali tolerance of GsPPCK3-SCMRP transgenic lines. Further investigation revealed that GsPPCK3-SCMRP co-overexpression promoted the PEPC activity, net photosynthetic rate and citric acid content of transgenic alfalfa under alkali stress. Moreover, we also observed the up-regulated expression of PEPC, CS (citrate synthase), H(+)-ATPase and NADP-ME genes in GsPPCK3-SCMRP transgenic alfalfa under alkali stress. As expected, we demonstrated that GsPPCK3-SCMRP transgenic lines displayed higher methionine content than wild type alfalfa. Taken together, results presented in this study supported the positive role of GsPPCK3 in plant response to alkali stress, and provided an effective way to simultaneously improve plant alkaline tolerance and methionine content, at least in legume crops.

  17. Ectopic Ptf1a expression in murine ESCs potentiates endocrine differentiation and models pancreas development in vitro.

    PubMed

    Nair, Gopika G; Vincent, Robert K; Odorico, Jon S

    2014-05-01

    Besides its role in exocrine differentiation, pancreas-specific transcription factor 1a (PTF1a) is required for pancreas specification from the foregut endoderm and ultimately for endocrine cell formation. Examining the early role of PTF1a in pancreas development has been challenging due to limiting amounts of embryonic tissue material for study. Embryonic stem cells (ESCs) which can be differentiated in vitro, and without limit to the amount of experimental material, can serve as a model system to study these early developmental events. To this end, we derived and characterized a mouse ESC line with tetracycline-inducible expression of PTF1a (tet-Ptf1a mESCs). We found that transient ectopic expression of PTF1a initiated the pancreatic program in differentiating ESCs causing cells to activate PDX1 expression in bud-like structures resembling pancreatic primordia in vivo. These bud-like structures also expressed progenitor markers characteristic of a developing pancreatic epithelium. The epithelium differentiated to generate a wave of NGN3+ endocrine progenitors, and further formed cells of all three pancreatic lineages. Notably, the insulin+ cells in the cultures were monohormonal, and expressed PDX1 and NKX6.1. PTF1a-induced cultures differentiated into significantly more endocrine and exocrine cells and the ratio of endocrine-to-exocrine cell differentiation could be regulated by retinoic acid (RA) and nicotinamide (Nic) signaling. Moreover, induced cultures treated with RA and Nic exhibited a modest glucose response. Thus, this tet-Ptf1a ESC-based in vitro system is a valuable new tool for interrogating the role of PTF1a in pancreas development and in directing differentiation of ESCs to endocrine cells.

  18. Deorphanization and characterization of the ectopically expressed olfactory receptor OR51B5 in myelogenous leukemia cells

    PubMed Central

    Manteniotis, S; Wojcik, S; Göthert, J R; Dürig, J; Dührsen, U; Gisselmann, G; Hatt, H

    2016-01-01

    The ectopic expression of olfactory receptors (ORs) in the human body has been of major interest in the past decade. Several studies have reported the expression of ORs not only in healthy tissues such as heart, sperm or skin cells, but also in cancerous tissues of the liver, prostate or intestine. In the present study, we detected the expression of OR51B5 in the chronic myelogenous leukemia (CML) cell line K562 and in white blood cell samples of clinically diagnosed acute myelogenous leukemia (AML) patients by reverse transcription-PCR and immunocytochemical staining. The known OR51B5 ligand isononyl alcohol increased the levels of intracellular Ca2+ in both AML patient blood cells and K562 cells. With calcium imaging experiments, we characterized in greater detail the OR51B5-mediated signaling pathway. Here, we observed an involvement of adenylate cyclase and the downstream L-type and T-type calcium channels. In addition, the activation of OR51B5 leads to an inhibition of cell proliferation in K562 cells. In western blot experiments, we found that incubation with isononyl alcohol led to a reduction in p38-MAPK (mitogen-activated protein kinase) phosphorylation that might be responsible for the decreased cell proliferation. In the present study, we characterized the OR51B5-mediated signaling pathway downstream of the activation with isononyl alcohol, which leads to reduced proliferation and therefore provide a novel pharmacological target for CML and AML, the latter of which remains difficult to treat. PMID:27551504

  19. Ectopic expression of the agouti gene in transgenic mice causes obesity, features of type II diabetes, and yellow fur

    SciTech Connect

    Klebig, M.L.; Woychik, R.P.; Wilkinson, J.E.; Geisler, J.G. |

    1995-05-23

    Mice that carry the lethal yellow (A{sup y}) or viable yellow (A{sup vy}) mutation, two dominant mutations of the agouti (a) gene in mouse chromosome 2, exhibit a phenotype that includes yellow fur, marked obesity, a form of type II diabetes associated with insulin resistance, and an increased susceptibility to tumor development. Molecular analyses of these and several other dominant {open_quotes}obese yellow{close_quotes} a-locus mutations suggested that ectopic expression of the normal agouti protein gives rise to this complex pleiotropic phenotype. We have now tested this hypothesis directly by generating transgenic mice that ectopically express an agouti cDNA clone encoding the normal agouti protein in all tissues examined. Transgenic mice of both sexes have yellow fur, become obese, and develop hyperinsulinemia. In addition, male transgenic mice develop hyperglycemia by 12-20 weeks of age. These results demonstrate conclusively that the ectopic agouti expression is responsible for most, if not all, of the phenotypic traits of the dominant, obese yellow mutants. 42 refs., 5 figs.

  20. The Drosophila melanogaster seminal fluid protein Acp62F is a protease inhibitor that is toxic upon ectopic expression.

    PubMed Central

    Lung, Oliver; Tram, Uyen; Finnerty, Casey M; Eipper-Mains, Marcie A; Kalb, John M; Wolfner, Mariana F

    2002-01-01

    Drosophila melanogaster seminal fluid proteins stimulate sperm storage and egg laying in the mated female but also cause a reduction in her life span. We report here that of eight Drosophila seminal fluid proteins (Acps) and one non-Acp tested, only Acp62F is toxic when ectopically expressed. Toxicity to preadult male or female Drosophila occurs upon one exposure, whereas multiple exposures are needed for toxicity to adult female flies. Of the Acp62F received by females during mating, approximately 10% enters the circulatory system while approximately 90% remains in the reproductive tract. We show that in the reproductive tract, Acp62F localizes to the lumen of the uterus and the female's sperm storage organs. Analysis of Acp62F's sequence, and biochemical assays, reveals that it encodes a trypsin inhibitor with sequence and structural similarities to extracellular serine protease inhibitors from the nematode Ascaris. In light of previous results demonstrating entry of Acp62F into the mated female's hemolymph, we propose that Acp62F is a candidate for a molecule to contribute to the Acp-dependent decrease in female life span. We propose that Acp62F's protease inhibitor activity exerts positive protective functions in the mated female's reproductive tract but that entry of a small amount of this protein into the female's hemolymph could contribute to the cost of mating. PMID:11805057

  1. Ectopic expression of ceramide synthase 2 in neurons suppresses neurodegeneration induced by ceramide synthase 1 deficiency

    PubMed Central

    Spassieva, Stefka D.; Ji, Xiaojie; Liu, Ye; Gable, Kenneth; Bielawski, Jacek; Dunn, Teresa M.; Bieberich, Erhard; Zhao, Lihong

    2016-01-01

    Sphingolipids exhibit extreme functional and chemical diversity that is in part determined by their hydrophobic moiety, ceramide. In mammals, the fatty acyl chain length variation of ceramides is determined by six (dihydro)ceramide synthase (CerS) isoforms. Previously, we and others showed that mutations in the major neuron-specific CerS1, which synthesizes 18-carbon fatty acyl (C18) ceramide, cause elevation of long-chain base (LCB) substrates and decrease in C18 ceramide and derivatives in the brain, leading to neurodegeneration in mice and myoclonus epilepsy with dementia in humans. Whether LCB elevation or C18 ceramide reduction leads to neurodegeneration is unclear. Here, we ectopically expressed CerS2, a nonneuronal CerS producing C22–C24 ceramides, in neurons of Cers1-deficient mice. Surprisingly, the Cers1 mutant pathology was almost completely suppressed. Because CerS2 cannot replenish C18 ceramide, the rescue is likely a result of LCB reduction. Consistent with this hypothesis, we found that only LCBs, the substrates common for all of the CerS isoforms, but not ceramides and complex sphingolipids, were restored to the wild-type levels in the Cers2-rescued Cers1 mutant mouse brains. Furthermore, LCBs induced neurite fragmentation in cultured neurons at concentrations corresponding to the elevated levels in the CerS1-deficient brain. The strong association of LCB levels with neuronal survival both in vivo and in vitro suggests high-level accumulation of LCBs is a possible underlying cause of the CerS1 deficiency-induced neuronal death. PMID:27162368

  2. Wnt activation downregulates olfactomedin-1 in Fallopian tubal epithelial cells: a microenvironment predisposed to tubal ectopic pregnancy.

    PubMed

    Kodithuwakku, Suranga P; Pang, Ronald T K; Ng, Ernest H Y; Cheung, Annie N Y; Horne, Andrew W; Ho, Pak-Chung; Yeung, William S B; Lee, Kai-Fai

    2012-02-01

    Ectopic pregnancy (EP) occurs when the embryo fails to transit to the uterus and attach to the luminal epithelium of the Fallopian tube (FT). Tubal EP is a common gynecological emergency and more than 95% of EP occurs in the ampullary region of the FT. In humans, Wnt activation and downregulation of olfactomedin-1 (Olfm-1) occur in the receptive endometrium and coincided with embryo implantation in vivo. Whether similar molecular changes happen in the FT leading to EP remains unclear. We hypothesized that activation of Wnt signaling downregulates Olfm-1 expression predisposes to EP. We investigated the spatiotemporal expression of Olfm-1 in FT from non-pregnant women and women with EP, and used a novel trophoblastic spheroid (embryo surrogate)-FT epithelial cell co-culture model (JAr and OE-E6/E7 cells) to study the role of Olfm-1 on spheroid attachment. Olfm-1 mRNA expression in the ampullary region of non-pregnant FT was higher (P<0.05) in the follicular phase than in the luteal phase. Ampullary tubal Olfm-1 expression was lower in FT from women with EP compared to normal controls at the luteal phase (histological scoring (H-SCORE)=1.3±0.2 vs 2.4±0.5; P<0.05). Treatment of OE-E6/E7 with recombinant Olfm-1 (0.2-5 μg/ml) suppressed spheroid attachment to OE-E6/E7 cells, while activation of Wnt-signaling pathway by Wnt3a or LiCl reduced endogenous Olfm-1 expression and increased spheroid attachment. Conversely, suppression of Olfm-1 expression by RNAi increased spheroid attachment to OE-E6/E7 cells. Taken together, Wnt activation suppresses Olfm-1 expression, and this may predispose a favorable microenvironment of the retained embryo in the FT, leading to EP in humans.

  3. Endometrial Gene Expression in Early Pregnancy: Lessons From Human Ectopic Pregnancy

    PubMed Central

    Savaris, Ricardo F.; Hamilton, Amy E.; Lessey, Bruce A.; Giudice, Linda C.

    2010-01-01

    Human endometrium undergoes modifications in preparation for embryonic implantation. This study investigated in vivo the endocrine effects of pregnancy on the endometrium, using the model of ectopic pregnancy. Endometrial biopsies from 9 subjects with ectopic pregnancy (Preg) were compared with 8 and 6 samples of mid and late secretory endometrium, respectively. After hybridizing with Affymetrix HGU133 Plus 2 chips, data were analyzed using GeneSpring GX and Ingenuity Pathways Analysis. From 54 675 genes, 3021 genes were significantly differentiated when mid-secretory endometrium was compared with the Preg (Volcano plot; P < .05, ≥2-fold change). The complement and coagulation cascade, phospholid degradation, glycosphingolipid biosynthesis (globoseries), retinol metabolism, antigen presentation pathway, glycosphingolipid biosynthesis, and O-glycan biosynthesis were main significant canonical pathways found in Preg samples. Validation was done with reverse transcriptase polymerase chain reaction. In conclusion, the ectopic embryo has a significant impact, by an endocrine mechanism, on endometrium, when compared with the window of implantation. PMID:18591649

  4. Noise-induced ectopic activity in a simple cardiac cell model

    NASA Astrophysics Data System (ADS)

    Hastings, Harold

    2005-03-01

    Ectopic activity in the form of premature ventricular contractions (PVCs) is relatively common in the normal heart. Although PVCs are normally harmless, sometimes but rarely PVCs can generate spiral waves of activation through interaction with other waves of activation, potentially progressing to ventricular tachycardia, followed by ventricular fibrillation and sudden cardiac death. Clusters of PVCs have been found to be significantly more dangerous than isolated PVCs. We model PVC generation by applying triggers (noise) to the generic FitzHugh-Nagumo model as substrate, and study the effects the noise level and excitability. We find: exponential waiting time behavior at fixed parameter levels; no evidence of clustering at fixed parameter levels; and a sharp increase in PVCs as excitability approaches the auto-oscillatory threshold or noise increases beyond a similar threshold. This produces sharp increases in theoretical rates of PVC-induced fibrillation, consistent with results of A Gelzer et al. in animal models. Partially supported by the NSF and NIH.

  5. Ectopic Expression of Transcription Factor AP-2δ in Developing Retina: Effect on PSA-NCAM and Axon Routing

    PubMed Central

    Li, Xiaodong; Monckton, Elizabeth A.; Godbout, Roseline

    2015-01-01

    Retinal ganglion cells transmit the visual signal from the retina to the brain. We have previously shown that the AP-2δ (TFAP2D) transcription factor is expressed in one-third of ganglion cells in developing retina suggesting a specialized role for these AP-2δ-expressing cells. Here, we address the role of AP-2δ in retina by in ovo electroporation of RCAS/AP-2δ retroviral constructs into the eyes of chick embryos at day 2 of gestation. Ectopic expression of AP-2δ does not affect lineage differentiation in the developing retina. However, immunostaining of retinal tissue with markers associated with axonal growth such as GAP43 and PSA-NCAM demonstrates axonal misrouting and abnormal axonal bundling. Treatment of AP-2δ-misexpressing retinal cell cultures with Endo-N, an enzyme that removes PSA from NCAM, decreases AP-2δ-induced axonal bundling. Our data suggest a role for AP-2δ in polysialylation of NCAM, with ectopic expression of AP-2δ resulting in premature bundling of emerging axons and misrouting of axons. We propose that expression of AP-2δ in a subset of ganglion cells contributes to the fine-tuning of axonal growth in the developing retina. PMID:24188130

  6. MiRNA profile associated with replicative senescence, extended cell culture, and ectopic telomerase expression in human foreskin fibroblasts.

    PubMed

    Bonifacio, Laura N; Jarstfer, Michael B

    2010-09-01

    Senescence is a highly regulated process that limits cellular replication by enforcing a G1 arrest in response to various stimuli. Replicative senescence occurs in response to telomeric DNA erosion, and telomerase expression can offset replicative senescence leading to immortalization of many human cells. Limited data exists regarding changes of microRNA (miRNA) expression during senescence in human cells and no reports correlate telomerase expression with regulation of senescence-related miRNAs. We used miRNA microarrays to provide a detailed account of miRNA profiles for early passage and senescent human foreskin (BJ) fibroblasts as well as early and late passage immortalized fibroblasts (BJ-hTERT) that stably express the human telomerase reverse transcriptase subunit hTERT. Selected miRNAs that were differentially expressed in senescence were assayed for expression in quiescent cells to identify miRNAs that are specifically associated with senescence-associated growth arrest. From this group of senescence-associated miRNAs, we confirmed the ability of miR-143 to induce growth arrest after ectopic expression in young fibroblasts. Remarkably, miR-143 failed to induce growth arrest in BJ-hTERT cells. Importantly, the comparison of late passage immortalized fibroblasts to senescent wild type fibroblasts reveals that miR-146a, a miRNA with a validated role in regulating the senescence associated secretory pathway, is also regulated during extended cell culture independently of senescence. The discovery that miRNA expression is impacted by expression of ectopic hTERT as well as extended passaging in immortalized fibroblasts contributes to a comprehensive understanding of the connections between telomerase expression, senescence and processes of cellular aging.

  7. Ectopic over-expression of oncogene Pim-2 induce malignant transformation of nontumorous human liver cell line L02.

    PubMed

    Ren, Ke; Duan, Wentao; Shi, Yujun; Li, Bo; Liu, Zuojin; Gong, Jiangping

    2010-07-01

    In order to prove that ectopic over-expression of Pim-2 could induce malignant transformation of human liver cell line L02, three groups of cells were set up including human liver cell line L02 (L02), L02 cells transfected with Pim-2 gene (L02/Pim-2) and L02 cells transfected with empty-vector (L02/Vector). Pim-2 expression levels were detected. The morphology, proliferation level, apoptosis rate and migration ability of the cells were detected respectively. Then the cells were subcutaneously inoculated into athymic mice and the microstructures of the neoplasm were observed. Compared with the controls, Pim-2 expression levels were significantly higher in L02/Pim-2 cells (P<0.05), and their morphology had obvious malignant changes. They also showed a significantly increased proliferation rate (P<0.05) and migration capacity (P<0.05), as well as a significantly decreased apoptosis rate (P<0.05). Only the athymic mice inoculated with L02/Pim-2 cells could generate neoplasm, and the morphology of the neoplasm coincided with that of the hepatoma. The results manifest that ectopic Pim-2 gene could be stably expressed in L02/Pim-2 cells. Both the morphological and biological changes of L02/Pim-2 cells demonstrate the trend of malignant transformation. L02/Pim-2 cells could generate hepatoma in athymic mice. In conclusion, Pim-2 could induce malignant transformation of human liver cell line L02.

  8. Constitutive activation of ectodermal β-catenin induces ectopic outgrowths at various positions in mouse embryo and affects abdominal ventral body wall closure.

    PubMed

    Zhu, Xuming; Huang, Sixia; Zhang, Lingling; Wu, Yumei; Chen, Yingwei; Tao, Yixin; Wang, Yushu; He, Shigang; Shen, Sanbing; Wu, Ji; Li, Baojie; Guo, Xizhi; He, Lin; Ma, Gang

    2014-01-01

    Vertebrate limbs originate from the lateral plate mesoderm (LPM) and the overlying ectoderm. While normal limb formation in defined regions has been well studied, the question of whether other positions retain limb-forming potential has not been fully investigated in mice. By ectopically activating β-catenin in the ectoderm with Msx2-cre, we observed that local tissue outgrowths were induced, which either progressed into limb-like structure within the inter-limb flank or formed extra tissues in other parts of the mouse embryo. In the presumptive abdominal region of severely affected embryos, ectopic limb formation was coupled with impaired abdominal ventral body wall (AVBW) closure, which indicates the existence of a potential counterbalance of limb formation and AVBW closure. At the molecular level, constitutive β-catenin activation was sufficient to trigger, but insufficient to maintain the ectopic expression of a putative limb-inducing factor, Fgf8, in the ectoderm. These findings provide new insight into the mechanism of limb formation and AVBW closure, and the crosstalk between the Wnt/β-catenin pathway and Fgf signal.

  9. Phenotypic alterations of petal and sepal by ectopic expression of a rice MADS box gene in tobacco.

    PubMed

    Kang, H G; Noh, Y S; Chung, Y Y; Costa, M A; An, K; An, G

    1995-10-01

    Floral organ development is controlled by a group of regulatory factors containing the MADS domain. In this study, we have isolated and characterized a cDNA clone from rice, OsMADS3, which encodes a MADS-domain containing protein. The OsMADS3 amino acid sequence shows over 60% identity to AG of Arabidopsis, PLE of Antirrhinum majus, and AG/PLE homologues of petunia, tobacco, tomato, Brassica napus, and maize. Homology in the MADS box region is most conserved. RNA blot analysis indicated that the rice MADS gene was preferentially expressed in reproductive organs, especially in stamen and carpel. In situ localization studies showed that the transcript was present primarily in stamen and carpel. The function of the rice OsMADS3 was elucidated by ectopic expression of the gene under the control of the CaMV 35S promoter in a heterologous tobacco plant system. Transgenic plants exhibited an altered morphology and coloration of the perianth organs. Sepals were pale green and elongated. Limbs of the corolla were split into sections which in some plants became antheroid structures attached to tubes that resembled filaments. The phenotypes mimic the results of ectopic expression of dicot AG gene or AG homologues. These results indicate that the OsMADS3 gene is possibly an AG homologue and that the AG genes appear to be structurally and functionally conserved between dicot and monocot.

  10. In vitro reprogramming of pancreatic alpha cells towards a beta cell phenotype following ectopic HNF4α expression.

    PubMed

    Sangan, Caroline B; Jover, Ramiro; Heimberg, Harry; Tosh, David

    2015-01-05

    There is currently a shortage of organ donors available for pancreatic beta cell transplantation into diabetic patients. An alternative source of beta cells is pre-existing pancreatic cells. While we know that beta cells can arise directly from alpha cells during pancreatic regeneration we do not understand the molecular basis for the switch in phenotype. The aim of the present study was to investigate if hepatocyte nuclear factor 4 alpha (HNF4α), a transcription factor essential for a normal beta cell phenotype, could induce the reprogramming of alpha cells towards potential beta cells. We utilised an in vitro model of pancreatic alpha cells, the murine αTC1-9 cell line. We initially characterised the αTC1-9 cell line before and following adenovirus-mediated ectopic expression of HNF4α. We analysed the phenotype at transcript and protein level and assessed its glucose-responsiveness. Ectopic HNF4α expression in the αTC1-9 cell line induced a change in morphology (1.7-fold increase in size), suppressed glucagon expression, induced key beta cell-specific markers (insulin, C-peptide, glucokinase, GLUT2 and Pax4) and pancreatic polypeptide (PP) and enabled the cells to secrete insulin in a glucose-regulated manner. In conclusion, HNF4α reprograms alpha cells to beta-like cells.

  11. A Regional Reduction in Ito and IKACh in the Murine Posterior Left Atrial Myocardium Is Associated with Action Potential Prolongation and Increased Ectopic Activity

    PubMed Central

    Tull, Samantha; Syeda, Fahima; Kuhlmann, Stefan M.; O’Brien, Sian-Marie; Patel, Pushpa; Brain, Keith L.; Pavlovic, Davor; Brown, Nigel A.; Fabritz, Larissa; Kirchhof, Paulus

    2016-01-01

    Background The left atrial posterior wall (LAPW) is potentially an important area for the development and maintenance of atrial fibrillation. We assessed whether there are regional electrical differences throughout the murine left atrial myocardium that could underlie regional differences in arrhythmia susceptibility. Methods We used high-resolution optical mapping and sharp microelectrode recordings to quantify regional differences in electrical activation and repolarisation within the intact, superfused murine left atrium and quantified regional ion channel mRNA expression by Taqman Low Density Array. We also performed selected cellular electrophysiology experiments to validate regional differences in ion channel function. Results Spontaneous ectopic activity was observed during sustained 1Hz pacing in 10/19 intact LA and this was abolished following resection of LAPW (0/19 resected LA, P<0.001). The source of the ectopic activity was the LAPW myocardium, distinct from the pulmonary vein sleeve and LAA, determined by optical mapping. Overall, LAPW action potentials (APs) were ca. 40% longer than the LAA and this region displayed more APD heterogeneity. mRNA expression of Kcna4, Kcnj3 and Kcnj5 was lower in the LAPW myocardium than in the LAA. Cardiomyocytes isolated from the LAPW had decreased Ito and a reduced IKACh current density at both positive and negative test potentials. Conclusions The murine LAPW myocardium has a different electrical phenotype and ion channel mRNA expression profile compared with other regions of the LA, and this is associated with increased ectopic activity. If similar regional electrical differences are present in the human LA, then the LAPW may be a potential future target for treatment of atrial fibrillation. PMID:27149380

  12. Endogenous and ectopic expression of telomere regulating genes in chicken embryonic fibroblasts

    SciTech Connect

    Michailidis, Georgios; Saretzki, Gabriele; Hall, Judith , E-Mail: Judith.hall@ncl.ac.uk

    2005-09-16

    In this study, we compared the endogenous expression of genes encoding telomere regulating proteins in cultured chicken embryonic fibroblasts (CEFs) and 10-day-old chicken embryos. CEFs maintained in vitro senesced and senescence was accompanied by reduced telomere length, telomerase activity, and expression of the chicken (c) TRF1 gene. There was no change in TRF2 gene expression although the major TRF2 transcript identified in 10-day-old chicken embryos encoded a truncated TRF2 protein (TRF2'), containing an N-terminal dimerisation domain but lacking a myb-related DNA binding domain and nuclear localisation signal. Senescence of the CEFs in vitro was associated with the loss of the TRF2' transcript, indicative of a novel function for the encoded protein. Senescence was also coupled with decreased expression of RAD51, but increased RAD52 expression. These data support that RAD51 independent recombination mechanisms do not function in vitro to maintain chicken telomeres. To attempt to rescue the CEFs from replicative senescence, we stably transfected passage 3 CEFs with the human telomerase reverse transcriptase (hTERT) catalytic subunit. While hTERT expression was detected in the stable transfectants neither telomerase activity nor the stabilisation of telomere length was observed, and the transfectant cells senesced at the same passage number as the untransfected cells. These data indicate that the human TERT is incompatible with the avian telomere maintenance apparatus and suggest the functioning of a species specific telomere system in the avian.

  13. Functional characterization of the apple MhGAI1 gene through ectopic expression and grafting experiments in tomatoes.

    PubMed

    Wang, Shuang-Shuang; Liu, Ze-Zhou; Sun, Chao; Shi, Qing-Hua; Yao, Yu-Xin; You, Chun-Xiang; Hao, Yu-Jin

    2012-02-15

    DELLA proteins are essential components of GA signal transduction. MhGAI1 was isolated from the tea crabapple (Malus hupehensis Redh. var. pingyiensis), and it was found to encode a DELLA protein. Mhgai1 is a GA-insensitive allele that was artificially generated via a bridge-PCR approach. Ectopic expression of Mhgai1 reduced plant stature, decreased spontaneous fruit-set-ratio and enhanced drought-tolerance in transgenic tomatoes. In addition, we examined the long-distance movement of Mhgai1 mRNAs by grafting experiments and SqRT-PCR analysis. It was found that the wild-type scions accumulated Mhgai1 transcripts trafficked from the transgenic rootstocks and therefore exhibited dwarf phenotypes. Furthermore, transgenic tomato plants produced more soluble solids, sugars and organic acids compared to wild-type tomatoes, suggesting an involvement of GA signaling in the regulation of fruit quality. Despite noticeable accumulation in the leaves and stems of WT scions, Mhgai1 transcripts were undetectable in flowers and fruit. Therefore, fruit quality was less influenced by the grafting of WT scions onto transgenic rootstocks than they were by the ectopic expression of Mhgai1 in transgenic rootstocks. Taken together, MhGAI1, which functions as a repressor in the GA signaling pathway, and its GA-insensitive allele, Mhgai1, could turn out to be useful targets for the genetic improvement of dwarfing rootstocks in apples.

  14. The developmental dismantling of pluripotency is reversed by ectopic Oct4 expression

    PubMed Central

    Osorno, Rodrigo; Tsakiridis, Anestis; Wong, Frederick; Cambray, Noemí; Economou, Constantinos; Wilkie, Ronald; Blin, Guillaume; Scotting, Paul J.; Chambers, Ian; Wilson, Valerie

    2012-01-01

    The transcription factors Nanog and Oct4 regulate pluripotency in the pre-implantation epiblast and in derivative embryonic stem cells. During post-implantation development, the precise timing and mechanism of the loss of pluripotency is unknown. Here, we show that in the mouse, pluripotency is extinguished at the onset of somitogenesis, coincident with reduced expression and chromatin accessibility of Oct4 and Nanog regulatory regions. Prior to somitogenesis expression of both Nanog and Oct4 is regionalized. We show that pluripotency tracks the in vivo level of Oct4 and not Nanog by assessing the ability to reactivate or maintain Nanog expression in cell culture. Enforced Oct4 expression in somitogenesis-stage tissue provokes rapid reopening of Oct4 and Nanog chromatin, Nanog re-expression and resuscitates moribund pluripotency. Our data suggest that decreasing Oct4 expression is converted to a sudden drop in competence to maintain pluripotency gene regulatory network activity that is subsequently stabilized by epigenetic locks. PMID:22669820

  15. Ectopic pregnancy

    MedlinePlus

    Tubal pregnancy; Cervical pregnancy; Tubal ligation - ectopic pregnancy ... In most pregnancies, the fertilized egg travels through the fallopian tube to the womb (uterus). If the movement of the egg ...

  16. ESCRT-0 is not required for ectopic Notch activation and tumor suppression in Drosophila.

    PubMed

    Tognon, Emiliana; Wollscheid, Nadine; Cortese, Katia; Tacchetti, Carlo; Vaccari, Thomas

    2014-01-01

    Multivesicular endosome (MVE) sorting depends on proteins of the Endosomal Sorting Complex Required for Transport (ESCRT) family. These are organized in four complexes (ESCRT-0, -I, -II, -III) that act in a sequential fashion to deliver ubiquitylated cargoes into the internal luminal vesicles (ILVs) of the MVE. Drosophila genes encoding ESCRT-I, -II, -III components function in sorting signaling receptors, including Notch and the JAK/STAT signaling receptor Domeless. Loss of ESCRT-I, -II, -III in Drosophila epithelia causes altered signaling and cell polarity, suggesting that ESCRTs genes are tumor suppressors. However, the nature of the tumor suppressive function of ESCRTs, and whether tumor suppression is linked to receptor sorting is unclear. Unexpectedly, a null mutant in Hrs, encoding one of the components of the ESCRT-0 complex, which acts upstream of ESCRT-I, -II, -III in MVE sorting is dispensable for tumor suppression. Here, we report that two Drosophila epithelia lacking activity of Stam, the other known components of the ESCRT-0 complex, or of both Hrs and Stam, accumulate the signaling receptors Notch and Dome in endosomes. However, mutant tissue surprisingly maintains normal apico-basal polarity and proliferation control and does not display ectopic Notch signaling activation, unlike cells that lack ESCRT-I, -II, -III activity. Overall, our in vivo data confirm previous evidence indicating that the ESCRT-0 complex plays no crucial role in regulation of tumor suppression, and suggest re-evaluation of the relationship of signaling modulation in endosomes and tumorigenesis.

  17. Excessive Supraventricular Ectopic Activity Is Indicative of Paroxysmal Atrial Fibrillation in Patients with Cerebral Ischemia

    PubMed Central

    Weber-Krüger, Mark; Gröschel, Klaus; Mende, Meinhard; Seegers, Joachim; Lahno, Rosine; Haase, Beatrice; Niehaus, Cord-Friedrich; Edelmann, Frank; Hasenfuß, Gerd

    2013-01-01

    Background Detecting paroxysmal atrial fibrillation (PAF) in patients with cerebral ischemia is challenging. Frequent premature atrial complexes (PAC/h) and the longest supraventricular run on 24-h-Holter (SV-run24 h), summarised as excessive supraventricular ectopic activity (ESVEA), may help selecting patients for extended ECG-monitoring, especially in combination with echocardiographic marker LAVI/a’ (left atrial volume index/late diastolic tissue Doppler velocity). Methods Retrospective analysis from the prospective monocentric observational trial Find-AF (ISRCTN-46104198). Patients with acute stroke or TIA were enrolled at the University Hospital Göttingen, Germany. Those with sinus rhythm at presentation received 7-day Holter-monitoring. ESVEA was quantified in one 24-hour interval free from PAF. Echocardiographic parameters were assessed prospectively. Results PAF was detected in 23/208 patients (11.1%). The median was 4 [IQR 1; 22] for PAC/h and 5 [IQR 0; 9] for SV-run24 h. PAF was more prevalent in patients with ESVEA: 19.6% vs. 2.8% for PAC/h >4 vs. ≤4 (p<0.001); 17.0% vs. 4.9% for SV-run24 h >5 vs. ≤5 beats (p = 0.003). Patients with PAF showed more supraventricular ectopic activity: 29 PAC/h [IQR 9; 143] vs. 4 PAC/h [1]; [14] and longest SV-run24 h = 10 [5]; [21] vs. 0 [0; 8] beats (both p<0.001). Both markers discriminated between the PAF- and the Non-PAF-group (area under receiver-operator-characteristics-curve 0.763 [95% CI 0.667; 0.858] and 0.716 [0.600; 0.832]). In multivariate analyses log(PAC/h) and log(SV-run24 h) were independently indicative of PAF. In Patients with PAC/h ≤4 and normal LAVI/a’ PAF was excluded, whereas those with PAC/h >4 and abnormal LAVI/a’ showed high PAF-rates. Conclusions ESVEA discriminated PAF from non-PAF beyond clinical factors including LAVI/a’ in patients with cerebral ischemia. Normal LAVI/a’+PAC/h ≤4 ruled out PAF, while prevalence was high in those with abnormal LAVI/a’+PAC/h >4. PMID

  18. Novel phosphate-activated macrophages prevent ectopic calcification by increasing extracellular ATP and pyrophosphate

    PubMed Central

    Villa-Bellosta, Ricardo; Hamczyk, Magda R.; Andrés, Vicente

    2017-01-01

    Purpose Phosphorus is an essential nutrient involved in many pathobiological processes. Less than 1% of phosphorus is found in extracellular fluids as inorganic phosphate ion (Pi) in solution. High serum Pi level promotes ectopic calcification in many tissues, including blood vessels. Here, we studied the effect of elevated Pi concentration on macrophage polarization and calcification. Macrophages, present in virtually all tissues, play key roles in health and disease and display remarkable plasticity, being able to change their physiology in response to environmental cues. Methods and results High-throughput transcriptomic analysis and functional studies demonstrated that Pi induces unpolarized macrophages to adopt a phenotype closely resembling that of alternatively-activated M2 macrophages, as revealed by arginine hydrolysis and energetic and antioxidant profiles. Pi-induced macrophages showed an anti-calcifying action mediated by increased availability of extracellular ATP and pyrophosphate. Conclusion We conclude that the ability of Pi-activated macrophages to prevent calcium-phosphate deposition is a compensatory mechanism protecting tissues from hyperphosphatemia-induced pathologic calcification. PMID:28362852

  19. Ectopic expression of HLA-DO in mouse dendritic cells diminishes MHC class II antigen presentation.

    PubMed

    Fallas, Jennifer L; Tobin, Helen M; Lou, Olivia; Guo, Donglin; Sant'Angelo, Derek B; Denzin, Lisa K

    2004-08-01

    The MHC class II-like molecule HLA-DM (DM) (H-2M in mice) catalyzes the exchange of CLIP for antigenic peptides in the endosomes of APCs. HLA-DO (DO) (H-2O in mice) is another class II-like molecule that is expressed in B cells, but not in other APCs. Studies have shown that DO impairs or modifies the peptide exchange activity of DM. To further evaluate the role of DO in Ag processing and presentation, we generated transgenic mice that expressed the human HLA-DOA and HLA-DOB genes under the control of a dendritic cell (DC)-specific promoter. Our analyses of DCs from these mice showed that as DO levels increased, cell surface levels of A(b)-CLIP also increased while class II-peptide levels decreased. The presentation of some, but not all, exogenous Ags to T cells or T hybridomas was significantly inhibited by DO. Surprisingly, H-2M accumulated in DO-expressing DCs and B cells, suggesting that H-2O/DO prolongs the half-life of H-2M. Overall, our studies showed that DO expression impaired H-2M function, resulting in Ag-specific down-modulation of class II Ag processing and presentation.

  20. The Expression and Cellular Localization of Galectin-1 and Galectin-3 in the Fallopian Tube Are Altered in Women with Tubal Ectopic Pregnancy.

    PubMed

    Nio-Kobayashi, Junko; Abidin, Hazirah B Z; Brown, Jeremy K; Iwanaga, Toshihiko; Horne, Andrew W; Duncan, W Colin

    2015-01-01

    Galectin-1 and galectin-3 are abundantly expressed at implantation sites in the uterus, suggesting their involvement in the establishment of pregnancy. In this study, we examined the expression and localization of galectin-1 and galectin-3 in fallopian tubes from nonpregnant women, and in those presenting with tubal ectopic pregnancy. There was no significant difference in the expression of either galectin-1 (LGALS1) or galectin-3 (LGALS3) transcripts in the fallopian tube across the menstrual cycle. Their expressions in the fallopian tube were inversely correlated to each other (r = -0.5134, p < 0.0001) and differentially localized. Galectin-1 protein was abundant in the stroma of nonpregnant fallopian tubes, whereas galectin-3 was mainly localized to the epithelium, notably to the cilia of ciliated cells and the apical cytoplasm of secretory cells. In ectopic pregnancies, LGALS3 expression was significantly reduced (p < 0.0001), but LGALS1 expression did not change when compared to nonpregnant fallopian tubes collected during the mid-secretory phase. The percentage of fallopian tube epithelial cells expressing galectin-3 in cilia tended to be reduced (p = 0.0685), with an accompanying loss of a normal ciliary structure, while nuclear galectin-3 increased (p < 0.05) in ectopic pregnancies. Epithelial immunostaining for galectin-1 tended to be elevated in fallopian tubes from women with ectopic pregnancy. Coculture of human trophoblast origin SW71 cells significantly increased LGALS1 expression in human fallopian tube epithelial OE-E6/E7 cells, suggesting that trophoblast-derived products regulate LGALS1 expression in the oviductal epithelium. These findings imply a differential contribution of galectin-1 and galectin-3 in the homeostasis of human fallopian tubes and in the pathophysiology of ectopic pregnancy.

  1. Ectopic expression of SOX9 in osteoblasts alters bone mechanical propertie

    PubMed Central

    Liang, Bojian; Cotter, Meghan M.; Chen, Dongxing; Hernandez, Christopher J.; Zhou, Guang

    2011-01-01

    Osteoporosis is a common skeletal disease characterized by low bone mass and micro-architectural deterioration of bone tissue, with a consequent increase in bone fragility and susceptibility to fracture. We previously demonstrated that Col1a1-SOX9 transgenic mice, in which SOX9 specifically expresses in osteoblasts driven by a 2.3kb Col1a1 promoter, display osteopenia during the early postnatal stage. In this study, to further analyze the osteopenia phenotype and especially the effect of the osteoblast-specific expression of SOX9 on bone mechanical properties, we performed bone geometry and mechanical property analysis of long bones from Col1a1-SOX9 transgenic mice (TG) and wild type littermates (WT) at different time points. Interestingly, after body weight adjustment, TG mice have similar whole-bone strength as WT mice, but exhibit significantly thinner cortical bone, lower elastic modulus, and higher moment of inertia. Thus, osteoblast-specific SOX9 expression results in altered bone structure and material properties. Furthermore, the expression levels of Pcna, Col1a1, Osteocalcin, and the Opg/Rankl ratio in TG mice are significantly lower until 4 months of age compared with WT mice, suggesting that TG mice have dysregulated bone homeostasis. Finally, bone marrow stromal cells (MSCs) isolated from TG mice display enhanced adipocyte differentiation and decreased osteoblast differentiation in vitro, suggesting that osteoblast-specific expression of SOX9 can lead to altered mesenchymal stem cell differentiation potentials. In conclusion, our study implies that SOX9 activity has to be tightly regulated in adult skeleton to ensure optimal bone quality. PMID:22143895

  2. Ectopic Expression of OsSta2 Enhances Salt Stress Tolerance in Rice

    PubMed Central

    Kumar, Manu; Choi, Juyoung; An, Gynheung; Kim, Seong-Ryong

    2017-01-01

    Salt stress can severely reduce crop yields. To understand how rice (Oryza sativa) plants respond to this environmental challenge, we investigated the genes involved in conferring salt tolerance by screening T-DNA tagging lines and identified OsSta2-D (Oryza sativa Salt tolerance activation 2-Dominant). In that line, expression of OsSta2 was enhanced by approximately eightfold when compared with the non-transformed wild type (WT). This gene was highly expressed in the callus, roots, and panicles. To confirm its role in stress tolerance, we generated transgenic rice that over-expresses OsSta2 under a maize ubiquitin promoter. The OsSta2-Ox plants were salt-tolerant at the vegetative stage, based on our calculations of chlorophyll fluorescence (Fv/Fm), fresh and dry weights, chlorophyll concentrations, and survival rates. Under normal paddy field conditions, the Ox plants were somewhat shorter than the WT control but had improved agronomic traits such as higher total grain yield. They were also more tolerant to osmotic stress and hypersensitive to abscisic acid. Based on all of these results, we suggest that OsSta2 has important roles in determining yields as well as in conferring tolerance to salt stresses. PMID:28344585

  3. Ectopic expression of Arabidopsis FD and FD PARALOGUE in rice results in dwarfism with size reduction of spikelets

    PubMed Central

    Jang, Seonghoe; Li, Hsing-Yi; Kuo, Mei-Lin

    2017-01-01

    Key flowering genes, FD and FD PARALOGUE (FDP) encoding bZIP transcription factors that interact with a FLOWERING LOCUS T (FT) in Arabidopsis were ectopically expressed in rice since we found AtFD and AtFDP also interact with HEADING DATE 3a (Hd3a) and RICE FLOWERING LOCUS T 1 (RFT1). Transgenic rice plants overexpressing AtFD and AtFDP caused reduction in plant height and spikelet size with decreased expression of genes involved in cell elongation without significant flowering time alteration in spite of increased expression of OsMADS14 and OsMADS15, rice homologues of APETALA1 (AP1) in the leaves. Simultaneous overexpression of AtFD and AtFDP enhanced phenotypes seen with overexpression of either single gene while transgenic rice plants expressing AtFD or AtFDP under the control of phloem-specific Hd3a promoter were indistinguishable from wild-type rice. Candidate genes responsible for the phenotypes were identified by comparison of microarray hybridization and their expression pattern was also examined in WT and transgenic rice plants. It has so far not been reported that AtFD and AtFDP affect cell elongation in plants, and our findings provide novel insight into the possible roles of AtFD and AtFDP in the mesophyll cells of plants, and potential genetic tools for manipulation of crop architecture. PMID:28290557

  4. Spatial distribution of the RABBIT EARS protein and effects of its ectopic expression in Arabidopsis thaliana flowers.

    PubMed

    Takeda, Seiji; Noguchi, Mariko; Hamamura, Yuki; Higashiyama, Tetsuya

    2014-03-01

    In many flowering plants, flowers consist of two peripheral organs, sepals and petals, occurring in outer two whorls, and two inner reproductive organs, stamens and carpels. These organs are arranged in a concentric pattern in a floral meristem, and the organ identity is established by the combined action of floral homeotic genes expressed along the whorls. Floral organ primordia arise at fixed positions in the floral meristem within each whorl. The RABBIT EARS (RBE) gene is transcribed in the petal precursor cells and primordia, and regulates petal initiation and early growth in Arabidopsis thaliana. We investigated the spatial and temporal expression pattern of a RBE protein fused to the green fluorescent protein (GFP). Expression of the GFP:RBE fusion gene under the RBE cis-regulatory genomic fragment rescues the rbe petal defects, indicating that the fusion protein is functional. The GFP signal is located to the cells where RBE is transcribed, suggesting that RBE function is cell-autonomous. Ectopic expression of GFP:RBE under the APETALA1 promoter causes the homeotic conversion of floral organs, resulting in sterile flowers. In these plants, the class B homeotic genes APETALA3 and PISTILLATA are down-regulated, suggesting that the restriction of the RBE expression to the petal precursor cells is crucial for flower development.

  5. Identification of a Copper-Inducible Promoter for Use in Ectopic Expression in the Fungal Pathogen Histoplasma capsulatum†

    PubMed Central

    Gebhart, Dana; Bahrami, Adam K.; Sil, Anita

    2006-01-01

    Despite the existence of a number of genetic tools to study the fungal pathogen Histoplasma capsulatum, strategies for conditional gene expression have not been developed. We used microarray analysis to identify genes that are transcriptionally induced or repressed by the addition of copper sulfate (CuSO4) to H. capsulatum yeast cultures. One of these genes, CRP1, encodes a putative copper efflux pump that is significantly induced in the presence of CuSO4. The upstream regulatory region of CRP1 was sufficient to drive copper-regulated expression of two reporter genes, lacZ and the gene encoding green fluorescent protein. Microarray experiments were performed to determine a copper concentration that triggers accumulation of the CRP1 transcript without significant perturbation of global gene expression. These studies show that the CRP1 upstream regulatory region can be used for ectopic expression of heterologous genes in H. capsulatum. Furthermore, they demonstrate the strategic use of microarrays to identify conditional promoters that confer induction in the absence of large-scale shifts in gene expression. PMID:16757741

  6. Ectopic expression of DREF induces DNA synthesis, apoptosis, and unusual morphogenesis in the Drosophila eye imaginal disc: possible interaction with Polycomb and trithorax group proteins.

    PubMed

    Hirose, F; Ohshima, N; Shiraki, M; Inoue, Y H; Taguchi, O; Nishi, Y; Matsukage, A; Yamaguchi, M

    2001-11-01

    The promoters of Drosophila genes encoding DNA replication-related proteins contain transcription regulatory element DRE (5'-TATCGATA) in addition to E2F recognition sites. A specific DRE-binding factor, DREF, positively regulates DRE-containing genes. In addition, it has been reported that DREF can bind to a sequence in the hsp70 scs' chromatin boundary element that is also recognized by boundary element-associated factor, and thus DREF may participate in regulating insulator activity. To examine DREF function in vivo, we established transgenic flies in which ectopic expression of DREF was targeted to the eye imaginal discs. Adult flies expressing DREF exhibited a severe rough eye phenotype. Expression of DREF induced ectopic DNA synthesis in the cells behind the morphogenetic furrow, which are normally postmitotic, and abolished photoreceptor specifications of R1, R6, and R7. Furthermore, DREF expression caused apoptosis in the imaginal disc cells in the region where commitment to R1/R6 cells takes place, suggesting that failure of differentiation of R1/R6 photoreceptor cells might cause apoptosis. The DREF-induced rough eye phenotype was suppressed by a half-dose reduction of the E2F gene, one of the genes regulated by DREF, indicating that the DREF overexpression phenotype is useful to screen for modifiers of DREF activity. Among Polycomb/trithorax group genes, we found that a half-dose reduction of some of the trithorax group genes involved in determining chromatin structure or chromatin remodeling (brahma, moira, and osa) significantly suppressed and that reduction of Distal-less enhanced the DREF-induced rough eye phenotype. The results suggest a possibility that DREF activity might be regulated by protein complexes that play a role in modulating chromatin structure. Genetic crosses of transgenic flies expressing DREF to a collection of Drosophila deficiency stocks allowed us to identify several genomic regions, deletions of which caused enhancement or

  7. Soaking RNAi-mediated modification of Sf9 cells for baculovirus expression system by ectopic expression of Caenorhabditis elegans SID-1.

    PubMed

    Xu, Jian; Nagata, Yudai; Mon, Hiroaki; Li, Zhiqing; Zhu, Li; Iiyama, Kazuhiro; Kusakabe, Takahiro; Lee, Jae Man

    2013-07-01

    RNA interference (RNAi) is a biological phenomenon that silences the expression of genes of interest. Passive double-stranded RNA (dsRNA) uptake has been uniquely observed in Caenorhabditis elegans due to the expression of systemic RNAi defective-1 (SID-1). We report that ectopic expression of CeSID-1 endows the Sf9 cells with a capacity for soaking RNAi. Soaking the Sf9-SID1 with dsRNA corresponding to either exogenous or endogenous target genes induced a significant decrease in the amount of mRNA or protein. These results enabled us to modify the target proteins of baculovirus expression vector system in both quantities and posttranslational modifications. The current low-cost and high-efficiency RNAi system is useful for high-throughput gene function analysis and mass production of recombinant protein.

  8. Identification and molecular characterization of an IDA-like gene from litchi, LcIDL1, whose ectopic expression promotes floral organ abscission in Arabidopsis

    PubMed Central

    Ying, Peiyuan; Li, Caiqin; Liu, Xuncheng; Xia, Rui; Zhao, Minglei; Li, Jianguo

    2016-01-01

    Unexpected abscission of flowers or fruits is a major limiting factor for crop productivity. Key genes controlling abscission in plants, especially in popular fruit trees, are largely unknown. Here we identified a litchi (Litchi chinensis Sonn.) IDA-like (INFLORESCENCE DEFICIENT IN ABSCISSION-like) gene LcIDL1 as a potential key regulator of abscission. LcIDL1 encodes a peptide that shows the closest homology to Arabidopsis IDA, and is localized in cell membrane and cytoplasm. Real-time PCR analysis showed that the expression level of LcIDL1 accumulated gradually following flower abscission, and it was obviously induced by fruit abscission-promoting treatments. Transgenic plants expressing LcIDL1 in Arabidopsis revealed a role of LcIDL1 similar to IDA in promoting floral organ abscission. Moreover, ectopic expression of LcIDL1 in Arabidopsis activated the expression of abscission-related genes. Taken together, our findings provide evidence that LcIDL1 may act as a key regulator in control of abscission. PMID:27845425

  9. Ectopic expression of Hrf1 enhances bacterial resistance via regulation of diterpene phytoalexins, silicon and reactive oxygen species burst in rice.

    PubMed

    Li, Wenqi; Shao, Min; Zhong, Weigong; Yang, Jie; Okada, Kazunori; Yamane, Hisakazu; Zhang, Lei; Wang, Guang; Wang, Dong; Xiao, Shanshan; Chang, Shanshan; Qian, Guoliang; Liu, Fengquan

    2012-01-01

    Harpin proteins as elicitor derived from plant gram negative bacteria such as Xanthomonas oryzae pv. oryzae (Xoo), Erwinia amylovora induce disease resistance in plants by activating multiple defense responses. However, it is unclear whether phytoalexin production and ROS burst are involved in the disease resistance conferred by the expression of the harpin(Xoo) protein in rice. In this article, ectopic expression of hrf1 in rice enhanced resistance to bacterial blight. Accompanying with the activation of genes related to the phytoalexin biosynthesis pathway in hrf1-transformed rice, phytoalexins quickly and consistently accumulated concurrent with the limitation of bacterial growth rate. Moreover, the hrf1-transformed rice showed an increased ability for ROS scavenging and decreased hydrogen peroxide (H(2)O(2)) concentration. Furthermore, the localization and relative quantification of silicon deposition in rice leaves was detected by scanning electron microscopy (SEM) and energy-dispersive X-ray spectrometer (EDS). Finally, the transcript levels of defense response genes increased in transformed rice. These results show a correlation between Xoo resistance and phytoalexin production, H(2)O(2), silicon deposition and defense gene expression in hrf1-transformed rice. These data are significant because they provide evidence for a better understanding the role of defense responses in the incompatible interaction between bacterial disease and hrf1-transformed plants. These data also supply an opportunity for generating nonspecific resistance to pathogens.

  10. Ectopic expression of Tsi1 in transgenic hot pepper plants enhances host resistance to viral, bacterial, and oomycete pathogens.

    PubMed

    Shin, Ryoung; Park, Jeong Mee; An, Jong-Min; Paek, Kyung-Hee

    2002-10-01

    In many plants, including hot pepper plants, productivity is greatly affected by pathogen attack. We reported previously that tobacco stress-induced gene 1 (Tsi1) may play an important role in regulating stress responsive genes and pathogenesis-related (PR) genes. In this study, we demonstrated that overexpression of Tsi1 gene in transgenic hot pepper plants induced constitutive expression of several PR genes in the absence of stress or pathogen treatment. The transgenic hot pepper plants expressing Tsi1 exhibited resistance to Pepper mild mottle virus (PMMV) and Cucumber mosaic virus (CMV). Furthermore, these transgenic plants showed increased resistance to a bacterial pathogen, Xanthomonas campestris pv. vesicatoria and also an oomycete pathogen, Phytophthora capsici. These results suggested that ectopic expression of Tsi1 in transgenic hot pepper plants enhanced the resistance of the plants to various pathogens, including viruses, bacteria, and oomycete. These results suggest that using transcriptional regulatory protein genes may contribute to developing broad-spectrum resistance in crop plants.

  11. Association between increased expression of endothelial isoform of nitric oxide synthase in the human fallopian tube and tubal ectopic pregnancy

    PubMed Central

    Fath Bayati, Leyla; Ghaffari Novin, Marefat; Fadaei Fathabadi, Fatemeh; Piryaei, Abbas; Heidari, Mohammad Hasan; Bandehpour, Mozhgan; Norouzian, Mohsen; Alizadeh Parhizgar, Mahdi; Shakooriyan Fard, Mahmood

    2014-01-01

    Background: Tubal ectopic pregnancy (tEP) is the most common type of extra-uterine pregnancy and the most common cause of maternal mortality. Nitric oxide (NO) is a molecule that incorporates in many physiological processes of female reproductive system. Recent studies have demonstrated the possible role of endothelial isoform of nitric oxide synthase (eNOS) enzyme in the regulation of many reproductive events that occur in the fallopian tube (FT). Objective: The aim of this study was to evaluate the expression of eNOS in the FTs of women with tEP. Materials and Methods: In this case-control study, a total number of 30FTs samples were obtained from three groups including: 10 FTs of women that bearing an EP, 10 FTs from the non-pregnant women at luteal phase of the menstrual cycle, and 10 FTs of healthy pregnant women (n=10). Samples were fixed in 10% buffered formalin and then were evaluated by immunohistochemistry. Results: Localization of eNOS was seen in secretory and ciliated luminal epithelium and vascular endothelium of all groups. However, we did not observed the expression of eNOS in smooth muscle cells of all groups. Expression of eNOS in luminal epithelium of women with EP compared to non-pregnant women at luteal phase of menstrual cycle and healthy pregnant group showed statistically significant increase (p=0.00). Significant difference in expression of eNOS was not observed in luminal epithelium of FTs of women at luteal phase compared to healthy pregnant groups (p=0.78). Conclusion: This study indicates that changes in expression of eNOS in luminal epithelium of FT may lead to development of EP. This article extracted from M.Sc. thesis. (Leyla Fath Bayati) PMID:24799858

  12. The cellular phenotype of Roberts syndrome fibroblasts as revealed by ectopic expression of ESCO2.

    PubMed

    van der Lelij, Petra; Godthelp, Barbara C; van Zon, Wouter; van Gosliga, Djoke; Oostra, Anneke B; Steltenpool, Jûrgen; de Groot, Jan; Scheper, Rik J; Wolthuis, Rob M; Waisfisz, Quinten; Darroudi, Firouz; Joenje, Hans; de Winter, Johan P

    2009-09-07

    Cohesion between sister chromatids is essential for faithful chromosome segregation. In budding yeast, the acetyltransferase Eco1/Ctf7 establishes cohesion during DNA replication in S phase and in response to DNA double strand breaks in G2/M phase. In humans two Eco1 orthologs exist: ESCO1 and ESCO2. Both proteins are required for proper sister chromatid cohesion, but their exact function is unclear at present. Since ESCO2 has been identified as the gene defective in the rare autosomal recessive cohesinopathy Roberts syndrome (RBS), cells from RBS patients can be used to elucidate the role of ESCO2. We investigated for the first time RBS cells in comparison to isogenic controls that stably express V5- or GFP-tagged ESCO2. We show that the sister chromatid cohesion defect in the transfected cell lines is rescued and suggest that ESCO2 is regulated by proteasomal degradation in a cell cycle-dependent manner. In comparison to the corrected cells RBS cells were hypersensitive to the DNA-damaging agents mitomycin C, camptothecin and etoposide, while no particular sensitivity to UV, ionizing radiation, hydroxyurea or aphidicolin was found. The cohesion defect of RBS cells and their hypersensitivity to DNA-damaging agents were not corrected by a patient-derived ESCO2 acetyltransferase mutant (W539G), indicating that the acetyltransferase activity of ESCO2 is essential for its function. In contrast to a previous study on cells from patients with Cornelia de Lange syndrome, another cohesinopathy, RBS cells failed to exhibit excessive chromosome aberrations after irradiation in G2 phase of the cell cycle. Our results point at an S phase-specific role for ESCO2 in the maintenance of genome stability.

  13. Ectopic expression of RASSF2 and its prognostic role for gastric adenocarcinoma patients.

    PubMed

    Luo, Deng; Ye, Ting; Li, Tian-Qian; Tang, Peng; Min, Sha-Dong; Zhao, Gong-Fang; Huang, Hua; Chang, Jiang; Wang, Yan; Lv, Lin; Lu, Ming-Liang; Zheng, Meng-Yao

    2012-03-01

    RASSF2 has recently been identified as a potential tumor suppressor that serves as a Ras effector in various types of human cancers. However, there have been few reports detailing this in gastric cancer. Samples of gastric adenocarcinoma from 276 Chinese patients with follow-up were analyzed for RASSF2 protein expression by immunohistochemistry. RASSF2 was expressed in up to 31.2% (86/276) of this group of gastric carcinoma. The expression of RASSF2 was significantly lower in carcinomas than in normal mucosas (P<0.05). RASSF2 corresponded positively with patient age, histological differentiation, depth of tumor invasion, regional lymph node and distant metastasis, and TNM stage (all P<0.05). Further multivariate analysis revealed that patient gender, depth of tumor invasion, distant metastasis, TNM stage and the expression of RASSF2 were independent prognostic factors for patients with gastric cancer. The Kaplan-Meier plot showed that the overall mean survival time of the patients with RASSF2-negative expression was shorter than that of patients with positive expression (χ(2)=156.874, P<0.0001). Moreover, RASSF2-negative expression had a much more significant effect on the survival of those patients with early stage tumors (χ(2)=127.167, P<0.0001), highlighted by a >50.9% reduction in 3-year survival compared to that of patients with RASSF2-positive expression. In late stages, the difference was also significant (χ(2)=6.246, P=0.019), with a 35.5% reduction in 3-year survival. It is suggested that RASSF2 plays an important role in the evolution of gastric adenocarcinoma and should be considered as a potential marker for its prognosis.

  14. Ectopic Expression of a Microbial-Type Rhodopsin Restores Visual Responses in Mice with Photoreceptor Degeneration

    PubMed Central

    Bi, Anding; Cui, Jinjuan; Ma, Yu-Ping; Olshevskaya, Elena; Pu, Mingliang; Dizhoor, Alexander M.; Pan, Zhuo-Hua

    2006-01-01

    Summary The death of photoreceptor cells caused by retinal degenerative diseases often results in a complete loss of retinal responses to light. We explore the feasibility of converting inner retinal neurons to photosensitive cells as a possible strategy for imparting light sensitivity to retinas lacking rods and cones. Using delivery by an adeno-associated viral vector, here, we show that long-term expression of a microbial-type rhodopsin, channelrhodopsin-2 (ChR2), can be achieved in rodent inner retinal neurons in vivo. Furthermore, we demonstrate that expression of ChR2 in surviving inner retinal neurons of a mouse with photoreceptor degeneration can restore the ability of the retina to encode light signals and transmit the light signals to the visual cortex. Thus, expression of microbial-type channelrhodopsins, such as ChR2, in surviving inner retinal neurons is a potential strategy for the restoration of vision after rod and cone degeneration. PMID:16600853

  15. Ectopic expression of TrPI, a Taihangia rupestris (Rosaceae) PI ortholog, causes modifications of vegetative architecture in Arabidopsis.

    PubMed

    Lü, Shanhua; Fan, Yinglun; Liu, Like; Liu, Shujun; Zhang, Wenhui; Meng, Zheng

    2010-12-15

    In eudicotyledonous model plants, the B-function genes encode a pair of partner MADS-domain proteins, APETALA3 (AP3) and PISTILLATA (PI) in Arabidopsis and DEFICIENS (DEF) and GLOBOSA (GLO) in Antirrhinum. These proteins, which must form heterodimers to function, are required to specify petal and stamen identity during flower development. Here, we report cloning and characterization of TrPI (Taihangia rupestris PISTILLATA), a PI/GLO-like gene from the core eudicot species Taihangia rupestris (Rosaceae). DNA gel blot analysis showed that TrPI is a single copy gene in the T. rupestris genome. Quantitative RT-PCR and in situ hybridization analyses revealed that TrPI is transcribed in both the vegetative and reproductive organs at different levels. Ectopic expression of TrPI in Arabidopsis caused severe modifications in vegetative plant architecture, including rosette leaves and cauline leaves arranged in a non-spiral phyllotaxy, and a flattened primary inflorescence stem that produced two or three offshoots at the base, middle or top. Moreover, we show that the TrPI gene is capable of rescuing pi-1 mutant phenotypes. Yeast two-hybrid assays showed that TrPI forms homodimers. Taken together, these results show that TrPI might function in regulating plant architecture in addition to its function as a floral organ identity gene in T. rupestris, suggesting that the TrPI protein has biochemical features that distinguish it from the well-studied orthologs, PI and GLO.

  16. Ectopic Expression of Plant RNA Chaperone Offering Multiple Stress Tolerance in E. coli.

    PubMed

    Jabeen, Bushra; Naqvi, S M Saqlan; Mahmood, Tariq; Sultana, Tasawar; Arif, Madiha; Khan, Fariha

    2017-03-01

    Members of the plant glycine-rich RNA-binding proteins (GR-RBPs) family have been reported in flowering, development, circadian rhythms, biotic and abiotic stresses. Particularly, GR-RBPs are reported to function as RNA chaperones, promoting growth and acclimation during cold shock. It is indispensable to further question the efficacy and mechanism of GR-RBPs under various environmental strains. Monitoring the expression of stress-regulated proteins under stress conditions has been a beneficial strategy to study their functional roles. In an effort to elucidate the NtGR-RBP1 function, stress markers such as salinity, drought, low temperature and heat stresses were studied. The NtGR-RBP1 gene was expressed in E. coli followed by the exposure to stress conditions. Recombinant E. coli expressing NtGR-RBP1 were more tolerant to stresses, e.g., salinity, drought, cold and heat shock. Recombinants exhibited higher growth rates compared to control in spot assays. The tolerance was further confirmed by monitoring the growth in liquid culture assays. Cells expressing NtGR-RBP1 under salt (500 mM NaCl), drought (20% PEG), cold (4 and 20 °C) and heat stresses (50 °C) had enhanced growing ability and better endurance. Our study supports the notion that the protective role of NtGR-RBP1 may contribute to growth and survival during diverse environmental stresses.

  17. Ectopic expression of syndecan-1 in basal epidermis affects keratinocyte proliferation and wound re-epithelialization.

    PubMed

    Ojeh, Nkemcho; Hiilesvuo, Katri; Wärri, Anni; Salmivirta, Markku; Henttinen, Tiina; Määttä, Arto

    2008-01-01

    Epidermal proliferation and differentiation can be regulated by soluble morphogens and growth factors. Heparan sulfate proteoglycans (HSPGs) modulate the action of several of these effector molecules, such as members of the fibroblast growth factor (FGF) and Wnt families. Syndecan-1 is a cell-surface proteoglycan that is expressed in differentiating keratinocytes and transiently upregulated in all layers of the epidermis upon tissue injury. To address the role of syndecan-1 in the regulation of keratinocyte proliferation and differentiation, we generated transgenic mice that overexpress syndecan-1 under K14 keratin promoter in the basal layer of the epidermis. We observed epidermal hyperproliferation in newborn transgenic mice, as evidenced by increased number of suprabasal cell layers, elevated proliferating cell nuclear antigen (PCNA) expression in both basal and suprabasal cell layers and by expression of keratin 6 in the interfollicular epidermis. Compared to both wild-type and syndecan-1-null animals, the transgene expression interfered with skin wound healing in adult mice by decreasing cell proliferation in the re-epithelialized epidermis. Thus, syndecan-1 regulates keratinocyte proliferation differently during skin development and in healing wounds.

  18. Adenomatous Polyposis Coli Tumor Suppressor Protein Has Signaling Activity in Xenopus laevis Embryos Resulting in the Induction of an Ectopic Dorsoanterior Axis

    PubMed Central

    Vleminckx, Kris; Wong, Ellen; Guger, Kathy; Rubinfeld, Bonnee; Polakis, Paul; Gumbiner, Barry M.

    1997-01-01

    Mutations in the adenomatous polyposis coli (APC) tumor suppressor gene are linked to both familial and sporadic human colon cancer. So far, a clear biological function for the APC gene product has not been determined. We assayed the activity of APC in the early Xenopus embryo, which has been established as a good model for the analysis of the signaling activity of the APC-associated protein β-catenin. When expressed in the future ventral side of a four-cell embryo, full-length APC induced a secondary dorsoanterior axis and the induction of the homeobox gene Siamois. This is similar to the phenotype previously observed for ectopic β-catenin expression. In fact, axis induction by APC required the availability of cytosolic β-catenin. These results indicate that APC has signaling activity in the early Xenopus embryo. Signaling activity resides in the central domain of the protein, a part of the molecule that is missing in most of the truncating APC mutations in colon cancer. Signaling by APC in Xenopus embryos is not accompanied by detectable changes in expression levels of β-catenin, indicating that it has direct positive signaling activity in addition to its role in β-catenin turnover. From these results we propose a model in which APC acts as part of the Wnt/β-catenin signaling pathway, either upstream of, or in conjunction with, β-catenin. PMID:9015311

  19. Novel DLK-independent neuronal regeneration in Caenorhabditis elegans shares links with activity-dependent ectopic outgrowth

    PubMed Central

    Awal, Mehraj R.; Shay, James; McLoed, Melissa M.; Mazur, Eric; Gabel, Christopher V.

    2016-01-01

    During development, a neuron transitions from a state of rapid growth to a stable morphology, and neurons within the adult mammalian CNS lose their ability to effectively regenerate in response to injury. Here, we identify a novel form of neuronal regeneration, which is remarkably independent of DLK-1/DLK, KGB-1/JNK, and other MAPK signaling factors known to mediate regeneration in Caenorhabditis elegans, Drosophila, and mammals. This DLK-independent regeneration in C. elegans has direct genetic and molecular links to a well-studied form of endogenous activity-dependent ectopic axon outgrowth in the same neuron type. Both neuron outgrowth types are triggered by physical lesion of the sensory dendrite or mutations disrupting sensory activity, calcium signaling, or genes that restrict outgrowth during neuronal maturation, such as SAX-1/NDR kinase or UNC-43/CaMKII. These connections suggest that ectopic outgrowth represents a powerful platform for gene discovery in neuronal regeneration. Moreover, we note numerous similarities between C. elegans DLK-independent regeneration and lesion conditioning, a phenomenon producing robust regeneration in the mammalian CNS. Both regeneration types are triggered by lesion of a sensory neurite via reduction of neuronal activity and enhanced by disrupting L-type calcium channels or elevating cAMP. Taken as a whole, our study unites disparate forms of neuronal outgrowth to uncover fresh molecular insights into activity-dependent control of the adult nervous system’s intrinsic regenerative capacity. PMID:27078101

  20. Usefulness of ventricular endocardial electric reconstruction from body surface potential maps to noninvasively localize ventricular ectopic activity in patients

    NASA Astrophysics Data System (ADS)

    Lai, Dakun; Sun, Jian; Li, Yigang; He, Bin

    2013-06-01

    As radio frequency (RF) catheter ablation becomes increasingly prevalent in the management of ventricular arrhythmia in patients, an accurate and rapid determination of the arrhythmogenic site is of important clinical interest. The aim of this study was to test the hypothesis that the inversely reconstructed ventricular endocardial current density distribution from body surface potential maps (BSPMs) can localize the regions critical for maintenance of a ventricular ectopic activity. Patients with isolated and monomorphic premature ventricular contractions (PVCs) were investigated by noninvasive BSPMs and subsequent invasive catheter mapping and ablation. Equivalent current density (CD) reconstruction (CDR) during symptomatic PVCs was obtained on the endocardial ventricular surface in six patients (four men, two women, years 23-77), and the origin of the spontaneous ectopic activity was localized at the location of the maximum CD value. Compared with the last (successful) ablation site (LAS), the mean and standard deviation of localization error of the CDR approach were 13.8 and 1.3 mm, respectively. In comparison, the distance between the LASs and the estimated locations of an equivalent single moving dipole in the heart was 25.5 ± 5.5 mm. The obtained CD distribution of activated sources extending from the catheter ablation site also showed a high consistency with the invasively recorded electroanatomical maps. The noninvasively reconstructed endocardial CD distribution is suitable to predict a region of interest containing or close to arrhythmia source, which may have the potential to guide RF catheter ablation.

  1. Ectopic expression of a WRKY homolog from Glycine soja alters flowering time in Arabidopsis.

    PubMed

    Luo, Xiao; Sun, Xiaoli; Liu, Baohui; Zhu, Dan; Bai, Xi; Cai, Hua; Ji, Wei; Cao, Lei; Wu, Jing; Wang, Mingchao; Ding, Xiaodong; Zhu, Yanming

    2013-01-01

    Flowering is a critical event in the life cycle of plants; the WRKY-type transcription factors are reported to be involved in many developmental processes sunch as trichome development and epicuticular wax loading, but whether they are involved in flowering time regulation is still unknown. Within this study, we provide clear evidence that GsWRKY20, a member of WRKY gene family from wild soybean, is involved in controlling plant flowering time. Expression of GsWRKY20 was abundant in the shoot tips and inflorescence meristems of wild soybean. Phenotypic analysis showed that GsWRKY20 over-expression lines flowered earlier than the wild-type plants under all conditions: long-day and short-day photoperiods, vernalization, or exogenous GA3 application, indicating that GsWRKY20 may mainly be involved in an autonomous flowering pathway. Further analyses by qRT-PCR and microarray suggests that GsWRKY20 accelerating plant flowering might primarily be through the regulation of flowering-related genes (i.e., FLC, FT, SOC1 and CO) and floral meristem identity genes (i.e., AP1, SEP3, AP3, PI and AG). Our results provide the evidence demonstrating the effectiveness of manipulating GsWRKY20 for altering plant flowering time.

  2. Protruding structures on caterpillars are controlled by ectopic Wnt1 expression.

    PubMed

    Edayoshi, Mina; Yamaguchi, Junichi; Fujiwara, Haruhiko

    2015-01-01

    Spine-like or protruding structures, which may be aposematic for predators, are often observed in multiple segments of lepidopteran larvae (caterpillars). For example, the larvae of the Chinese wheel butterfly, Byasa alcinous, display many protrusions on their backs as a warning that they are toxic. Although these protrusions are formed by an integument lined with single-layered epidermal cells, the molecular mechanisms underlying their formation have remained unclear. In this study, we focused on a spontaneous mutant of the silkworm, Bombyx mori, Knobbed, which shows similar protrusions to B. alcinous and demonstrates that Wnt1 plays a crucial role in the formation of protrusion structures. Using both transgene expression and RNAi-based knockdown approaches, we showed that Wnt1 designates the position where epidermal cells excessively proliferate, leading to the generation of knobbed structures. Furthermore, in the B. alcinous larvae, Wnt1 was also specifically expressed in association with the protrusions. Our results suggest that Wnt1 plays a role in the formation of protrusions on the larval body, and is conserved broadly among diverse species in Lepidoptera.

  3. Clomiphene citrate causes aberrant tubal apoptosis and estrogen receptor activation in rat fallopian tube: implications for tubal ectopic pregnancy.

    PubMed

    Shao, Ruijin; Nutu, Magdalena; Weijdegård, Birgitta; Egecioglu, Emil; Fernandez-Rodriguez, Julia; Karlsson-Lindahl, Linda; Gemzell-Danielsson, Kristina; Bergh, Christina; Billig, Håkan

    2009-06-01

    Clomiphene citrate (CC) therapy for disorders of anovulatory infertility has been linked to an increased frequency of tubal ectopic pregnancy. Although CC enhances apoptotic processes in the ovaries, villi, and decidual tissues, its effect on apoptosis in the fallopian tube is unknown. Here, we show that chronic treatment with CC induces tubal apoptosis, but not necrosis, through an intrinsic mitochondria-dependent signaling pathway in vivo. The apoptosis was specific to epithelial cells in the isthmus, and the damage was reversed with 17beta-estradiol (E2); however, pretreatment or concomitant treatment with E2 did not protect against tubal apoptosis induced by chronic treatment with CC. Chronic treatment activated estrogen receptors (ESRs), particularly cilia-localized ESR2A (formerly ERbeta2). In contrast to E2, acute treatment of superovulating rats with a high dose of CC or the ESR2-selective agonist 2,3-bis (4-hydroxyphenyl)-propionitrile (DPN) significantly delayed the transport of oocyte-cumulus complexes through the fallopian tube. Our findings suggest that in response to chronic CC therapy, isthmus-specific apoptosis of epithelial cells and activation of cilia-ESR2A act in parallel to block gamete and embryo passage through the fallopian tube, eventually resulting in tubal ectopic pregnancy.

  4. Ectopic expression of a tobacco vacuolar invertase inhibitor in guard cells confers drought tolerance in Arabidopsis.

    PubMed

    Chen, Su-Fen; Liang, Ke; Yin, Dong-Mei; Ni, Di-An; Zhang, Zhi-Guo; Ruan, Yong-Ling

    2016-12-01

    There are several hypotheses that explain stomatal behavior. These include the concept of osmoregulation mediated by potassium and its counterions malate and chlorine and the more recent starch-sugar hypothesis. We have previously reported that the activity of the sucrose cleavage enzyme, vacuolar invertase (VIN), is significantly higher in guard cells than in other leaf epidermal cells and its activity is correlated with stomatal aperture. Here, we examined whether VIN indeed controls stomatal movement under normal and drought conditions by transforming Arabidopsis with a tobacco vacuolar invertase inhibitor homolog (Nt-inhh) under the control of an abscisic acid-sensitive and guard cell-specific promoter (AtRab18). The data obtained showed that guard cells of transgenic Arabidopsis plants had lower VIN activity, stomatal aperture and conductance than that of wild-type plants. Moreover, the transgenic plants also displayed higher drought tolerance than wild-type plants. The data indicate that VIN is a promising target for manipulating stomatal function to increase drought tolerance.

  5. Ectopic Expression of JcWRKY Transcription Factor Confers Salinity Tolerance via Salicylic Acid Signaling

    PubMed Central

    Agarwal, Parinita; Dabi, Mitali; Sapara, Komal K.; Joshi, Priyanka S.; Agarwal, Pradeep K.

    2016-01-01

    Plants, being sessile, have developed intricate signaling network to specifically respond to the diverse environmental stress. The plant-specific WRKY TFs form one of the largest TF family and are involved in diverse plant processes, involving growth, development and stress signaling through auto and cross regulation with different genes and TFs. Here, we report the functional characterization of a salicylic acid -inducible JcWRKY TF. The JcWRKY overexpression confers salinity tolerance in transgenic tobacco, as was evident by increased chlorophyll content and seed germination potential. The transgenic plants showed increased soluble sugar, membrane stability, reduced electrolyte leakage and generation of reactive oxygen species (H2O2 and O2•-) as compared to the wild type. Furthermore, the low SA treatment along with salinity improved the tolerance potential of the transgenics by maintaining ROS homeostasis and high K+/Na+ ratio. The transcript expression of SA biosynthetic gene ICS1 and antioxidative enzymes (CAT and SOD) showed upregulation during stress. Thus, the present study reflects that JcWRKY is working in co-ordination with SA signaling to orchestrate the different biochemical and molecular pathways to maneuvre salt stress tolerance of the transgenic plants. PMID:27799936

  6. Ectopic expression of inhibitors of protein phosphatase type 1 (PP1) can be used to analyze roles of PP1 in Drosophila development.

    PubMed Central

    Bennett, Daimark; Szöor, Balázs; Gross, Sascha; Vereshchagina, Natalia; Alphey, Luke

    2003-01-01

    We have identified two proteins that bind with high specificity to type 1 serine/threonine protein phosphatase (PP1) and have exploited their inhibitory properties to develop an efficient and flexible strategy for conditional inactivation of PP1 in vivo. We show that modest overexpression of Drosophila homologs of I-2 and NIPP1 (I-2Dm and NIPP1Dm) reduces the level of PP1 activity and phenotypically resembles known PP1 mutants. These phenotypes, which include lethality, abnormal mitotic figures, and defects in muscle development, are suppressed by coexpression of PP1, indicating that the effect is due specifically to loss of PP1 activity. Reactivation of I-2Dm:PP1c complexes suggests that inhibition of PP1 activity in vivo does not result in a compensating increase in synthesis of active PP1. PP1 mutants enhance the wing overgrowth phenotype caused by ectopic expression of the type II TGF beta superfamily signaling receptor Punt. Using I-2Dm, which has a less severe effect than NIPP1Dm, we show that lowering the level of PP1 activity specifically in cells overexpressing Punt is sufficient for wing overgrowth and that the interaction between PP1 and Punt requires the type I receptor Thick-veins (Tkv) but is not strongly sensitive to the level of the ligand, Decapentaplegic (Dpp), nor to that of the other type I receptors. This is consistent with a role for PP1 in antagonizing Punt by preventing phosphorylation of Tkv. These studies demonstrate that inhibitors of PP1 can be used in a tissue- and developmental-specific manner to examine the developmental roles of PP1. PMID:12750335

  7. Tomato plants ectopically expressing Arabidopsis CBF1 show enhanced resistance to water deficit stress.

    PubMed

    Hsieh, Tsai-Hung; Lee, Jent-turn; Charng, Yee-yung; Chan, Ming-Tsair

    2002-10-01

    A DNA cassette containing an Arabidopsis C repeat/dehydration-responsive element binding factor 1 (CBF1) cDNA and a nos terminator, driven by a cauliflower mosaic virus 35S promoter, was transformed into the tomato (Lycopersicon esculentum) genome. These transgenic tomato plants were more resistant to water deficit stress than the wild-type plants. The transgenic plants exhibited growth retardation by showing dwarf phenotype, and the fruit and seed numbers and fresh weight of the transgenic tomato plants were apparently less than those of the wild-type plants. Exogenous gibberellic acid treatment reversed the growth retardation and enhanced growth of transgenic tomato plants, but did not affect the level of water deficit resistance. The stomata of the transgenic CBF1 tomato plants closed more rapidly than the wild type after water deficit treatment with or without gibberellic acid pretreatment. The transgenic tomato plants contained higher levels of Pro than those of the wild-type plants under normal or water deficit conditions. Subtractive hybridization was used to isolate the responsive genes to heterologous CBF1 in transgenic tomato plants and the CAT1 (CATALASE1) was characterized. Catalase activity increased, and hydrogen peroxide concentration decreased in transgenic tomato plants compared with the wild-type plants with or without water deficit stress. These results indicated that the heterologous Arabidopsis CBF1 can confer water deficit resistance in transgenic tomato plants.

  8. A retrotransposon insertion in the 5' regulatory domain of Ptf1a results in ectopic gene expression and multiple congenital defects in Danforth's short tail mouse.

    PubMed

    Lugani, Francesca; Arora, Ripla; Papeta, Natalia; Patel, Ami; Zheng, Zongyu; Sterken, Roel; Singer, Ruth A; Caridi, Gianluca; Mendelsohn, Cathy; Sussel, Lori; Papaioannou, Virginia E; Gharavi, Ali G

    2013-01-01

    Danforth's short tail mutant (Sd) mouse, first described in 1930, is a classic spontaneous mutant exhibiting defects of the axial skeleton, hindgut, and urogenital system. We used meiotic mapping in 1,497 segregants to localize the mutation to a 42.8-kb intergenic segment on chromosome 2. Resequencing of this region identified an 8.5-kb early retrotransposon (ETn) insertion within the highly conserved regulatory sequences upstream of Pancreas Specific Transcription Factor, 1a (Ptf1a). This mutation resulted in up to tenfold increased expression of Ptf1a as compared to wild-type embryos at E9.5 but no detectable changes in the expression levels of other neighboring genes. At E9.5, Sd mutants exhibit ectopic Ptf1a expression in embryonic progenitors of every organ that will manifest a developmental defect: the notochord, the hindgut, and the mesonephric ducts. Moreover, at E 8.5, Sd mutant mice exhibit ectopic Ptf1a expression in the lateral plate mesoderm, tail bud mesenchyme, and in the notochord, preceding the onset of visible defects such as notochord degeneration. The Sd heterozygote phenotype was not ameliorated by Ptf1a haploinsufficiency, further suggesting that the developmental defects result from ectopic expression of Ptf1a. These data identify disruption of the spatio-temporal pattern of Ptf1a expression as the unifying mechanism underlying the multiple congenital defects in Danforth's short tail mouse. This striking example of an enhancer mutation resulting in profound developmental defects suggests that disruption of conserved regulatory elements may also contribute to human malformation syndromes.

  9. Drosophila Crumbs prevents ectopic Notch activation in developing wings by inhibiting ligand-independent endocytosis.

    PubMed

    Nemetschke, Linda; Knust, Elisabeth

    2016-12-01

    Many signalling components are apically restricted in epithelial cells, and receptor localisation and abundance is key for morphogenesis and tissue homeostasis. Hence, controlling apicobasal epithelial polarity is crucial for proper signalling. Notch is a ubiquitously expressed, apically localised receptor, which performs a plethora of functions; therefore, its activity has to be tightly regulated. Here, we show that Drosophila Crumbs, an evolutionarily conserved polarity determinant, prevents Notch endocytosis in developing wings through direct interaction between the two proteins. Notch endocytosis in the absence of Crumbs results in the activation of the ligand-independent, Deltex-dependent Notch signalling pathway, and does not require the ligands Delta and Serrate or γ-secretase activity. This function of Crumbs is not due to general defects in apicobasal polarity, as localisation of other apical proteins is unaffected. Our data reveal a mechanism to explain how Crumbs directly controls localisation and trafficking of the potent Notch receptor, and adds yet another aspect of Crumbs regulation in Notch pathway activity. Furthermore, our data highlight a close link between the apical determinant Crumbs, receptor trafficking and tissue homeostasis.

  10. Ectopic Terpene Synthase Expression Enhances Sesquiterpene Emission in Nicotiana attenuata without Altering Defense or Development of Transgenic Plants or Neighbors1[W

    PubMed Central

    Schuman, Meredith C.; Palmer-Young, Evan C.; Schmidt, Axel; Gershenzon, Jonathan; Baldwin, Ian T.

    2014-01-01

    Sesquiterpenoids, with approximately 5,000 structures, are the most diverse class of plant volatiles with manifold hypothesized functions in defense, stress tolerance, and signaling between and within plants. These hypotheses have often been tested by transforming plants with sesquiterpene synthases expressed behind the constitutively active 35S promoter, which may have physiological costs measured as inhibited growth and reduced reproduction or may require augmentation of substrate pools to achieve enhanced emission, complicating the interpretation of data from affected transgenic lines. Here, we expressed maize (Zea mays) terpene synthase10 (ZmTPS10), which produces (E)-α-bergamotene and (E)-β-farnesene, or a point mutant ZmTPS10M, which produces primarily (E)-β-farnesene, under control of the 35S promoter in the ecological model plant Nicotiana attenuata. Transgenic N. attenuata plants had specifically enhanced emission of target sesquiterpene(s) with no changes detected in their emission of any other volatiles. Treatment with herbivore or jasmonate elicitors induces emission of (E)-α-bergamotene in wild-type plants and also tended to increase emission of (E)-α-bergamotene and (E)-β-farnesene in transgenics. However, transgenics did not differ from the wild type in defense signaling or chemistry and did not alter defense chemistry in neighboring wild-type plants. These data are inconsistent with within-plant and between-plant signaling functions of (E)-β-farnesene and (E)-α-bergamotene in N. attenuata. Ectopic sesquiterpene emission was apparently not costly for transgenics, which were similar to wild-type plants in their growth and reproduction, even when forced to compete for common resources. These transgenics would be well suited for field experiments to investigate indirect ecological effects of sesquiterpenes for a wild plant in its native habitat. PMID:25187528

  11. Hoxc8 initiates an ectopic mammary program by regulating Fgf10 and Tbx3 expression and Wnt/β-catenin signaling

    PubMed Central

    Carroll, Lara S.; Capecchi, Mario R.

    2015-01-01

    The role of Hox genes in the formation of cutaneous accessory organs such as hair follicles and mammary glands has proved elusive, a likely consequence of overlapping function and expression among various homeobox factors. Lineage and immunohistochemical analysis of Hoxc8 in mice revealed that this midthoracic Hox gene has transient but strong regional expression in ventrolateral surface ectoderm at E10.5, much earlier than previously reported. Targeted mice were generated to conditionally misexpress Hoxc8 from the Rosa locus using select Cre drivers, which significantly expanded the domain of thoracic identity in mutant embryos. Accompanying this expansion was the induction of paired zones of ectopic mammary development in the cervical region, which generated between three and five pairs of mammary placodes anterior to the first wild-type mammary rudiment. These rudiments expressed the mammary placode markers Wnt10b and Tbx3 and were labeled by antibodies to the mammary mesenchyme markers ERα and androgen receptor. Somitic Fgf10 expression, which is required for normal mammary line formation, was upregulated in mutant cervical somites, and conditional ablation of ectodermal Tbx3 expression eliminated all normally positioned and ectopic mammary placodes. We present evidence that Hoxc8 participates in regulating the initiation stages of mammary placode morphogenesis, and suggest that this and other Hox genes are likely to have important roles during regional specification and initiation of these and other cutaneous accessory organs. PMID:26459221

  12. Ectopic Expression in Arabidopsis thaliana of an NB-ARC Encoding Putative Disease Resistance Gene from Wild Chinese Vitis pseudoreticulata Enhances Resistance to Phytopathogenic Fungi and Bacteria

    PubMed Central

    Wen, Zhifeng; Yao, Liping; Wan, Ran; Li, Zhi; Liu, Chonghuai; Wang, Xiping

    2015-01-01

    Plant resistance proteins mediate pathogen recognition and activate innate immune responses to restrict pathogen proliferation. One common feature of these proteins is an NB-ARC domain. In this study, we characterized a gene encoding a protein with an NB-ARC domain from wild Chinese grapevine Vitis pseudoreticulata accession “Baihe-35-1,” which was identified in a transcriptome analysis of the leaves following inoculation with Erysiphe necator (Schw.), a causal agent of powdery mildew. Transcript levels of this gene, designated VpCN (GenBank accession number KT265084), increased strongly after challenge of grapevine leaves with E. necator. The deduced amino acid sequence was predicted to contain an NB-ARC domain in the C-terminus and an RxCC-like domain similar to CC domain of Rx protein in the N-terminus. Ectopic expression of VpCN in Arabidopsis thaliana resulted in either a wild-type phenotype or a dwarf phenotype. The phenotypically normal transgenic A. thaliana showed enhance resistance to A. thaliana powdery mildew Golovinomyces cichoracearum, as well as to a virulent bacterial pathogen Pseudomonas syringae pv. tomato DC3000. Moreover, promoter::GUS (β-glucuronidase) analysis revealed that powdery mildew infection induced the promoter activity of VpCN in grapevine leaves. Finally, a promoter deletion analysis showed that TC rich repeat elements likely play an important role in the response to E. necator infection. Taken together, our results suggest that VpCN contribute to powdery mildew disease resistant in grapevine. PMID:26697041

  13. Ectopic expression of class 1 KNOX genes induce adventitious shoot regeneration and alter growth and development of tobacco (Nicotiana tabacum L) and European plum (Prunus domestica L).

    PubMed

    Srinivasan, C; Liu, Zongrang; Scorza, Ralph

    2011-04-01

    Transgenic plants of tobacco (Nicotiana tabacum L) and European plum (Prunus domestica L) were produced by transforming with the apple class 1 KNOX genes (MdKN1 and MdKN2) or corn KNOX1 gene. Transgenic tobacco plants were regenerated in vitro from transformed leaf discs cultured in a medium lacking cytokinin. Ectopic expression of KNOX genes retarded shoot growth by suppressing elongation of internodes in transgenic tobacco plants. Expression of each of the three KNOX1 genes induced malformation and extensive lobbing in tobacco leaves. In situ regeneration of adventitious shoots was observed from leaves and roots of transgenic tobacco plants expressing each of the three KNOX genes. In vitro culture of leaf explants and internode sections excised from in vitro grown MdKN1 expressing tobacco shoots regenerated adventitious shoots on MS (Murashige and Skoog 1962) basal medium in the absence of exogenous cytokinin. Transgenic plum plants that expressed the MdKN2 or corn KNOX1 gene grew normally but MdKN1 caused a significant reduction in plant height, leaf shape and size and produced malformed curly leaves. A high frequency of adventitious shoot regeneration (96%) was observed in cultures of leaf explants excised from corn KNOX1-expressing transgenic plum shoots. In contrast to KNOX1-expressing tobacco, leaf and internode explants of corn KNOX1-expressing plum required synthetic cytokinin (thidiazuron) in the culture medium to induce adventitious shoot regeneration. The induction of high-frequency regeneration of adventitious shoots in vitro from leaves and stem internodal sections of plum through the ectopic expression of a KNOX1 gene is the first such report for a woody perennial fruit trees.

  14. Inflammation in mice ectopically expressing human Pyogenic Arthritis, Pyoderma Gangrenosum, and Acne (PAPA) Syndrome-associated PSTPIP1 A230T mutant proteins.

    PubMed

    Wang, Donghai; Höing, Susanne; Patterson, Heide Christine; Ahmad, Umtul M; Rathinam, Vijay A K; Rajewsky, Klaus; Fitzgerald, Katherine A; Golenbock, Douglas T

    2013-02-15

    Pyogenic Arthritis, Pyoderma Gangrenosum, and Acne Syndrome (PAPA syndrome) is an autoinflammatory disease caused by aberrant production of the proinflammatory cytokine interleukin-1. Mutations in the gene encoding proline serine threonine phosphatase-interacting protein-1 (PSTPIP1) have been linked to PAPA syndrome. PSTPIP1 is an adaptor protein that interacts with PYRIN, the protein encoded by the Mediterranean Fever (MEFV) gene whose mutations cause Familial Mediterranean Fever (FMF). However, the pathophysiological function of PSTPIP1 remains to be elucidated. We have generated mouse strains that either are PSTPIP1 deficient or ectopically express mutant PSTPIP1. Results from analyzing these mice suggested that PSTPIP1 is not an essential regulator of the Nlrp3, Aim2, or Nlrc4 inflammasomes. Although common features of human PAPA syndrome such as pyogenic arthritis and skin inflammation were not recapitulated in the mouse model, ectopic expression of the mutant but not the wild type PSTPIP1 in mice lead to partial embryonic lethality, growth retardation, and elevated level of circulating proinflammatory cytokines.

  15. Ectopic pregnancy (image)

    MedlinePlus

    An ectopic pregnancy is one in which the fertilized egg implants in tissue outside of the uterus and the placenta ... common site is within a Fallopian tube, however, ectopic pregnancies can occur in the ovary, the abdomen, and ...

  16. Ectopic expression of the HAM59 gene causes homeotic transformations of reproductive organs in sunflower (Helianthus annuus L.).

    PubMed

    Shulga, O A; Neskorodov, Ya B; Shchennikova, A V; Gaponenko, A K; Skryabin, K G

    2015-01-01

    The function of the HAM59 MADS-box gene in sunflower (Helianthus annuus L.) was studied to clarify homeotic C activity in the Asteraceae plant family. For the first time, transgenic sunflower plants with a modified pattern of HAM59 expression were obtained. It was shown that the HAM59 MADS-box transcription factor did mediate C activity in sunflower. In particular, it participated in termination of the floral meristem, repression of the cadastral function of A-activity, and together with other C-type sunflower protein HAM45-in the specification of the identity of stamens and pistils.

  17. The ectopic expression of a pectin methyl esterase inhibitor increases pectin methyl esterification and limits fungal diseases in wheat.

    PubMed

    Volpi, Chiara; Janni, Michela; Lionetti, Vincenzo; Bellincampi, Daniela; Favaron, Francesco; D'Ovidio, Renato

    2011-09-01

    Cell wall pectin methyl esterification can influence plant resistance because highly methyl-esterified pectin can be less susceptible to the hydrolysis by pectic enzymes such as fungal endopolygalacturonases (PG). Pectin is secreted into the cell wall in a highly methyl-esterified form and, here, is de-methyl esterified by pectin methyl esterase (PME). The activity of PME is controlled by specific protein inhibitors called PMEI; consequently, an increased inhibition of PME by PMEI might modify the pectin methyl esterification. In order to test the possibility of improving wheat resistance by modifying the methyl esterification of pectin cell wall, we have produced durum wheat transgenic lines expressing the PMEI from Actinidia chinensis (AcPMEI). The expression of AcPMEI endows wheat with a reduced endogenous PME activity, and transgenic lines expressing a high level of the inhibitor showed a significant increase in the degree of methyl esterification. These lines showed a significant reduction of disease symptoms caused by the fungal pathogens Bipolaris sorokiniana or Fusarium graminearum. This increased resistance was related to the impaired ability of these fungal pathogens to grow on methyl-esterified pectin and to a reduced activity of the fungal PG to hydrolyze methyl-esterified pectin. In addition to their importance for wheat improvement, these results highlight the primary role of pectin despite its low content in the wheat cell wall.

  18. A high affinity RIM-binding protein/Aplip1 interaction prevents the formation of ectopic axonal active zones

    PubMed Central

    Siebert, Matthias; Böhme, Mathias A; Driller, Jan H; Babikir, Husam; Mampell, Malou M; Rey, Ulises; Ramesh, Niraja; Matkovic, Tanja; Holton, Nicole; Reddy-Alla, Suneel; Göttfert, Fabian; Kamin, Dirk; Quentin, Christine; Klinedinst, Susan; Andlauer, Till FM; Hell, Stefan W; Collins, Catherine A; Wahl, Markus C; Loll, Bernhard; Sigrist, Stephan J

    2015-01-01

    Synaptic vesicles (SVs) fuse at active zones (AZs) covered by a protein scaffold, at Drosophila synapses comprised of ELKS family member Bruchpilot (BRP) and RIM-binding protein (RBP). We here demonstrate axonal co-transport of BRP and RBP using intravital live imaging, with both proteins co-accumulating in axonal aggregates of several transport mutants. RBP, via its C-terminal Src-homology 3 (SH3) domains, binds Aplip1/JIP1, a transport adaptor involved in kinesin-dependent SV transport. We show in atomic detail that RBP C-terminal SH3 domains bind a proline-rich (PxxP) motif of Aplip1/JIP1 with submicromolar affinity. Pointmutating this PxxP motif provoked formation of ectopic AZ-like structures at axonal membranes. Direct interactions between AZ proteins and transport adaptors seem to provide complex avidity and shield synaptic interaction surfaces of pre-assembled scaffold protein transport complexes, thus, favouring physiological synaptic AZ assembly over premature assembly at axonal membranes. DOI: http://dx.doi.org/10.7554/eLife.06935.001 PMID:26274777

  19. Cancer-related ectopic expression of the bone-related transcription factor RUNX2 in non-osseous metastatic tumor cells is linked to cell proliferation and motility

    PubMed Central

    2010-01-01

    Introduction Metastatic breast cancer cells frequently and ectopically express the transcription factor RUNX2, which normally attenuates proliferation and promotes maturation of osteoblasts. RUNX2 expression is inversely regulated with respect to cell growth in osteoblasts and deregulated in osteosarcoma cells. Methods Here, we addressed whether the functional relationship between cell growth and RUNX2 gene expression is maintained in breast cancer cells. We also investigated whether the aberrant expression of RUNX2 is linked to phenotypic parameters that could provide a selective advantage to cells during breast cancer progression. Results We find that, similar to its regulation in osteoblasts, RUNX2 expression in MDA-MB-231 breast adenocarcinoma cells is enhanced upon growth factor deprivation, as well as upon deactivation of the mitogen-dependent MEK-Erk pathway or EGFR signaling. Reduction of RUNX2 levels by RNAi has only marginal effects on cell growth and expression of proliferation markers in MDA-MB-231 breast cancer cells. Thus, RUNX2 is not a critical regulator of cell proliferation in this cell type. However, siRNA depletion of RUNX2 in MDA-MB-231 cells reduces cell motility, while forced exogenous expression of RUNX2 in MCF7 cells increases cell motility. Conclusions Our results support the emerging concept that the osteogenic transcription factor RUNX2 functions as a metastasis-related oncoprotein in non-osseous cancer cells. PMID:21029421

  20. Ectopic expression of vaccinia virus E3 and K3 cannot rescue ectromelia virus replication in rabbit RK13 cells.

    PubMed

    Hand, Erin S; Haller, Sherry L; Peng, Chen; Rothenburg, Stefan; Hersperger, Adam R

    2015-01-01

    As a group, poxviruses have been shown to infect a wide variety of animal species. However, there is individual variability in the range of species able to be productively infected. In this study, we observed that ectromelia virus (ECTV) does not replicate efficiently in cultured rabbit RK13 cells. Conversely, vaccinia virus (VACV) replicates well in these cells. Upon infection of RK13 cells, the replication cycle of ECTV is abortive in nature, resulting in a greatly reduced ability to spread among cells in culture. We observed ample levels of early gene expression but reduced detection of virus factories and severely blunted production of enveloped virus at the cell surface. This work focused on two important host range genes, named E3L and K3L, in VACV. Both VACV and ECTV express a functional protein product from the E3L gene, but only VACV contains an intact K3L gene. To better understand the discrepancy in replication capacity of these viruses, we examined the ability of ECTV to replicate in wild-type RK13 cells compared to cells that constitutively express E3 and K3 from VACV. The role these proteins play in the ability of VACV to replicate in RK13 cells was also analyzed to determine their individual contribution to viral replication and PKR activation. Since E3L and K3L are two relevant host range genes, we hypothesized that expression of one or both of them may have a positive impact on the ability of ECTV to replicate in RK13 cells. Using various methods to assess virus growth, we did not detect any significant differences with respect to the replication of ECTV between wild-type RK13 compared to versions of this cell line that stably expressed VACV E3 alone or in combination with K3. Therefore, there remain unanswered questions related to the factors that limit the host range of ECTV.

  1. Ectopic expression in the giant fiber system of Drosophila reveals distinct roles for roundabout (Robo), Robo2, and Robo3 in dendritic guidance and synaptic connectivity.

    PubMed

    Godenschwege, Tanja A; Simpson, Julie H; Shan, Xiaoliang; Bashaw, Greg J; Goodman, Corey S; Murphey, Rodney K

    2002-04-15

    The Roundabout (Robo) receptors have been intensively studied for their role in regulating axon guidance in the embryonic nervous system, whereas a role in dendritic guidance has not been explored. In the adult giant fiber system of Drosophila, we have revealed that ectopic Robo expression can regulate the growth and guidance of specific motor neuron dendrites, whereas Robo2 and Robo3 have no effect. We also show that the effect of Robo on dendritic guidance can be suppressed by Commissureless coexpression. Although we confirmed a role for all three Robo receptors in giant fiber axon guidance, the strong axon guidance alterations caused by overexpression of Robo2 or Robo3 have no effect on synaptic connectivity. In contrast, Robo overexpression in the giant fiber seems to directly interfere with synaptic function. We conclude that axon guidance, dendritic guidance, and synaptogenesis are separable processes and that the different Robo family members affect them distinctly.

  2. Ectopic expression of RNF168 and 53BP1 increases mutagenic but not physiological non-homologous end joining

    PubMed Central

    Zong, Dali; Callén, Elsa; Pegoraro, Gianluca; Lukas, Claudia; Lukas, Jiri; Nussenzweig, André

    2015-01-01

    DNA double strand breaks (DSBs) formed during S phase are preferentially repaired by homologous recombination (HR), whereas G1 DSBs, such as those occurring during immunoglobulin class switch recombination (CSR), are repaired by non-homologous end joining (NHEJ). The DNA damage response proteins 53BP1 and BRCA1 regulate the balance between NHEJ and HR. 53BP1 promotes CSR in part by mediating synapsis of distal DNA ends, and in addition, inhibits 5’ end resection. BRCA1 antagonizes 53BP1 dependent DNA end-blocking activity during S phase, which would otherwise promote mutagenic NHEJ and genome instability. Recently, it was shown that supra-physiological levels of the E3 ubiquitin ligase RNF168 results in the hyper-accumulation of 53BP1/BRCA1 which accelerates DSB repair. Here, we ask whether increased expression of RNF168 or 53BP1 impacts physiological versus mutagenic NHEJ. We find that the anti-resection activities of 53BP1 are rate-limiting for mutagenic NHEJ but not for physiological CSR. As heterogeneity in the expression of RNF168 and 53BP1 is found in human tumors, our results suggest that deregulation of the RNF168/53BP1 pathway could alter the chemosensitivity of BRCA1 deficient tumors. PMID:25916843

  3. The tripartite leader sequence is required for ectopic expression of HAdV-B and HAdV-E E3 CR1 genes.

    PubMed

    Bair, Camden R; Kotha Lakshmi Narayan, Poornima; Kajon, Adriana E

    2017-05-01

    The unique repertoire of genes that characterizes the early region 3 (E3) of the different species of human adenovirus (HAdV) likely contributes to their distinct pathogenic traits. The function of many E3 CR1 proteins remains unknown possibly due to unidentified intrinsic properties that make them difficult to express ectopically. This study shows that the species HAdV-B- and HAdV-E-specific E3 CR1 genes can be expressed from vectors carrying the HAdV tripartite leader (TPL) sequence but not from traditional mammalian expression vectors. Insertion of the TPL sequence upstream of the HAdV-B and HAdV-E E3 CR1 open reading frames was sufficient to rescue protein expression from pCI-neo constructs in transfected 293T cells. The detection of higher levels of HAdV-B and HAdV-E E3 CR1 transcripts suggests that the TPL sequence may enhance gene expression at both the transcriptional and translational levels. Our findings will facilitate the characterization of additional AdV E3 proteins.

  4. Resistance through inhibition: ectopic expression of serine protease inhibitor offers stress tolerance via delayed senescence in yeast cell.

    PubMed

    Joshi, Rakesh S; Tanpure, Rahul S; Singh, Rajan Kumar; Gupta, Vidya S; Giri, Ashok P

    2014-09-26

    Protease inhibitors have been known to confer multiple stress tolerance in transgenic plants. We have assessed growth of yeast (Pichia pastoris GS115) strains expressing inhibitory repeat domains (PpIRD(+)) of previously characterized Capsicum annuum protease inhibitors under high salt, heavy metal and oxidative stress. PpIRD(+) strains exhibited multiple stress tolerance and showed differential molecular responses at transcriptional and translational level on exposure to stress inducing agents like heavy metal, high salt and H2O2. PpIRD(+) strains display significant reduction in metacaspase (Yca1) activity, the key enzyme in apoptosis, indicates the possibility of cross reactivity of IRDs (serine protease inhibitor) with cysteine proteases. PpIRD(+) and Saccharomyces cerevisiae knockout with Yca1 (ΔYca1) strain showed similar growth characteristics under stress, which indicated the delayed senescence due to cellular metacaspase inhibition. Molecular docking study showed a close proximity of IRDs reactive site and the active site of metacaspase in the complex that signified their strong interactions. Maintenance of GAPDH activity, primary target of metacaspase, in PpIRD(+) strain evidenced the inhibition of metacaspase activity and survival of these cells under stress. This report demonstrates a potential molecular mechanism of protease inhibitor-based multiple stress tolerance in yeast strains.

  5. The double-corolla phenotype in the Hawaiian lobelioid genus Clermontia involves ectopic expression of PISTILLATA B-function MADS box gene homologs

    PubMed Central

    2012-01-01

    Abstract Background The Hawaiian endemic genus Clermontia (Campanulaceae) includes 22 species, 15 of which, the double-corolla species, are characterized by an extra whorl of organs that appear to be true petals occupying what is normally the sepal whorl. Previous research has shown that the presence of homeotic petaloid organs in some other plant groups correlates with ectopic expression of B-function MADS box genes, but similar core eudicot examples of apparent groundplan divergence remain unstudied. B-function genes, which are not normally expressed in the sepal whorl, are required for determination and maintenance of petal identity. Here, we investigate the potential role of altered B-function gene expression contributing to the morphological diversity of this island genus. Results We examined the morphology and developmental genetics of two different species of Clermontia, one of which, C. arborescens, has normal sepals while the other, C. parviflora, has two whorls of petal-like organs. Scanning electron microscopy of cell surface morphologies of first and second whorl organs in the double-corolla species C. parviflora revealed conical epidermal cells on the adaxial surfaces of both first and second whorl petaloid organs, strongly suggesting a homeotic conversion in the former. Phylogenetic analysis of Clermontia species based on 5S ribosomal DNA non-transcribed spacer sequences indicated a probable single and geologically recent origin of the double-corolla trait within the genus, with numerous potential reversals to the standard sepal-petal format. Quantitative polymerase chain reaction analysis of homologs of the B-function genes PISTILLATA (PI), APETALA3 and TOMATO MADS 6 indicated ectopic expression of two PI paralogs in the first whorl of C. parviflora; no such homeotic expression was observed for the other two genes, nor for several other MADS box genes involved in various floral and non-floral functions. In the standard sepal-petal species C

  6. Identification of Genes in the Phenylalanine Metabolic Pathway by Ectopic Expression of a MYB Transcription Factor in Tomato Fruit[W

    PubMed Central

    Dal Cin, Valeriano; Tieman, Denise M.; Tohge, Takayuki; McQuinn, Ryan; de Vos, Ric C.H.; Osorio, Sonia; Schmelz, Eric A.; Taylor, Mark G.; Smits-Kroon, Miriam T.; Schuurink, Robert C.; Haring, Michel A.; Giovannoni, James; Fernie, Alisdair R.; Klee, Harry J.

    2011-01-01

    Altering expression of transcription factors can be an effective means to coordinately modulate entire metabolic pathways in plants. It can also provide useful information concerning the identities of genes that constitute metabolic networks. Here, we used ectopic expression of a MYB transcription factor, Petunia hybrida ODORANT1, to alter Phe and phenylpropanoid metabolism in tomato (Solanum lycopersicum) fruits. Despite the importance of Phe and phenylpropanoids to plant and human health, the pathway for Phe synthesis has not been unambiguously determined. Microarray analysis of ripening fruits from transgenic and control plants permitted identification of a suite of coregulated genes involved in synthesis and further metabolism of Phe. The pattern of coregulated gene expression facilitated discovery of the tomato gene encoding prephenate aminotransferase, which converts prephenate to arogenate. The expression and biochemical data establish an arogenate pathway for Phe synthesis in tomato fruits. Metabolic profiling and 13C flux analysis of ripe fruits further revealed large increases in the levels of a specific subset of phenylpropanoid compounds. However, while increased levels of these human nutrition-related phenylpropanoids may be desirable, there were no increases in levels of Phe-derived flavor volatiles. PMID:21750236

  7. Ectopic expression of anti-HIV-1 shRNAs protects CD8{sup +} T cells modified with CD4ζ CAR from HIV-1 infection and alleviates impairment of cell proliferation

    SciTech Connect

    Kamata, Masakazu; Kim, Patrick Y.; Ng, Hwee L.; Ringpis, Gene-Errol E.; Kranz, Emiko; Chan, Joshua; O'Connor, Sean; Yang, Otto O.; Chen, Irvin S.Y.

    2015-07-31

    Chimeric antigen receptors (CARs) are artificially engineered receptors that confer a desired specificity to immune effector T cells. As an HIV-1-specific CAR, CD4ζ CAR has been extensively tested in vitro as well as in clinical trials. T cells modified with this CAR mediated highly potent anti-HIV-1 activities in vitro and were well-tolerated in vivo, but exerted limited effects on viral load and reservoir size due to poor survival and/or functionality of the transduced cells in patients. We hypothesize that ectopic expression of CD4ζ on CD8{sup +} T cells renders them susceptible to HIV-1 infection, resulting in poor survival of those cells. To test this possibility, highly purified CD8{sup +} T cells were genetically modified with a CD4ζ-encoding lentiviral vector and infected with HIV-1. CD8{sup +} T cells were vulnerable to HIV-1 infection upon expression of CD4ζ as evidenced by elevated levels of p24{sup Gag} in cells and culture supernatants. Concurrently, the number of CD4ζ-modified CD8{sup +} T cells was reduced relative to control cells upon HIV-1 infection. To protect these cells from HIV-1 infection, we co-expressed two anti-HIV-1 shRNAs previously developed by our group together with CD4ζ. This combination vector was able to suppress HIV-1 infection without impairing HIV-1-dependent effector activities of CD4ζ. In addition, the number of CD4ζ-modified CD8{sup +} T cells maintained similar levels to that of the control even under HIV-1 infection. These results suggest that protecting CD4ζ-modified CD8{sup +} T cells from HIV-1 infection is required for prolonged HIV-1-specific immune surveillance. - Highlights: • Ectopic expression of CD4ζ CAR in CD8{sup +} T cells renders them susceptible to HIV-1 infection. • Co-expression of two anti-HIV-1 shRNAs protects CD4ζ CAR-modified CD8{sup +} T cells from HIV-1 infection. • Protecting CD4ζ CAR-modified CD8{sup +} T cells from HIV-1 infection suppresses its cytopathic effect.

  8. Ectopic expression of EbFIE from apomictic Eulaliopsis binata in rice results in pleiotropic phenotypes likely due to interaction with OsCLF.

    PubMed

    Ma, Kai; Zhang, Dongliang; Liu, Yaqin; Ouyang, Yidan; Li, Jiajia; Hu, Chungen; Yao, Jialing

    2015-05-01

    FERTILIZATION INDEPENDENT ENDOSPERM (FIE) is a core component of PcG complexes and functions in plant phase transition and seed generation. However, understanding in its function of apomictic monocot plants remains blank. Here an FIE homology EbFIE, has been isolated from apomictic Graminae species Eulaliopsis binata. EbFIE shares higher homology to OsFIE2 than OsFIE1, and has been classified into the monocot FIE2 clade. In addition, the broad expression pattern of EbFIE is also similar to OsFIE2. While, ectopic expression of EbFIE in rice resulted in pleiotropic phenotypes similar to that of OsFIE1 over-expressing plants. Meanwhile, EbFIE could bind OsCLF in vitro as OsFIE1 but different with OsFIE2. Molecular models comparison indicated that both EbFIE and OsFIE1 had a smaller E(z) protein binding groove than OsFIE2. Further site-directed mutagenesis analysis revealed that single amino acid substitution of I194F in OsFIE2 could improve its OsCLF binding capacity. Taken together, our results suggested that EbFIE was a conserved FIE homolog belonging to monocot FIE2 clade, but due to the similarity in protein conformation with FIE1, EbFIE might play a broad role in vegetative and reproductive development regulation by interaction with CLF homolog.

  9. Ectopic expression of an apple apomixis-related gene MhFIE induces co-suppression and results in abnormal vegetative and reproductive development in tomato.

    PubMed

    Liu, Dan-Dan; Dong, Qing-Long; Fang, Mou-Jing; Chen, Ke-Qin; Hao, Yu-Jin

    2012-12-15

    It has been well documented that FERTILIZATION-INDEPENDENT ENDOSPERM (FIE) plays important regulatory roles in diverse developmental processes in model plant Arabidopsis thaliana. However, it is largely unknown how FIE genes function in economically important crops. In this study, MhFIE gene, which was previously isolated from apomictic tea crabapple (Malus hupehensis Redh. var. pingyiensis), was introduced into tomato. The hemizygous transgenic tomato lines produced curly leaves and decreased in seed germination. In addition, the co-suppression of the transgenic MhFIE and endogenous (SlFIE) genes occurred in homozygous transgenic tomatoes. As a result, FIE silencing brought about abnormal phenotypes during reproductive development in tomato, such as increased sepal and petal numbers in flower, a fused ovule and pistil and parthenocarpic fruit formation. A yeast two-hybrid assay and bimolecular fluorescence complementation (BiFC) demonstrated that MhFIE interacted with a tomato protein, EZ2 (SlEZ2). Its ectopic expression and SlFIE co-suppression notably influenced the expression of genes associated with leaf, flower, and fruit development. Therefore, together with other PcG proteins, FIE was involved in the regulation of vegetative and reproductive development by modulating the expression of related genes in plants.

  10. Ectopic Expression of a Glycine soja myo-Inositol Oxygenase Gene (GsMIOX1a) in Arabidopsis Enhances Tolerance to Alkaline Stress

    PubMed Central

    Chen, Chen; Sun, Xiaoli; Duanmu, Huizi; Yu, Yang; Liu, Ailin; Xiao, Jialei; Zhu, Yanming

    2015-01-01

    Myo-inositol participates in various aspects of plant physiology, and myo-inositol oxygenase is the key enzyme of the myo-inositol oxygenation pathway. Previous studies indicated that myo-inositol oxygenase may play a role in plant responses to abiotic stresses. In this study, we focused on the functional characterization of GsMIOX1a, a remarkable alkaline stress-responsive gene of Glycine soja 07256, based on RNA-seq data. Using quantitative real-time PCR, we demonstrated that GsMIOX1a is rapidly induced by alkaline stress and expressed predominantly in flowers. We also elucidated the positive function of GsMIOX1a in the alkaline response in the wild type, atmiox1 mutant as well as GsMIOX1a-overexpressing Arabidopsis. We determined that atmiox1 mutant decreased Arabidopsis tolerance to alkaline stress, whereas GsMIOX1a overexpression increased tolerance. Moreover, the expression levels of some alkaline stress-responsive and inducible marker genes, including H+-Ppase, NADP-ME, KIN1 and RD29B, were also up-regulated in GsMIOX1a overexpression lines compared with the wild type and atmiox1 mutant. Together, these results suggest that the GsMIOX1a gene positively regulates plant tolerance to alkaline stress. This is the first report to demonstrate that ectopic expression of myo-inositol oxygenase improves alkaline tolerance in plants. PMID:26091094

  11. Ectopic Expression of MiR-125a Inhibits the Proliferation and Metastasis of Hepatocellular Carcinoma by Targeting MMP11 and VEGF

    PubMed Central

    Du, Rui; Fan, Rui; Gao, Liucun; Jin, Jiang; Liang, Shuhui; Chen, Zheng; Xu, Guanghui; Nie, Yongzhan; Wu, Kaichun; Liu, Jie; Shi, Yongquan; Ding, Jie; Fan, Daiming

    2012-01-01

    Background Studies have been shown that miR-125a plays an important role in carcinogenesis, however, the role of miR-125a in hepatocellular carcinoma (HCC) remains elusive. Methodology/Principal Real time-PCR (qRT-PCR) was performed to test the significance of miR-125a in HCC. Ectopic expression of miR-125a was used to test the influences of miR-125a on proliferation and metastasis of HCC cells in vitro and in vivo. Predicted target genes of miR-125a were determined by dual-luciferase reporting, qRT-PCR, and western blot (WB) analyses. Then immunohistochemical staining (IHC) was used to detect the expression of target genes, and the correlations and prognostic values of miR-125a and its target genes were also investigated. Conclusions/Significance Decreased miR-125a was observed in both HCC tissues and cell lines, and associated with patients’ aggressive pathologic features. Up-regulating miR-125a significantly inhibited the malignant phenotypes by repressing the expression of matrix metalloproteinase 11 (MMP11) and vascular endothelial growth factor A (VEGF-A) both in vitro and in vivo. Furthermore, miR-125a expression was inversely correlated with both MMP11 and VEGF-A expression in HCC tissues. Inhibiting miR-125a could increase both MMP11 and VEGF-A expression, and RNA interference targeting MMP11 or VEGF-A mRNA could rescue the loss of miR-125a functions. MiR-125a inhibits the proliferation and metastasis of HCC by targeting MMP11 and VEGF-A. Up-regulation of miR-125a might be a promising approach and a prognostic marker for HCC. PMID:22768249

  12. The Flexibility of Ectopic Lipids.

    PubMed

    Loher, Hannah; Kreis, Roland; Boesch, Chris; Christ, Emanuel

    2016-09-14

    In addition to the subcutaneous and the visceral fat tissue, lipids can also be stored in non-adipose tissue such as in hepatocytes (intrahepatocellular lipids; IHCL), skeletal (intramyocellular lipids; IMCL) or cardiac muscle cells (intracardiomyocellular lipids; ICCL). Ectopic lipids are flexible fuel stores that can be depleted by physical exercise and repleted by diet. They are related to obesity and insulin resistance. Quantification of IMCL was initially performed invasively, using muscle biopsies with biochemical and/or histological analysis. ¹H-magnetic resonance spectroscopy (¹H-MRS) is now a validated method that allows for not only quantifying IMCL non-invasively and repeatedly, but also assessing IHCL and ICCL. This review summarizes the current available knowledge on the flexibility of ectopic lipids. The available evidence suggests a complex interplay between quantitative and qualitative diet, fat availability (fat mass), insulin action, and physical exercise, all important factors that influence the flexibility of ectopic lipids. Furthermore, the time frame of the intervention on these parameters (short-term vs. long-term) appears to be critical. Consequently, standardization of physical activity and diet are critical when assessing ectopic lipids in predefined clinical situations.

  13. The Flexibility of Ectopic Lipids

    PubMed Central

    Loher, Hannah; Kreis, Roland; Boesch, Chris; Christ, Emanuel

    2016-01-01

    In addition to the subcutaneous and the visceral fat tissue, lipids can also be stored in non-adipose tissue such as in hepatocytes (intrahepatocellular lipids; IHCL), skeletal (intramyocellular lipids; IMCL) or cardiac muscle cells (intracardiomyocellular lipids; ICCL). Ectopic lipids are flexible fuel stores that can be depleted by physical exercise and repleted by diet. They are related to obesity and insulin resistance. Quantification of IMCL was initially performed invasively, using muscle biopsies with biochemical and/or histological analysis. 1H-magnetic resonance spectroscopy (1H-MRS) is now a validated method that allows for not only quantifying IMCL non-invasively and repeatedly, but also assessing IHCL and ICCL. This review summarizes the current available knowledge on the flexibility of ectopic lipids. The available evidence suggests a complex interplay between quantitative and qualitative diet, fat availability (fat mass), insulin action, and physical exercise, all important factors that influence the flexibility of ectopic lipids. Furthermore, the time frame of the intervention on these parameters (short-term vs. long-term) appears to be critical. Consequently, standardization of physical activity and diet are critical when assessing ectopic lipids in predefined clinical situations. PMID:27649157

  14. Cotinine exposure increases Fallopian tube PROKR1 expression via nicotinic AChRalpha-7: a potential mechanism explaining the link between smoking and tubal ectopic pregnancy.

    PubMed

    Shaw, Julie L V; Oliver, Elizabeth; Lee, Kai-Fai; Entrican, Gary; Jabbour, Henry N; Critchley, Hilary O D; Horne, Andrew W

    2010-11-01

    Tubal ectopic pregnancy (EP) is the most common cause of maternal mortality in the first trimester of pregnancy; however, its etiology is uncertain. In EP, embryo retention within the Fallopian tube (FT) is thought to be due to impaired smooth muscle contractility (SMC) and alterations in the tubal microenvironment. Smoking is a major risk factor for EP. FTs from women with EP exhibit altered prokineticin receptor-1 (PROKR1) expression, the receptor for prokineticins (PROK). PROK1 is angiogenic, regulates SMC, and is involved in intrauterine implantation. We hypothesized that smoking predisposes women to EP by altering tubal PROKR1 expression. Sera/FT were collected at hysterectomy (n=21). Serum levels of the smoking metabolite, cotinine, were measured by enzyme-linked immunosorbent assay. FTs were analyzed by q-RT-PCR, immunohistochemistry, and Western blotting for expression of PROKR1 and the predicted cotinine receptor, nicotinic acetylcholine receptor α-7 (AChRα-7). FT explants (n=4) and oviductal epithelial cells (cell line OE-E6/E7) were treated with cotinine and an nAChRα-7 antagonist. PROKR1 transcription was higher in FTs from smokers (P<0.01). nAChRα-7 expression was demonstrated in FT epithelium. Cotinine treatment of FT explants and OE-E6/E7 cells increased PROKR1 expression (P<0.05), which was negated by cotreatment with nAChRα-7 antagonist. Smoking targets human FTs via nAChRα-7 to increase tubal PROKR1, leading to alterations in the tubal microenvironment that could predispose to EP.

  15. RUNX2 correlates with subtype-specific breast cancer in a human tissue microarray, and ectopic expression of Runx2 perturbs differentiation in the mouse mammary gland.

    PubMed

    McDonald, Laura; Ferrari, Nicola; Terry, Anne; Bell, Margaret; Mohammed, Zahra M; Orange, Clare; Jenkins, Alma; Muller, William J; Gusterson, Barry A; Neil, James C; Edwards, Joanne; Morris, Joanna S; Cameron, Ewan R; Blyth, Karen

    2014-05-01

    RUNX2, a master regulator of osteogenesis, is oncogenic in the lymphoid lineage; however, little is known about its role in epithelial cancers. Upregulation of RUNX2 in cell lines correlates with increased invasiveness and the capacity to form osteolytic disease in models of breast and prostate cancer. However, most studies have analysed the effects of this gene in a limited number of cell lines and its role in primary breast cancer has not been resolved. Using a human tumour tissue microarray, we show that high RUNX2 expression is significantly associated with oestrogen receptor (ER)/progesterone receptor (PR)/HER2-negative breast cancers and that patients with high RUNX2 expression have a poorer survival rate than those with negative or low expression. We confirm RUNX2 as a gene that has a potentially important functional role in triple-negative breast cancer. To investigate the role of this gene in breast cancer, we made a transgenic model in which Runx2 is specifically expressed in murine mammary epithelium under the control of the mouse mammary tumour virus (MMTV) promoter. We show that ectopic Runx2 perturbs normal development in pubertal and lactating animals, delaying ductal elongation and inhibiting lobular alveolar differentiation. We also show that the Runx2 transgene elicits age-related, pre-neoplastic changes in the mammary epithelium of older transgenic animals, suggesting that elevated RUNX2 expression renders such tissue more susceptible to oncogenic changes and providing further evidence that this gene might have an important, context-dependent role in breast cancer.

  16. Monoacylglycerols Are Components of Root Waxes and Can Be Produced in the Aerial Cuticle by Ectopic Expression of a Suberin-Associated Acyltransferase1[W][OA

    PubMed Central

    Li, Yonghua; Beisson, Fred; Ohlrogge, John; Pollard, Mike

    2007-01-01

    The interface between plants and the environment is provided for aerial organs by epicuticular waxes that have been extensively studied. By contrast, little is known about the nature, biosynthesis, and role of waxes at the root-rhizosphere interface. Waxes isolated by rapid immersion of Arabidopsis (Arabidopsis thaliana) roots in organic solvents were rich in saturated C18-C22 alkyl esters of p-hydroxycinnamic acids, but also contained significant amounts of both α- and β-isomers of monoacylglycerols with C22 and C24 saturated acyl groups and the corresponding free fatty acids. Production of these compounds in root waxes was positively correlated to the expression of sn-glycerol-3-P acyltransferase5 (GPAT5), a gene encoding an acyltransferase previously shown to be involved in aliphatic suberin synthesis. This suggests a direct metabolic relationship between suberin and some root waxes. Furthermore, when ectopically expressed in Arabidopsis, GPAT5 produced very-long-chain saturated monoacylglycerols and free fatty acids as novel components of cuticular waxes. The crystal morphology of stem waxes was altered and the load of total stem wax compounds was doubled, although the major components typical of the waxes found on wild-type plants decreased. These results strongly suggest that GPAT5 functions in vivo as an acyltransferase to a glycerol-containing acceptor and has access to the same pool of acyl intermediates and/or may be targeted to the same membrane domain as that of wax synthesis in aerial organs. PMID:17496107

  17. Ectopic expression of a polyalanine expansion mutant of poly(A)-binding protein N1 in muscle cells in culture inhibits myogenesis

    SciTech Connect

    Wang Qishan; Bag, Jnanankur . E-mail: jbag@uoguelph.ca

    2006-02-17

    Oculopharyngeal muscular dystrophy (OPMD) is an adult-onset dominant genetic disease caused by the expansion of a GCG trinucleotide repeat that encodes the polyalanine tract at the N-terminus of the nuclear poly(A)-binding protein (PABPN1). Presence of intranuclear inclusions (INIs) containing PABPN1 aggregates in the skeletal muscles is the hallmark of OPMD. Here, we show that ectopic expression of the mutant PABPN1 produced INIs in a muscle cell culture model and reduced expression of several muscle-specific proteins including {alpha}-actin, slow troponin C, muscle creatine kinase, and two myogenic transcription factors, myogenin and MyoD. However, the levels of two upstream regulators of the MyoD gene, the Myf-5 and Pax3/7, were not affected, but both proteins co-localized with the PABPN1 aggregates in the mutant PABPN1 overexpressing cells. In these cells, although myogenin and MyoD levels were reduced, these two transcription factors did not co-localize with the mutant PABPN1 aggregates. Therefore, sequestration of Myf5 and Pax3/7 by the mutant PABPN1 aggregates was a specific effect on these factors. Our results suggest that trapping of these two important myogenic determinants may interfere with an early step in myogenesis.

  18. The silver lining of a viral agent: increasing seed yield and harvest index in Arabidopsis by ectopic expression of the potato leaf roll virus movement protein.

    PubMed

    Kronberg, Kristin; Vogel, Florian; Rutten, Twan; Hajirezaei, Mohammed-Reza; Sonnewald, Uwe; Hofius, Daniel

    2007-11-01

    Ectopic expression of viral movement proteins (MPs) has previously been shown to alter plasmodesmata (PD) function and carbon partitioning in transgenic plants, giving rise to the view of PD being dynamic and highly regulated structures that allow resource allocation to be adapted to environmental and developmental needs. However, most work has been restricted to solanaceous species and the potential use of MP expression to improve biomass and yield parameters has not been addressed in detail. Here we demonstrate that MP-mediated modification of PD function can substantially alter assimilate allocation, biomass production, and reproductive growth in Arabidopsis (Arabidopsis thaliana). These effects were achieved by constitutive expression of the potato leaf roll virus 17-kD MP (MP17) fused to green fluorescent protein (GFP) in different Arabidopsis ecotypes. The resulting transgenic plants were analyzed for PD localization of the MP17:GFP fusion protein and different lines with low to high expression levels were selected for further analysis. Low-level accumulation of MP17 resulted in enhanced sucrose efflux from source leaves and a considerably increased vegetative biomass production. In contrast, high MP17 levels impaired sucrose export, resulting in source leaf-specific carbohydrate accumulation and a strongly reduced vegetative growth. Surprisingly, later during development the MP17-mediated inhibition of resource allocation was reversed, and final seed yield increased in average up to 30% in different transgenic lines as compared to wild-type plants. This resulted in a strongly improved harvest index. The release of the assimilate export block was paralleled by a reduced PD binding of MP17 in senescing leaves, indicating major structural changes of PD during leaf senescence.

  19. A Complex Structural Variation on Chromosome 27 Leads to the Ectopic Expression of HOXB8 and the Muffs and Beard Phenotype in Chickens

    PubMed Central

    Wang, Yanqiang; Luo, Chenglong; Liu, Ranran; Qu, Hao; Shu, Dingming; Wen, Jie; Crooijmans, Richard P. M. A.; Zhao, Yiqiang; Hu, Xiaoxiang; Li, Ning

    2016-01-01

    Muffs and beard (Mb) is a phenotype in chickens where groups of elongated feathers gather from both sides of the face (muffs) and below the beak (beard). It is an autosomal, incomplete dominant phenotype encoded by the Muffs and beard (Mb) locus. Here we use genome-wide association (GWA) analysis, linkage analysis, Identity-by-Descent (IBD) mapping, array-CGH, genome re-sequencing and expression analysis to show that the Mb allele causing the Mb phenotype is a derived allele where a complex structural variation (SV) on GGA27 leads to an altered expression of the gene HOXB8. This Mb allele was shown to be completely associated with the Mb phenotype in nine other independent Mb chicken breeds. The Mb allele differs from the wild-type mb allele by three duplications, one in tandem and two that are translocated to that of the tandem repeat around 1.70 Mb on GGA27. The duplications contain total seven annotated genes and their expression was tested during distinct stages of Mb morphogenesis. A continuous high ectopic expression of HOXB8 was found in the facial skin of Mb chickens, strongly suggesting that HOXB8 directs this regional feather-development. In conclusion, our results provide an interesting example of how genomic structural rearrangements alter the regulation of genes leading to novel phenotypes. Further, it again illustrates the value of utilizing derived phenotypes in domestic animals to dissect the genetic basis of developmental traits, herein providing novel insights into the likely role of HOXB8 in feather development and differentiation. PMID:27253709

  20. The onset of human ectopic pregnancy demonstrates a differential expression of miRNAs and their cognate targets in the Fallopian tube.

    PubMed

    Feng, Yi; Zou, Shien; Weijdegård, Birgitta; Chen, Jie; Cong, Qing; Fernandez-Rodriguez, Julia; Wang, Lei; Billig, Håkan; Shao, Ruijin

    2014-01-01

    Human ectopic pregnancy (EP) is a leading cause of pregnancy-related death, but the molecular basis underlying the onset of tubal EP is largely unknown. Female Dicer1 conditional knockout mice are infertile with dysfunctional Fallopian tube and have a different miRNA expression profile compared to wild-type mice, and we speculated that Dicer-mediated regulation of miRNA expression and specific miRNA-controlled targets might contribute to the onset of tubal EP. In the present study, we used microarray analysis and quantitative RT-PCR to examine the expression of miRNAs and core miRNA regulatory components in Fallopian tube tissues from women with EP. We found that the levels of DICER1, four miRNAs (let-7i, miR-149, miR-182, and miR-424), and estrogen receptor α distinguished the tubal implantation site from the non-implantation site. Computational algorithms and screening for interactions with the estrogen and progesterone receptor signaling pathways showed that the four miRNAs were predicted to target ten genes, including NEDD4, TAF15, and SPEN. Subsequent experiments showed differences in NEDD4 mRNA and protein levels between the implantation and non-implantation sites. Finally, we revealed that increases in smooth muscle cell NEDD4 and stromal cell TAF15, in parallel with a decrease in epithelial cell SPEN, were associated with tubal implantation. Our study suggests that changes in miRNA levels by the DICER-mediated miRNA-processing machinery result in aberrant expression of cell type-specific proteins that are potentially involved in the onset of tubal EP.

  1. Ectopic molar pregnancy: a case report.

    PubMed

    Bousfiha, Najoua; Erarhay, Sanaa; Louba, Adnane; Saadi, Hanan; Bouchikhi, Chahrazad; Banani, Abdelaziz; El Fatemi, Hind; Sekkal, Med; Laamarti, Afaf

    2012-01-01

    The incidence of hydatidiform moles is 1 per 1,000 pregnancies. Ectopic pregnancy occurs in 20 per 1,000 pregnancies. Thus, the incidence of the ectopic molar gestation is very rare. We report a case of tubal molar pregnancy diagnosed at the systematic histology exam of an ectopic pregnancy. We report the case of 32 years old nulliparus women who presented a vaginal bleeding, lower abdominal pain and 6 weeks amenorrhea corresponding to the last menstrual period. At the clinical examination, the arterial pressure was 100/60 mmHG. The gynecological examination was difficult because of lower abdominal pain. Serum gonadotropin activity was 3454 ui/l. Pelvic ultrasound revealed an irregular echogenic mass in the left adnexa. Diagnostic laparoscopy revealed a left-sided unruptured ampullary ectopic pregnancy. A left laparoscopic salpingectomy was performed. The systematic histologic test identified an ectopic partial molar pregnancy, which was confirmed by DNA ploidy image analysis. The patient was followed with weekly quantitative B-hCG titers until three successive B-hCG levels were negative. It is pertinent that clinicians take routine histological examination of tubal specimens in ectopic pregnancy very seriously in order to diagnose cases of ectopic molar gestations early and mount appropriate post treatment surveillance.

  2. [Transverse ectopic testis].

    PubMed

    Jouini, Riadh; Lefi, Mounir; Sami, Chelly; Manef, Gesmi; Mohsen, Belguith; Nouri, Abdellatif

    2002-09-01

    Transverse ectopic testis (TET) is a rare form of ectopic testis. The authors report the case of a 2-month-old infant presenting with right inguinoscrotal hernia and ectopic left testis with an impalpable testis. Opening of the hernia sac revealed two testes with two distally fused vasa deferentes. The contralateral testis was easily descended by translocation through the other inguinal canal. A favourable result was obtained with two testes situated in a normal position. In the light of this case, the authors emphasize the clinical and therapeutic features of this anomaly.

  3. Activation of the Hedgehog Pathway in the Mouse Fetal Ovary Leads to Ectopic Appearance of Fetal Leydig Cells and Female Pseudohermaphroditism

    PubMed Central

    Barsoum, Ivraym B.; Bingham, Nathan C.; Parker, Keith L.; Jorgensen, Joan S.; Yao, Humphrey H-C

    2009-01-01

    Proper cell fate determination in mammalian gonads is critical for the establishment of sexual identity. The Hedgehog (Hh) pathway has been implicated in cell fate decision for various organs, including gonads. Desert Hedgehog (Dhh), one of the three mammalian Hh genes, has been implicated with other genes in the establishment of mouse fetal Leydig cells. To investigate whether Hh alone is sufficient to induce fetal Leydig cell differentiation, we ectopically activated the Hh pathway in Steroidogenic factor 1 (SF1)-positive somatic cell precursors of fetal ovaries. Hh activation transformed SF1-positive somatic ovarian cells into functional fetal Leydig cells. These ectopic fetal Leydig cells produced androgens and insulin-like growth factor 3 (INLS3) that cause virilization of female embryos and ovarian descent. However, the female reproductive system remained intact, indicating a typical example of female pseudohermaphroditism. The appearance of fetal Leydig cells was a direct consequence of Hh activation as evident by the absence of other testicular components in the affected ovary. This study provides not only insights into mechanisms of cell lineage specification in gonads, but also a model to understand defects in sexual differentiation. PMID:19268447

  4. Ectopic expressed miR-203 contributes to chronic obstructive pulmonary disease via targeting TAK1 and PIK3CA.

    PubMed

    Shi, Liang; Xin, Qinghong; Chai, Ruonan; Liu, Lei; Ma, Zhuang

    2015-01-01

    MiRNA is a group of powerful short non-coding RNAs that suppress the expression of protein coding genes by targeting to the 3'UTRs of mRNAs. Some researchers have detected the miRNAs expression profile in tissue and blood samples of chronic obstructive pulmonary disease (COPD) patients recently. Several disturbed miRNAs were found to be related to COPD; however, the mechanisms were still well understood. In this study, we first detected the expression of 11 candidate miRNAs in the lung samples of COPD patients, non-COPD smokers and non-smock controls. We found that the expression of miR-181a, miR-203, miR-338, miR-1 and miR-199a was altered compared with control. Subsequently, we detected these five miRNAs expression in the blood samples of the participants. A significant higher expression of miR-203 was found in the blood samples of smokers and COPD patients. Predicted by bioinformatics tools and confirmed by luciferase assay and western blot, we demonstrated that TAK1 and PIK3CA are two direct targets of miR-203. Furthermore, we detected a lower p-IκBα and p-p65 level in the bronchial/tracheal epithelial cells from COPD patients compared with the cells from healthy controls, when stimulated by LPS. The concentration of TNF-α and IL-6 in the medium from bronchial/tracheal epithelial cells from COPD patients is also lower. Meanwhile, the miR-203 level was down-regulated significantly in the control cells, but non-significant change in the cells from COPD patients. miR-203 represses NF-κB signaling via targeting TAK1 and PI3KCA and miR-203 overexpression may contribute to the COPD initiation.

  5. Ectopic Expression of an Atypical Hydrophobic Group 5 LEA Protein from Wild Peanut, Arachis diogoi Confers Abiotic Stress Tolerance in Tobacco

    PubMed Central

    Sharma, Akanksha; Kumar, Dilip; Kumar, Sumit; Rampuria, Sakshi; Reddy, Attipalli R.; Kirti, Pulugurtha Bharadwaja

    2016-01-01

    Late embryogenesis abundant (LEA) proteins are a group of hydrophilic proteins, which accumulate in plants under varied stress conditions like drought, salinity, extreme temperatures and oxidative stress suggesting their role in the protection of plants against these stresses. A transcript derived fragment (TDF) corresponding to LEA gene, which got differentially expressed in wild peanut, Arachis diogoi against the late leaf spot pathogen, Phaeoisariopsis personata was used in this study. We have cloned its full length cDNA by RACE-PCR, which was designated as AdLEA. AdLEA belongs to the atypical Group 5C of LEA protein family as confirmed by sequence analysis. Group 5C LEA protein subfamily contains Pfam LEA_2 domain and is highly hydrophobic. In native conditions, expression of AdLEA was upregulated considerably upon hormonal and abiotic stress treatments emphasizing its role in abiotic stress tolerance. Subcellular localization studies showed that AdLEA protein is distributed in both nucleus and cytosol. Ectopic expression of AdLEA in tobacco resulted in enhanced tolerance of plants to dehydration, salinity and oxidative stress with the transgenic plants showing higher chlorophyll content and reduced lipid peroxidation as compared to wild type plants. Overexpressed AdLEA tobacco plants maintained better photosynthetic efficiency under drought conditions as demonstrated by chlorophyll fluorescence measurements. These plants showed enhanced transcript accumulation of some stress-responsive genes. Our study also elucidates that ROS levels were significantly reduced in leaves and stomatal guard cells of transgenic plants upon stress treatments. These results suggest that AdLEA confers multiple stress tolerance to plants, which make it a potential gene for genetic modification in plants. PMID:26938884

  6. Ectopic Expression of an Atypical Hydrophobic Group 5 LEA Protein from Wild Peanut, Arachis diogoi Confers Abiotic Stress Tolerance in Tobacco.

    PubMed

    Sharma, Akanksha; Kumar, Dilip; Kumar, Sumit; Rampuria, Sakshi; Reddy, Attipalli R; Kirti, Pulugurtha Bharadwaja

    2016-01-01

    Late embryogenesis abundant (LEA) proteins are a group of hydrophilic proteins, which accumulate in plants under varied stress conditions like drought, salinity, extreme temperatures and oxidative stress suggesting their role in the protection of plants against these stresses. A transcript derived fragment (TDF) corresponding to LEA gene, which got differentially expressed in wild peanut, Arachis diogoi against the late leaf spot pathogen, Phaeoisariopsis personata was used in this study. We have cloned its full length cDNA by RACE-PCR, which was designated as AdLEA. AdLEA belongs to the atypical Group 5C of LEA protein family as confirmed by sequence analysis. Group 5C LEA protein subfamily contains Pfam LEA_2 domain and is highly hydrophobic. In native conditions, expression of AdLEA was upregulated considerably upon hormonal and abiotic stress treatments emphasizing its role in abiotic stress tolerance. Subcellular localization studies showed that AdLEA protein is distributed in both nucleus and cytosol. Ectopic expression of AdLEA in tobacco resulted in enhanced tolerance of plants to dehydration, salinity and oxidative stress with the transgenic plants showing higher chlorophyll content and reduced lipid peroxidation as compared to wild type plants. Overexpressed AdLEA tobacco plants maintained better photosynthetic efficiency under drought conditions as demonstrated by chlorophyll fluorescence measurements. These plants showed enhanced transcript accumulation of some stress-responsive genes. Our study also elucidates that ROS levels were significantly reduced in leaves and stomatal guard cells of transgenic plants upon stress treatments. These results suggest that AdLEA confers multiple stress tolerance to plants, which make it a potential gene for genetic modification in plants.

  7. Cellular localization and effects of ectopically expressed hepatitis A virus proteins 2B, 2C, 3A and their intermediates 2BC, 3AB and 3ABC.

    PubMed

    Seggewiß, Nicole; Kruse, Hedi Verena; Weilandt, Rebecca; Domsgen, Erna; Dotzauer, Andreas; Paulmann, Dajana

    2016-04-01

    In the course of hepatitis A virus (HAV) infections, the seven nonstructural proteins and their intermediates are barely detectable. Therefore, little is known about their functions and mechanisms of action. Ectopic expression of the presumably membrane-associated proteins 2B, 2C, 3A and their intermediates 2BC, 3AB and 3ABC allowed the intracellular localization of these proteins and their possible function during the replication cycle of HAV to be investigated. In this study, we used rhesus monkey kidney cells, which are commonly used for cell culture experiments, and human liver cells, which are the natural target cells. We detected specific associations of these proteins with distinct membrane compartments and the cytoskeleton, different morphological alterations of the respective structures, and specific effects on cellular functions. Besides comparable findings in both cell lines used with regard to localization and effects of the proteins examined, we also found distinct differences. The data obtained identify so far undocumented interactions with and effects of the HAV proteins investigated on cellular components, which may reflect unknown aspects of the interaction of HAV with the host cell, for example the modification of the ERGIC (ER-Golgi intermediate compartment) structure, an interaction with lipid droplets and lysosomes, and inhibition of the classical secretory pathway.

  8. Complementation of Seed Maturation Phenotypes by Ectopic Expression of ABSCISIC ACID INSENSITIVE3, FUSCA3 and LEAFY COTYLEDON2 in Arabidopsis.

    PubMed

    Roscoe, Thomas T; Guilleminot, Jocelyne; Bessoule, Jean-Jacques; Berger, Frédéric; Devic, Martine

    2015-06-01

    ABSCISIC ACID INSENSITIVE3 (ABI3), FUSCA3 (FUS3) and LEAFY COTYLEDON2 (LEC2), collectively the AFL, are master regulators of seed maturation processes. This study examined the role of AFL in the production of seed reserves in Arabidopsis. Quantification of seed reserves and cytological observations of afl mutant embryos show that protein and lipid but not starch reserves are spatially regulated by AFL. Although AFL contribute to a common regulation of reserves, ABI3 exerts a quantitatively greater control over storage protein content whereas FUS3 controls lipid content to a greater extent. Although ABI3 controls the reserve content throughout the embryo, LEC2 and FUS3 regulate reserves in distinct embryonic territories. By analyzing the ability of an individual ectopically expressed AFL to suppress afl phenotypes genetically, we show that conserved domains common to each component of the AFL are sufficient for the initiation of storage product synthesis and the establishment of embryo morphology. This confirms redundancy among the AFL and indicates a threshold necessary for function within the AFL pool. Since no individual AFL was able to suppress the tolerance to desiccation, mid- and late-maturation programs were uncoupled.

  9. Early B-cell-specific inactivation of ATM synergizes with ectopic CyclinD1 expression to promote pre-germinal center B-cell lymphomas in mice.

    PubMed

    Yamamoto, K; Lee, B J; Li, C; Dubois, R L; Hobeika, E; Bhagat, G; Zha, S

    2015-06-01

    Ataxia telangiectasia-mutated (ATM) kinase is a master regulator of the DNA damage response. ATM is frequently inactivated in human B-cell non-Hodgkin lymphomas, including ~50% of mantle cell lymphomas (MCLs) characterized by ectopic expression of CyclinD1. Here we report that early and robust deletion of ATM in precursor/progenitor B cells causes cell autonomous, clonal mature B-cell lymphomas of both pre- and post-germinal center (GC) origins. Unexpectedly, naive B-cell-specific deletion of ATM is not sufficient to induce lymphomas in mice, highlighting the important tumor suppressor function of ATM in immature B cells. Although EμCyclinD1 is not sufficient to induce lymphomas, EμCyclinD1 accelerates the kinetics and increases the incidence of clonal lymphomas in ATM-deficient B-cells and skews the lymphomas toward pre-GC-derived small lymphocytic neoplasms, sharing morphological features of human MCL. This is in part due to CyclinD1-driven expansion of ATM-deficient naive B cells with genomic instability, which promotes the deletions of additional tumor suppressor genes (i.e. Trp53, Mll2, Rb1 and Cdkn2a). Together these findings define a synergistic function of ATM and CyclinD1 in pre-GC B-cell proliferation and lymphomagenesis and provide a prototypic animal model to study the pathogenesis of human MCL.

  10. [The laparoscopic management of ectopic pregnancy].

    PubMed

    Solar, O; Gómez, E

    1992-01-01

    Twenty-five ectopics pregnancies were treated successfully by laparoscopic approach from november 1990 to april 1992. The technics used were: salpingostomy, salpingectomy and taking out trophoblastic tissue exclusively. At the present time, two out of eight women who have wanted become pregnant, have delivered a healthy babies. We concluded that the ectopic pregnancy is efficiently managed by laparoscopy with advantages such as: minimal scar, short time in the hospital, quickly return to total activity, low costs, etc.

  11. [Ectopic pregnancy in Senegal].

    PubMed

    Cissé, Cheikh A Tidiane; Bernis, Luc De; Faye, El Hadj Ousseynou; Diadhiou, Fadel

    2002-01-01

    The objective of this prospective study was to analyse the epidemiology and prognosis of ectopic pregnancy in Senegal. From January 1 to December 31, 1996, 255 ectopic pregnancies were registered. The national rate of ectopic pregnancy was 0.6%. of expected pregnancies. However, rates differed greatly between areas in Senegal, with extremes ranging from 0.85%. in Dakar to 0.32%. in Thiès. The epidemiological profile was that of a young woman-mean age: 23 years old, mean parity=3, admitted with broken ectopic pregnancy (95%). A salpingectomy was performed in all cases. The maternal mortality rate was 1.20%, while morbidity, mainly due to post-operative infection, was found in 2.7% of the cases.

  12. Ectopic expression of a cyanobacterial flavodoxin in creeping bentgrass impacts plant development and confers broad abiotic stress tolerance.

    PubMed

    Li, Zhigang; Yuan, Shuangrong; Jia, Haiyan; Gao, Fangyuan; Zhou, Man; Yuan, Ning; Wu, Peipei; Hu, Qian; Sun, Dongfa; Luo, Hong

    2016-09-16

    Flavodoxin (Fld) plays a pivotal role in photosynthetic microorganisms as an alternative electron carrier flavoprotein under adverse environmental conditions. Cyanobacterial Fld has been demonstrated to be able to substitute ferredoxin of higher plants in most electron transfer processes under stressful conditions. We have explored the potential of Fld for use in improving plant stress response in creeping bentgrass (Agrostis stolonifera L.). Overexpression of Fld altered plant growth and development. Most significantly, transgenic plants exhibited drastically enhanced performance under oxidative, drought and heat stress as well as nitrogen (N) starvation, which was associated with higher water retention and cell membrane integrity than wild-type controls, modified expression of heat-shock protein genes, production of more reduced thioredoxin, elevated N accumulation and total chlorophyll content as well as up-regulated expression of nitrite reductase and N transporter genes. Further analysis revealed that the expression of other stress-related genes was also impacted in Fld-expressing transgenics. Our data establish a key role of Fld in modulating plant growth and development and plant response to multiple sources of adverse environmental conditions in crop species. This demonstrates the feasibility of manipulating Fld in crop species for genetic engineering of plant stress tolerance.

  13. Ectopic expression of AtPAD4 broadens resistance of soybean to soybean cyst and root-knot nematodes

    PubMed Central

    2013-01-01

    Background The gene encoding PAD4 (PHYTOALEXIN-DEFICIENT4) is required in Arabidopsis for expression of several genes involved in the defense response to Pseudomonas syringae pv. maculicola. AtPAD4 (Arabidopsis thaliana PAD4) encodes a lipase-like protein that plays a regulatory role mediating salicylic acid signaling. Results We expressed the gene encoding AtPAD4 in soybean roots of composite plants to test the ability of AtPAD4 to deter plant parasitic nematode development. The transformed roots were challenged with two different plant parasitic nematode genera represented by soybean cyst nematode (SCN; Heterodera glycines) and root-knot nematode (RKN; Meloidogyne incognita). Expression of AtPAD4 in soybean roots decreased the number of mature SCN females 35 days after inoculation by 68 percent. Similarly, soybean roots expressing AtPAD4 exhibited 77 percent fewer galls when challenged with RKN. Conclusions Our experiments show that AtPAD4 can be used in an economically important crop, soybean, to provide a measure of resistance to two different genera of nematodes. PMID:23617694

  14. Ectopic expression of cytosolic superoxide dismutase and ascorbate peroxidase leads to salt stress tolerance in transgenic plums.

    PubMed

    Diaz-Vivancos, Pedro; Faize, Mohamed; Barba-Espin, Gregorio; Faize, Lydia; Petri, Cesar; Hernández, José Antonio; Burgos, Lorenzo

    2013-10-01

    To fortify the antioxidant capacity of plum plants, genes encoding cytosolic antioxidants ascorbate peroxidase (cytapx) and Cu/Zn-superoxide dismutase (cytsod) were genetically engineered in these plants. Transgenic plum plants expressing the cytsod and/or cytapx genes in cytosol have been generated under the control of the CaMV35S promoter. High levels of cytsod and cytapx gene transcripts suggested that the transgenes were constitutively and functionally expressed. We examined the potential functions of cytSOD and cytAPX in in vitro plum plants against salt stress (100 mm NaCl). Several transgenic plantlets expressing cytsod and/or cytapx showed an enhanced tolerance to salt stress, mainly lines C5-5 and J8-1 (expressing several copies of sod and apx, respectively). Transformation as well as NaCl treatments influenced the antioxidative metabolism of plum plantlets, including enzymatic and nonenzymatic antioxidants. Transgenic plantlets exhibited higher contents of nonenzymatic antioxidants glutathione and ascorbate than nontransformed control, which correlated with lower accumulation of hydrogen peroxide. Overall, our results suggest that transformation of plum plants with genes encoding antioxidant enzymes enhances the tolerance to salinity.

  15. Ki-1/57 and CGI-55 ectopic expression impact cellular pathways involved in proliferation and stress response regulation.

    PubMed

    Costa, Fernanda C; Saito, Angela; Gonçalves, Kaliandra A; Vidigal, Pedro M; Meirelles, Gabriela V; Bressan, Gustavo C; Kobarg, Jörg

    2014-12-01

    Ki-1/57 (HABP4) and CGI-55 (SERBP1) are regulatory proteins and paralogs with 40.7% amino acid sequence identity and 67.4% similarity. Functionally, they have been implicated in the regulation of gene expression on both the transcriptional and mRNA metabolism levels. A link with tumorigenesis is suggested, since both paralogs show altered expression levels in tumor cells and the Ki-1/57 gene is found in a region of chromosome 9q that represents a haplotype for familiar colon cancer. However, the target genes regulated by Ki-1/57 and CGI-55 are unknown. Here, we analyzed the alterations of the global transcriptome profile after Ki-1/57 or CGI-55 overexpression in HEK293T cells by DNA microchip technology. We were able to identify 363 or 190 down-regulated and 50 or 27 up-regulated genes for Ki-1/57 and CGI-55, respectively, of which 20 were shared between both proteins. Expression levels of selected genes were confirmed by qRT-PCR both after protein overexpression and siRNA knockdown. The majority of the genes with altered expression were associated to proliferation, apoptosis and cell cycle control processes, prompting us to further explore these contexts experimentally. We observed that overexpression of Ki-1/57 or CGI-55 results in reduced cell proliferation, mainly due to a G1 phase arrest, whereas siRNA knockdown of CGI-55 caused an increase in proliferation. In the case of Ki-1/57 overexpression, we found protection from apoptosis after treatment with the ER-stress inducer thapsigargin. Together, our data give important new insights that may help to explain these proteins putative involvement in tumorigenic events.

  16. Ectopic expression of Dahlia merckii defensin DmAMP1 improves papaya resistance to Phytophthora palmivora by reducing pathogen vigor.

    PubMed

    Zhu, Yun J; Agbayani, Ricelle; Moore, Paul H

    2007-06-01

    Phytophthora spp., some of the more important casual agents of plant diseases, are responsible for heavy economic losses worldwide. Plant defensins have been introduced as transgenes into a range of species to increase host resistance to pathogens to which they were originally susceptible. However, the effectiveness and mechanism of interaction of the defensins with Phytophthora spp. have not been clearly characterized in planta. In this study, we expressed the Dahlia merckii defensin, DmAMP1, in papaya (Carica papaya L.), a plant highly susceptible to a root, stem, and fruit rot disease caused by Phytophthora palmivora. Extracts of total leaf proteins from transformed plants inhibited growth of Phytophthora in vitro and discs cut from the leaves of transformed plants inhibited growth of Phytophthora in a bioassay. Results from our greenhouse inoculation experiments demonstrate that expressing the DmAMP1 gene in papaya plants increased resistance against P. palmivora and that this increased resistance was associated with reduced hyphae growth of P. palmivora at the infection sites. The inhibitory effects of DmAMP1 expression in papaya suggest this approach has good potential to impart transgenic resistance against Phytophthora in papaya.

  17. The ectopic expression of the wheat Puroindoline genes increase germ size and seed oil content in transgenic corn.

    PubMed

    Zhang, Jinrui; Martin, John M; Beecher, Brian; Lu, Chaofu; Hannah, L Curtis; Wall, Michael L; Altosaar, Illimar; Giroux, Michael J

    2010-11-01

    Plant oil content and composition improvement is a major goal of plant breeding and biotechnology. The Puroindoline a and b (PINA and PINB) proteins together control whether wheat seeds are soft or hard textured and share a similar structure to that of plant non-specific lipid-transfer proteins. Here we transformed corn (Zea mays L.) with the wheat (Triticum aestivum L.) puroindoline genes (Pina and Pinb) to assess their effects upon seed oil content and quality. Pina and Pinb coding sequences were introduced into corn under the control of a corn Ubiquitin promoter. Three Pina/Pinb expression positive transgenic events were evaluated over two growing seasons. The results showed that Pin expression increased germ size significantly without negatively impacting seed size. Germ yield increased 33.8% while total seed oil content was increased by 25.23%. Seed oil content increases were primarily the result of increased germ size. This work indicates that higher oil content corn hybrids having increased food or feed value could be produced via puroindoline expression.

  18. Ectopic expression of SlAGO7 alters leaf pattern and inflorescence architecture and increases fruit yield in tomato.

    PubMed

    Lin, Dongbo; Xiang, Ya; Xian, Zhiqiang; Li, Zhengguo

    2016-08-01

    ARGONAUTE7 (AGO7), a key regulator of the trans-acting small interfering RNAs (ta-siRNA) pathway, plays a conserved role in controlling leaf pattern among species. However, little is known about the ta-siRNA pathway in regulating inflorescence architecture and fruit yield. In this study, we characterized the expression pattern, subcellular localization and developmental functions of SlAGO7 in tomato (Solanum lycopersicum). Overexpressing SlAGO7 in tomato exhibited pleiotropic phenotypes, including improved axillary bud formation, altered leaf morphology and inflorescence architecture, and increased fruit yield. Cross-sectioning of leaves showed that the number of vascular bundles was significantly increased in 35:SlAGO7 lines. Overexpression of SlAGO7 increased the production of ta-siRNA, and repressed the expression ta-siRNA-targeted genes (SlARF2a, SlARF2b, SlARF3 and SlARF4). Further analysis showed that overexpression of SlAGO7 alters the expression of key genes implicated in leaf morphology, inflorescence architecture, auxin transport and signaling. In addition, the altered auxin response of 35:SlAGO7 lines were also investigated. These results suggested that SlAGO7 plays a positive role in determining inflorescence architecture and fruit yield though the ta-siRNA pathway. Therefore, SlAGO7 represents a useful gene that can be incorporated in tomato breeding programs for developing cultivars with yield potential.

  19. The Salmonella effector SseJ disrupts microtubule dynamics when ectopically expressed in normal rat kidney cells

    PubMed Central

    Raines, Sally A.; Hodgkinson, Michael R.; Dowle, Adam A.

    2017-01-01

    Salmonella effector protein SseJ is secreted by Salmonella into the host cell cytoplasm where it can then modify host cell processes. Whilst host cell small GTPase RhoA has previously been shown to activate the acyl-transferase activity of SseJ we show here an un-described effect of SseJ protein production upon microtubule dynamism. SseJ prevents microtubule collapse and this is independent of SseJ’s acyl-transferase activity. We speculate that the effects of SseJ on microtubules would be mediated via its known interactions with the small GTPases of the Rho family. PMID:28235057

  20. Human Cytomegalovirus Nuclear Egress Proteins Ectopically Expressed in the Heterologous Environment of Plant Cells are Strictly Targeted to the Nuclear Envelope.

    PubMed

    Lamm, Christian E; Link, Katrin; Wagner, Sabrina; Milbradt, Jens; Marschall, Manfred; Sonnewald, Uwe

    2016-03-10

    In all eukaryotic cells, the nucleus forms a prominent cellular compartment containing the cell's nuclear genome. Although structurally similar, animal and plant nuclei differ substantially in details of their architecture. One example is the nuclear lamina, a layer of tightly interconnected filament proteins (lamins) underlying the nuclear envelope of metazoans. So far no orthologous lamin genes could be detected in plant genomes and putative lamin-like proteins are only poorly described in plants. To probe for potentially conserved features of metazoan and plant nuclear envelopes, we ectopically expressed the core nuclear egress proteins of human cytomegalovirus pUL50 and pUL53 in plant cells. pUL50 localizes to the inner envelope of metazoan nuclei and recruits the nuclear localized pUL53 to it, forming heterodimers. Upon expression in plant cells, a very similar localization pattern of both proteins could be determined. Notably, pUL50 is specifically targeted to the plant nuclear envelope in a rim-like fashion, a location to which coexpressed pUL53 becomes strictly corecruited from its initial nucleoplasmic distribution. Using pUL50 as bait in a yeast two-hybrid screening, the cytoplasmic re-initiation supporting protein RISP could be identified. Interaction of pUL50 and RISP could be confirmed by coexpression and coimmunoprecipitation in mammalian cells and by confocal laser scanning microscopy in plant cells, demonstrating partial pUL50-RISP colocalization in areas of the nuclear rim and other intracellular compartments. Thus, our study provides strong evidence for conserved structural features of plant and metazoan nuclear envelops and identifies RISP as a potential pUL50-interacting plant protein.

  1. Prevalence of cytomegalovirus, and its effect on the expression of inducible and endothelial nitric oxide synthases in Fallopian tubes collected from women with and without ectopic pregnancy.

    PubMed

    Batwa, S A; Ashshi, A M; Kamfar, F F; Ahmad, J; Idris, S; Khojah, A; Al-Qadi, N M; Refaat, B

    2016-01-01

    To measure the prevalence of cytomegalovirus (CMV) infection in ectopic pregnancy (EP) and its effect on the expression of inducible and endothelial nitric oxide synthases (iNOS, eNOS) by Fallopian tubes (FT) bearing an EP. This was a prospective case-control study. Blood and tubal samples were collected from 84 Eps and 51 controls (20 total abdominal hysterectomy (TAH) during the luteal phase and another 31 tubal ligations). CMV IgM and IgG antibodies were measured by ELISA, and an IVD CE PCR kit was used to detect CMV in the FTs. iNOS and eNOS were measured by immunohistochemistry and quantitative RT-PCR in FTs obtained from CMV-positive EP (n = 12), and the results were compared with those obtained from CMV-negative EP (n = 11) and TAH (n = 8). The frequencies of CMV IgM (51.2 % vs 17.6 %), IgG (77.4 % vs 52.9 %) or both antibodies (41.6 % vs 11.7 %) were significantly higher in EP compared with control. CMV was more common by PCR in FTs from EP (21.4 %) than controls (5.9 %). Twelve women from the PCR positive EP cases (66.6 %) were also simultaneously positive for both CMV IgM & IgG antibodies and had higher expression of eNOS and iNOS at the protein and gene levels compared with negative EP and TAH. Tubal infection with CMV may lead to EP by increasing the production of endothelial and inducible NOS by the FT epithelial cells. Further studies are required to illustrate the role of CMV in the pathogenesis of EP.

  2. Ectopic expression of anti-HIV-1 shRNAs protects CD8(+) T cells modified with CD4ζ CAR from HIV-1 infection and alleviates impairment of cell proliferation.

    PubMed

    Kamata, Masakazu; Kim, Patrick Y; Ng, Hwee L; Ringpis, Gene-Errol E; Kranz, Emiko; Chan, Joshua; O'Connor, Sean; Yang, Otto O; Chen, Irvin S Y

    2015-07-31

    Chimeric antigen receptors (CARs) are artificially engineered receptors that confer a desired specificity to immune effector T cells. As an HIV-1-specific CAR, CD4ζ CAR has been extensively tested in vitro as well as in clinical trials. T cells modified with this CAR mediated highly potent anti-HIV-1 activities in vitro and were well-tolerated in vivo, but exerted limited effects on viral load and reservoir size due to poor survival and/or functionality of the transduced cells in patients. We hypothesize that ectopic expression of CD4ζ on CD8(+) T cells renders them susceptible to HIV-1 infection, resulting in poor survival of those cells. To test this possibility, highly purified CD8(+) T cells were genetically modified with a CD4ζ-encoding lentiviral vector and infected with HIV-1. CD8(+) T cells were vulnerable to HIV-1 infection upon expression of CD4ζ as evidenced by elevated levels of p24(Gag) in cells and culture supernatants. Concurrently, the number of CD4ζ-modified CD8(+) T cells was reduced relative to control cells upon HIV-1 infection. To protect these cells from HIV-1 infection, we co-expressed two anti-HIV-1 shRNAs previously developed by our group together with CD4ζ. This combination vector was able to suppress HIV-1 infection without impairing HIV-1-dependent effector activities of CD4ζ. In addition, the number of CD4ζ-modified CD8(+) T cells maintained similar levels to that of the control even under HIV-1 infection. These results suggest that protecting CD4ζ-modified CD8(+) T cells from HIV-1 infection is required for prolonged HIV-1-specific immune surveillance.

  3. Ectopic expression of Jatropha curcas APETALA1 (JcAP1) caused early flowering in Arabidopsis, but not in Jatropha

    PubMed Central

    Tang, Mingyong; Tao, Yan-Bin

    2016-01-01

    Jatropha curcas is a promising feedstock for biofuel production because Jatropha oil is highly suitable for the production of biodiesel and bio-jet fuels. However, Jatropha exhibits a low seed yield as a result of unreliable and poor flowering. APETALA1 (AP1) is a floral meristem and organ identity gene in higher plants. The flower meristem identity genes of Jatropha have not yet been identified or characterized. To better understand the genetic control of flowering in Jatropha, an AP1 homolog (JcAP1) was isolated from Jatropha. An amino acid sequence analysis of JcAP1 revealed a high similarity to the AP1 proteins of other perennial plants. JcAP1 was expressed in inflorescence buds, flower buds, sepals and petals. The highest expression level was observed during the early developmental stage of the flower buds. The overexpression of JcAP1 using the cauliflower mosaic virus (CaMV) 35S promoter resulted in extremely early flowering and abnormal flowers in transgenic Arabidopsis plants. Several flowering genes downstream of AP1 were up-regulated in the JcAP1-overexpressing transgenic plant lines. Furthermore, JcAP1 overexpression rescued the phenotype caused by the Arabidopsis AP1 loss-of-function mutant ap1-11. Therefore, JcAP1 is an ortholog of AtAP1, which plays a similar role in the regulation of flowering in Arabidopsis. However, the overexpression of JcAP1 in Jatropha using the same promoter resulted in little variation in the flowering time and floral organs, indicating that JcAP1 may be insufficient to regulate flowering by itself in Jatropha. This study helps to elucidate the function of JcAP1 and contributes to the understanding of the molecular mechanisms of flower development in Jatropha. PMID:27168978

  4. Improvement of enzymatic saccharification yield in Arabidopsis thaliana by ectopic expression of the rice SUB1A-1 transcription factor

    PubMed Central

    Núñez-López, Lizeth; Aguirre-Cruz, Andrés

    2015-01-01

    Saccharification of polysaccharides releases monosaccharides that can be used by ethanol-producing microorganisms in biofuel production. To improve plant biomass as a raw material for saccharification, factors controlling the accumulation and structure of carbohydrates must be identified. Rice SUB1A-1 is a transcription factor that represses the turnover of starch and postpones energy-consuming growth processes under submergence stress. Arabidopsis was employed to test if heterologous expression of SUB1A-1 or SUB1C-1 (a related gene) can be used to improve saccharification. Cellulolytic and amylolytic enzymatic treatments confirmed that SUB1A-1 transgenics had better saccharification yield than wild-type (Col-0), mainly from accumulated starch. This improved saccharification yield was developmentally controlled; when compared to Col-0, young transgenic vegetative plants yielded 200–300% more glucose, adult vegetative plants yielded 40–90% more glucose and plants in reproductive stage had no difference in yield. We measured photosynthetic parameters, starch granule microstructure, and transcript abundance of genes involved in starch degradation (SEX4, GWD1), juvenile transition (SPL3-5) and meristematic identity (FUL, SOC1) but found no differences to Col-0, indicating that starch accumulation may be controlled by down-regulation of CONSTANS and FLOWERING LOCUS T by SUB1A-1 as previously reported. SUB1A-1 transgenics also offered less resistance to deformation than wild-type concomitant to up-regulation of AtEXP2 expansin and BGL2 glucan-1,3,-beta-glucosidase. We conclude that heterologous SUB1A-1 expression can improve saccharification yield and softness, two traits needed in bioethanol production. PMID:25780769

  5. Ectopic expression of Jatropha curcas APETALA1 (JcAP1) caused early flowering in Arabidopsis, but not in Jatropha.

    PubMed

    Tang, Mingyong; Tao, Yan-Bin; Xu, Zeng-Fu

    2016-01-01

    Jatropha curcas is a promising feedstock for biofuel production because Jatropha oil is highly suitable for the production of biodiesel and bio-jet fuels. However, Jatropha exhibits a low seed yield as a result of unreliable and poor flowering. APETALA1 (AP1) is a floral meristem and organ identity gene in higher plants. The flower meristem identity genes of Jatropha have not yet been identified or characterized. To better understand the genetic control of flowering in Jatropha, an AP1 homolog (JcAP1) was isolated from Jatropha. An amino acid sequence analysis of JcAP1 revealed a high similarity to the AP1 proteins of other perennial plants. JcAP1 was expressed in inflorescence buds, flower buds, sepals and petals. The highest expression level was observed during the early developmental stage of the flower buds. The overexpression of JcAP1 using the cauliflower mosaic virus (CaMV) 35S promoter resulted in extremely early flowering and abnormal flowers in transgenic Arabidopsis plants. Several flowering genes downstream of AP1 were up-regulated in the JcAP1-overexpressing transgenic plant lines. Furthermore, JcAP1 overexpression rescued the phenotype caused by the Arabidopsis AP1 loss-of-function mutant ap1-11. Therefore, JcAP1 is an ortholog of AtAP1, which plays a similar role in the regulation of flowering in Arabidopsis. However, the overexpression of JcAP1 in Jatropha using the same promoter resulted in little variation in the flowering time and floral organs, indicating that JcAP1 may be insufficient to regulate flowering by itself in Jatropha. This study helps to elucidate the function of JcAP1 and contributes to the understanding of the molecular mechanisms of flower development in Jatropha.

  6. ANA deficiency enhances bone morphogenetic protein-induced ectopic bone formation via transcriptional events.

    PubMed

    Miyai, Kentaro; Yoneda, Mitsuhiro; Hasegawa, Urara; Toita, Sayaka; Izu, Yayoi; Hemmi, Hiroaki; Hayata, Tadayoshi; Ezura, Yoichi; Mizutani, Shuki; Miyazono, Kohei; Akiyoshi, Kazunari; Yamamoto, Tadashi; Noda, Masaki

    2009-04-17

    Ectopic bone formation after joint replacement or brain injury in humans is a serious complication that causes immobility of joints and severe pain. However, mechanisms underlying such ectopic bone formation are not fully understood. Bone morphogenetic protein (BMPs) are defined as inducers of ectopic bone formation, and they are regulated by several types of inhibitors. ANA is an antiproliferative molecule that belongs to Tob/BTG family, but its activity in bone metabolism has not been known. Here, we examined the role of ANA on ectopic bone formation activity of BMP. In ANA-deficient and wild-type mice, BMP2 was implanted to induce ectopic bone formation in muscle. ANA deficiency increased mass of newly formed bone in vivo compared with wild-type based on 3D-muCT analyses. ANA mRNA was expressed in bone in vivo as well as in osteoblastic cells in vitro. Such ANA mRNA levels were increased by BMP2 treatment in MC3T3-E1 osteoblastic cells. Overexpression of ANA suppressed BMP-induced expression of luciferase reporter gene linked to BMP response elements in these cells. Conversely, ANA mRNA knockdown by small interference RNA enhanced the BMP-dependent BMP response element reporter expression. It also enhanced BMP-induced osteoblastic differentiation in muscle-derived C2C12 cells. Immunoprecipitation assay indicated that ANA interacts with Smad8. Thus, ANA is a suppressor of ectopic bone formation induced by BMP, and this inhibitory ANA activity is a part of the negative feedback regulation of BMP function.

  7. Leaf-Like Sepals Induced by Ectopic Expression of a SHORT VEGETATIVE PHASE (SVP)-Like MADS-Box Gene from the Basal Eudicot Epimedium sagittatum

    PubMed Central

    Li, Zhineng; Zeng, Shaohua; Li, Yanbang; Li, Mingyang; Souer, Erik

    2016-01-01

    Epimedium L. (Berberidaceae, Ranales), a perennial traditional Chinese medicinal herb, has become a new popular landscape plant for ground cover and pot culture in many countries based on its excellent ornamental characteristics and, distinctive and diverse floral morphology. However, little is known about the molecular genetics of flower development in Epimedium sagittatum. Here, we describe the characterization of EsSVP that encodes a protein sharing 68, 54, and 35% similarity with SVP, AGAMOUS-like 24 (AGL24) and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1) in Arabidopsis, respectively. Quantitative RT-PCR (qRT-PCR) indicated that EsSVP transcripts were principally found in petiole and leaf tissues, with little expression in roots and flowers and no in fruits. The highest EsSVP expression was observed in leaves. The flowering time of 35S::EsSVP in most Arabidopsis thaliana and in all petunia plants was not affected in both photoperiod conditions, but 35S::EsSVP 5# and 35S::EsSVP 1# Arabidopsis lines induced late and early flowering under long day (LD, 14 h light/10 h dark) and short day (SD, 10 h light/14 h dark) conditions, respectively. The 35S::EsSVP Arabidopsis produced extra secondary inflorescence or floral meristems in the axils of the leaf-like sepals with excrescent trichomes, and leaf-like sepals not able to enclose the inner three whorls completely. Moreover, almost all transgenic Arabidopsis plants showed persistent sepals around the completely matured fruits. Upon ectopic expression of 35S::EsSVP in Petunia W115, sepals were enlarged, sometimes to the size of leaves; corollas were greenish and did not fully open. These results suggest that EsSVP is involved in inflorescence meristem identity and flowering time regulation in some conditions. Although, the SVP homologs might have suffered functional diversification among diverse species between core and basal eudicots, the protein functions are conserved between Arabidopsis/Petunia and Epimedium

  8. Ectopically hTERT expressing adult human mesenchymal stem cells are less radiosensitive than their telomerase negative counterpart

    SciTech Connect

    Serakinci, Nedime . E-mail: nserakinci@health.sdu.dk; Christensen, Rikke; Graakjaer, Jesper; Cairney, Claire J.; Keith, W. Nicol; Alsner, Jan; Saretzki, Gabriele; Kolvraa, Steen

    2007-03-10

    During the past several years increasing evidence indicating that the proliferation capacity of mammalian cells is highly radiosensitive, regardless of the species and the tissue of origin of the cells, has accumulated. It has also been shown that normal bone marrow cells of mice have a similar radiosensitivity to other mammalian cells so far tested. In this study, we investigated the genetic effects of ionizing radiation (2.5-15 Gy) on normal human mesenchymal stem cells and their telomerised counterpart hMSC-telo1. We evaluated overall genomic integrity, DNA damage/repair by applying a fluorescence-detected alkaline DNA unwinding assay together with Western blot analyses for phosphorylated H2AX and Q-FISH was applied for investigation of telomeric damage. Our results indicate that hMSC and TERT-immortalized hMSCs can cope with relatively high doses of {gamma}-rays and that overall DNA repair is similar in the two cell lines. The telomeres were extensively destroyed after irradiation in both cell types suggesting that telomere caps are especially sensitive to radiation. The TERT-immortalized hMSCs showed higher stability at telomeric regions than primary hMSCs indicating that cells with long telomeres and high telomerase activity have the advantage of re-establishing the telomeric caps.

  9. Ectopic expression of gamma interferon in the eye protects transgenic mice from intraocular herpes simplex virus type 1 infections.

    PubMed Central

    Geiger, K; Howes, E L; Sarvetnick, N

    1994-01-01

    Transgenic (rho gamma) mice provide a model for studying the influence of gamma interferon (IFN-gamma) produced in the eye on ocular and cerebral viral infection. To establish this model, we injected BALB/c- and C57BL/6-derived transgenic and nontransgenic mice of different ages intravitreally with herpes simplex virus type 1 (HSV-1) strain F. Eye and brain tissues of these mice were assessed for pathological and immunocytochemical changes. HSV-1 infection induced severe retinitis of the injected eyes and infection of the brain in all mice. In transgenic mice inoculated with HSV-1, the left, nontreated eyes were protected from retinitis, whereas nontransgenic mice developed bilateral retinitis. Additional intravitreal injection of IFN-gamma with the virus protected the noninoculated eyes of nontransgenic mice. Three-week-old nontransgenic mice died from HSV-1 infection, whereas transgenic mice of the same age and nontransgenic mice intravitreally treated with IFN-gamma survived. Ocular IFN-gamma production increased the extent of inflammation in transgenic mice but did not have a significant influence on the growth of HSV-1 until day 3 after inoculation and did not influence the neuroinvasion of this virus. Thus, the effects of IFN-gamma were not caused by an early block of viral replication. Possible mechanisms of IFN-gamma action include activation of the immune response, alteration of the properties of the virus, and direct protection of neurons. Images PMID:8057437

  10. Ectopic Hidradenoma Papilliferum

    PubMed Central

    Rosmaninho, Aristóteles David Neiva; de Almeida, Maria Teresa Duarte Pinto; Costa, Vírgilio; Sanches, Maria Madalena Vasconcelos; Lopes, Carlos; Selores Gomes Meirinhos, Maria Manuela

    2010-01-01

    Hidradenoma papilliferum is a rare tumor that occurs almost exclusively in females on the anogenital area. Rare cases of ectopic (nongenital) hidradenoma papilliferum have been described. The lesions usually present as an asymptomatic slow-growing, red, firm, mobile, well-delimitated nodule that grows for a long time before resection. We describe a case of an 26-year-old man that presented with an enlarging nodule on his right eyelid. The histological findings revealed a hidradenoma papilliferum. So far, among the very few reports of ectopic hidradenoma papilliferum, only a very small number were localized to the eyelid. PMID:21197082

  11. Ectopic phytocystatin expression leads to enhanced drought stress tolerance in soybean (Glycine max) and Arabidopsis thaliana through effects on strigolactone pathways and can also result in improved seed traits.

    PubMed

    Quain, Marian D; Makgopa, Matome E; Márquez-García, Belén; Comadira, Gloria; Fernandez-Garcia, Nieves; Olmos, Enrique; Schnaubelt, Daniel; Kunert, Karl J; Foyer, Christine H

    2014-09-01

    Ectopic cystatin expression has long been used in plant pest management, but the cysteine protease, targets of these inhibitors, might also have important functions in the control of plant lifespan and stress tolerance that remain poorly characterized. We therefore characterized the effects of expression of the rice cystatin, oryzacystatin-I (OCI), on the growth, development and stress tolerance of crop (soybean) and model (Arabidopsis thaliana) plants. Ectopic OCI expression in soybean enhanced shoot branching and leaf chlorophyll accumulation at later stages of vegetative development and enhanced seed protein contents and decreased the abundance of mRNAs encoding strigolactone synthesis enzymes. The OCI-expressing A. thaliana showed a slow-growth phenotype, with increased leaf numbers and enhanced shoot branching at flowering. The OCI-dependent inhibition of cysteine proteases enhanced drought tolerance in soybean and A. thaliana, photosynthetic CO2 assimilation being much less sensitive to drought-induced inhibition in the OCI-expressing soybean lines. Ectopic OCI expression or treatment with the cysteine protease inhibitor E64 increased lateral root densities in A. thaliana. E64 treatment also increased lateral root densities in the max2-1 mutants that are defective in strigolactone signalling, but not in the max3-9 mutants that are defective in strigolactone synthesis. Taken together, these data provide evidence that OCI-inhibited cysteine proteases participate in the control of growth and stress tolerance through effects on strigolactones. We conclude that cysteine proteases are important targets for manipulation of plant growth, development and stress tolerance, and also seed quality traits.

  12. Mutations in TCF8 Cause Posterior Polymorphous Corneal Dystrophy and Ectopic Expression of COL4A3 by Corneal Endothelial Cells

    PubMed Central

    Krafchak, Charles M. ; Pawar, Hemant ; Moroi, Sayoko E. ; Sugar, Alan ; Lichter, Paul R. ; Mackey, David A. ; Mian, Shahzad ; Nairus, Theresa ; Elner, Victor ; Schteingart, Miriam T. ; Downs, Catherine A. ; Kijek, Theresa Guckian ; Johnson, Jenae M. ; Trager, Edward H. ; Rozsa, Frank W. ; Ali Mandal, Md Nawajes ; Epstein, Michael P. ; Vollrath, Douglas ; Ayyagari, Radha ; Boehnke, Michael ; Richards, Julia E. 

    2005-01-01

    Posterior polymorphous corneal dystrophy (PPCD, also known as PPMD) is a rare disease involving metaplasia and overgrowth of corneal endothelial cells. In patients with PPCD, these cells manifest in an epithelial morphology and gene expression pattern, produce an aberrant basement membrane, and, sometimes, spread over the iris and nearby structures in a way that increases the risk for glaucoma. We previously mapped PPCD to a region (PPCD3) on chromosome 10 containing the gene that encodes the two-handed zinc-finger homeodomain transcription factor TCF8. Here, we report a heterozygous frameshift mutation in TCF8 that segregates with PPCD in the family used to map PPCD3 and four different heterozygous nonsense and frameshift mutations in TCF8 in four other PPCD probands. Family reports of inguinal hernia, hydrocele, and possible bone anomalies in affected individuals suggest that individuals with TCF8 mutations should be examined for nonocular anomalies. We detect transcripts of all three identified PPCD genes (VSX1, COL8A2, and TCF8) in the cornea. We show presence of a complex (core plus secondary) binding site for TCF8 in the promoter of Alport syndrome gene COL4A3, which encodes collagen type IV α3, and we present immunohistochemical evidence of ectopic expression of COL4A3 in corneal endothelium of the proband of the original PPCD3 family. Identification of TCF8 as the PPCD3 gene provides a valuable tool for the study of critical gene regulation events in PPCD pathology and suggests a possible role for TCF8 mutations in altered structure and function of cells lining body cavities other than the anterior chamber of the eye. Thus, this study has identified TCF8 as the gene responsible for approximately half of the cases of PPCD, has implicated TCF8 mutations in developmental abnormalities outside the eye, and has presented the TCF8 regulatory target, COL4A3, as a key, shared molecular component of two different diseases, PPCD and Alport syndrome. PMID:16252232

  13. Ectopic Lignification in the Flax lignified bast fiber1 Mutant Stem Is Associated with Tissue-Specific Modifications in Gene Expression and Cell Wall Composition[C][W

    PubMed Central

    Chantreau, Maxime; Portelette, Antoine; Dauwe, Rebecca; Kiyoto, Shingo; Crônier, David; Morreel, Kris; Arribat, Sandrine; Neutelings, Godfrey; Chabi, Malika; Boerjan, Wout; Yoshinaga, Arata; Mesnard, François; Grec, Sebastien; Chabbert, Brigitte; Hawkins, Simon

    2014-01-01

    Histochemical screening of a flax ethyl methanesulfonate population led to the identification of 93 independent M2 mutant families showing ectopic lignification in the secondary cell wall of stem bast fibers. We named this core collection the Linum usitatissimum (flax) lbf mutants for lignified bast fibers and believe that this population represents a novel biological resource for investigating how bast fiber plants regulate lignin biosynthesis. As a proof of concept, we characterized the lbf1 mutant and showed that the lignin content increased by 350% in outer stem tissues containing bast fibers but was unchanged in inner stem tissues containing xylem. Chemical and NMR analyses indicated that bast fiber ectopic lignin was highly condensed and rich in G-units. Liquid chromatography-mass spectrometry profiling showed large modifications in the oligolignol pool of lbf1 inner- and outer-stem tissues that could be related to ectopic lignification. Immunological and chemical analyses revealed that lbf1 mutants also showed changes to other cell wall polymers. Whole-genome transcriptomics suggested that ectopic lignification of flax bast fibers could be caused by increased transcript accumulation of (1) the cinnamoyl-CoA reductase, cinnamyl alcohol dehydrogenase, and caffeic acid O-methyltransferase monolignol biosynthesis genes, (2) several lignin-associated peroxidase genes, and (3) genes coding for respiratory burst oxidase homolog NADPH-oxidases necessary to increase H2O2 supply. PMID:25381351

  14. Distinct function of COAR and B3 domains of maize VP1 in induction of ectopic gene expression and plant developmental phenotypes in Arabidopsis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Maize VP1 enhancement of ABA sensitivity in roots is B3 independent. ABA and VP1 mediated suppression of auxin induced lateral root development is B3 independent. Arabidopsis transgenic system to delineate B3 dependent and COAR domain dependent regulatory functions of VP1. Analyses of ectopic ABA re...

  15. Ectopic lignification in the flax lignified bast fiber1 mutant stem is associated with tissue-specific modifications in gene expression and cell wall composition.

    PubMed

    Chantreau, Maxime; Portelette, Antoine; Dauwe, Rebecca; Kiyoto, Shingo; Crônier, David; Morreel, Kris; Arribat, Sandrine; Neutelings, Godfrey; Chabi, Malika; Boerjan, Wout; Yoshinaga, Arata; Mesnard, François; Grec, Sebastien; Chabbert, Brigitte; Hawkins, Simon

    2014-11-01

    Histochemical screening of a flax ethyl methanesulfonate population led to the identification of 93 independent M2 mutant families showing ectopic lignification in the secondary cell wall of stem bast fibers. We named this core collection the Linum usitatissimum (flax) lbf mutants for lignified bast fibers and believe that this population represents a novel biological resource for investigating how bast fiber plants regulate lignin biosynthesis. As a proof of concept, we characterized the lbf1 mutant and showed that the lignin content increased by 350% in outer stem tissues containing bast fibers but was unchanged in inner stem tissues containing xylem. Chemical and NMR analyses indicated that bast fiber ectopic lignin was highly condensed and rich in G-units. Liquid chromatography-mass spectrometry profiling showed large modifications in the oligolignol pool of lbf1 inner- and outer-stem tissues that could be related to ectopic lignification. Immunological and chemical analyses revealed that lbf1 mutants also showed changes to other cell wall polymers. Whole-genome transcriptomics suggested that ectopic lignification of flax bast fibers could be caused by increased transcript accumulation of (1) the cinnamoyl-CoA reductase, cinnamyl alcohol dehydrogenase, and caffeic acid O-methyltransferase monolignol biosynthesis genes, (2) several lignin-associated peroxidase genes, and (3) genes coding for respiratory burst oxidase homolog NADPH-oxidases necessary to increase H2O2 supply.

  16. Primary ectopic frontotemporal craniopharyngioma

    PubMed Central

    Ortega-Porcayo, Luis Alberto; Ponce-Gómez, Juan Antonio; Martínez-Moreno, Mauricio; Portocarrero-Ortíz, Lesly; Tena-Suck, Martha Lilia; Gómez-Amador, Juan Luis

    2015-01-01

    Introduction Primary ectopic craniopharyngiomas have only rarely been reported. Craniopharyngiomas involve usually the sellar and suprasellar region, but can be originated from cell remnants of the obliterated craniopharyngeal duct or metaplastic change of andenohypophyseal cells. We present the first case of a primary ectopic frontotemporal craniopharyngioma. Presentation of case A 35-year old woman presented with a one-year history of headache and diplopia. MRI showed a large frontotemporal cystic lesion. Tumor resection was performed with a keyhole endoscopic frontal lateral approach. The pathological features showed an adamantinomatous craniopharyngioma with a cholesterol granuloma reaction. Discussion There have been reported different localizations for primary ectopic craniopharyngioma. Our case presented a lobulated frontotemporal cystic mass formed by a dense eosinophilic proteinaceous material dystrophic calcifications and cholesterol crystals, with epithelial remnants. No tumor regrowth was observed in the magnetic resonance image 27 months postoperatively. Conclusion Primary ectopic craniopharyngioma is a rare entity with a pathogenesis that remains uncertain. This is an unusual anatomic location associated with unique clinical findings. PMID:25725331

  17. Ectopic expression of a maize calreticulin mitigates calcium deficiency-like disorders in "sCAX1"-expressing tobacco and tomato

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Deregulated expression of an Arabidopsis H(+)/Ca(2+) antiporter (sCAX1) in agricultural crops increases total calcium (Ca(2+)) but may result in yield losses due to Ca(2+) deficiency-like symptoms. Here we demonstrate that co-expression of a maize calreticulin (CRT, a Ca(2+) binding protein located ...

  18. Ectopic expression of CsCTR1, a cucumber CTR-like gene, attenuates constitutive ethylene signaling in an Arabidopsis ctr1-1 mutant and expression pattern analysis of CsCTR1 in cucumber (Cucumis sativus).

    PubMed

    Bie, Beibei; Sun, Jin; Pan, Junsong; He, Huanle; Cai, Run

    2014-09-15

    The gaseous plant hormone ethylene regulates many aspects of plant growth, development and responses to the environment. Constitutive triple response 1 (CTR1) is a central regulator involved in the ethylene signal transduction pathway. To obtain a better understanding of this particular pathway in cucumber, the cDNA-encoding CTR1 (designated CsCTR1) was isolated from cucumber. A sequence alignment and phylogenetic analyses revealed that CsCTR1 has a high degree of homology with other plant CTR1 proteins. The ectopic expression of CsCTR1 in the Arabidopsis ctr1-1 mutant attenuates constitutive ethylene signaling of this mutant, suggesting that CsCTR1 indeed performs its function as negative regulator of the ethylene signaling pathway. CsCTR1 is constitutively expressed in all of the examined cucumber organs, including roots, stems, leaves, shoot apices, mature male and female flowers, as well as young fruits. CsCTR1 expression gradually declined during male flower development and increased during female flower development. Additionally, our results indicate that CsCTR1 can be induced in the roots, leaves and shoot apices by external ethylene. In conclusion, this study provides a basis for further studies on the role of CTR1 in the biological processes of cucumber and on the molecular mechanism of the cucumber ethylene signaling pathway.

  19. Ectopic Expression of CsCTR1, a Cucumber CTR-Like Gene, Attenuates Constitutive Ethylene Signaling in an Arabidopsis ctr1-1 Mutant and Expression Pattern Analysis of CsCTR1 in Cucumber (Cucumis sativus)

    PubMed Central

    Bie, Beibei; Sun, Jin; Pan, Junsong; He, Huanle; Cai, Run

    2014-01-01

    The gaseous plant hormone ethylene regulates many aspects of plant growth, development and responses to the environment. Constitutive triple response 1 (CTR1) is a central regulator involved in the ethylene signal transduction pathway. To obtain a better understanding of this particular pathway in cucumber, the cDNA-encoding CTR1 (designated CsCTR1) was isolated from cucumber. A sequence alignment and phylogenetic analyses revealed that CsCTR1 has a high degree of homology with other plant CTR1 proteins. The ectopic expression of CsCTR1 in the Arabidopsis ctr1-1 mutant attenuates constitutive ethylene signaling of this mutant, suggesting that CsCTR1 indeed performs its function as negative regulator of the ethylene signaling pathway. CsCTR1 is constitutively expressed in all of the examined cucumber organs, including roots, stems, leaves, shoot apices, mature male and female flowers, as well as young fruits. CsCTR1 expression gradually declined during male flower development and increased during female flower development. Additionally, our results indicate that CsCTR1 can be induced in the roots, leaves and shoot apices by external ethylene. In conclusion, this study provides a basis for further studies on the role of CTR1 in the biological processes of cucumber and on the molecular mechanism of the cucumber ethylene signaling pathway. PMID:25226540

  20. Ovarian tumor-derived ectopic hyperprolactinemia.

    PubMed

    Elms, Autumn F; Carlan, S J; Rich, Amy E; Cerezo, Lizardo

    2012-12-01

    To describe extreme hyperprolactinemia originating from a pituitary adenoma in the wall of an ovarian dermoid. This is a description of an unusual case and a review of ectopic prolactin production. Ectopic production of prolactin is a rare condition that has been reported in isolated organ system pathology including ovaries. An ovarian dermoid is a benign neoplasm that has the potential for active unregulated endocrine function. Hyperprolactinemia can result from functioning lactotrophs found in ovarian dermoids and can lead to clinical sequelae. Definitive treatment of the condition requires surgical removal of the functioning endocrine tissue. Extreme hyperprolactinemia caused by a pituitary tumor located within a dermoid has not been reported before. We present a case of profound hyperprolactinemia originating from a pituitary adenoma found in the wall of an ovarian dermoid and give a broad overview of the condition and literature. Ectopic prolactin production should always be considered in symptomatic patients found to have elevated serum levels and no findings on brain imaging.

  1. Addition of an N-terminal epitope tag significantly increases the activity of plant fatty acid desaturases expressed in yeast cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Saccharomyces cerevisiae shows great potential for development of bioreactor systems geared towards the production of high-value lipids such as polyunsaturated omega-3 fatty acids, the yields of which are largely dependent on the activity of ectopically-expressed enzymes. Here we show that the addit...

  2. The pan-neural bHLH proteins DEADPAN and ASENSE regulate mitotic activity and cdk inhibitor dacapo expression in the Drosophila larval optic lobes.

    PubMed

    Wallace, K; Liu, T H; Vaessin, H

    2000-01-01

    Developmental regulators and cell cycle regulators have to interface in order to ensure appropriate cell proliferation during organogenesis. Our analysis of the roles of the pan-neural genes deadpan and asense defines critical roles for these genes in regulation of mitotic activities in the larval optic lobes. Loss of deadpan results in reduced cell proliferation, while ectopic deadpan expression causes over-proliferation. In contrast, loss of asense results in increased proliferation, while ectopic asense expression causes reduced proliferation. Consistent with these observations endogenous Deadpan is expressed in mitotic areas of the optic lobes, and endogenous Asense is expressed in cells that will become quiescent. Altered Deadpan or Asense expression results in altered expression of the cyclin dependent kinase inhibitor gene dacapo. Thus, regulation of mitotic activity during optic lobe development may, at least in part, involve deadpan and asense mediated regulation of the cyclin dependent kinase inhibitor gene dacapo. genesis 26:77-85, 2000.

  3. Intra-cholecystic ectopic liver.

    PubMed

    Natori, T; Hawkin, S; Aizawa, M; Asai, T; Kameda, Y; Ikuyohashi, K

    1986-08-01

    A 56-year-old Japanese woman who had an ectopic liver attached to the inner surface of the gallbladder is reported. The ectopic nodule measuring 1.2 X 0.5 X 0.8 cm was found incidentally when her gallbladder was resected because of cholelithiasis. Ectopic liver tissue in reported cases war located on the serosal surface, or in the wall of the gallbladder. This report described, to our knowledge, the first case of an ectopic liver nodule attached to the mucosal surface of the gallbladder with a thin pedicle.

  4. Ectopic expression of ubiquitin-conjugating enzyme gene from wild rice, OgUBC1, confers resistance against UV-B radiation and Botrytis infection in Arabidopsis thaliana

    SciTech Connect

    Jeon, En Hee; Pak, Jung Hun; Kim, Mi Jin; Kim, Hye Jeong; Shin, Sang Hyun; Lee, Jai Heon; Kim, Doh Hoon; Oh, Ju Sung; Oh, Boung-Jun; Jung, Ho Won; Chung, Young Soo

    2012-10-19

    Highlights: Black-Right-Pointing-Pointer We isolated a novel E2 ubiquitin-conjugating enzyme from leaves of wild rice plants. Black-Right-Pointing-Pointer The OgUBC1 was highly expressed in leaves treated with SA and UV-B radiation. Black-Right-Pointing-Pointer The recombinant OgUBC1 has an enzymatic activity of E2 in vitro. Black-Right-Pointing-Pointer The OgUBC1 could protect disruption of plant cells by UV-B radiation. Black-Right-Pointing-Pointer OgUBC1 confers disease resistance and UV-B tolerance in transgenic Arabidopsis plants. -- Abstract: A previously unidentified gene encoding ubiquitin-conjugating enzyme was isolated from leaves of wild rice plant treated with wounding and microbe-associated molecular patterns. The OgUBC1 gene was composed of 148 amino acids and contained a typical active site and 21 ubiquitin thioester intermediate interaction residues and 4 E3 interaction residues. Both exogenous application of salicylic acid and UV-B irradiation triggered expression of OgUBC1 in leaves of wild rice. Recombinant OgUBC1 proteins bound to ubiquitins in vitro, proposing that the protein might act as E2 enzyme in planta. Heterologous expression of the OgUBC1 in Arabidopsis thaliana protected plants from cellular damage caused by an excess of UV-B radiation. A stable expression of chalcone synthase gene was detected in leaves of OgUBC1-expressing Arabidopsis, resulting in producing higher amounts of anthocyanin than those in wild-type Col-0 plants. Additionally, both pathogenesis-related gene1 and 5 were transcribed in the transgenic Arabidopsis in the absence of pathogen infection. The OgUBC1-expressing plants were resistant to the infection of Botrytis cinerea. Taken together, we suggested that the OgUBC1 is involved in ubiquitination process important for cellular response against biotic and abiotic stresses in plants.

  5. Functional Conservation and Divergence of Four Ginger AP1/AGL9 MADS–Box Genes Revealed by Analysis of Their Expression and Protein–Protein Interaction, and Ectopic Expression of AhFUL Gene in Arabidopsis

    PubMed Central

    Song, Juanjuan; Sun, Wei; Xia, Kuaifei; Liao, Jingping; Zhang, Mingyong

    2014-01-01

    Alpinia genus are known generally as ginger–lilies for showy flowers in the ginger family, Zingiberaceae, and their floral morphology diverges from typical monocotyledon flowers. However, little is known about the functions of ginger MADS–box genes in floral identity. In this study, four AP1/AGL9 MADS–box genes were cloned from Alpinia hainanensis, and protein–protein interactions (PPIs) and roles of the four genes in floral homeotic conversion and in floral evolution are surveyed for the first time. AhFUL is clustered to the AP1lineage, AhSEP4 and AhSEP3b to the SEP lineage, and AhAGL6–like to the AGL6 lineage. The four genes showed conserved and divergent expression patterns, and their encoded proteins were localized in the nucleus. Seven combinations of PPI (AhFUL–AhSEP4, AhFUL–AhAGL6–like, AhFUL–AhSEP3b, AhSEP4–AhAGL6–like, AhSEP4–AhSEP3b, AhAGL6–like–AhSEP3b, and AhSEP3b–AhSEP3b) were detected, and the PPI patterns in the AP1/AGL9 lineage revealed that five of the 10 possible combinations are conserved and three are variable, while conclusions cannot yet be made regarding the other two. Ectopic expression of AhFUL in Arabidopsis thaliana led to early flowering and floral organ homeotic conversion to sepal–like or leaf–like. Therefore, we conclude that the four A. hainanensis AP1/AGL9 genes show functional conservation and divergence in the floral identity from other MADS–box genes. PMID:25461565

  6. Cervical ectopic pregnancy.

    PubMed

    Samal, Sunil Kumar; Rathod, Setu

    2015-01-01

    Cervical pregnancy is a rare type of ectopic pregnancy and it represents <1% of all ectopic pregnancies. Early diagnosis and medical management with systemic or local administration of methotrexate is the treatment of choice. If the pregnancy is disturbed, it may lead to massive hemorrhage, which may require hysterectomy to save the patient. We report three cases of cervical pregnancy managed successfully with different approaches of management. Our first case, 28 years old G3P2L2 with previous two lower segment cesarean sections, presented with bleeding per vaginum following 6 weeks of amenorrhea. Clinical examination followed by transvaginal ultrasound confirmed the diagnosis of cervical pregnancy. Total abdominal hysterectomy was done in view of intractable bleeding to save the patient. The second case, a 26-year-old second gravida with previous normal vaginal delivery presented with pain abdomen and single episode of spotting per vaginum following 7 weeks of amenorrhea. Transvaginal ultrasound revealed empty endometrial cavity, closed internal os with gestational sac containing live fetus of 7 weeks gestational age in cervical canal and she was treated with intra-amniotic potassium chloride followed by systemic methotrexate. Follow up with serum beta human chorionic gonadotropin level revealed successful outcome. Our third case, a 27-year-old primigravida with history of infertility treatment admitted with complaints of bleeding per vaginum for 1 day following 8 weeks amenorrhea. She was diagnosed as cervical pregnancy by clinical examination, confirmed by transvaginal ultrasonography and subsequently managed by dilation and curettage with intracervical Foleys' ballon tamponade.

  7. Ectopic TBX1 suppresses thymic epithelial cell differentiation and proliferation during thymus organogenesis.

    PubMed

    Reeh, Kaitlin A G; Cardenas, Kim T; Bain, Virginia E; Liu, Zhijie; Laurent, Micheline; Manley, Nancy R; Richie, Ellen R

    2014-08-01

    The thymus and parathyroid glands arise from a shared endodermal primordium in the third pharyngeal pouch (3rd pp). Thymus fate is specified in the ventral 3rd pp between E9.5 and E11, whereas parathyroid fate is specified in the dorsal domain. The molecular mechanisms that specify fate and regulate thymus and parathyroid development are not fully delineated. Previous reports suggested that Tbx1 is required for thymus organogenesis because loss of Tbx1 in individuals with DiGeorge syndrome and in experimental Tbx1 deletion mutants is associated with thymus aplasia or hypoplasia. However, the thymus phenotype is likely to be secondary to defects in pharyngeal pouch formation. Furthermore, the absence of Tbx1 expression in the thymus-fated domain of the wild-type 3rd pp suggested that Tbx1 is instead a negative regulator of thymus organogenesis. To test this hypothesis, we generated a novel mouse strain in which expression of a conditional Tbx1 allele was ectopically activated in the thymus-fated domain of the 3rd pp. Ectopic Tbx1 expression severely repressed expression of Foxn1, a transcription factor that marks the thymus-fated domain and is required for differentiation and proliferation of thymic epithelial cell (TEC) progenitors. By contrast, ectopic Tbx1 did not alter the expression pattern of Gcm2, a transcription factor restricted to the parathyroid-fated domain and required for parathyroid development. Ectopic Tbx1 expression impaired TEC proliferation and arrested TEC differentiation at an early progenitor stage. The results support the hypothesis that Tbx1 negatively regulates TEC growth and differentiation, and that extinction of Tbx1 expression in 3rd pp endoderm is a prerequisite for thymus organogenesis.

  8. Comprehensive Analysis of Rice Laccase Gene (OsLAC) Family and Ectopic Expression of OsLAC10 Enhances Tolerance to Copper Stress in Arabidopsis

    PubMed Central

    Liu, Qingquan; Luo, Le; Wang, Xiaoxiao; Shen, Zhenguo; Zheng, Luqing

    2017-01-01

    Laccases are encoded by a multigene family and widely distributed in plant genomes where they play roles oxidizing monolignols to produce higher-order lignin involved in plant development and stress responses. We identified 30 laccase genes (OsLACs) from rice, which can be divided into five subfamilies, mostly expressed during early development of the endosperm, growing roots, and stems. OsLACs can be induced by hormones, salt, drought, and heavy metals stresses. The expression level of OsLAC10 increased 1200-fold after treatment with 20 μM Cu for 12 h. The laccase activities of OsLAC10 were confirmed in an Escherichia coli expression system. Lignin accumulation increased in the roots of Arabidopsis over-expressing OsLAC10 (OsLAC10-OX) compared to wild-type controls. After growth on 1/2 Murashige and Skoog (MS) medium containing toxic levels of Cu for seven days, roots of the OsLAC10-OX lines were significantly longer than those of the wild type. Compared to control plants, the Cu concentration decreased significantly in roots of the OsLAC10-OX line under hydroponic conditions. These results provided insights into the evolutionary expansion and functional divergence of OsLAC family. In addition, OsLAC10 is likely involved in lignin biosynthesis, and reduces the uptake of Cu into roots required for Arabidopsis to develop tolerance to Cu. PMID:28146098

  9. Ectopic expression of the mycorrhiza-specific chitinase gene Mtchit 3-3 in Medicago truncatula root-organ cultures stimulates spore germination of glomalean fungi.

    PubMed

    Elfstrand, Malin; Feddermann, Nadja; Ineichen, Kurt; Nagaraj, Vinay Jantakahalli; Wiemken, Andres; Boller, Thomas; Salzer, Peter

    2005-08-01

    Expression of Mtchit 3-3, a class III chitinase gene, is specifically induced by arbuscular mycorrhizal (AM) fungi in roots of the model legume Medicago truncatula and its transcripts accumulate in cells containing arbuscules. Agrobacterium rhizogenes-transformed roots and root-organ cultures of M. truncatula were used to study effects of Mtchit 3-3 on AM fungi. * This work provides evidence for enzymatic activity of the Mtchit 3-3 gene product and shows with promoter:gus fusions that a 2 kb fragment located 5' upstream from the translational start codon of Mtchit 3-3 is sufficient to confer arbuscule-dependent gene expression. By fusing the Mtchit 3-3 coding region to the CaMV 35S promoter the expression pattern was disrupted. Surprisingly, disruption stimulated spore germination of Glomus intraradices and Glomus constrictum, and in the case of G. intraradices resulted in a higher probability of root colonization and spore formation. However, no effect on the abundance of arbuscules within colonized roots became apparent. These observations demonstrate that disruption of the tight arbuscule-dependent expression pattern of Mtchit 3-3 has effects on the early interaction between roots and AM fungi.

  10. Simulation of Ectopic Pacemakers in the Heart: Multiple Ectopic Beats Generated by Reentry inside Fibrotic Regions

    PubMed Central

    Gouvêa de Barros, Bruno; Weber dos Santos, Rodrigo; Lobosco, Marcelo; Alonso, Sergio

    2015-01-01

    The inclusion of nonconducting media, mimicking cardiac fibrosis, in two models of cardiac tissue produces the formation of ectopic beats. The fraction of nonconducting media in comparison with the fraction of healthy myocytes and the topological distribution of cells determines the probability of ectopic beat generation. First, a detailed subcellular microscopic model that accounts for the microstructure of the cardiac tissue is constructed and employed for the numerical simulation of action potential propagation. Next, an equivalent discrete model is implemented, which permits a faster integration of the equations. This discrete model is a simplified version of the microscopic model that maintains the distribution of connections between cells. Both models produce similar results when describing action potential propagation in homogeneous tissue; however, they slightly differ in the generation of ectopic beats in heterogeneous tissue. Nevertheless, both models present the generation of reentry inside fibrotic tissues. This kind of reentry restricted to microfibrosis regions can result in the formation of ectopic pacemakers, that is, regions that will generate a series of ectopic stimulus at a fast pacing rate. In turn, such activity has been related to trigger fibrillation in the atria and in the ventricles in clinical and animal studies. PMID:26583127

  11. Seed size plasticity in response to embryonic lethality conferred by ectopic CYCD activation is dependent on plant architecture

    PubMed Central

    Sornay, E.; Dewitte, W.; Murray, J. A. H.

    2016-01-01

    ABSTRACT The size of seeds is the result of cell proliferation and growth in the three seed compartments: the embryo, endosperm and integuments. Targeting expression of the D-type cyclin CYCD7;1 to the central cell and early endosperm (FWA:CYCD7;1) triggered nuclear divisions and partial ovule abortion, reducing seed number in each silique and leading to increased seed size. A similar effect on seed size was observed with other segregating embryo lethal mutations, suggesting caution is needed in interpreting apparent seed size phenotypes. Here, we show that the positive effect of FWA:CYCD7;1 on Arabidopsis seed size is modulated by the architecture of the mother plant. Larger seeds were produced in FWA:CYCD7;1 lines with unmodified inflorescences, and also upon removal of side branches and axillary stems. This phenotype was absent from inflorescences with increased axillary floral stems produced by pruning of the main stem. Given this apparent confounding influence of resource allocation on transgenes effect, we conclude that plant architecture is a further important factor to consider in appraising seed phenotypes. PMID:27286190

  12. Ectopic vesicular neurotransmitter release along sensory axons mediates neurovascular coupling via glial calcium signaling.

    PubMed

    Thyssen, Anne; Hirnet, Daniela; Wolburg, Hartwig; Schmalzing, Günther; Deitmer, Joachim W; Lohr, Christian

    2010-08-24

    Neurotransmitter release generally is considered to occur at active zones of synapses, and ectopic release of neurotransmitters has been demonstrated in a few instances. However, the mechanism of ectopic neurotransmitter release is poorly understood. We took advantage of the intimate morphological and functional proximity of olfactory receptor axons and specialized glial cells, olfactory ensheathing cells (OECs), to study ectopic neurotransmitter release. Axonal stimulation evoked purinergic and glutamatergic Ca(2+) responses in OECs, indicating ATP and glutamate release. In axons expressing synapto-pHluorin, stimulation evoked an increase in synapto-pHluorin fluorescence, indicative of vesicle fusion. Transmitter release was dependent on Ca(2+) and could be inhibited by bafilomycin A1 and botulinum toxin A. Ca(2+) transients in OECs evoked by ATP, axonal stimulation, and laser photolysis of NP-EGTA resulted in constriction of adjacent blood vessels. Our results indicate that ATP and glutamate are released ectopically by vesicles along axons and mediate neurovascular coupling via glial Ca(2+) signaling.

  13. Cervical ectopic pregnancy

    PubMed Central

    Samal, Sunil Kumar; Rathod, Setu

    2015-01-01

    Cervical pregnancy is a rare type of ectopic pregnancy and it represents <1% of all ectopic pregnancies. Early diagnosis and medical management with systemic or local administration of methotrexate is the treatment of choice. If the pregnancy is disturbed, it may lead to massive hemorrhage, which may require hysterectomy to save the patient. We report three cases of cervical pregnancy managed successfully with different approaches of management. Our first case, 28 years old G3P2L2 with previous two lower segment cesarean sections, presented with bleeding per vaginum following 6 weeks of amenorrhea. Clinical examination followed by transvaginal ultrasound confirmed the diagnosis of cervical pregnancy. Total abdominal hysterectomy was done in view of intractable bleeding to save the patient. The second case, a 26-year-old second gravida with previous normal vaginal delivery presented with pain abdomen and single episode of spotting per vaginum following 7 weeks of amenorrhea. Transvaginal ultrasound revealed empty endometrial cavity, closed internal os with gestational sac containing live fetus of 7 weeks gestational age in cervical canal and she was treated with intra-amniotic potassium chloride followed by systemic methotrexate. Follow up with serum beta human chorionic gonadotropin level revealed successful outcome. Our third case, a 27-year-old primigravida with history of infertility treatment admitted with complaints of bleeding per vaginum for 1 day following 8 weeks amenorrhea. She was diagnosed as cervical pregnancy by clinical examination, confirmed by transvaginal ultrasonography and subsequently managed by dilation and curettage with intracervical Foleys’ ballon tamponade. PMID:25810679

  14. Melanoma Expressed-CD70 Is Regulated by RhoA and MAPK Pathways without Affecting Vemurafenib Treatment Activity

    PubMed Central

    Sarrabayrouse, Guillaume; Gallardo, Franck; Gence, Rémi; Tilkin-Mariamé, Anne-Françoise

    2016-01-01

    CD70 is a costimulatory molecule member of the Tumor Necrosis Factor family that is expressed on activated immune cells. Its ectopic expression has been described in several types of cancer cells including lymphomas, renal cell carcinomas and glioblastomas. We have recently described its expression in a part of tumor cells from the vast majority of melanoma biopsies and human melanoma cell lines, and found that CD70 expression decreased over time as the disease progressed. Here, we show that RhoA, BRAF and Mitogen Activating Protein Kinase pathways are involved in the positive transcriptional regulation of CD70 expression in melanomas. Interestingly, the clinical inhibitor of the common BRAF V600E/D variants, Vemurafenib (PLX-4032), which is currently used to treat melanoma patients with BRAF V600E/D-mutated metastatic melanomas, decreased CD70 expression in human CD70+ melanoma cell lines. This decrease was seen in melanoma cells both with and without the BRAFV600E/D mutation, although was less efficient in those lacking the mutation. But interestingly, by silencing CD70 in CD70+ melanoma cell lines we show that PLX-4032-induced melanoma cell killing and its inhibitory effect on MAPK pathway activation are unaffected by CD70 expression. Consequently, our work demonstrates that CD70 ectopic expression in melanomas is not a valuable biomarker to predict tumor cells sensitivity to BRAF V600 inhibitors. PMID:26828592

  15. Melanoma Expressed-CD70 Is Regulated by RhoA and MAPK Pathways without Affecting Vemurafenib Treatment Activity.

    PubMed

    Pich, Christine; Teiti, Iotefa; Sarrabayrouse, Guillaume; Gallardo, Franck; Gence, Rémi; Tilkin-Mariamé, Anne-Françoise

    2016-01-01

    CD70 is a costimulatory molecule member of the Tumor Necrosis Factor family that is expressed on activated immune cells. Its ectopic expression has been described in several types of cancer cells including lymphomas, renal cell carcinomas and glioblastomas. We have recently described its expression in a part of tumor cells from the vast majority of melanoma biopsies and human melanoma cell lines, and found that CD70 expression decreased over time as the disease progressed. Here, we show that RhoA, BRAF and Mitogen Activating Protein Kinase pathways are involved in the positive transcriptional regulation of CD70 expression in melanomas. Interestingly, the clinical inhibitor of the common BRAF V600E/D variants, Vemurafenib (PLX-4032), which is currently used to treat melanoma patients with BRAF V600E/D-mutated metastatic melanomas, decreased CD70 expression in human CD70+ melanoma cell lines. This decrease was seen in melanoma cells both with and without the BRAFV600E/D mutation, although was less efficient in those lacking the mutation. But interestingly, by silencing CD70 in CD70+ melanoma cell lines we show that PLX-4032-induced melanoma cell killing and its inhibitory effect on MAPK pathway activation are unaffected by CD70 expression. Consequently, our work demonstrates that CD70 ectopic expression in melanomas is not a valuable biomarker to predict tumor cells sensitivity to BRAF V600 inhibitors.

  16. Arabidopsis thaliana Contains Both Ni2+ and Zn2+ Dependent Glyoxalase I Enzymes and Ectopic Expression of the Latter Contributes More towards Abiotic Stress Tolerance in E. coli

    PubMed Central

    Jain, Muskan; Batth, Rituraj; Kumari, Sumita; Mustafiz, Ananda

    2016-01-01

    The glyoxalase pathway is ubiquitously found in all the organisms ranging from prokaryotes to eukaryotes. It acts as a major pathway for detoxification of methylglyoxal (MG), which deleteriously affects the biological system in stress conditions. The first important enzyme of this system is Glyoxalase I (GLYI). It is a metalloenzyme which requires divalent metal ions for its activity. This divalent metal ion can be either Zn2+ as found in most of eukaryotes or Ni2+ as seen in prokaryotes. In the present study, we have found three active GLYI enzymes (AtGLYI2, AtGLYI3 and AtGLYI6) belonging to different metal activation classes coexisting in Arabidopsis thaliana. These enzymes have been found to efficiently complement the GLYI yeast mutants. These three enzymes have been characterized in terms of their activity, metal dependency, kinetic parameters and their role in conferring tolerance to multiple abiotic stresses in E. coli and yeast. AtGLYI2 was found to be Zn2+ dependent whereas AtGLYI3 and AtGLYI6 were Ni2+ dependent. Enzyme activity of Zn2+ dependent enzyme, AtGLYI2, was observed to be exceptionally high (~250–670 fold) as compared to Ni2+ dependent enzymes, AtGLYI3 and AtGLYI6. The activity of these GLYI enzymes correlated well to their role in stress tolerance. Heterologous expression of these enzymes in E. coli led to better tolerance against various stress conditions. This is the first report of a higher eukaryotic species having multiple active GLYI enzymes belonging to different metal activation classes. PMID:27415831

  17. AtMYB41 activates ectopic suberin synthesis and assembly in multiple plant species and cell types

    PubMed Central

    Kosma, Dylan K; Murmu, Jhadeswar; Razeq, Fakhria M; Santos, Patricia; Bourgault, Richard; Molina, Isabel; Rowland, Owen

    2014-01-01

    Suberin is a lipid and phenolic cell wall heteropolymer found in the roots and other organs of all vascular plants. Suberin plays a critical role in plant water relations and in protecting plants from biotic and abiotic stresses. Here we describe a transcription factor, AtMYB41 (At4g28110), that can activate the steps necessary for aliphatic suberin synthesis and deposition of cell wall-associated suberin-like lamellae in both Arabidopsis thaliana and Nicotiana benthamiana. Overexpression of AtMYB41 increased the abundance of suberin biosynthetic gene transcripts by orders of magnitude and resulted in the accumulation of up to 22 times more suberin-type than cutin-type aliphatic monomers in leaves. Overexpression of AtMYB41 also resulted in elevated amounts of monolignols in leaves and an increase in the accumulation of phenylpropanoid and lignin biosynthetic gene transcripts. Surprisingly, ultrastructural data indicated that overexpression led to the formation of suberin-like lamellae in both epidermal and mesophyll cells of leaves. We further implicate AtMYB41 in the production of aliphatic suberin under abiotic stress conditions. These results provide insight into the molecular-genetic mechanisms of the biosynthesis and deposition of a ubiquitous cell wall-associated plant structure and will serve as a basis for discovering the transcriptional network behind one of the most abundant lipid-based polymers in nature. PMID:25060192

  18. Sonography of Methotrexate for Ectopics

    NASA Astrophysics Data System (ADS)

    Urzicǎ, Denise; Dorohoi, Dana-Ortansa

    2007-04-01

    Treatment unruptured ectopic pregnancy with methotrexate (MTX) and citrovorum factor is now an established alternative to surgical therapy. Serial measurements of serum beta-HCG and early ultrasound examination have allowed detection of early and unruptured tubal ectopic pregnancies, permitting treatment without removal of the tube. It is believed that preserving the tube increases the chance of subsequent live births. Our findings suggest that outpatient transvaginal intratubal methorexate administration can provide a safe and effective alternative to surgical treatment for patients with early and unruptured tubal ectopic pregnancy.

  19. Ectopic expression of an EAR motif deletion mutant of SlERF3 enhances tolerance to salt stress and Ralstonia solanacearum in tomato.

    PubMed

    Pan, I-Chun; Li, Chia-Wen; Su, Ruey-Chih; Cheng, Chiu-Ping; Lin, Choun-Sea; Chan, Ming-Tsair

    2010-10-01

    Ethylene-responsive transcription factors (ERFs) bind specifically to cis-acting DNA regulatory elements such as GCC boxes and play an important role in the regulation of defense- and stress-related genes in plants. In contrast to other ERFs, class II ERFs contain an ERF-associated amphiphilic repression (EAR) domain and act as GCC-mediated transcriptional repressors. In this study, SlERF3, a class II ERF was isolated from tomato and characterized. To examine whether the EAR motif of class II ERF proteins participates in ERF-mediated functions in plants, the EAR domain was deleted to generate SlERF3ΔRD. We show that SlERF3ΔRD protein retains the character of a transcription factor and becomes a GCC-mediated transcriptional activator. Constitutive expression of full-length SlERF3 in tomato severely suppressed growth and, as a result, no transgenic plants were obtained. However, no apparent effects on growth and development of SlERF3ΔRD transgenic plants were observed. Overexpression of SlERF3ΔRD in transgenic tomato induced expression of pathogenesis-related protein genes such as PR1, PR2 and PR5, and enhanced tolerance to Ralstonia solanacearum. Furthermore, transgenic Arabidopsis and tomatoes constitutively expressing SlERF3ΔRD exhibited reduced levels of membrane lipid peroxidation and enhanced tolerance to salt stress. In comparison with wild-type plants grown under stress conditions, transgenic SlERF3ΔRD tomatoes produced more flowers, fruits, and seeds. This study illustrates a gene-enhancing tolerance to both biotic and abiotic stresses in transgenic plants with the deletion of a repressor domain. Our findings suggest that class II ERF proteins may find important use in crop improvement or genetic engineering to increase stress tolerance in plants.

  20. Ectopic Expression of the Wild Grape WRKY Transcription Factor VqWRKY52 in Arabidopsis thaliana Enhances Resistance to the Biotrophic Pathogen Powdery Mildew But Not to the Necrotrophic Pathogen Botrytis cinerea

    PubMed Central

    Wang, Xianhang; Guo, Rongrong; Tu, Mingxing; Wang, Dejun; Guo, Chunlei; Wan, Ran; Li, Zhi; Wang, Xiping

    2017-01-01

    WRKY transcription factors are known to play important roles in plant responses to biotic stresses. We previously showed that the expression of the WRKY gene, VqWRKY52, from Chinese wild Vitis quinquangularis was strongly induced 24 h post inoculation with powdery mildew. In this study, we analyzed the expression levels of VqWRKY52 following treatment with the defense related hormones salicylic acid (SA) and methyl jasmonate, revealing that VqWRKY52 was strongly induced by SA but not JA. We characterized the VqWRKY52 gene, which encodes a WRKY III gene family member, and found that ectopic expression in Arabidopsis thaliana enhanced resistance to powdery mildew and Pseudomonas syringae pv. tomato DC3000, but increased susceptibility to Botrytis cinerea, compared with wild type (WT) plants. The transgenic A. thaliana lines displayed strong cell death induced by the biotrophic powdery mildew pathogen, the hemibiotrophic P. syringe pathogen and the necrotrophic pathogen B. cinerea. In addition, the relative expression levels of various defense-related genes were compared between the transgenic A. thaliana lines and WT plants following the infection by different pathogens. Collectively, the results indicated that VqWRKY52 plays essential roles in the SA dependent signal transduction pathway and that it can enhance the hypersensitive response cell death triggered by microbial pathogens. PMID:28197166

  1. Ectopic Expression of the Wild Grape WRKY Transcription Factor VqWRKY52 in Arabidopsis thaliana Enhances Resistance to the Biotrophic Pathogen Powdery Mildew But Not to the Necrotrophic Pathogen Botrytis cinerea.

    PubMed

    Wang, Xianhang; Guo, Rongrong; Tu, Mingxing; Wang, Dejun; Guo, Chunlei; Wan, Ran; Li, Zhi; Wang, Xiping

    2017-01-01

    WRKY transcription factors are known to play important roles in plant responses to biotic stresses. We previously showed that the expression of the WRKY gene, VqWRKY52, from Chinese wild Vitis quinquangularis was strongly induced 24 h post inoculation with powdery mildew. In this study, we analyzed the expression levels of VqWRKY52 following treatment with the defense related hormones salicylic acid (SA) and methyl jasmonate, revealing that VqWRKY52 was strongly induced by SA but not JA. We characterized the VqWRKY52 gene, which encodes a WRKY III gene family member, and found that ectopic expression in Arabidopsis thaliana enhanced resistance to powdery mildew and Pseudomonas syringae pv. tomato DC3000, but increased susceptibility to Botrytis cinerea, compared with wild type (WT) plants. The transgenic A. thaliana lines displayed strong cell death induced by the biotrophic powdery mildew pathogen, the hemibiotrophic P. syringe pathogen and the necrotrophic pathogen B. cinerea. In addition, the relative expression levels of various defense-related genes were compared between the transgenic A. thaliana lines and WT plants following the infection by different pathogens. Collectively, the results indicated that VqWRKY52 plays essential roles in the SA dependent signal transduction pathway and that it can enhance the hypersensitive response cell death triggered by microbial pathogens.

  2. Ectopic expression of a Ve homolog VvVe gene from Vitis vinifera enhances defense response to Verticillium dahliae infection in tobacco.

    PubMed

    Tang, Juan; Lin, Jing; Yang, Yuwen; Chen, Tianzi; Ling, Xitie; Zhang, Baolong; Chang, Youhong

    2016-01-15

    Verticillium wilt is a soil borne disease that can cause devastating losses to the production of many economically important crops. A Ve1 homologous gene responding to Verticillium dahliae infection was identified in Vitis vinifera cv. "HeiFeng" by semi-quantitative reverse transcription polymerase chain reaction and was designated as VvVe. The overexpression of VvVe in transgenic Nicotiana benthamiana plants significantly enhanced the resistance to isolate V991 of V. dahliae when compared with the wild type plants. The expressions of defense-related genes including the salicylic acid regulated gene pathogen-related 1 (PR1) but not PR2, the ethylene- and jasmonic acid-regulated genes ethylene response factor 1 (ERF1) and lipoxygenase (LOX) were significantly increased due to over expression of VvVe. And greater accumulation of active oxygen, callose and phenylalanine-ammonia lyase were observed in the leaves of transgenic VvVe tobacco plants than the wild type when under infection by V. dahliae. Moreover, the hypersensitive response mimicking cell death was exclusively occurred in the transgenic VvVe tobacco plants but not in the wild type. Taken together, the VvVe gene is a Ve1 like gene which involves in the signal cascade of salicylic acid, jasmonate, and ethylene defense pathways and enhances defense response to V. dahliae infection in the transgenic tobacco.

  3. [Early diagnosis of ectopic pregnancy].

    PubMed

    Belics, Zoran; Gérecz, Balázs; Csákány, M György

    2014-07-20

    Ectopic pregnancy is a high-risk condition that occurs in 2% of reported pregnancies. This percentage is fivefold higher than that registered in the 1970s. Since 1970 there has been a two-fold increase in the ratio of ectopic pregnancies to all reported pregnancies in Hungary and in 2012 7.4 ectopic pregnancies per thousand registered pregnancies were reported. Recently, the majority (80%) of cases can be diagnosed in early stage, and the related mortality objectively decreased in the past few decades to 3.8/10,000 ectopic pregnancies. If a woman with positive pregnancy test has abdominal pain and/or vaginal bleeding the physician should perform a work-up to safely exclude the possibility of ectopic pregnancy. The basis of diagnosis is ultrasonography, especially vaginal ultrasound examination and measurement of the β-subunit of human chorionic gonadotropin. The ultrasound diagnosis is based on the visualization of an ectopic mass rather than the inability to visualize an intrauterine pregnancy. In some questionable cases the diagnostic uterine curettage or laparoscopy may be useful. The actuality of this topic is justified by practical difficulties in obtaining correct diagnosis, especially in the early gestational time.

  4. Ectopic expression of MCAM/MUC18 increases in vitro motility and invasiveness, but decreases in vivo tumorigenesis and metastasis of a mouse melanoma K1735-9 subline in a syngeneic mouse model.

    PubMed

    Wu, Guang-Jer

    2016-12-01

    Ectopic expression of MCAM/MUC18, a cell adhesion molecule in the immunoglobulin-like gene superfamily, induces two moMCAM/MUC18-minus, non-metastatic mouse melanoma K1735 sublines, K3 (tumor(+)/met(low)) and K10 (tumor(-)/met(low)), to metastasize to lungs in a syngeneic C3H mouse model. In this report, we extended investigation of effects of moMCAM/MUC18 expression on tumorigenesis and metastasis in another lowly metastatic, however highly tumorigenic moMCAM/MUC18-minus mouse melanoma K1735 subline, K9 (tumor(+++)/met(low)). We transfected this subline with the moMCAM/MUC18 cDNA, selected for G418-resistant clones with different expression levels of moMCAM/MUC18, and used them for testing effects of MCAM/MUC18 expression on in vitro growth rate, motility, and invasiveness, in vivo subcutaneous tumor growth, and pulmonary metastasis in syngeneic C3H brown mice. Similar to K3 and K10 cells, increased expression of MCAM/MUC18 in K9 cells did not significantly affect in vitro growth rate, but increased in vitro motility and invasiveness. Surprisingly, increased expression of MCAM/MUC18 in K9 cells decreased their induction of tumorigenesis and suppressed their establishment of pulmonary nodules in syngeneic C3H brown mice. We concluded that increased MCAM/MUC18 expression in K9 subline increased in vitro epithelial-to-mesenchymal transition; however, it suppressed in vivo tumorigenicity and metastasis. Thus MCAM/MUC18 acts as a tumor and metastasis suppressor for the K9 subline, different from its role in other K1735 sublines, K3 and K10. Different intrinsic co-factors in different K1735 sublines, which modulate the functions of MCAM/MUC18 in the cells that interact differently to the tumor microenvironment, may render sublines manifest differently in tumorigenicity and metastasis in vivo.

  5. Spontaneous live unilateral twin ectopic pregnancy – A case presentation

    PubMed Central

    Mahsood, Shazia; Shelton, Hannah; Zaedi, Khaled; Economides, DL

    2014-01-01

    The incidence of ectopic pregnancy has increased in recent years and now is around one in 100 pregnancies. However, the incidence of live twin ectopic pregnancy in a spontaneous conception is still quite rare. A 34-year-old gravida 3, para 0 presented in the Early Pregnancy Unit with a positive pregnancy test, lower abdominal pain and vaginal spotting. Her quantitative serum Beta hCG was high, and the transvaginal scan revealed an empty uterine cavity with a twin ectopic pregnancy in the left adnexa with cardiac activity in both embryos. The patient was taken for laparoscopic surgery and a left ampullary twin pregnancy was confirmed. She underwent a left salpingectomy and is well on a one-year follow-up. This case report discusses the incidence, diagnoses and treatment of ectopic pregnancies in general. PMID:27433227

  6. Two IIIf Clade-bHLHs from Freesia hybrida Play Divergent Roles in Flavonoid Biosynthesis and Trichome Formation when Ectopically Expressed in Arabidopsis

    PubMed Central

    Li, Yueqing; Shan, Xiaotong; Gao, Ruifang; Yang, Song; Wang, Shucai; Gao, Xiang; Wang, Li

    2016-01-01

    The MBW complex, comprised by R2R3-MYB, basic helix-loop-helix (bHLH) and WD40, is a single regulatory protein complex that drives the evolution of multiple traits such as flavonoid biosynthesis and epidermal cell differentiation in plants. In this study, two IIIf Clade-bHLH regulator genes, FhGL3L and FhTT8L, were isolated and functionally characterized from Freesia hybrida. Different spatio-temporal transcription patterns were observed showing diverse correlation with anthocyanin and proanthocyanidin accumulation. When overexpressed in Arabidopsis, FhGL3L could enhance the anthocyanin accumulation through up-regulating endogenous regulators and late structural genes. Unexpectedly, trichome formation was inhibited associating with the down-regulation of AtGL2. Comparably, only the accumulation of anthocyanins and proanthocyanidins was strengthened in FhTT8L transgenic lines. Furthermore, transient expression assays demonstrated that FhGL3L interacted with AtPAP1, AtTT2 and AtGL1, while FhTT8L only showed interaction with AtPAP1 and AtTT2. In addition, similar activation of the AtDFR promoter was found between AtPAP1-FhGL3L/FhTT8L and AtPAP1- AtGL3/AtTT8 combinations. When FhGL3L was fused with a strong activation domain VP16, it could activate the AtGL2 promoter when co-transfected with AtGL1. Therefore, it can be concluded that the functionality of bHLH factors may have diverged, and a sophisticated interaction and hierarchical network might exist in the regulation of flavonoid biosynthesis and trichome formation. PMID:27465838

  7. Ectopic expression of MdSPDS1 in sweet orange (Citrus sinensis Osbeck) reduces canker susceptibility: involvement of H2O2 production and transcriptional alteration

    PubMed Central

    2011-01-01

    Background Enormous work has shown that polyamines are involved in a variety of physiological processes, but information is scarce on the potential of modifying disease response through genetic transformation of a polyamine biosynthetic gene. Results In the present work, an apple spermidine synthase gene (MdSPDS1) was introduced into sweet orange (Citrus sinensis Osbeck 'Anliucheng') via Agrobacterium-mediated transformation of embryogenic calluses. Two transgenic lines (TG4 and TG9) varied in the transgene expression and cellular endogenous polyamine contents. Pinprick inoculation demonstrated that the transgenic lines were less susceptible to Xanthomonas axonopodis pv. citri (Xac), the causal agent of citrus canker, than the wild type plants (WT). In addition, our data showed that upon Xac attack TG9 had significantly higher free spermine (Spm) and polyamine oxidase (PAO) activity when compared with the WT, concurrent with an apparent hypersensitive response and the accumulation of more H2O2. Pretreatment of TG9 leaves with guazatine acetate, an inhibitor of PAO, repressed PAO activity and reduced H2O2 accumulation, leading to more conspicuous disease symptoms than the controls when both were challenged with Xac. Moreover, mRNA levels of most of the defense-related genes involved in synthesis of pathogenesis-related protein and jasmonic acid were upregulated in TG9 than in the WT regardless of Xac infection. Conclusion Our results demonstrated that overexpression of the MdSPDS1 gene prominently lowered the sensitivity of the transgenic plants to canker. This may be, at least partially, correlated with the generation of more H2O2 due to increased production of polyamines and enhanced PAO-mediated catabolism, triggering hypersensitive response or activation of defense-related genes. PMID:21439092

  8. Ectopic expression of a novel peach (Prunus persica) CBF transcription factor in apple (Malus × domestica) results in short-day induced dormancy and increased cold hardiness.

    PubMed

    Wisniewski, Michael; Norelli, John; Bassett, Carole; Artlip, Timothy; Macarisin, Dumitru

    2011-05-01

    Low, non-freezing temperatures and/or short daylength (SD) regulates cold acclimation and dormancy in fruit trees. Regarding cold acclimation, C-repeat binding factor (CBF/DREB) transcriptional activator genes have the well-documented ability to induce the expression of a suite of genes associated with increased cold tolerance. We isolated a full-length cDNA of a peach CBF gene, designated PpCBF1 (GenBank Accession HM992943), and constitutively expressed it using an enhanced 35S promoter in apple. Unexpectedly, constitutive overexpression of the PpCBF1 in apple resulted in strong sensitivity to short daylength. Growth cessation and leaf senescence were induced in transgenic lines exposed to SD and optimal growth temperatures of 25°C over a 4-week period. Following 1-4 weeks of SD and 25°C trees were returned to LD and 25°C in the greenhouse. Control (untransformed) plants continued to grow while transgenic lines receiving two or more weeks of SD remained dormant and began to drop leaves. Constitutive overexpression of the PpCBF1 in apple resulted in a 4-6°C increase in freezing tolerance in both the non-acclimated and acclimated states, respectively, compared with untransformed M.26 trees. This is the first instance that constitutive overexpression of a CBF gene has resulted in SD-induction of dormancy and to our knowledge the first time apple has been shown to strongly respond to short daylength as a result of the insertion of a transgene.

  9. SWI/SNF enzymes promote SOX10- mediated activation of myelin gene expression.

    PubMed

    Marathe, Himangi G; Mehta, Gaurav; Zhang, Xiaolu; Datar, Ila; Mehrotra, Aanchal; Yeung, Kam C; de la Serna, Ivana L

    2013-01-01

    SOX10 is a Sry-related high mobility (HMG)-box transcriptional regulator that promotes differentiation of neural crest precursors into Schwann cells, oligodendrocytes, and melanocytes. Myelin, formed by Schwann cells in the peripheral nervous system, is essential for propagation of nerve impulses. SWI/SNF complexes are ATP dependent chromatin remodeling enzymes that are critical for cellular differentiation. It was recently demonstrated that the BRG1 subunit of SWI/SNF complexes activates SOX10 expression and also interacts with SOX10 to activate expression of OCT6 and KROX20, two transcriptional regulators of Schwann cell differentiation. To determine the requirement for SWI/SNF enzymes in the regulation of genes that encode components of myelin, which are downstream of these transcriptional regulators, we introduced SOX10 into fibroblasts that inducibly express dominant negative versions of the SWI/SNF ATPases, BRM or BRG1. Dominant negative BRM and BRG1 have mutations in the ATP binding site and inhibit gene activation events that require SWI/SNF function. Ectopic expression of SOX10 in cells derived from NIH 3T3 fibroblasts led to the activation of the endogenous Schwann cell specific gene, myelin protein zero (MPZ) and the gene that encodes myelin basic protein (MBP). Thus, SOX10 reprogrammed these cells into myelin gene expressing cells. Ectopic expression of KROX20 was not sufficient for activation of these myelin genes. However, KROX20 together with SOX10 synergistically activated MPZ and MBP expression. Dominant negative BRM and BRG1 abrogated SOX10 mediated activation of MPZ and MBP and synergistic activation of these genes by SOX10 and KROX20. SOX10 was required to recruit BRG1 to the MPZ locus. Similarly, in immortalized Schwann cells, BRG1 recruitment to SOX10 binding sites at the MPZ locus was dependent on SOX10 and expression of dominant negative BRG1 inhibited expression of MPZ and MBP in these cells. Thus, SWI/SNF enzymes cooperate with SOX10 to

  10. The paracrinology of tubal ectopic pregnancy.

    PubMed

    Shaw, Julie L V; Horne, Andrew W

    2012-07-25

    As part of successful human reproduction, the Fallopian tube must provide a suitable environment for pre-implantation development of the embryo and for efficient transport of the embryo to the uterus for implantation. These functions are coordinated by paracrine interactions between tubal epithelial, smooth muscle and immune cells and the cells of the developing embryo. Alterations in these signals can lead to a tubal microenvironment encouraging of embryo implantation and to dysregulated tubal motility, ultimately resulting in inappropriate and early implantation of the embryo in the Fallopian tube. Here, we highlight novel and emerging concepts in tubal physiology and pathobiology, such as the induction of a receptive phenotype within the Fallopian tube, leading to ectopic implantation. Chlamydia trachomatis infection is a risk factor for tubal ectopic pregnancy. Activation of toll-like receptor 2 (TLR-2) in the Fallopian tube epithelium, by C. trachomatis has recently been demonstrated, leading to the dysregulation of factors involved in implantation and smooth muscle contractility, such as prokineticins (PROK), activin A and interleukin 1 (IL-1). The Fallopian tube has also recently been shown to harbour a unique population of immune cells, compared to the endometrium. In addition, the complement of immune cells in the Fallopian tube has been reported to be altered in Fallopian tube from women with ectopic pregnancy. There are increasing data suggesting that vascularisation of the Fallopian tube, by the embryo during ectopic pregnancy, differs from that initiated in the uterus during normal pregnancy. This too, is likely the result of paracrine signals between the embryo and the tubal microenvironment.

  11. Ectopic overexpression of castor bean LEAFY COTYLEDON2 (LEC2) in Arabidopsis triggers the expression of genes that encode regulators of seed maturation and oil body proteins in vegetative tissues.

    PubMed

    Kim, Hyun Uk; Jung, Su-Jin; Lee, Kyeong-Ryeol; Kim, Eun Ha; Lee, Sang-Min; Roh, Kyung Hee; Kim, Jong-Bum

    2013-01-01

    The LEAFY COTYLEDON2 (LEC2) gene plays critically important regulatory roles during both early and late embryonic development. Here, we report the identification of the LEC2 gene from the castor bean plant (Ricinus communis), and characterize the effects of its overexpression on gene regulation and lipid metabolism in transgenic Arabidopsis plants. LEC2 exists as a single-copy gene in castor bean, is expressed predominantly in embryos, and encodes a protein with a conserved B3 domain, but different N- and C-terminal domains to those found in LEC2 from Arabidopsis. Ectopic overexpression of LEC2 from castor bean under the control of the cauliflower mosaic virus (CaMV) 35S promoter in Arabidopsis plants induces the accumulation of transcripts that encodes five major transcription factors (the LEAFY COTYLEDON1 (LEC1), LEAFY COTYLEDON1-LIKE (L1L), FUSCA3 (FUS3), and ABSCISIC ACID INSENSITIVE 3 (ABI3) transcripts for seed maturation, and WRINKELED1 (WRI1) transcripts for fatty acid biosynthesis), as well as OLEOSIN transcripts for the formation of oil bodies in vegetative tissues. Transgenic Arabidopsis plants that express the LEC2 gene from castor bean show a range of dose-dependent morphological phenotypes and effects on the expression of LEC2-regulated genes during seedling establishment and vegetative growth. Expression of castor bean LEC2 in Arabidopsis increased the expression of fatty acid elongase 1 (FAE1) and induced the accumulation of triacylglycerols, especially those containing the seed-specific fatty acid, eicosenoic acid (20:1(Δ11)), in vegetative tissues.

  12. Ectopic expression of wheat expansin gene TaEXPA2 improved the salt tolerance of transgenic tobacco by regulating Na(+) /K(+) and antioxidant competence.

    PubMed

    Chen, Yanhui; Han, Yangyang; Kong, Xiangzhu; Kang, Hanhan; Ren, Yuanqing; Wang, Wei

    2017-02-01

    High salinity is one of the most serious environmental stresses that limit crop growth. Expansins are cell wall proteins that regulate plant development and abiotic stress tolerance by mediating cell wall expansion. We studied the function of a wheat expansin gene, TaEXPA2, in salt stress tolerance by overexpressing it in tobacco. Overexpression of TaEXPA2 enhanced the salt stress tolerance of transgenic tobacco plants as indicated by the presence of higher germination rates, longer root length, more lateral roots, higher survival rates and more green leaves under salt stress than in the wild type (WT). Further, when leaf disks of WT plants were incubated in cell wall protein extracts from the transgenic tobacco plants, their chlorophyll content was higher under salt stress, and this improvement from TaEXPA2 overexpression in transgenic tobacco was inhibited by TaEXPA2 protein antibody. The water status of transgenic tobacco plants was improved, perhaps by the accumulation of osmolytes such as proline and soluble sugar. TaEXPA2-overexpressing tobacco lines exhibited lower Na(+) but higher K(+) accumulation than WT plants. Antioxidant competence increased in the transgenic plants because of the increased activity of antioxidant enzymes. TaEXPA2 protein abundance in wheat was induced by NaCl, and ABA signaling was involved. Gene expression regulation was involved in the enhanced salt stress tolerance of the TaEXPA2 transgenic plants. Our results suggest that TaEXPA2 overexpression confers salt stress tolerance on the transgenic plants, and this is associated with improved water status, Na(+) /K(+) homeostasis, and antioxidant competence. ABA signaling participates in TaEXPA2-regulated salt stress tolerance.

  13. Ectopic expression of miR160 results in auxin hypersensitivity, cytokinin hyposensitivity, and inhibition of symbiotic nodule development in soybean.

    PubMed

    Turner, Marie; Nizampatnam, Narasimha Rao; Baron, Mathieu; Coppin, Stéphanie; Damodaran, Suresh; Adhikari, Sajag; Arunachalam, Shivaram Poigai; Yu, Oliver; Subramanian, Senthil

    2013-08-01

    Symbiotic root nodules in leguminous plants result from interaction between the plant and nitrogen-fixing rhizobia bacteria. There are two major types of legume nodules, determinate and indeterminate. Determinate nodules do not have a persistent meristem, while indeterminate nodules have a persistent meristem. Auxin is thought to play a role in the development of both these types of nodules. However, inhibition of rootward auxin transport at the site of nodule initiation is crucial for the development of indeterminate nodules but not determinate nodules. Using the synthetic auxin-responsive DR5 promoter in soybean (Glycine max), we show that there is relatively low auxin activity during determinate nodule initiation and that it is restricted to the nodule periphery subsequently during development. To examine if and what role auxin plays in determinate nodule development, we generated soybean composite plants with altered sensitivity to auxin. We overexpressed microRNA393 to silence the auxin receptor gene family, and these roots were hyposensitive to auxin. These roots nodulated normally, suggesting that only minimal/reduced auxin signaling is required for determinate nodule development. We overexpressed microRNA160 to silence a set of repressor auxin response factor transcription factors, and these roots were hypersensitive to auxin. These roots were not impaired in epidermal responses to rhizobia but had significantly reduced nodule primordium formation, suggesting that auxin hypersensitivity inhibits nodule development. These roots were also hyposensitive to cytokinin and had attenuated expression of key nodulation-associated transcription factors known to be regulated by cytokinin. We propose a regulatory feedback loop involving auxin and cytokinin during nodulation.

  14. Conserved Overlapping Gene Arrangement, Restricted Expression, and Biochemical Activities of DNA Polymerase ν (POLN)*

    PubMed Central

    Takata, Kei-ichi; Tomida, Junya; Reh, Shelley; Swanhart, Lisa M.; Takata, Minoru; Hukriede, Neil A.; Wood, Richard D.

    2015-01-01

    DNA polymerase ν (POLN) is one of 16 DNA polymerases encoded in vertebrate genomes. It is important to determine its gene expression patterns, biological roles, and biochemical activities. By quantitative analysis of mRNA expression, we found that POLN from the zebrafish Danio rerio is expressed predominantly in testis. POLN is not detectably expressed in zebrafish embryos or in mouse embryonic stem cells. Consistent with this, injection of POLN-specific morpholino antisense oligonucleotides did not interfere with zebrafish embryonic development. Analysis of transcripts revealed that vertebrate POLN has an unusual gene expression arrangement, sharing a first exon with HAUS3, the gene encoding augmin-like complex subunit 3. HAUS3 is broadly expressed in embryonic and adult tissues, in contrast to POLN. Differential expression of POLN and HAUS3 appears to arise by alternate splicing of transcripts in mammalian cells and zebrafish. When POLN was ectopically overexpressed in human cells, it specifically coimmunoprecipitated with the homologous recombination factors BRCA1 and FANCJ, but not with previously suggested interaction partners (HELQ and members of the Fanconi anemia core complex). Purified zebrafish POLN protein is capable of thymine glycol bypass and strand displacement, with activity dependent on a basic amino acid residue known to stabilize the primer-template. These properties are conserved with the human enzyme. Although the physiological function of pol ν remains to be clarified, this study uncovers distinctive aspects of its expression control and evolutionarily conserved properties of this DNA polymerase. PMID:26269593

  15. Connexin 43 contributes to ectopic orofacial pain following inferior alveolar nerve injury

    PubMed Central

    Shinoda, Masamichi; Honda, Kuniya; Unno, Syumpei; Shimizu, Noriyoshi; Iwata, Koichi

    2016-01-01

    Background Clinically, it is well known that injury of mandibular nerve fiber induces persistent ectopic pain which can spread to a wide area of the orofacial region innervated by the uninjured trigeminal nerve branches. However, the exact mechanism of such persistent ectopic orofacial pain is not still known. The present study was undertaken to determine the role of connexin 43 in the trigeminal ganglion on mechanical hypersensitivity in rat whisker pad skin induced by inferior alveolar nerve injury. Here, we examined changes in orofacial mechanical sensitivity following inferior alveolar nerve injury. Furthermore, changes in connexin 43 expression in the trigeminal ganglion and its localization in the trigeminal ganglion were also examined. In addition, we investigated the functional significance of connexin 43 in relation to mechanical allodynia by using a selective gap junction blocker (Gap27). Results Long-lasting mechanical allodynia in the whisker pad skin and the upper eyelid skin, and activation of satellite glial cells in the trigeminal ganglion, were induced after inferior alveolar nerve injury. Connexin 43 was expressed in the activated satellite glial cells encircling trigeminal ganglion neurons innervating the whisker pad skin, and the connexin 43 protein expression was significantly increased after inferior alveolar nerve injury. Administration of Gap27 in the trigeminal ganglion significantly reduced satellite glial cell activation and mechanical hypersensitivity in the whisker pad skin. Moreover, the marked activation of satellite glial cells encircling trigeminal ganglion neurons innervating the whisker pad skin following inferior alveolar nerve injury implies that the satellite glial cell activation exerts a major influence on the excitability of nociceptive trigeminal ganglion neurons. Conclusions These findings indicate that the propagation of satellite glial cell activation throughout the trigeminal ganglion via gap junctions, which are

  16. Ectopic testis: a rare case.

    PubMed

    Ebrahimi, Ali

    2010-01-01

    Congenital undescending testis is a common anomaly of testis, but we had a rare case of ectopic testis. A 15-month-old infant was operated emergently because of left incarcerate inguinal hernia. Intraoperative exploration of hernial sac revealed two ectopic testes with one spermatic cord proximally but in the middle divided to two spermatic cords in a 8 shape. There was an important point about vas deferens as it was single proximal to the chord, but divided into two in the middle of the chord. Vessels showed a similar condition about. We released both testes and brought down both of them into scrotum. This is a rare case of ectopic testis transectopia with partially common vas and vessels.

  17. Spontaneous triplet, tubal ectopic gestation.

    PubMed Central

    Nwanodi, Oroma; Berry, Robert

    2006-01-01

    Only six cases of spontaneous, unilateral, triplet ectopic gestations have previously been reported. We now present a seventh case. The patient's prior obstetrical history was significant for a term stillbirth and a term cesarean section for breech. Quantitative betahCG was normal for gestational age; however, the increased trophoblastic mass of an inappropriately implanted multiple gestation may produce sufficient betahCG to mimic an intrauterine singleton gestation. Resolution was achieved via salpingostomy. This case is significant for being spontaneously conceived and not the result of assisted reproductive technologies. Furthermore, this case supports an association between prior cesarean section and ectopic gestation. Images Figure 1 PMID:16775922

  18. Spontaneous bilateral tubal ectopic pregnancy.

    PubMed

    Marasinghe, Jeevan P; Condous, George; Amarasinghe, W I

    2009-03-01

    A 28-year-old woman presented at eight weeks and four days of gestation, according to her menstrual dates, complaining of painless vaginal bleeding for three days. Her urinary pregnancy test was positive. Initial transvaginal ultrasound demonstrated an irregular complex structure with a fluid filled centre in the right adnexum. Despite the diagnosis of a possible underlying unruptured right tubal ectopic pregnancy, she declined surgical intervention and was managed expectantly as an inpatient. When she complained of increasing abdominal pain with haemodynamic instability, an emergency laparotomy was performed and a diagnosis of bilateral tubal ectopic pregnancy was made.

  19. Surgical management of ectopic pregnancy.

    PubMed

    Stock, Laura; Milad, Magdy

    2012-06-01

    Surgery remains an acceptable, and sometimes necessary, modality for the treatment of ectopic pregnancy. Laparoscopy is the preferred method of access, yet controversy remains regarding the optimal procedure and postoperative management. Generally, salpingostomy is employed with the goal of maintaining fertility, although data to support this tenet are lacking. In most cases, the decision to perform conservative versus radical surgery is on the basis of the patient's history, her desire for future fertility, and surgical findings. The procedures of salpingostomy and salpingectomy, techniques to prevent and control blood loss at the time of surgery, and surgical options for nontubal ectopic pregnancies are reviewed.

  20. Heterochronic bilateral ectopic pregnancy after ovulation induction.

    PubMed

    Zhu, Bo; Xu, Gu-feng; Liu, Yi-feng; Qu, Fan; Yao, Wei-miao; Zhu, Yi-min; Gao, Hui-juan; Zhang, Dan

    2014-08-01

    Ectopic pregnancy is identified with the widely-applied assisted reproductive technology (ART). Bilateral ectopic pregnancy is a rare form of ectopic pregnancy which is difficult to be diagnosed at the pre-operation stage. In this paper, we presented an unusual case of heterochronic bilateral ectopic pregnancy after stimulated intrauterine insemination (IUI), where there has been a delay of 22 d between the diagnoses of the two ectopic pregnancies. Literature was reviewed on the occurrence of bilateral ectopic pregnancy during the past four years in the MEDLINE database. We found 16 cases of bilateral ectopic pregnancy reported since 2008, and analyzed the characteristics of those cases of bilateral ectopic pregnancy. We emphasize that ovulation induction and other ARTs may increase the risk of bilateral ectopic pregnancy. Because of the difficulty in identification of bilateral ectopic pregnancy by ultrasonography, the clinician should be aware that the treatment of one ectopic pregnancy does not preclude the occurrence of a second ectopic pregnancy in the same patient and should pay attention to the intra-operation inspection of both side fallopian tubes in any ectopic pregnancy case.

  1. Ectopic expression of a hot pepper bZIP-like transcription factor in potato enhances drought tolerance without decreasing tuber yield.

    PubMed

    Moon, Seok-Jun; Han, Se-Youn; Kim, Dool-Yi; Yoon, In Sun; Shin, Dongjin; Byun, Myung-Ok; Kwon, Hawk-Bin; Kim, Beom-Gi

    2015-11-01

    Over-expression of group A bZIP transcription factor genes in plants improves abiotic stress tolerance but usually reduces yields. Thus, there have been several efforts to overcome yield penalty in transgenic plants. In this study, we characterized that expression of the hot pepper (Capsicum annuum) gene CaBZ1, which encodes a group S bZIP transcription factor, was induced by salt and osmotic stress as well as abscisic acid (ABA). Transgenic potato (Solanum tuberosum) plants over-expressing CaBZ1 exhibited reduced rates of water loss and faster stomatal closure than non transgenic potato plants under drought and ABA treatment conditions. CaBZ1 over-expression in transgenic potato increased the expression of ABA- and stress-related genes (such as CYP707A1, CBF and NAC-like genes) and improved drought stress tolerance. Interestingly, over-expression of CaBZ1 in potato did not produce undesirable growth phenotypes in major agricultural traits such as plant height, leaf size and tuber formation under normal growth conditions. The transgenic potato plants also had higher tuber yields than non transgenic potato plants under drought stress conditions. Thus, CaBZ1 may be useful for improving drought tolerance in tuber crops. This might be the first report of the production of transgenic potato with improved tuber yields under drought conditions.

  2. Possible link between ectopic pancreas and holoprosencephaly.

    PubMed

    Kin, Tatsuya; Korbutt, Gregory S; Shapiro, A M James

    2012-01-01

    We report on the incidental observation of ectopic pancreas in a donor for islet cell transplantation. The donor's clinical and imaging presentation was definitive for holoprosencephaly. This case report discusses a possible link between ectopic pancreas and holoprosencephaly.

  3. [Ectopic breast fibroadenoma. Case report].

    PubMed

    Senatore, G; Zanotti, S; Cambrini, P; Montroni, I; Pellegrini, A; Montanari, E; Santini, D; Taffurelli, M

    2010-03-01

    Among the rare anomalies of the breast development, polythelia is the most common, between 1% and 5% of women and men present supernumerary nipples. Polymastia, usually presenting as ectopic breast tissue without areola-nipple complex, is seen mostly along the milk line, extending from the axilla to the pubic region. Ectopic breast tissue is functionally analogous to mammary gland and it is subjected to the same alterations and diseases, whether benign or malignant, that affect normal breast tissue. We report the case of a 21 years-old female evaluated by the medical staff after founding a solid nodular mass by suspect axillary lymphadenopathy. Differential diagnosis with lymphoma is the major problem in these cases. The mass was removed and the intraoperative histological examination showed fibroadenoma in axillary supernumerary breast. Presence of ectopic breast tissue is a rare condition; development of benign mass or malignant degeneration is possible, but it is very unusual. In case of polymastia diagnosis is simple; in case of isolated nodule, without local inflammation or infection, there are greater difficulties. Ultrasonography is diagnostic in case of breast fibroadenoma, but it might be inadequate in ectopic localizations owing to the shortage of mammary tissue around the mass. Preoperative diagnosis is important to plan an adequate surgical treatment; lumpectomy is indicated in case of benign tissue; in case of malignancy, therapy is based on the standard treatment used for breast cancer (surgery, chemotherapy and radiation therapy).

  4. MRI atlas of ectopic endometriosis.

    PubMed

    Dallaudière, B; Salut, C; Hummel, V; Pouquet, M; Piver, P; Rouanet, J-P; Maubon, A

    2013-03-01

    Ectopic endometriosis is a common condition which is often underdiagnosed, where MRI can help make a diagnosis simply, non-invasively and without irradiation. However, imagery signs of it are enormously polymorphic with a wide range of possible locations. In this paper, we have tried to illustrate comprehensively all its MRI appearances depending on the different locations where it occurs.

  5. Aberrant, ectopic expression of VEGF and VEGF receptors 1 and 2 in malignant colonic epithelial cells. Implications for these cells growth via an autocrine mechanism

    SciTech Connect

    Ahluwalia, Amrita; Jones, Michael K.; Szabo, Sandor; Tarnawski, Andrzej S.

    2013-08-09

    Highlights: •Malignant colonic epithelial cells express VEGF and its receptors. •Cultured colon cancer cells secrete VEGF into the medium. •Inhibition of VEGF receptor significantly decreases colon cancer cell proliferation. •VEGF is critical for colon cancer cell growth. -- Abstract: Vascular endothelial growth factor A (referred to as VEGF) is implicated in colon cancer growth. Currently, the main accepted mechanism by which VEGF promotes colon cancer growth is via the stimulation of angiogenesis, which was originally postulated by late Judah Folkman. However, the cellular source of VEGF in colon cancer tissue; and, the expression of VEGF and its receptors VEGF-R1 and VEGF-R2 in colon cancer cells are not fully known and are subjects of controversy. Material and methods: We examined and quantified expression of VEGF, VEGF-R1 and VEGF-R2 in three different human colonic tissue arrays containing sections of adenocarcinoma (n = 43) and normal mucosa (n = 41). In human colon cancer cell lines HCT116 and HT29 and normal colon cell lines NCM356 and NCM460, we examined expression of VEGF, VEGF-R1 and VEGF-R2 mRNA and protein, VEGF production and secretion into the culture medium; and, the effect of a potent, selective inhibitor of VEGF receptors, AL-993, on cell proliferation. Results: Human colorectal cancer specimens had strong expression of VEGF in cancer cells and also expressed VEGF-R1 and VEGF-R2.In vitro studies showed that human colon cancer cell lines, HCT116 and HT29, but not normal colonic cell lines, express VEGF, VEGF-R1 and VEGF-R2 and secrete VEGF into the medium up to a concentration 2000 pg/ml within 48 h. Furthermore, we showed that inhibition of VEGF receptors using a specific VEGF-R inhibitor significantly reduced proliferation (by >50%) of cultured colon cancer cell lines. Conclusions: Our findings support the contention that VEGF generated by colon cancer cells stimulates their growth directly through an autocrine mechanism that is

  6. Use of a stress inducible promoter to drive ectopic AtCBF expression improves potato freezing tolerance while minimizing negative effects on tuber yield.

    PubMed

    Pino, María-Teresa; Skinner, Jeffrey S; Park, Eung-Jun; Jeknić, Zoran; Hayes, Patrick M; Thomashow, Michael F; Chen, Tony H H

    2007-09-01

    Solanum tuberosum is a frost-sensitive species incapable of cold acclimation. A brief exposure to frost can significantly reduce its yields, while hard frosts can completely destroy entire crops. Thus, gains in freezing tolerance of even a few degrees would be of considerable benefit relative to frost damage. The S. tuberosum cv. Umatilla was transformed with three Arabidopsis CBF genes (AtCBF1-3) driven by either a constitutive CaMV35S or a stress-inducible Arabidopsis rd29A promoter. AtCBF1 and AtCBF3 over-expression via the 35S promoter increased freezing tolerance about 2 degrees C, whereas AtCBF2 over-expression failed to increase freezing tolerance. Transgenic plants of AtCBF1 and AtCBF3 driven by the rd29A promoter reached the same level of freezing tolerance as the 35S versions within a few hours of exposure to low but non-freezing temperatures. Constitutive expression of AtCBF genes was associated with negative phenotypes, including smaller leaves, stunted plants, delayed flowering, and reduction or lack of tuber production. While imparting the same degree of freezing tolerance, control of AtCBF expression via the stress-inducible promoter ameliorated these negative phenotypic effects and restored tuber production to levels similar to wild-type plants. These results suggest that use of a stress-inducible promoter to direct CBF transgene expression can yield significant gains in freezing tolerance without negatively impacting agronomically important traits in potato.

  7. Ectopic expression of a cecropin transgene in the human malaria vector mosquito Anopheles gambiae (Diptera: Culicidae): effects on susceptibility to Plasmodium.

    PubMed

    Kim, Won; Koo, Hyeyoung; Richman, Adam M; Seeley, Douglas; Vizioli, Jacopo; Klocko, Andrew D; O'Brochta, David A

    2004-05-01

    Genetically altering the disease vector status of insects using recombinant DNA technologies is being considered as an alternative to eradication efforts. Manipulating the endogenous immune response of mosquitoes such as the temporal and special expression of antimicrobial peptides like cecropin may result in a refractory phenotype. Using transgenic technology a unique pattern of expression of cecropin A (cecA) in Anopheles gambiae was created such that cecA was expressed beginning 24 h after a blood meal in the posterior midgut. Two independent lines of transgenic An. gambiae were created using a piggyBac gene vector containing the An. gambiae cecA cDNA under the regulatory control of the Aedes aegypti carboxypeptidase promoter. Infection with Plasmodium berghei resulted in a 60% reduction in the number of oocysts in transgenic mosquitoes compared with nontransgenic mosquitoes. Manipulating the innate immune system of mosquitoes can negatively affect their capacity to serve as hosts for the development of disease-causing microbes.

  8. Ectopic expression of a grape aspartic protease gene, AP13, in Arabidopsis thaliana improves resistance to powdery mildew but increases susceptibility to Botrytis cinerea.

    PubMed

    Guo, Rongrong; Tu, Mingxing; Wang, Xianhang; Zhao, Jiao; Wan, Ran; Li, Zhi; Wang, Yuejin; Wang, Xiping

    2016-07-01

    The grape aspartic protease gene, AP13 was previously reported to be responsive, in Chinese wild Vitis quinquangularis cv. 'Shang-24', to infection by Erysiphe necator, the causal agent of powdery mildew disease, as well as to treatment with salicylic acid in V. labrusca×V. vinifera cv. 'Kyoho'. In the current study, we evaluated the expression levels of AP13 in 'Shang-24' in response to salicylic acid (SA), methyl jasmonate (MeJA) and ethylene (ET) treatments, as well as to infection by the necrotrophic fungus, Botrytis cinerea, and the transcript levels of VqAP13 decreased after B. cinerea infection and MeJA treatment, but increased following ET and SA treatments. Transgenic Arabidopsis thaliana lines over-expressing VqAP13 under the control of a constitutive promoter showed enhanced resistance to powdery mildew and to the bacterium Pseudomonas syringae pv. tomato DC3000, and accumulated more callose than wild type plants, while the resistance of transgenic A. thaliana lines to B. cinerea inoculation was reduced. In addition, the expression profiles of various disease resistance- related genes in the transgenic A. thaliana lines following infection by different pathogens were compared to the equivalent profiles in the wild type plants. The results suggest that VqAP13 action promotes the SA dependent signal transduction pathway, but suppresses the JA signal transduction pathway.

  9. Adenovirus-mediated ectopic expression of Msx2 in even-numbered rhombomeres induces apoptotic elimination of cranial neural crest cells in ovo.

    PubMed

    Takahashi, K; Nuckolls, G H; Tanaka, O; Semba, I; Takahashi, I; Dashner, R; Shum, L; Slavkin, H C

    1998-05-01

    Distinct cranial neural crest-derived cell types (a number of neuronal as well as non-neuronal cell lineages) are generated at characteristic times and positions in the rhombomeres of the hindbrain in developing vertebrate embryos. To examine this developmental process, we developed a novel strategy designed to test the efficacy of gain-of-function Msx2 expression within rhombomeres in ovo prior to the emigration of cranial neural crest cells (CNCC). Previous studies indicate that CNCC from odd-numbered rhombomeres (r3 and r5) undergo apoptosis in response to exogenous BMP4. We provide evidence that targeted infection in ovo using adenovirus containing Msx2 and a reporter molecule indicative of translation can induce apoptosis in either even- or odd-numbered rhombomeres. Furthermore, infected lacZ-control explants indicated that CNCC emigrated, and that 20% of these cells were double positive for crest cell markers HNK-1 and beta-gal. In contrast, there were no HNK-1 and Msx2 double positive cells emigrating from Msx2 infected explants. These results support the hypothesis that apoptotic elimination of CNCC can be induced by 'gain-of-function' Msx2 expression in even-numbered rhombomeres. These inductive interactions involve qualitative, quantitative, positional and temporal differences in TGF-beta-related signals, Msx2 expression and other transcriptional control.

  10. Promotion of flowering and reduction of a generation time in apple seedlings by ectopical expression of the Arabidopsis thaliana FT gene using the Apple latent spherical virus vector.

    PubMed

    Yamagishi, Noriko; Sasaki, Shintaro; Yamagata, Kousuke; Komori, Sadao; Nagase, Momoyo; Wada, Masato; Yamamoto, Toshiya; Yoshikawa, Nobuyuki

    2011-01-01

    Tree crops have a long juvenile period which is a serious constraint for genetic improvement and experimental research. For example, apple remains in a juvenile phase for more than five years after seed germination. Here, we report about induction of rapid flowering in apple seedlings using the Apple latent spherical virus (ALSV) vector expressing a FLOWERING LOCUS T (FT) gene from Arabidopsis thaliana. Apple seedlings could be flowered at 1.5-2 months after inoculation to cotyledons of seeds just after germination with ALSV expressing the FT gene. A half of precocious flowers was normal in appearance with sepals, petals, stamens, and pistils. Pollen from a precocious flower successfully pollinated flowers of 'Fuji' apple from which fruits developed normally and next-generation seeds were produced. Our system using the ALSV vector promoted flowering time of apple seedlings within two months after germination and shortened the generation time from seed germination to next-generation seed maturation to within 7 months when pollen from precocious flowers was used for pollination.

  11. Widening the mutation spectrum of EVC and EVC2: ectopic expression of Weyer variants in NIH 3T3 fibroblasts disrupts Hedgehog signaling.

    PubMed

    Valencia, Maria; Lapunzina, Pablo; Lim, Derek; Zannolli, Raffaella; Bartholdi, Deborah; Wollnik, Bernd; Al-Ajlouni, Othman; Eid, Suhair S; Cox, Helen; Buoni, Sabrina; Hayek, Joseph; Martinez-Frias, Maria L; Antonio, Perez-Aytes; Temtamy, Samia; Aglan, Mona; Goodship, Judith A; Ruiz-Perez, Victor L

    2009-12-01

    Autosomal recessive Ellis-van Creveld syndrome and autosomal dominant Weyer acrodental dysostosis are allelic conditions caused by mutations in EVC or EVC2. We performed a mutation screening study in 36 EvC cases and 3 cases of Weyer acrodental dysostosis, and identified pathogenic changes either in EVC or in EVC2 in all cases. We detected 40 independent EVC/EVC2 mutations of which 29 were novel changes in Ellis-van Creveld cases and 2 were novel mutations identified in Weyer pedigrees. Of interest one EvC patient had a T>G nucleotide substitution in intron 7 of EVC (c.940-150T>G), which creates a new donor splice site and results in the inclusion of a new exon. The T>G substitution is at nucleotide +5 of the novel 5' splice site. The three Weyer mutations occurred in the final exon of EVC2 (exon 22), suggesting that specific residues encoded by this exon are a key part of the protein. Using murine versions of EVC2 exon 22 mutations we demonstrate that the expression of a Weyer variant, but not the expression of a truncated protein that mimics an Ellis-van Creveld syndrome mutation, impairs Hedgehog signal transduction in NIH 3T3 cells in keeping with its dominant effect.

  12. Ectopic expression of a LEA protein gene TsLEA1 from Thellungiella salsuginea confers salt-tolerance in yeast and Arabidopsis.

    PubMed

    Zhang, Yiyue; Li, Yin; Lai, Jianbin; Zhang, Huawei; Liu, Yuanyuan; Liang, Liming; Xie, Qi

    2012-04-01

    Thellungiella salsuginea is a valuable halophytic genetic model plant in the Brassicaceae family. Based on previous construction of a salt treated Thellungiella cDNA library carried by pGAD-GH shuttle vector which could directly express in Saccharomyces cerevisiae, a putative salt-tolerance gene TsLEA1 was identified by large-scale stress-tolerance screen in salt sensitive yeast strain G19. The longest 483 bp ORF of TsLEA1 cDNA coding a 160 amino acids protein with a predicted conserved pfam domain shared an 89% amino acid sequence similarity to Arabidopsis LEA group 4 proteins. The transcription level of TsLEA1 gene in T. salsuginea seedlings increased upon salt treatment and its transcript accumulated more in roots than in aerial parts. The ability of the TsLEA1 to facilitate salinity tolerance was analyzed in yeast and transgenic Arabidopsis. It was confirmed that TsLEA1 exhibits conserved salt tolerance in plant as well as in yeast. The results suggested that the TsLEA1 may participate in response to stresses in over expressed circumstance, protecting yeast and plant cells under stress conditions.

  13. Ectopic Expression of Aeluropus littoralis Plasma Membrane Protein Gene AlTMP1 Confers Abiotic Stress Tolerance in Transgenic Tobacco by Improving Water Status and Cation Homeostasis.

    PubMed

    Ben Romdhane, Walid; Ben-Saad, Rania; Meynard, Donaldo; Verdeil, Jean-Luc; Azaza, Jalel; Zouari, Nabil; Fki, Lotfi; Guiderdoni, Emmanuel; Al-Doss, Abdullah; Hassairi, Afif

    2017-03-24

    We report here the isolation and functional analysis of AlTMP1 gene encoding a member of the PMP3 protein family. In Aeluropus littoralis, AlTMP1 is highly induced by abscisic acid (ABA), cold, salt, and osmotic stresses. Transgenic tobacco expressing AlTMP1 exhibited enhanced tolerance to salt, osmotic, H₂O₂, heat and freezing stresses at the seedling stage. Under greenhouse conditions, the transgenic plants showed a higher level of tolerance to drought than to salinity. Noteworthy, AlTMP1 plants yielded two- and five-fold more seeds than non-transgenic plants (NT) under salt and drought stresses, respectively. The leaves of AlTMP1 plants accumulated lower Na⁺ but higher K⁺ and Ca(2+) than those of NT plants. Tolerance to osmotic and salt stresses was associated with higher membrane stability, low electrolyte leakage, and improved water status. Finally, accumulation of AlTMP1 in tobacco altered the regulation of some stress-related genes in either a positive (NHX1, CAT1, APX1, and DREB1A) or negative (HKT1 and KT1) manner that could be related to the observed tolerance. These results suggest that AlTMP1 confers stress tolerance in tobacco through maintenance of ion homeostasis, increased membrane integrity, and water status. The observed tolerance may be due to a direct or indirect effect of AlTMP1 on the expression of stress-related genes which could stimulate an adaptive potential not present in NT plants.

  14. The long non-coding RNA maternally expressed gene 3 activates p53 and is downregulated in esophageal squamous cell cancer.

    PubMed

    Lv, Desheng; Sun, Run; Yu, Qian; Zhang, Xuefei

    2016-10-24

    Esophageal squamous cell cancer (ESCC) is an aggressive malignancy with poor survival. Long non-coding RNAs (lncRNAs) play important roles in tumorigenesis and cancer progression; hence, lncRNAs are also involved in the development and progression of ESCC. In this study, we used quantitative real-time polymerase chain reaction (qRT-PCR) to investigate expression of lncRNA, maternally expressed gene 3 (MEG3) in ESCC. Ectopic expression of MEG3 was performed in ESCC cell lines. Proliferation and apoptosis of ESCC cell lines were analyzed after ectopic expression of MEG3. We found MEG3 was significantly downregulated in ESCC tissues compared with normal tissues by qRT-PCR. Low expression of MEG3 was correlated with lymph node metastasis and advanced TNM stages of ESCC patients and indicated shorter survival (HR = 0.471, 95 % CI 0.234-0.950, P = 0.035), which was confirmed by The Cancer Genome Atlas (TCGA) esophageal cancer dataset. DNA-demethylating agent (5-aza-2-deoxy-cytidine (5-aza-CdR)) treatment significantly increased MEG3 expression level in ESCC cells, and TCGA esophageal cancer dataset also showed that DNA methylation of MEG3 predicted survival. Ectopic expression of MEG3 in ESCC cells inhibited cell proliferation, promoted apoptosis, and suppressed metastasis. Further investigation showed enforced expression of MEG3 activated p53 and its target genes by downregulation of mouse double minute 2 homolog (MDM2). Overall, our study indicated that MEG3 expression loss is common in ESCC and MEG3 could activate p53 and predict prognosis in ESCC.

  15. Ectopic ACTH syndrome: clinicopathological correlations.

    PubMed Central

    Singer, W; Kovacs, K; Ryan, N; Horvath, E

    1978-01-01

    Ten out of 164 cases of bronchogenic carcinoma showed pathological evidence at necropsy of the ectopic ACTH syndrome. All occurred in association with oat-cell carcinoma, constituting 19% of that group. The pathological features consisted of adrenocortical hyperplasia confined to the zona fasciculata and Crooke's hyaline change in the pituitary. Immunoperoxidase stainable ACTH was detected in the pituitary but not in the carcinoma tissue, a surprising finding, which may be due to the different nature of ACTH present in tumour tissue. The ectopic ACTH syndrome was diagnosed ante mortem in only four out of 10 patients on the basis of hypokalaemia and metabolic alkalosis. The lack of clinical pointers in all but terminal cases is discussed as well as possible measures for earlier diagnosis. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:209063

  16. Ectopic expression of apple fruit homogentisate phytyltransferase gene (MdHPT1) increases tocopherol in transgenic tomato (Solanum lycopersicum cv. Micro-Tom) leaves and fruits.

    PubMed

    Seo, Young Sam; Kim, Soo Jin; Harn, Chee Hark; Kim, Woo Taek

    2011-04-01

    Homogentisate phytyltransferase (HPT) is an important enzyme in the biosynthesis of tocopherols (vitamin E). Herein, an HPT homolog (MdHPT1) was isolated from apple (Malus domestica Borkh. cv. Fuji) fruits, whose gene expression level gradually decreased during fruit ripening, reaching a background level in ripened apple fruits. The amounts of α- and γ-tocopherols, two major tocopherols in plant organs, were 5- to 14-fold lower in the fruits than in the leaves and flowers of apple plants. Transgenic tomato plants (Solanum lycopersicum cv. Micro-Tom) overexpressing MdHPT1 were next constructed. Transgenic independent T(1) leaves contained ∼1.8- to 3.6-fold and ∼1.6- to 2.9-fold higher levels of α-tocopherol and γ-tocopherol, respectively, than those in control plants. In addition, the levels of α-tocopherol and γ-tocopherol in 35S:MdHPT1 T(1) fruits increased up to 1.7-fold and 3.1-fold, respectively, as compared to the control fruits, indicating that an increase in α-tocopherol in fruits (maximal 1.7-fold) was less evident than that in leaves (maximal 3.6-fold). This finding suggests that the apple MdHPT1 plays a role in tocopherol production in transgenic tomatoes.

  17. Ectopic Expression of Capsicum-Specific Cell Wall Protein Capsicum annuum Senescence-Delaying 1 (CaSD1) Delays Senescence and Induces Trichome Formation in Nicotiana benthamiana

    PubMed Central

    Seo, Eunyoung; Yeom, Seon-In; Jo, SungHwan; Jeong, Heejin; Kang, Byoung-Cheorl; Choi, Doil

    2012-01-01

    Secreted proteins are known to have multiple roles in plant development, metabolism, and stress response. In a previous study to understand the roles of secreted proteins, Capsicum annuum secreted proteins (CaS) were isolated by yeast secretion trap. Among the secreted proteins, we further characterized Capsicum annuum senescence-delaying 1 (CaSD1), a gene encoding a novel secreted protein that is present only in the genus Capsicum. The deduced CaSD1 contains multiple repeats of the amino acid sequence KPPIHNHKPTDYDRS. Interestingly, the number of repeats varied among cultivars and species in the Capsicum genus. CaSD1 is constitutively expressed in roots, and Agrobacterium-mediated transient overexpression of CaSD1 in Nicotiana benthamiana leaves resulted in delayed senescence with a dramatically increased number of trichomes and enlarged epidermal cells. Furthermore, senescence- and cell division-related genes were differentially regulated by CaSD1-overexpressing plants. These observations imply that the pepper-specific cell wall protein CaSD1 plays roles in plant growth and development by regulating cell division and differentiation. PMID:22441673

  18. Ectopic expression of Capsicum-specific cell wall protein Capsicum annuum senescence-delaying 1 (CaSD1) delays senescence and induces trichome formation in Nicotiana benthamiana.

    PubMed

    Seo, Eunyoung; Yeom, Seon-In; Jo, Sunghwan; Jeong, Heejin; Kang, Byoung-Cheorl; Choi, Doil

    2012-04-01

    Secreted proteins are known to have multiple roles in plant development, metabolism, and stress response. In a previous study to understand the roles of secreted proteins, Capsicum annuum secreted proteins (CaS) were isolated by yeast secretion trap. Among the secreted proteins, we further characterized Capsicum annuum senescence-delaying 1 (CaSD1), a gene encoding a novel secreted protein that is present only in the genus Capsicum. The deduced CaSD1 contains multiple repeats of the amino acid sequence KPPIHNHKPTDYDRS. Interestingly, the number of repeats varied among cultivars and species in the Capsicum genus. CaSD1 is constitutively expressed in roots, and Agrobacterium-mediated transient overexpression of CaSD1 in Nicotiana benthamiana leaves resulted in delayed senescence with a dramatically increased number of trichomes and enlarged epidermal cells. Furthermore, senescence- and cell division-related genes were differentially regulated by CaSD1-overexpressing plants. These observations imply that the pepper-specific cell wall protein CaSD1 plays roles in plant growth and development by regulating cell division and differentiation.

  19. Activation of FXR protects against renal fibrosis via suppressing Smad3 expression

    PubMed Central

    Zhao, Kai; He, Jialin; Zhang, Yan; Xu, Zhizhen; Xiong, Haojun; Gong, Rujun; Li, Song; Chen, Shan; He, Fengtian

    2016-01-01

    Renal fibrosis is the common pathway of most chronic kidney disease progression to end-stage renal failure. The nuclear receptor FXR (farnesoid X receptor), a multiple functional transcription factor, plays an important role in protecting against fibrosis. The TGFβ-Smad signaling has a central role in kidney fibrosis. However, it remains unclear whether FXR plays direct anti-fibrotic effect in renal fibrosis via regulating TGFβ-Smad pathway. In this study, we found that the level of FXR was negatively correlated with that of Smad3 and fibronectin (a marker of fibrosis) in human fibrotic kidneys. Activation of FXR suppressed kidney fibrosis and downregulated Smad3 expression, which was markedly attenuated by FXR antagonist. Moreover, the FXR-mediated repression of fibrosis was significantly alleviated by ectopic expression of Smad3. Luciferase reporter assay revealed that FXR activation inhibited the transcriptional activity of Smad3 gene promoter. The in vivo experiments showed that FXR agonist protected against renal fibrosis and downregulated Smad3 expression in UUO mice. These results suggested that FXR may serve as an important negative regulator for manipulating Smad3 expression, and the FXR/Smad3 pathway may be a novel target for the treatment of renal fibrosis. PMID:27853248

  20. Ectopic Expression of a Basic Helix-Loop-Helix Gene Transactivates Parallel Pathways of Proanthocyanidin Biosynthesis. Structure, Expression Analysis, and Genetic Control of Leucoanthocyanidin 4-Reductase and Anthocyanidin Reductase Genes in Lotus corniculatus1[W

    PubMed Central

    Paolocci, Francesco; Robbins, Mark P.; Madeo, Laura; Arcioni, Sergio; Martens, Stefan; Damiani, Francesco

    2007-01-01

    Proanthocyanidins (PAs) are plant secondary metabolites and are composed primarily of catechin and epicatechin units in higher plant species. Due to the ability of PAs to bind reversibly with plant proteins to improve digestion and reduce bloat, engineering this pathway in leaves is a major goal for forage breeders. Here, we report the cloning and expression analysis of anthocyanidin reductase (ANR) and leucoanthocyanidin 4-reductase (LAR), two genes encoding enzymes committed to epicatechin and catechin biosynthesis, respectively, in Lotus corniculatus. We show the presence of two LAR gene families (LAR1 and LAR2) and that the steady-state levels of ANR and LAR1 genes correlate with the levels of PAs in leaves of wild-type and transgenic plants. Interestingly, ANR and LAR1, but not LAR2, genes produced active proteins following heterologous expression in Escherichia coli and are affected by the same basic helix-loop-helix transcription factor that promotes PA accumulation in cells of palisade and spongy mesophyll. This study provides direct evidence that the same subclass of transcription factors can mediate the expression of the structural genes of both branches of PA biosynthesis. PMID:17098849

  1. Ectopic expression of a basic helix-loop-helix gene transactivates parallel pathways of proanthocyanidin biosynthesis. structure, expression analysis, and genetic control of leucoanthocyanidin 4-reductase and anthocyanidin reductase genes in Lotus corniculatus.

    PubMed

    Paolocci, Francesco; Robbins, Mark P; Madeo, Laura; Arcioni, Sergio; Martens, Stefan; Damiani, Francesco

    2007-01-01

    Proanthocyanidins (PAs) are plant secondary metabolites and are composed primarily of catechin and epicatechin units in higher plant species. Due to the ability of PAs to bind reversibly with plant proteins to improve digestion and reduce bloat, engineering this pathway in leaves is a major goal for forage breeders. Here, we report the cloning and expression analysis of anthocyanidin reductase (ANR) and leucoanthocyanidin 4-reductase (LAR), two genes encoding enzymes committed to epicatechin and catechin biosynthesis, respectively, in Lotus corniculatus. We show the presence of two LAR gene families (LAR1 and LAR2) and that the steady-state levels of ANR and LAR1 genes correlate with the levels of PAs in leaves of wild-type and transgenic plants. Interestingly, ANR and LAR1, but not LAR2, genes produced active proteins following heterologous expression in Escherichia coli and are affected by the same basic helix-loop-helix transcription factor that promotes PA accumulation in cells of palisade and spongy mesophyll. This study provides direct evidence that the same subclass of transcription factors can mediate the expression of the structural genes of both branches of PA biosynthesis.

  2. Lkb1 deletion promotes ectopic lipid accumulation in muscle progenitor cells and mature muscles.

    PubMed

    Shan, Tizhong; Zhang, Pengpeng; Bi, Pengpeng; Kuang, Shihuan

    2015-05-01

    Excessive intramyocellular triglycerides (muscle lipids) are associated with reduced contractile function, insulin resistance, and Type 2 diabetes, but what governs lipid accumulation in muscle is unclear. Here we report a role of Lkb1 in regulating lipid metabolism in muscle stem cells and their descendent mature muscles. We used Myod(Cre) and Lkb1(flox/flox) mice to specifically delete Lkb1 in myogenic cells including stem and differentiated cells, and examined the lipid accumulation and gene expression of myoblasts cultured from muscle stem cells (satellite cells). Genetic deletion of Lkb1 in myogenic progenitors led to elevated expression of lipogenic genes and ectopic lipid accumulation in proliferating myoblasts. Interestingly, the Lkb1-deficient myoblasts differentiated into adipocyte-like cells upon adipogenic induction. However, these adipocyte-like cells maintained myogenic gene expression with reduced ability to form myotubes efficiently. Activation of AMPK by AICAR prevented ectopic lipid formation in the Lkb1-null myoblasts. Notably, Lkb1-deficient muscles accumulated excessive lipids in vivo in response to high-fat diet feeding. These results demonstrate that Lkb1 acts through AMPK to limit lipid deposition in muscle stem cells and their derivative mature muscles, and point to the possibility of controlling muscle lipid content using AMPK activating drugs.

  3. RAS–Mitogen-Activated Protein Kinase Signal Is Required for Enhanced PD-L1 Expression in Human Lung Cancers

    PubMed Central

    Sumimoto, Hidetoshi; Takano, Atsushi; Teramoto, Koji; Daigo, Yataro

    2016-01-01

    Ectopic programmed cell death ligand 1 (PD-L1) expression in non-small cell lung cancers (NSCLCs) is related to immune evasion by cancer, and it is a molecular target of immune checkpoint therapies. Although some altered signals in NSCLCs are responsible for ectopic PD-L1 expression, the precise mechanisms remain obscure. Because we found a higher frequency of EGFR/KRAS mutations in NSCLC cell lines with high PD-L1 expression (p < 0.001), we evaluated the relationships between downstream signals and PD-L1 expression, particularly in three KRAS-mutant adenocarcinoma cell lines. The MEK inhibitor U0126 (20 μM) significantly decreased the surface PD-L1 levels by 50–60% compared with dimethyl sulfoxide (p < 0.0001). Phorbol 12-myristate 13-acetate stimulation (100 nM, 15 min) increased (p < 0.05) and two ERK2 siRNAs as well as KRAS siRNAs decreased (p < 0.05) PD-L1 expression. The transcriptional activity of the potential AP-1 site (+4785 to +5056 from the transcription start site) in the PD-L1 gene was demonstrated by luciferase assays, which was inhibited by U0126. The chromatin immunoprecipitation assay demonstrated the binding of cJUN to the AP-1 site. Two STAT3 siRNAs decreased PD-L1 expression by 10–32% in two of the three KRAS-mutant lung adenocarcinoma cell lines (p < 0.05), while the PI3K inhibitor LY294002 (40 μM) did not change the expression level. Supervised cluster analysis and gene set enrichment analysis between the PD-L1-high and -low NSCLCs revealed a correlation between PD-L1 expression and genes/pathways related to cell motility/adhesion. These results indicate that MAPK signaling is the dominant downstream signal responsible for ectopic PD-L1 expression, in which STAT3 is also involved to some extent. Furthermore, MAPK signaling may control the expression of PD-L1 and several genes related to enhanced cell motility. Our findings suggest that MAPK, along with STAT3, is important for determining PD-L1 expression, which could be useful for

  4. Inferior ectopic bone formation of mesenchymal stromal cells from adipose tissue compared to bone marrow: rescue by chondrogenic pre-induction.

    PubMed

    Brocher, J; Janicki, P; Voltz, P; Seebach, E; Neumann, E; Mueller-Ladner, U; Richter, W

    2013-11-01

    Human mesenchymal stromal cells derived from bone marrow (BMSC) and adipose tissue (ATSC) represent a valuable source of progenitor cells for cell therapy and tissue engineering. While ectopic bone formation is a standard activity of human BMSC on calcium phosphate ceramics, the bone formation capacity of human ATSC has so far been unclear. The objectives of this study were to assess the therapeutic potency of ATSC for bone formation in an ectopic mouse model and determine molecular differences by standardized comparison with BMSC. Although ATSC contained less CD146(+) cells, exhibited better proliferation and displayed similar alkaline phosphatase activity upon osteogenic in vitro differentiation, cells did not develop into bone-depositing osteoblasts on β-TCP after 8weeks in vivo. Additionally, ATSC expressed less BMP-2, BMP-4, VEGF, angiopoietin and IL-6 and more adiponectin mRNA, altogether suggesting insufficient osteochondral commitment and reduced proangiogenic activity. Chondrogenic pre-induction of ATSC/β-TCP constructs with TGF-β and BMP-6 initiated ectopic bone formation in >75% of samples. Both chondrogenic pre-induction and the osteoconductive microenvironment of β-TCP were necessary for ectopic bone formation by ATSC pointing towards a need for inductive conditions/biomaterials to make this more easily accessible cell source attractive for future applications in bone regeneration.

  5. Different Sonographic Faces of Ectopic Pregnancy

    PubMed Central

    Chanana, Charu; Gupta, Nishant; Bansal, Itisha; Hooda, Kusum; Sharma, Pranav; Gupta, Mohit; Gandhi, Darshan; Kumar, Yogesh

    2017-01-01

    Vaginal bleeding in the first trimester has wide differential diagnoses, the most common being a normal early intrauterine pregnancy, with other potential causes including spontaneous abortion and ectopic pregnancy. The incidence of ectopic pregnancy is approximately 2% of all reported pregnancies and is one of the leading causes of maternal mortality worldwide. Clinical signs and symptoms of ectopic pregnancy are often nonspecific. History of pelvic pain with bleeding and positive β-human chorionic gonadotropin should raise the possibility of ectopic pregnancy. Knowledge of the different locations of ectopic pregnancy is of utmost importance, in which ultrasound imaging plays a crucial role. This pictorial essay depicts sonographic findings and essential pitfalls in diagnosing ectopic pregnancy. PMID:28299234

  6. Different Sonographic Faces of Ectopic Pregnancy.

    PubMed

    Chanana, Charu; Gupta, Nishant; Bansal, Itisha; Hooda, Kusum; Sharma, Pranav; Gupta, Mohit; Gandhi, Darshan; Kumar, Yogesh

    2017-01-01

    Vaginal bleeding in the first trimester has wide differential diagnoses, the most common being a normal early intrauterine pregnancy, with other potential causes including spontaneous abortion and ectopic pregnancy. The incidence of ectopic pregnancy is approximately 2% of all reported pregnancies and is one of the leading causes of maternal mortality worldwide. Clinical signs and symptoms of ectopic pregnancy are often nonspecific. History of pelvic pain with bleeding and positive β-human chorionic gonadotropin should raise the possibility of ectopic pregnancy. Knowledge of the different locations of ectopic pregnancy is of utmost importance, in which ultrasound imaging plays a crucial role. This pictorial essay depicts sonographic findings and essential pitfalls in diagnosing ectopic pregnancy.

  7. Ectopic Premolar Tooth in the Sigmoid Notch

    PubMed Central

    Akgül, H. M.; Bayrakdar, I. S.

    2016-01-01

    Impaction of a mandibular premolar is relatively uncommon. Ectopic placement is more unusual and there has been no discussion in the literature of an ectopic mandibular premolar in the coronoid process. In this case report, we present an impacted ectopic mandibular permanent premolar in the sigmoid notch (incisura mandibulae) region. Etiology of the tooth and treatment options are discussed and illustrated by Cone Beam Computed Tomography (CBCT) images. PMID:27547475

  8. Cutaneous ectopic schistosomiasis: diagnostic challenge*

    PubMed Central

    Barros, Cláudia Renata Castro do Rêgo; Maia, Daniela Cristina Caetano; dos Santos, Josemir Belo; Medeiros, Camila Carolina Queiroz; de Araújo, Jessica Guido

    2016-01-01

    Cutaneous schistosomiasis is a rare clinical manifestation of schistosomiasis, an infectious and parasitic disease, caused in Brazil by the trematode Schistosoma mansoni. The lesions are due to the deposition of eggs or, rarely, adult worms, usually involving the genital and groin areas. Extra-genital lesions occur mainly on the torso as papules of zosteriform appearance. The case of a patient with ectopic cutaneous schistosomiasis is reported in this article, due to the rarity of its occurrence and its difficult clinical diagnosis. PMID:26982792

  9. Ectopic activation of the rice NLR heteropair RGA4/RGA5 confers resistance to bacterial blight and bacterial leaf streak diseases.

    PubMed

    Hutin, Mathilde; Césari, Stella; Chalvon, Véronique; Michel, Corinne; Tran, Tuan Tu; Boch, Jens; Koebnik, Ralf; Szurek, Boris; Kroj, Thomas

    2016-10-01

    Bacterial blight (BB) and bacterial leaf streak (BLS) are important diseases in Oryza sativa caused by Xanthomonas oryzae pv. oryzae (Xoo) and Xanthomonas oryzae pv. oryzicola (Xoc), respectively. In both bacteria, transcription activator-like (TAL) effectors are major virulence determinants that act by transactivating host genes downstream of effector-binding elements (EBEs) bound in a sequence-specific manner. Resistance to Xoo is mostly related to the action of TAL effectors, either by polymorphisms that prevent the induction of susceptibility (S) genes or by executor (R) genes with EBEs embedded in their promoter, and that induce cell death and resistance. For Xoc, no resistance sources are known in rice. Here, we investigated whether the recognition of effectors by nucleotide binding and leucine-rich repeat domain immune receptors (NLRs), the most widespread resistance mechanism in plants, is also able to stop BB and BLS. In one instance, transgenic rice lines harboring the AVR1-CO39 effector gene from the rice blast fungus Magnaporthe oryzae, under the control of an inducible promoter, were challenged with transgenic Xoo and Xoc strains carrying a TAL effector designed to transactivate the inducible promoter. This induced AVR1-CO39 expression and triggered BB and BLS resistance when the corresponding Pi-CO39 resistance locus was present. In a second example, the transactivation of an auto-active NLR by Xoo-delivered designer TAL effectors resulted in BB resistance, demonstrating that NLR-triggered immune responses efficiently control Xoo. This forms the foundation for future BB and BLS disease control strategies, whereupon endogenous TAL effectors will target synthetic promoter regions of Avr or NLR executor genes.

  10. Phosphatase activities analyzed by in vivo expressions.

    PubMed

    Schweighofer, Alois; Ayatollahi, Zahra; Meskiene, Irute

    2009-01-01

    Protein phosphatases act to reverse phosphorylation-related modifications induced by protein kinases. Type 2C protein phosphatases (PP2C) are monomeric Ser/Thr phosphatases that require a metal for their activity and are abundant in prokaryotes and eukaryotes. In plants, such as Medicago and Arabidopsis PP2Cs control several essential processes, including ABA signaling, development, and wound-induced mitogen-activated protein kinase (MAPK) pathways. In vitro assays with recombinant proteins and yeast two-hybrid systems usually provide initial information about putative PP2C substrates; however, these observations have to be verified in vivo. Therefore, a method for transient expression in isolated Arabidopsis suspension cell protoplasts was developed to assay PP2C action in living cells. This system has proven to be very useful in producing active enzymes and their substrates and in performing enzymatic reactions in vivo. Transient gene expression in isolated cells enabled assembly of functional protein kinase cascades and the creation of phosphorylated targets for PP2Cs. The method is based on the co-transformation and transient co-expression of different PP2C proteins with MAPK. It shows that epitope-tagged PP2C and MAPK proteins exhibit high enzymatic activities and produce substantial protein amounts easily monitored by Western blot analysis. Additionally, PP2C phosphatase activities can be directly tested in protein extracts from protoplasts, suggesting a possibility for analysis of activities of new PP2C family members.

  11. Autotransplantation of Ectopic Permanent Maxillary Incisors

    PubMed Central

    Abd Jalil, Laila; Muhd Noor, Nurhidayah

    2017-01-01

    The report presents examples of successful cases of autotransplantation of ectopic teeth as donor in the treatment of clinically missing maxillary anterior teeth in young patients. The transplanted teeth were either severely ectopic, inverted, rotated or in an unfavourable position that they are commonly sacrificed as a result. Details of surgical technique as well as clinical and radiographic assessments were discussed. PMID:28352481

  12. The Lin28/Let-7 system in early human embryonic tissue and ectopic pregnancy.

    PubMed

    Lozoya, Teresa; Domínguez, Francisco; Romero-Ruiz, Antonio; Steffani, Liliana; Martínez, Sebastián; Monterde, Mercedes; Ferri, Blanca; Núñez, Maria Jose; AinhoaRomero-Espinós; Zamora, Omar; Gurrea, Marta; Sangiao-Alvarellos, Susana; Vega, Olivia; Simón, Carlos; Pellicer, Antonio; Tena-Sempere, Manuel

    2014-01-01

    Our objective was to determine the expression of the elements of the Lin28/Let-7 system, and related microRNAs (miRNAs), in early stages of human placentation and ectopic pregnancy, as a means to assess the potential role of this molecular hub in the pathogenesis of ectopic gestation. Seventeen patients suffering from tubal ectopic pregnancy (cases) and forty-three women with normal on-going gestation that desired voluntary termination of pregnancy (VTOP; controls) were recruited for the study. Embryonic tissues were subjected to RNA extraction and quantitative PCR analyses for LIN28B, Let-7a, miR-132, miR-145 and mir-323-3p were performed. Our results demonstrate that the expression of LIN28B mRNA was barely detectable in embryonic tissue from early stages of gestation and sharply increased thereafter to plateau between gestational weeks 7-9. In contrast, expression levels of Let-7, mir-132 and mir-145 were high in embryonic tissue from early gestations (≤ 6-weeks) and abruptly declined thereafter, especially for Let-7. Opposite trends were detected for mir-323-3p. Embryonic expression of LIN28B mRNA was higher in early stages (≤ 6-weeks) of ectopic pregnancy than in normal gestation. In contrast, Let-7a expression was significantly lower in early ectopic pregnancies, while miR-132 and miR-145 levels were not altered. Expression of mir-323-3p was also suppressed in ectopic embryonic tissue. We are the first to document reciprocal changes in the expression profiles of the gene encoding the RNA-binding protein, LIN28B, and the related miRNAs, Let-7a, mir-132 and mir-145, in early stages of human placentation. This finding suggests the potential involvement of LIN28B/Let-7 (de)regulated pathways in the pathophysiology of ectopic pregnancy in humans.

  13. Revisiting Ectopic Pregnancy: A Pictorial Essay

    PubMed Central

    Petrides, Artemis; Dinglas, Cheryl; Chavez, Martin; Taylor, Sharon; Mahboob, Sabrina

    2014-01-01

    Ectopic pregnancies occur in approximately 1.4% of all pregnancies and account for 15% of pregnancy-related deaths. Considering the high degree of mortality, recognizing an ectopic pregnancy is important. Signs and symptoms of an ectopic pregnancy are nonspecific and include pain, vaginal bleeding, and an adnexal mass. Therefore, imaging can play a critical role in diagnosis. There are different types of ectopic pregnancies, which are tubal, cornual, cesarean scar, cervical, heterotopic, abdominal, and ovarian. Initial imaging evaluation of pregnant patients with pelvic symptoms is by ultrasonography, transabdominal, transvaginal or both. We review the sonographic appearance of different types of ectopic pregnancies that will aid in accurate and prompt diagnosis. PMID:25161806

  14. Ovarian ectopic pregnancy: A rare case

    PubMed Central

    Ghasemi Tehrani, Hatav; Hamoush, Zaynab; Ghasemi, Mojdeh; Hashemi, Leila

    2014-01-01

    Background: Ovarian pregnancy is a rare form of the non-tubal ectopic pregnancy. It ends with rupture before the end of the first trimester. One of the important risk factors for ovarian pregnancy is in the use of Intra uterine devices (IUD). Case: We report here one such uncommon case of ovarian ectopic pregnancy. Our patient is a 30 years old multiparous woman with two previous cesarean sections with severe hypogastric abdominal pain. During laparotomy, ruptured ovarian ectopic pregnancy was diagnosed, and wedge resection of the ovary was only done. Histopathological examination confirmed it to be an ovarian ectopic pregnancy. Conclusion: IUD is one of contraceptive methods which prevents intra-uterine implantation in 99.5%, if implant occurs with IUD, it is tubal implantation in 95% of cases, and it is very rare in other places such as ovary. The most important risk factor of ovarian ectopic pregnancy is IUD as in this study it was showed. PMID:24976824

  15. Expression and Activity of Metalloproteinases in Depression

    PubMed Central

    Bobińska, Kinga; Szemraj, Janusz; Czarny, Piotr; Gałecki, Piotr

    2016-01-01

    Background Depression is one of the most common mental disorders and often co-exists with somatic diseases. The most probable cause of comorbidity is a generalized inflammatory process that occurs in both depression and somatic diseases. Matrix metalloproteinases MMPs play a role in modulating inflammation and their impact in many inflammatory diseases has been investigated. The purpose of this study was to evaluate gene expression for selected polymorphisms of MMP-2 (C-735T), MMP-7 (A-181G), and MMP-9 (T-1702A, C1562T), which have been confirmed to participate in development of depression, and TIMP-2 (G-418C, tissue inhibitor of MMP). Activity variability of pro-MMP-2 and pro-MMP-9 was measured in a group of people with depression and a group of healthy individuals. Material/Methods The examined population comprised 142 individuals suffering from depression and 100 individuals who formed a control group (CG). Designations were carried out for MMP-2 (C-735T), MMP-7 (A-181G), MMP-9 (T-1702A, C1562T), and TIMP-2 (G-418C). Results For all examined and tested MMPs and for TIMP-2, gene expression at the mRNA level was higher in patients with depression than in the CG. Similar results were recorded for gene expression at the protein level, while expression on the protein level for TIMP-2 was higher in the CG. Change in activity of MMP-2 and pro-MMP-2 was statistically more significant in the group with depression. The opposite result was recorded for MMP-9 and pro-MMP-9, in which the change in activity was statistically more significant in the CG. Conclusions Changes in MMPs and TIMP expression may be a common element in, or perhaps even a marker for, recurrent depressive disorders and somatic diseases. PMID:27098106

  16. [Ectopic Decidualization: A Forgotten Entity].

    PubMed

    Mendes, Joana; Costa, Antónia

    2016-01-01

    Introdução: Apesar da decidualização ectópica ser uma entidade frequentemente subdiagnosticada, pode ter impacto clínico adverso na morbimortalidade materno-fetal. O objetivo deste trabalho foi rever a evidência científica relativa a etiopatogenia, clínica, abordagem diagnóstica e terapêutica sobre esta temática. Material e Métodos: A pesquisa bibliográfica foi realizada na PubMed, Web of Science e Scopus, através da query ('deciduosis' OR 'ectopic decidualization' OR 'ectopic decidua' OR 'ectopic decidua reaction'), incluindo-se artigos publicados até 31/6/2014 e de todos os níveis de evidência. Resultados: A decidualização ectópica, geralmente, representa uma condição benigna, assintomática e sem necessidade de intervenção terapêutica. Encontra-se, maioritariamente, associada à gravidez, com regressão completa no período pós-parto. A frequência do seu diagnóstico depende da suspeição clínica, bem como do local onde surge, sendo o omento e o ovário os locais mais comuns. Quando sintomática, as principais manifestações clínicas são quadros hemorrágicos, nomeadamente hemorragia genital e hemoperitoneu. Os diagnósticos diferenciais incluem patologia maligna, sendo essencial, nestas situações, a confirmação histopatológica. O baixo índice de suspeição clínica pode levar à realização de biópsia, que pode acarretar impacto adverso grave devido à elevada friabilidade destas lesões. Discussão e Conclusão: O reconhecimento desta entidade e das suas características clínicas torna-se essencial na conduta destas doentes. Tal permite por um lado a abordagem médica precoce e adequada nos casos graves, e por outro lado (na maioria dos casos) manter a atitude expectante minimizando a iatrogenia, mantendo o desfecho favorável da decidualização ectópica.

  17. Erk5 Controls Slug Expression and Keratinocyte Activation during Wound Healing

    PubMed Central

    Arnoux, Valerie; Nassour, Mayssaa; L'Helgoualc'h, Annie; Hipskind, Robert A.

    2008-01-01

    Reepithelialization during cutaneous wound healing involves numerous signals that result in basal keratinocyte activation, spreading, and migration, all linked to a loosening of cell–cell adhesion structures. The transcription factor Slug is required for this process, and EGF treatment of human keratinocytes induced activating phosphorylation of Erk5 that coincides with slug transcription. Accordingly, ectopic activation of Erk5 led to increased Slug mRNA levels and faster wound healing, whereas keratinocyte migration was totally blocked by Erk5 pathway inhibition. Expression of a shRNA specific for Erk5 strongly diminished Erk5 levels in keratinocytes and significantly decreased their motility response to EGF, along with induction of Slug expression. These Erk5-deprived keratinocytes showed an altered, more compact morphology, along with disruption of desmosome organization. Accordingly, they displayed an altered ability to form cell aggregates. These results implicate a novel EGFR/Erk5/Slug pathway in the control of cytoskeleton organization and cell motility in keratinocytes treated with EGF. PMID:18716062

  18. Polyphenol Oxidase Activity Expression in Ralstonia solanacearum

    PubMed Central

    Hernández-Romero, Diana; Solano, Francisco; Sanchez-Amat, Antonio

    2005-01-01

    Sequencing of the genome of Ralstonia solanacearum revealed several genes that putatively code for polyphenol oxidases (PPOs). To study the actual expression of these genes, we looked for and detected all kinds of PPO activities, including laccase, cresolase, and catechol oxidase activities, in cellular extracts of this microorganism. The conditions for the PPO assays were optimized for the phenolic substrate, pH, and sodium dodecyl sulfate concentration used. It was demonstrated that three different PPOs are expressed. The genes coding for the enzymes were unambiguously correlated with the enzymatic activities detected by generation of null mutations in the genes by using insertional mutagenesis with a suicide plasmid and estimating the changes in the levels of enzymatic activities compared to the levels in the wild-type strain. The protein encoded by the RSp1530 locus is a multicopper protein with laccase activity. Two other genes, RSc0337 and RSc1501, code for nonblue copper proteins exhibiting homology to tyrosinases. The product of RSc0337 has strong tyrosine hydroxylase activity, and it has been shown that this enzyme is involved in melanin synthesis by R. solanacearum. The product of the RSc1501 gene is an enzyme that shows a clear preference for oxidation of o-diphenols. Preliminary characterization of the mutants obtained indicated that PPOs expressed by R. solanacearum may participate in resistance to phenolic compounds since the mutants exhibited higher sensitivity to l-tyrosine than the wild-type strain. These results suggest a possible role in the pathogenic process to avoid plant resistance mechanisms involving the participation of phenolic compounds. PMID:16269713

  19. Ectopic trypsin in the myocardium promotes dilated cardiomyopathy after influenza A virus infection.

    PubMed

    Pan, Hai-Yan; Sun, Hua-Mei; Xue, Lu-Jing; Pan, Min; Wang, Yi-Ping; Kido, Hiroshi; Zhu, Jian-Hua

    2014-09-15

    We have previously reported that ectopic trypsin in the myocardium triggers acute myocarditis after influenza A virus (IAV) infection. As myocarditis is a common precursor to dilated cardiomyopathy (DCM), the aim of the present study was to investigate the influence of trypsin on the progression of DCM after IAV infection. IAV-infected mice treated with saline or trypsin inhibitor were euthanized on days 0, 9, 20, 40 and 60 postinfection. Trypsin expression colocalized with myocardial inflammatory loci and IAV-induced myocarditis peaked on day 9 postinfection and alleviated by day 20 but persisted until day 60 postinfection, even though replication of IAV was not detected from day 20 postinfection. Similar time courses were observed for the activation of pro-matrix metalloproteinase (pro-MMP)-9 and expression of the proinflammatory cytokines IL-6, IL-1β, and TNF-α. Degradation of collagen type I, proliferation of ventricular interstitial collagen, and expression of collagen type I and III mRNA increased significantly during acute and chronic phases; collagen type III mRNA increased more significantly than collagen type I mRNA. Cardiac function progressively deteriorated with progressive left ventricular dilation. The trypsin inhibitor aprotinin suppressed pro-MMP-9 activation and cytokine release, alleviated myocardial inflammation, and restored collagen metabolism during acute and chronic phases of myocarditis. This effectively prevented ventricular dilation and improved cardiac function. These results suggest that ectopic trypsin in the myocardium promoted DCM through chronic activation of pro-MMP-9, persistent induction of cytokines, and mediation of collagen remodeling. Pharmacological inhibition of trypsin activity might be a promising approach for the prevention of viral cardiomyopathy.

  20. Aquaporin 5 Plays a Role in Estrogen-Induced Ectopic Implantation of Endometrial Stromal Cells in Endometriosis.

    PubMed

    Jiang, Xiu Xiu; Fei, Xiang Wei; Zhao, Li; Ye, Xiao Lei; Xin, Liao Bin; Qu, Yang; Xu, Kai Hong; Wu, Rui Jin; Lin, Jun

    2015-01-01

    Aquaporin 5 (AQP5) participates in the migration of endometrial cells. Elucidation of the molecular mechanisms associated with AQP5-mediated, migration of endometrial cells may contribute to a better understanding of endometriosis. Our objectives included identifying the estrogen-response element (ERE) in the promoter region of the AQP5 gene, and, investigating the effects of AQP5 on ectopic implantation of endometrial cells. Luciferase reporter assays and electrophoretic mobility shift assay (EMSA) identified the ERE-like motif in the promoter region of the AQP5 gene. After blocking and up-regulating estradiol (E2) levels, we analysed the expression of AQP5 in endometrial stromal (ES) cells. After blocking E2 /or phosphatidylinositol 3 kinase(PI3K), we analysed the role of AQP5 in signaling pathways. We constructed an AQP5, shRNA, lentiviral vector to knock out the AQP5 gene in ES cells. After knock-out of the AQP5 gene, we studied the role of AQP5 in cell invasion, proliferation, and the formation of ectopic endometrial implants in female mice. We identified an estrogen-response element in the promoter region of the AQP5 gene. Estradiol (E2) increased AQP5 expression in a dose-dependent fashion, that was blocked by ICI182,780(an estrogen receptor inhibitor). E2 activated PI3K /protein kinase B(AKT) pathway (PI3K/AKT), that, in turn, increased AQP5 expression. LY294002(PI3K inhibitor) attenuated estrogen-enhanced, AQP5 expression. Knock-out of the AQP5 gene with AQP5 shRNA lentiviral vector significantly inhibited E2-enhanced invasion, proliferation of ES cells and formation of ectopic implants. Estrogen induces AQP5 expression by activating ERE in the promoter region of the AQP5gene, activates the PI3K/AKT pathway, and, promotes endometrial cell invasion and proliferation. These results provide new insights into some of the mechanisms that may underpin the development of deposits of ectopic endometrium.

  1. Formation of ectopic osteogenesis in weightlessness

    NASA Technical Reports Server (NTRS)

    1977-01-01

    An ectopic osteogenesis experiment aboard the Cosmos-936 biosatellite is described. Decalcified, lyophilized femur and tibia were implanted under the fascia or in the anterior wall of the abdomen in rats. Bone formation before and after the tests is described and illustrated. The extent of formation of ectopic bone in weightlessness did not differ significantly from that in the ground controls, but the bone marrow of the ectopic bone of the flight rats consisted exclusively of fat cells. The deficit of support-muscle loading was considered to cause the disturbance in skeletal bone tissue development.

  2. DNA damage induces N-acetyltransferase NAT10 gene expression through transcriptional activation.

    PubMed

    Liu, Haijing; Ling, Yun; Gong, Yilei; Sun, Ying; Hou, Lin; Zhang, Bo

    2007-06-01

    NAT10 (N-acetyltransferase 10) is a protein with histone acetylation activity and primarily identified to be involved in regulation of telomerase activity. The presented research shows its transcriptional activation by genotoxic agents and possible role in DNA damage. NAT10 mRNA could be markedly increased by using hydrogen peroxide (H2O2) or cisplatin in a dose- and time-dependent way, and the immunofluorescent staining revealed that the treatment of H2O2 or cisplatin induced focal accumulation of NAT10 protein in cellular nuclei. Both H2O2 and cisplatin could stimulate the transcriptional activity of the NAT10 promoter through the upstream sequences from -615 bp to +110 bp, with which some nuclear proteins interacted. Ectopic expression of NAT10 could enhance the number of survival cells in the presence of H2O2 or cisplatin. The above results suggested that NAT10 could be involved in DNA damage response and increased cellular resistance to genotoxicity.

  3. Ectopic pregnancy in Conakry, Guinea.

    PubMed Central

    Thonneau, Patrick; Hijazi, Yolande; Goyaux, Nathalie; Calvez, Thierry; Keita, Namory

    2002-01-01

    OBJECTIVE: To assess the incidence of ectopic pregnancy (EP) in hospitals in Conakry, the capital of Guinea, West Africa. Data on EP incidence in developing countries are rare and often out of date, particularly in Africa. METHODS: A retrospective study was carried out, examining all cases of EP registered in the medical files of two referral maternity units at the Donka and Ignace Deen university hospitals between 1995 and 1999. FINDINGS: The EP incidence at the two maternity units increased from 0.41% to 1.5% of annual deliveries over this period. Haemoperitoneum was observed in most women, with tubal rupture in 93%; only 6 women received conservative treatment. CONCLUSION: The results suggest that the hospital-based incidence of EP per delivery has increased over the last decade in this West African capital, and that health professionals and public health officials in developing countries, especially those in Africa, should consider EP as a major obstetric problem for maternal morbidity. PMID:12077611

  4. Molecular diagnostics and therapeutics for ectopic pregnancy.

    PubMed

    Tong, Stephen; Skubisz, Monika M; Horne, Andrew W

    2015-02-01

    Ectopic pregnancies are a serious gynaecological emergency that can be fatal. As such, prompt diagnosis and safe timely treatment is essential. Here, we review the literature on the development of molecularly targeted diagnostics and therapeutics for ectopic pregnancy. A blood-based biomarker that accurately identifies an ectopic pregnancy could be used to offer early diagnostic certainty in cases where ultrasound cannot determine the location of the embryo ('a pregnancy of unknown location'). Molecules examined so far can be broadly grouped into biological themes of relevance to reproduction: (i) Fallopian tube (dys)function, (ii) embryo/trophoblast growth, (iii) corpus luteum function, (iv) inflammation, (v) uterine function and (vi) angiogenesis. While a sensitive and specific biomarker for ectopic pregnancy has yet to be identified, it is possible that improvements in platform technologies or a multi-modal biomarker approach may yield an accurate diagnostic biomarker test. Furthermore, with the advent of better imaging technology, the need for a blood-based biomarker test may be superseded by improvements in ultrasound or magnetic resonance imaging technology. There have been some recent preclinical studies describing molecularly targeted therapeutic approaches for ectopic pregnancy. Notably, bench-to-bedside studies have examined the use of combination gefitinib (orally available epidermal growth factor receptor inhibitor) and methotrexate. Preclinical studies suggest that combination gefitinib and methotrexate is highly effective in inducing placental cell death, and is significantly more effective than methotrexate alone. In early human trials, encouraging preliminary efficacy data have shown that combination gefitinib and methotrexate can rapidly resolve tubal ectopic pregnancies, and large extra-tubal ectopic pregnancies. If a large clinical randomized controlled trial confirms these findings, combination gefitinib and methotrexate could become a new

  5. Expression and purification of active recombinant human bone morphogenetic 7-2 dimer fusion protein.

    PubMed

    Dang, Jianli; Jing, Lei; Shi, Weiwei; Qin, Ping; Li, Yuyin; Diao, Aipo

    2015-11-01

    Bone morphogenetic proteins (BMPs) have been applied in bone regeneration therapy due to their significant osteogenic activity, however, the complicated processing and high cost in producing recombinant BMP have limited their use in the clinic. In this study, we have developed a simple method to prepare recombinant human BMP7-BMP2 fusion protein with a flexible peptide linker (rhBMP7-2). The rhBMP7-2 protein is expressed efficiently in Escherichia coli, and the denatured protein purified by anion exchange chromatography then refolded by dialysis. The yield was about 6.8 mg per gram of wet cell weight. The bioactivity of re-folded rhBMP7-2 was measured by alkaline phosphatase assay and alizarin red staining using both C2C12 and MC3T3-E1 cells, and also using the rat subcutaneous ectopic bone formation model. High level osteogenic activity was found in all the assays tested demonstrating the production of corrected folded and active rhBMP7-2 protein.

  6. Diagnosis and management of ectopic pregnancy.

    PubMed

    Barash, Joshua H; Buchanan, Edward M; Hillson, Christina

    2014-07-01

    Ectopic pregnancy affects 1% to 2% of all pregnancies and is responsible for 9% of pregnancy-related deaths in the United States. When a pregnant patient presents with first-trimester bleeding or abdominal pain, physicians should consider ectopic pregnancy as a possible cause. The patient history, physical examination, and imaging with transvaginal ultrasonography can usually confirm the diagnosis. When ultrasonography does not clearly identify the pregnancy location, the physician must determine whether the pregnancy is intrauterine (either viable or failing) or ectopic. Use of the beta subunit of human chorionic gonadotropin (ß-hCG) discriminatory level, the ß-hCG value above which an intrauterine pregnancy should be visualized by transvaginal ultrasonography, may be helpful. Failure to visualize an intrauterine pregnancy when ß-hCG is above the discriminatory level suggests ectopic pregnancy. In addition to single measurements of ß-hCG levels, serial levels can be monitored to detect changes. ß-hCG values in approximately 99% of viable intrauterine pregnancies increase by about 50% in 48 hours. The remaining 1% of patients have a slower rate of increase; these patients may have pregnancies that are misdiagnosed as nonviable intrauterine or ectopic. After an ectopic pregnancy has been confirmed, treatment options include medical, surgical, or expectant management. For patients who are medically unstable or experiencing life-threatening hemorrhage, a surgical approach is indicated. For others, management should be based on patient preference after discussion of the risks, benefits, and monitoring requirements of all approaches.

  7. Ectopic recurrent craniopharyngioma of the frontal bone.

    PubMed

    Jakobs, Martin; Orakcioglu, Berk

    2012-09-01

    Ectopic recurrence of craniopharyngioma is a rare phenomenon after transcranial resection of the primary tumor. The authors present a case of ectopic recurrent adamantinomatous craniopharyngioma of the frontal bone resected 16 years after initial transcranial resection of the primary tumor. The lesion was first radiographically described 12 years after surgery and was adjacent to the osteosynthesis plate that had been implanted at the craniotomy site. The recurrent craniopharyngioma was totally resected via a lateral eyebrow approach. No infiltration of the meninges or the brain was detected. Only 50 cases of ectopic recurrent craniopharyngioma have been described to date, with the present case being the first one with recurrence located at the skull bone. So far 2 mechanisms have been described: contamination with tumor cells alongside the surgical tract and spreading via CSF and the subarachnoid space. The authors reviewed the literature, provided the largest collection of cases so far, and performed basic statistical analysis regarding ectopic recurrence. Pediatric and adult patients as well as male and female ones are affected equally by this phenomenon. The mean time of ectopic recurrence after initial surgery was 7.1 years. Ectopic recurrence, although rare, should always be considered in a patient with a newly diagnosed intracranial lesion who has undergone transcranial craniopharyngioma resection before.

  8. Effect of polyphenolic phytochemicals on ectopic oxidative phosphorylation in rod outer segments of bovine retina

    PubMed Central

    Calzia, Daniela; Oneto, Michele; Caicci, Federico; Bianchini, Paolo; Ravera, Silvia; Bartolucci, Martina; Diaspro, Alberto; Degan, Paolo; Manni, Lucia; Traverso, Carlo Enrico; Panfoli, Isabella

    2015-01-01

    Background and Purpose The rod outer segments (OS) of the retina are specialized organelles where phototransduction takes place. The mitochondrial electron transport complexes I–IV, cytochrome c and FoF1-ATP synthase are functionally expressed in the OS disks. Here, we have studied the effect of some polyphenolic compounds acting as inhibitors of mitochondrial ATPase/synthase activity on the OS ectopic FoF1- ATP synthase. The mechanism of apoptosis in the OS was also investigated studying the expression of cytochrome c, caspase 9 and 3 and Apaf-1. Experimental Approach We prepared OS from fresh bovine retinae. Semi-quantitative Western blotting, confocal and electron microscopy, and cytofluorimetry were used along with biochemical analyses such as oximetry, ATP synthesis and hydrolysis. Key Results Resveratrol and curcumin plus piperine inhibited ATP synthesis and oxygen consumption in the OS. Epigallocatechin gallate and quercetin inhibited ATP hydrolysis and oxygen consumption in the OS. Malondialdehyde and hydrogen peroxide were produced in respiring OS in the presence of substrates. Cytochrome c was located inside the disk membranes. Procaspase 9 and 3, as well as Apaf-1 were expressed in the OS. Conclusions and Implications These polyphenolic phytochemicals modulated the FoF1-ATP synthase activity of the the OS reducing production of reactive oxygen intermediates by the OS ectopic electron transport chain. Polyphenols decrease membrane peroxidation and cytochrome c release from disks, preventing the induction of caspase-dependent apoptosis in the OS Such effects are relevant in the design of protection against functional impairment of the OS following oxidative stress from exposure to intense illumination. PMID:25917043

  9. Management Strategies for Aggressive Cushing's Syndrome: From Macroadenomas to Ectopics

    PubMed Central

    Pozza, Carlotta; Graziadio, Chiara; Giannetta, Elisa; Lenzi, Andrea; Isidori, Andrea M.

    2012-01-01

    Cushing's syndrome (CS) is a rare but severe clinical condition represented by an excessive endogenous cortisol secretion and hence excess circulating free cortisol, characterized by loss of the normal feedback regulation and circadian rhythm of the hypothalamic-pituitary axis due to inappropriate secretion of ACTH from a pituitary tumor (Cushing's disease, CD) or an ectopic source (ectopic ACTH secretion, EAS). The remaining causes (20%) are ACTH independent. As soon as the diagnosis is established, the therapeutic goal is the removal of the tumor. Whenever surgery is not curative, management of patients with CS requires a major effort to control hypercortisolemia and associated symptoms. A multidisciplinary approach that includes endocrinologists, neurosurgeons, oncologists, and radiotherapists should be adopted. This paper will focus on traditional and novel medical therapy for aggressive ACTH-dependent CS. Several drugs are able to reduce cortisol levels. Their mechanism of action involves blocking adrenal steroidogenesis (ketoconazole, metyrapone, aminoglutethimide, mitotane, etomidate) or inhibiting the peripheral action of cortisol through blocking its receptors (mifepristone “RU-486”). Other drugs include centrally acting agents (dopamine agonists, somatostatin receptor agonists, retinoic acid, peroxisome proliferator-activated receptor γ “PPAR-γ” ligands) and novel chemotherapeutic agents (temozolomide and tyrosine kinase inhibitors) which have a significant activity against aggressive pituitary or ectopic tumors. PMID:22934113

  10. Expression of the epidermodysplasia verruciformis-associated genes EVER1 and EVER2 is activated by exogenous DNA and inhibited by LMP1 oncoprotein from Epstein-Barr virus.

    PubMed

    Frecha, Cecilia; Chevalier, Sébastien A; van Uden, Patrick; Rubio, Ivonne; Siouda, Maha; Saidj, Djamel; Cohen, Camille; Lomonte, Patrick; Accardi, Rosita; Tommasino, Massimo

    2015-01-15

    EVER1 and EVER2 are mutated in epidermodysplasia verruciformis patients, who are susceptible to human betapapillomavirus (HPV) infection. It is unknown whether their products control the infection of other viruses. Here, we show that the expression of both genes in B cells is activated immediately after Epstein-Barr virus (EBV) infection, whereas at later stages, it is strongly repressed via activation of the NF-κB signaling pathway by latent membrane protein 1 (LMP1). Ectopic expression of EVER1 impairs the ability of EBV to infect B cells.

  11. Dominant Suppression of β1 Integrin by Ectopic CD98-ICD Inhibits Hepatocellular Carcinoma Progression

    PubMed Central

    Wu, Bo; Zhou, Yang; Wang, Yu; Yang, Xiang-Min; Liu, Zhen-Yu; Li, Jiang-Hua; Feng, Fei; Chen, Zhi-Nan; Jiang, Jian-Li

    2016-01-01

    Hepatocellular carcinoma (HCC) is currently the third most common cause of cancer-related death in the Asia-Pacific region. Our previous work showed that knockdown of CD98 significantly inhibits malignant HCC cell phenotypes in vitro and in vivo. The level of CD98 in the membrane is tightly regulated to mediate complex processes associated with cell–cell communication and intracellular signaling. In addition, the intracellular domain of CD98 (CD98-ICD) seems to be of vital importance for recycling CD98 to the membrane after it is endocytosed. The intracellular and transmembrane domains of CD98 associate with β-integrins (primarily β1 but also β3), and this association is essential for CD98 mediation of integrin-like signaling and complements dominant suppression of β1-integrin. We speculated that isolated CD98-ICD would similarly suppress β1-integrin activation and inhibit the malignant behaviors of cancer cells. In particular, the exact role of CD98-ICD has not been studied independently in HCC. In this study, we found that ectopic expression of CD98-ICD inhibited the malignant phenotypes of HCC cells, and the mechanism possibly involves β1-integrin suppression. Moreover, the expression levels of CD98, β1-integrin-A (the activated form of β1-integrin) and Ki-67 were significantly increased in HCC tissues relative to those of normal liver tissues. Therefore, our preliminary study indicates that ectopic CD98-ICD has an inhibitory role in the malignant development of HCC, and shows that CD98-ICD acts as a dominant negative mutant of CD98 that attenuates β1-integrin activation. CD98-ICD may emerge as a promising candidate for antitumor treatment. PMID:27834933

  12. Nerve Growth Factor Promoter Activity Revealed in Mice Expressing Enhanced Green Fluorescent Protein

    PubMed Central

    Kawaja, Michael D.; Smithson, Laura J.; Elliott, Janet; Trinh, Gina; Crotty, Anne-Marie; Michalski, Bernadeta; Fahnestock, Margaret

    2012-01-01

    Nerve growth factor (NGF) and its precursor proNGF are perhaps the best described growth factors of the mammalian nervous system. There remains, however, a paucity of information regarding the precise cellular sites of proNGF/NGF synthesis. Here we report the generation of transgenic mice in which the NGF promoter controls the ectopic synthesis of enhanced green fluorescent protein (EGFP). These transgenic mice provide an unprecedented resolution of both neural cells (e.g., neocortical and hippocampal neurons) and non-neural cells (e.g., renal interstitial cells and thymic reticular cells) that display NGF promoter activity from postnatal development to adulthood. Moreover, the transgene is inducible by injury. At 2 days after sciatic nerve ligation, a robust population of EGFP-positive cells is seen in the proximal nerve stump. These transgenic mice offer novel insights into the cellular sites of NGF promoter activity and can be used as models for investigating the regulation of proNGF/NGF expression after injury. PMID:21456011

  13. [Conservative therapy of ectopic pregnancy with methotrexate].

    PubMed

    Molina Sosa, A; Morales Garcia, V E

    1993-07-01

    Ectopic pregnancy (EP) treated by traditional methods, signifies an approximate lose of fertility capacity of the patient, of 50%. The early diagnosis allows conservative treatment with metotrexate (MTXE), with the objective of preserving the affected tube (salpynx) and its function. Early detection of EP, intact, was done, in 11 cases, with the following parameters: clinical picture, ultrasonography (Us) (empty uterus, adnexial mass and free fluid; adnexial ring or extrauterine embryo); quantitative determination of beta fraction of chorionic gonadotropin (CGH b) and laparoscopic observation, when the following characteristics were met: tubal pregnancy, no greater than 30 mm, intact tubarian serosa, lack of active bleeding and visualization of all pelvic cavity. MTXE 12.5 mg, intrasacularly via laparoscopy in eight patients, and three by laparotomy. The ulterior control of the procedure was: careful surveillance during eight days, Us daily and CGH b in two occasions; ten patients evolutionated satisfactorily, two of them required additional dosis of MTXE, parenterally, 0.5 mg/kg and citovorum factor, 0.1 mg/kg/4 days. The obtained conclusions are: 1. Conservative treatment with MTXE may be carried out provided the conditions established before, are met. 2. The patient's follow up must be precise and because of the high cost of the determination every third day of CGH b, Us may be used. Hysterosalpingography was done in five patients after six months of the procedure, and tubal permeability was seen.(ABSTRACT TRUNCATED AT 250 WORDS)

  14. Bilateral tubal ectopic pregnancies: a report of two cases.

    PubMed

    Eze, Justus N; Obuna, Johnson A; Ejikeme, Brown N

    2012-01-01

    Bilateral tubal ectopic pregnancies are rare occurrences. Two recently managed cases are discussed. The first was a single, sexually active 23-year-old nullipara with family history of twinning who presented with eight weeks amenorrhea, positive pregnancy test, lower abdominal discomfort and other clinical and ultrasound findings suggestive of unruptured left tubal pregnancy. Intra-operatively, unruptured bilateral tubal pregnancies were found and bilateral salpingotomy performed with uneventful recovery. Histology of the specimens confirmed the intra-operative diagnosis. She was appropriately counseled. Case 2, a 30-year-old multiparous housewife who had been on clomid for secondary infertility, presented with signs and symptoms of ruptured tubal ectopic. Intra-operatively, ruptured left and unruptured right tubal pregnancies were found and salpingectomy and salpingotomy were done respectively, with uneventful recovery. The diagnosis was also confirmed histologically and counseling given as in case 1. Bilateral tubal ectopic pregnancies appear to be increasing with twin proneness and use of fertility drugs as risk factors. Whether spontaneous or induced, the hallmarks of good management include early presentation, high index of suspicion, meticulous ultrasound scanning, good case selection, judicious intra-operative inspection of the contralateral tube, histology of specimens and appropriate patient counseling.

  15. GADD45α induction by nickel negatively regulates JNKs/p38 activation via promoting PP2Cα expression.

    PubMed

    Yu, Yonghui; Li, Jingxia; Wan, Yu; Lu, Jianyi; Gao, Jimin; Huang, Chuanshu

    2013-01-01

    Growth arrest and DNA damage (GADD) 45α is a member of GADD inducible gene family, and is inducible in cell response to oxidative stress. GADD45α upregulation induces MKK4/JNK activation in some published experimental systems. However, we found here that the depletion of GADD45α (GADD45α-/-) in mouse embryonic fibroblasts (MEFs) resulted in an increase in the phosphorylation of MKK4/7, MKK3/6 and consequently specific up-regulated the activation of JNK/p38 and their downstream transcription factors, such as c-Jun and ATF2, in comparison to those in GADD45α+/+ MEFs cell following nickel exposure. This up-regulation of MKK-JNK/p38 pathway in GADD45α-/- cell could be rescued by the reconstitutional expression of HA-GADD45α in GADD45α-/- MEFs, GADD45α-/-(HA-GADD45α). Subsequent studies indicated that GADD45α deletion repressed expression of PP2Cα, the phosphotase of MKK3/6 and MKK4/7, whereas ectopic expression of HA-PP2Cα in GADD45α-/- cells attenuated activation of MKK3/6-p38 and MKK4/7-JNK pathways. Collectively, our results demonstrate a novel function and mechanism responsible for GADD45α regulation of MKK/MAPK pathway, further provides insight into understanding the big picture of GADD45α in the regulation of cellular responses to oxidative stress and environmental carcinogens.

  16. Interleukin-27 inhibits ectopic lymphoid-like structure development in early inflammatory arthritis

    PubMed Central

    Bombardieri, Michele; Greenhill, Claire J.; McLeod, Louise; Nerviani, Alessandra; Rocher-Ros, Vidalba; Cardus, Anna; Williams, Anwen S.; Pitzalis, Costantino; Jenkins, Brendan J.

    2015-01-01

    Ectopic lymphoid-like structures (ELSs) reminiscent of secondary lymphoid organs often develop at sites of chronic inflammation where they contribute to immune-mediated pathology. Through evaluation of synovial tissues from rheumatoid arthritis (RA) patients, we now show that low interleukin-27 (IL-27) expression corresponds with an increased incidence of ELS and gene signatures associated with their development and activity. The presence of synovial ELS was also noted in mice deficient in the IL-27 receptor (IL-27R) after the onset of inflammatory arthritis. Here, pathology was associated with increased synovial expression of pro-inflammatory cytokines, homeostatic chemokines, and transcriptional regulators linked with lymphoid neogenesis. In both clinical and experimental RA, synovial ELS coincided with the heightened local expression of cytokines and transcription factors of the Th17 and T follicular helper (Tfh) cell lineages, and included podoplanin-expressing T cells within lymphoid aggregates. IL-27 inhibited the differentiation of podoplanin-expressing Th17 cells, and an increased number of these cells were observed in IL-27R–deficient mice with inflammatory arthritis. Thus, IL-27 appears to negatively regulate ELS development in RA through control of effector T cells. These studies open new opportunities for patient stratification and treatment. PMID:26417004

  17. Characterization and Ectopic Expression of CoWRI1, an AP2/EREBP Domain-Containing Transcription Factor from Coconut (Cocos nucifera L.) Endosperm, Changes the Seeds Oil Content in Transgenic Arabidopsis thaliana and Rice (Oryza sativa L.)

    PubMed Central

    Sun, RuHao; Ye, Rongjian; Gao, Lingchao; Zhang, Lin; Wang, Rui; Mao, Ting; Zheng, Yusheng; Li, Dongdong; Lin, Yongjun

    2017-01-01

    Coconut (Cocos nucifera L.) is a key tropical crop and a member of the monocotyledonous family Arecaceae (Palmaceae). Few genes and related metabolic processes involved in coconut endosperm development have been investigated. In this study, a new member of the WRI1 gene family was isolated from coconut endosperm and was named CoWRI1. Its transcriptional activities and interactions with the acetyl-CoA carboxylase (BCCP2) promoter of CoWRI1 were confirmed by the yeast two-hybrid and yeast one-hybrid approaches, respectively. Functional characterization was carried out through seed-specific expression in Arabidopsis and endosperm-specific expression in rice. In transgenic Arabidopsis, high over-expressions of CoWRI1 in seven independent T2 lines were detected by quantitative real-time PCR. The relative mRNA accumulation of genes encoding enzymes involved in either fatty acid biosynthesis or triacylglycerols assembly (BCCP2, KASI, MAT, ENR, FATA, and GPDH) were also assayed in mature seeds. Furthermore, lipid and fatty acids C16:0 and C18:0 significantly increased. In two homozygous T2 transgenic rice lines (G5 and G2), different CoWRI1 expression levels were detected, but no CoWRI1 transcripts were detected in the wild type. Analyses of the seed oil content, starch content, and total protein content indicated that the two T2 transgenic lines showed a significant increase (P < 0.05) in seed oil content. The transgenic lines also showed a significant increase in starch content, whereas total protein content decreased significantly. Further analysis of the fatty acid composition revealed that palmitic acid (C16:0) and linolenic acid (C18:3) increased significantly in the seeds of the transgenic rice lines, but oleic acid (C18:1) levels significantly declined. PMID:28179911

  18. Characterization and Ectopic Expression of CoWRI1, an AP2/EREBP Domain-Containing Transcription Factor from Coconut (Cocos nucifera L.) Endosperm, Changes the Seeds Oil Content in Transgenic Arabidopsis thaliana and Rice (Oryza sativa L.).

    PubMed

    Sun, RuHao; Ye, Rongjian; Gao, Lingchao; Zhang, Lin; Wang, Rui; Mao, Ting; Zheng, Yusheng; Li, Dongdong; Lin, Yongjun

    2017-01-01

    Coconut (Cocos nucifera L.) is a key tropical crop and a member of the monocotyledonous family Arecaceae (Palmaceae). Few genes and related metabolic processes involved in coconut endosperm development have been investigated. In this study, a new member of the WRI1 gene family was isolated from coconut endosperm and was named CoWRI1. Its transcriptional activities and interactions with the acetyl-CoA carboxylase (BCCP2) promoter of CoWRI1 were confirmed by the yeast two-hybrid and yeast one-hybrid approaches, respectively. Functional characterization was carried out through seed-specific expression in Arabidopsis and endosperm-specific expression in rice. In transgenic Arabidopsis, high over-expressions of CoWRI1 in seven independent T2 lines were detected by quantitative real-time PCR. The relative mRNA accumulation of genes encoding enzymes involved in either fatty acid biosynthesis or triacylglycerols assembly (BCCP2, KASI, MAT, ENR, FATA, and GPDH) were also assayed in mature seeds. Furthermore, lipid and fatty acids C16:0 and C18:0 significantly increased. In two homozygous T2 transgenic rice lines (G5 and G2), different CoWRI1 expression levels were detected, but no CoWRI1 transcripts were detected in the wild type. Analyses of the seed oil content, starch content, and total protein content indicated that the two T2 transgenic lines showed a significant increase (P < 0.05) in seed oil content. The transgenic lines also showed a significant increase in starch content, whereas total protein content decreased significantly. Further analysis of the fatty acid composition revealed that palmitic acid (C16:0) and linolenic acid (C18:3) increased significantly in the seeds of the transgenic rice lines, but oleic acid (C18:1) levels significantly declined.

  19. Involvement of p300 in constitutive and HIV-1 Tat-activated expression of glial fibrillary acidic protein in astrocytes

    PubMed Central

    Zou, Wei; Wang, Zhenyuan; Liu, Ying; Fan, Yan; Zhou, Betty Y.; Yang, X. Frank; He, Johnny J.

    2010-01-01

    HIV-1 Tat protein is an important pathogenic factor in HIV-1-associated neurological diseases. One hallmark of HIV-1 infection of the central nervous system (CNS) is astrocytosis, which is characterized by elevated GFAP expression in astrocytes. We have shown that Tat activates GFAP expression in astrocytes (Zhou, et al., Mol. Cell. Neurosci. 27:296, 2004) and that GFAP is an important regulator of Tat neurotoxicity (Zou, et. al., Am. J. Pathol. 171:1293, 2007). However, the underlying mechanisms for Tat-mediated GFAP up-regulation are not understood. In the current study, we reported concurrent up-regulation of adenovirus E1a-associated 300 kDa protein p300 and GFAP in Tat-expressing human astroytoma cells and primary astrocytes. We showed that p300 was indeed induced by Tat expression and HIV-1 infection and that the induction occurred at the transcriptional level through the cis-acting elements of early growth response 1 (Egr-1) within its promoter. Using siRNA, we further showed that p300 regulated both constitutive and Tat-mediated GFAP expression. Moreover, we showed that ectopic expression of p300 potentiated Tat transactivation activity and increased proliferation of HIV-1-infected astrocytes, but had little effect on HIV-1 replication in these cells. Taken together, these results demonstrate for the first time that Tat is a positive regulator of p300 expression, which in turn regulates GFAP expression, and suggest that the Tat-Egr-1-p300-GFAP axis likely contributes to Tat neurotoxicity and predisposes astrocytes to be an HIV-1 sanctuary in the CNS. PMID:20578042

  20. What Is the Risk for a Second Ectopic Pregnancy?

    MedlinePlus

    ... Old What Is the Risk for a Second Ectopic Pregnancy? KidsHealth > For Parents > What Is the Risk for ... TOPIC Medical Care During Pregnancy Pregnancy & Newborn Center Pregnancy Precautions: FAQs Ectopic Pregnancy Contact Us Print Resources Send to a ...

  1. An Ectopic Network of Transcription Factors Regulated by Hippo Signaling Drives Growth and Invasion of a Malignant Tumor Model.

    PubMed

    Atkins, Mardelle; Potier, Delphine; Romanelli, Lucia; Jacobs, Jelle; Mach, Jana; Hamaratoglu, Fisun; Aerts, Stein; Halder, Georg

    2016-08-22

    Cancer cells have abnormal gene expression profiles; however, to what degree these are chaotic or driven by structured gene regulatory networks is often not known. Here we studied a model of Ras-driven invasive tumorigenesis in Drosophila epithelial tissues and combined in vivo genetics with next-generation sequencing and computational modeling to decipher the regulatory logic of tumor cells. Surprisingly, we discovered that the bulk of the tumor-specific gene expression is controlled by an ectopic network of a few transcription factors that are overexpressed and/or hyperactivated in tumor cells. These factors are Stat, AP-1, the bHLH proteins Myc and AP-4, the nuclear hormone receptor Ftz-f1, the nuclear receptor coactivator Taiman/SRC3, and Mef2. Notably, many of these transcription factors also are hyperactivated in human tumors. Bioinformatic analysis predicted that these factors directly regulate the majority of the tumor-specific gene expression, that they are interconnected by extensive cross-regulation, and that they show a high degree of co-regulation of target genes. Indeed, the factors of this network were required in multiple epithelia for tumor growth and invasiveness, and knockdown of several factors caused a reversion of the tumor-specific expression profile but had no observable effect on normal tissues. We further found that the Hippo pathway effector Yorkie was strongly activated in tumor cells and initiated cellular reprogramming by activating several transcription factors of this network. Thus, modeling regulatory networks identified an ectopic and ordered network of master regulators that control a large part of tumor cell-specific gene expression.

  2. Triplets—one ectopic, two intrauterine

    PubMed Central

    Khan, A. A.

    1973-01-01

    A case of ovarian ectopic conception, complicating a simultaneous twin intra-uterine pregnancy is described, in which the operative diagnosis was confirmed by histological examination. Such cases are extremely rare and comment is made upon this with references. PMID:4786440

  3. Ectopic pregnancy in an apparently healthy bitch.

    PubMed

    Eddey, Philip D

    2012-01-01

    This case describes an extrauterine fetus that was discovered in an apparently healthy bitch 5 mo after whelping. The extrauterine fetus was surgically removed, and the bitch made a complete recovery. The topic of canine ectopic pregnancy is discussed, and a review of previously reported cases is presented.

  4. Ectopic pregnancy comparison of different treatments

    PubMed Central

    Kopani, Fatmir; Rrugia, Arben; Manoku, Nikita

    2010-01-01

    Objectives: We would like to determine the best treatment option depending of ectopic pregnancy situation. Methods. This is a retrospective cohort study that registered all women admitted in Obstetrics and Gynecologic “Queen Geraldine” Hospital June 2003 until 2008 dicember. There were admitted 228 women diagnosed with Ectopic Pregnancy that were treated in our Hospital. Results: Unruptured ectopic pregnancy is diagnosed in 5,2 week of pregnancy and ruptured ectopic in an average of 6,4 weeks. Surgical intervention is registered in 170 patients and we did tubectomy. Success rate of Methotrexate application was more successful if β-hCG level was lower. If the β-hCG level is higher over 10 000 the success rate will decrease in 83 % and in β-hCG levels over 15 000 the success rate will be until 50%. Conclusions: The treatment will be determined by combination of clinical symptoms, ultrasound examination and β-hCG levels. MTX is recommended for all women without hemodinamic problems, unruptured pregnancy and low β-hCG level (β-hCG < 5000 mlU/mL). It is confirmed that the reduction of 15% of β-hCG in the fourth day after application of MTX is a success guide. PMID:22439058

  5. Leukemia inhibitory factor (LIF) is immunohistochemically localized in tubal ectopic pregnancy.

    PubMed

    Güney, Mehmet; Erdemoglu, Evrim; Oral, Baha; Karahan, Nermin; Mungan, Tamer

    2008-01-01

    Leukemia inhibitory factor (LIF) is essential for implantation of the embryo in the endometrium. It is not clear whether the blastocyst requires expression of LIF for implantation into tissues other than endometrium. Immunohistochemical localization of LIF was performed in the fallopian tube of 20 women with ectopic pregnancies, 7 women with normal pregnancies and 20 healthy non-pregnant women. Fallopian tubes were evaluated from specimens taken during tubal ligation in normal pregnancies and non-pregnant fertile women or at operation for tubal surgery in ectopic pregnancies. Biopsies were assayed by immunohistochemistry. Semi-quantitative immunohistochemical reaction scores (IRS) were used for immunohistochemical analyses. Immunolabeling of LIF was detected in the surface epithelium and stroma of fallopian tubes in all subjects. IRS score in the epithelium and stroma of non-pregnant women and women with intrauterine pregnancy were similar (p>0.05). However, women with ectopic pregnancy had significantly increased labeling of LIF compared to others (p<0.05). Immunohistochemical labeling of LIF in the fallopian tube was found to be increased in ectopic pregnancies compared to non-pregnant and healthy pregnant controls. This may indicate a role of LIF in the ectopic implantation of embryos.

  6. Cell cycle stage-specific differential expression of topoisomerase I in tobacco BY-2 cells and its ectopic overexpression and knockdown unravels its crucial role in plant morphogenesis and development.

    PubMed

    Singh, Badri Nath; Mudgil, Yashwanti; John, Riffat; Achary, V Mohan Murali; Tripathy, Manas Kumar; Sopory, Sudhir K; Reddy, Malireddy K; Kaul, Tanushri

    2015-11-01

    DNA topoisomerases catalyze the inter-conversion of different topological forms of DNA. Cell cycle coupled differential accumulation of topoisomerase I (Topo I) revealed biphasic expression maximum at S-phase and M/G1-phase of cultured synchronized tobacco BY-2 cells. This suggested its active role in resolving topological constrains during DNA replication (S-phase) and chromosome decondensation (M/G1 phase). Immuno-localization revealed high concentrations of Topo I in nucleolus. Propidium iodide staining and Br-UTP incorporation patterns revealed direct correlation between immunofluorescence intensity and rRNA transcription activity within nucleolus. Immuno-stained chromosomes during metaphase and anaphase suggested possible role of Topo I in resolving topological constrains during mitotic chromosome condensation. Inhibitor studies showed that in comparison to Topo I, Topo II was essential in resolving topological constrains during chromosome condensation. Probably, Topo II substituted Topo I functioning to certain extent during chromosome condensation, but not vice-versa. Transgenic Topo I tobacco lines revealed morphological abnormalities and highlighted its crucial role in plant morphogenesis and development.

  7. Aberrant FGF signaling, independent of ectopic hedgehog signaling, initiates preaxial polydactyly in Dorking chickens.

    PubMed

    Bouldin, Cortney M; Harfe, Brian D

    2009-10-01

    The formation of supernumerary digits, or polydactyly, is a common congenital malformation. Although mutations in a number of genes have been linked to polydactyly, the molecular etiology for a third of human disorders with polydactyly remains unknown. To increase our understanding of the potential causes for polydactyly, we characterized a spontaneous chicken mutant, known as Dorking. The hindlimbs of Dorkings form a preaxial supernumerary digit. During the early stages of limb development, ectopic expression of several genes, including Sonic Hedgehog (Shh) and Fibroblast Growth Factor 4 (Fgf4), was found in Dorking hindlimbs. In addition to ectopic gene expression, a decrease in cell death in the anterior of the developing Dorking hindlimb was observed. Further molecular investigation revealed that ectopic Fgf4 expression was initiated and maintained independent of ectopic Shh. Additionally, inhibition of Fgf signaling but not hedgehog signaling was capable of restoring the normal anterior domain of cell death in Dorking hindlimbs. Our data indicates that in Dorking chickens, preaxial polydactyly is initiated independent of Shh.

  8. Recurrent ectopic pregnancy after ipsilateral partial salpingectomy: a case report.

    PubMed

    Lee, D H

    2015-01-01

    Ectopic pregnancy is associated with maternal morbidity and mortality during early pregnancy. Ectopic pregnancy occurs in approximately 2% of all pregnancies, and the risk of ectopic pregnancy is increased by eight-fold in women with a history of eopic pregnancy. However, recurrent ectopic pregnancy after ipsilateral partial salpingectomy is quite rare. The authors experienced a case of recurrent ectopic pregnancy in the distal remnant after right partial salpingectomy. In this case report, they discuss this unusual case and provide a brief review of the literature.

  9. Impact of ectopic pregnancy for reproductive prognosis in next generation.

    PubMed

    Kårhus, Line Lund; Egerup, Pia; Skovlund, Charlotte Wessel; Lidegaard, Øjvind

    2014-04-01

    The impact of an ectopic pregnancy in the next generation is unknown. Our aim was to compare reproductive outcomes in daughters of women with and without ectopic pregnancy. Designed as a historical prospective controlled cohort study with data collected in four Danish registries from 1977-2009, women with ectopic pregnancy during 1977-1982 were age-matched to women without ectopic pregnancy. Daughters of these two cohorts were followed until 2009. We compared 5126 daughters of women with ectopic pregnancy with 19 928 daughters of women without ectopic pregnancy. The daughters of women with ectopic pregnancy had a 1.5-fold (95% confidence interval 1.2-1.9) increased risk of ectopic pregnancy, while for deliveries this was 1.0 (1.0-1.1), for miscarriages 1.1 (1.0-1.2), and for induced abortions 1.3 (1.2-1.4). Daughters of mothers with ectopic pregnancy have a 50% higher risk of ectopic pregnancy than daughters of women without an ectopic pregnancy, but a normal delivery rate.

  10. The Caenorhabditis elegans NK-2 homeobox gene ceh-22 activates pharyngeal muscle gene expression in combination with pha-1 and is required for normal pharyngeal development.

    PubMed

    Okkema, P G; Ha, E; Haun, C; Chen, W; Fire, A

    1997-10-01

    Pharyngeal muscle development in the nematode Caenorhabditis elegans appears to share similarities with cardiac muscle development in other species. We have previously described CEH-22, an NK-2 class homeodomain transcription factor similar to Drosophila tinman and vertebrate Nkx2-5, which is expressed exclusively in the pharyngeal muscles. In vitro, CEH-22 binds the enhancer from myo-2, a pharyngeal muscle-specific myosin heavy chain gene. In this paper, we examine the role CEH-22 plays in pharyngeal muscle development and gene activation by (a) ectopically expressing ceh-22 in transgenic C. elegans and (b) examining the phenotype of a ceh-22 loss-of-function mutant. These experiments indicate that CEH-22 is an activator of myo-2 expression and that it is required for normal pharyngeal muscle development. However, ceh-22 is necessary for neither formation of the pharyngeal muscles, nor for myo-2 expression. Our data suggest parallel and potentially compensating pathways contribute to pharyngeal muscle differentiation. We also examine the relationship between ceh-22 and the pharyngeal organ-specific differentiation gene pha-1. Mutations in ceh-22 and pha-1 have strongly synergistic effects on pharyngeal muscle gene expression; in addition, a pha-1 mutation enhances the lethal phenotype caused by a mutation in ceh-22. Wild-type pha-1 is not required for the onset of ceh-22 expression but it appears necessary for maintained expression of ceh-22.

  11. Wavelet variability of SA node originated P waves in atrial fibrillation and in signals with ectopic beats.

    PubMed

    Filos, D; Chouvarda, I; Dakos, G; Mantziari, L; Vassilikos, V; Maglaveras, N

    2012-01-01

    Atrial fibrillation (AF) is one of the most common cardiac arrhythmia. Electrical properties of the atrial myocardium may be related to the appearance of this type of arrhythmia. However ectopic beats, occurring normally in healthy people, disturb cardiac rhythm. Those beats arise from fiber outside the region of SA node. With this work we aim at highlighting differences in the atrial activity between healthy subjects, healthy subjects presenting many ectopic events and patients with AF. The X-Y-Z leads of vectorcardiography recordings are considered. Wavelet-based parameters are extracted from a window which represents atrial activity originated from SA node and compared between signals of the three groups. Results show differences potentially related to the conduction system of the atrium between healthy people and people with AF, as well as between healthy people and people with ectopic events. No difference was found from the analysis of SA node beats between people with AF and healthy with ectopic events.

  12. Correspondence between resting state activity and brain gene expression

    PubMed Central

    Wang, Guang-Zhong; Belgard, T. Grant; Mao, Deng; Chen, Leslie; Berto, Stefano; Preuss, Todd M.; Lu, Hanzhang; Geschwind, Daniel H.; Konopka, Genevieve

    2015-01-01

    SUMMARY The relationship between functional brain activity and gene expression has not been fully explored in the human brain. Here, we identify significant correlations between gene expression in the brain and functional activity by comparing fractional Amplitude of Low Frequency Fluctuations (fALFF) from two independent human fMRI resting state datasets to regional cortical gene expression from a newly generated RNA-seq dataset and two additional gene expression datasets to obtain robust and reproducible correlations. We find significantly more genes correlated with fALFF than expected by chance, and identify specific genes correlated with the imaging signals in multiple expression datasets in the default mode network. Together, these data support a population-level relationship between regional steady state brain gene expression and resting state brain activity. PMID:26590343

  13. Ectopic Lymphoid Structures: Powerhouse of Autoimmunity

    PubMed Central

    Corsiero, Elisa; Nerviani, Alessandra; Bombardieri, Michele; Pitzalis, Costantino

    2016-01-01

    Ectopic lymphoid structures (ELS) often develop at sites of inflammation in target tissues of autoimmune diseases, such as rheumatoid arthritis, Sjögren’s syndrome, multiple sclerosis, myasthenia gravis, and systemic lupus erythematosus. ELS are characterized by the formation of organized T/B cells aggregates, which can acquire follicular dendritic cells network supporting an ectopic germinal center response. In this review, we shall summarize the mechanisms that regulate the formation of ELS in tertiary lymphoid organs, with particular emphasis on the role of lymphoid chemokines in both formation and maintenance of ELS, the role of emerging positive and negative regulators of ELS development and function, including T follicular helper cells and IL-27, respectively. Finally, we shall discuss the main functions of ELS in supporting the affinity maturation, clonal selection, and differentiation of autoreactive B cells contributing to the maintenance and perpetuation of humoral autoimmunity. PMID:27799933

  14. Ectopic Epithelial Deaminase in IBD

    DTIC Science & Technology

    2014-05-01

    transducers and activators of transcription 3 (STAT3). We initially hypothesized the deleterious role of AID in colitis , but our new data rather...disease (IBD) including Crohn’s disease (CD) and ulcerative colitis (UC) is a chronic intestinal disorder that is caused by dysregulated host/microbial...after exposure to dextran sulfate sodium (DSS) that induces acute colitis . During the first budget year, we have successfully collected DNA from

  15. Perforation of jejunal diverticulum with ectopic pancreas.

    PubMed

    Shiratori, Hiroshi; Nishikawa, Takeshi; Shintani, Yukako; Murono, Koji; Sasaki, Kazuhito; Yasuda, Koji; Otani, Kensuke; Tanaka, Toshiaki; Kiyomatsu, Tomomichi; Hata, Keisuke; Kawai, Kazushige; Nozawa, Hiroaki; Ishihara, Soichiro; Fukayama, Masashi; Watanabe, Toshiaki

    2017-04-01

    Perforation of jejunal diverticulum is a rare complication. Here, we report a case of jejunal diverticulum penetration with surrounding ectopic pancreas. An 83-year-old female patient was admitted to our department with acute onset of severe abdominal pain lasting for half a day. Abdominal computed tomography showed outpouching of the small intestine that contained air/fluid, with multiple surrounding air bubbles in the mesentery of the small intestine. She was diagnosed with penetration of the small intestine, and an emergency laparotomy was indicated. The penetrated jejunal diverticulum was identified ~20-cm distal to the ligament of Treitz. Partial resection of the jejunum was performed, and her postoperative course was uneventful. The pathological findings confirmed diverticulum penetration into the mesentery and severe inflammation at the site, with surrounding ectopic pancreas. Furthermore, the pancreatic ducts were opened through the penetrated diverticulum. This rare case shows that the ectopic pancreas might have caused penetration of jejunal diverticulum owing to the pancreatic duct opening through the diverticulum.

  16. Laparoscopic Treatment of Cesarean Scar Ectopic Pregnancy

    PubMed Central

    Felsingerová, Zuzana; Felsinger, Michal; Jandakova, Eva

    2014-01-01

    Abstract Background: An ectopic pregnancy within a Cesarean scar represents a rare type of extrauterine pregnancy in which the fertilized egg nidates in the myometrium of the uterine wall within a scar left from a previous Cesarean delivery. An unrecognized growing Cesarian scar pregnancy may result in uterine rupture, uncontrollable metrorrhagia, and bleeding into the abdominal cavity; therefore, early diagnosis and therapy are necessary to prevent the development of severe complications. Case: A 34-year-old woman after a previous Cesarean delivery presented with amenorrhoa of 7 weeks' duration. Transvaginal ultrasonography revealed an ectopic pregnancy in the Cesarean scar, and a laparoscopic removal of the gestational sac was performed with no complications. Results: Three months later, another laparoscopy with chromopertubation showed no signs of penetration in the suture, both the Fallopian tubes being bilaterally passable. The patient was advised that she could try to achieve pregnancy through spontaneous conception, after which monitoring of the gestational development and a careful assessment of the nidation site would be needed. Conclusions: Laparoscopic surgical management of a viable ectopic pregnancy is technically simple, and is followed by a good recovery. (J GYNECOL SURG 30:309) PMID:25336858

  17. Salvage of impending replant failure by temporary ectopic replantation: a case report.

    PubMed

    Cavadas, Pedro C; Landin, Luis; Navarro-Monzones, Angel; Soler-Nomdedeu, Santiago

    2006-03-01

    Temporary ectopic replantation of amputated parts has been reported previously as an alternative to orthotopic replantation in difficult cases. We report a case in which the left arm initially was replanted orthotopically with subsequent development of extensive infection. The impending vascular failure of the replanted arm was salvaged by reamputation and temporary ectopic transfer of the arm to the groin region. Nine days later the arm was transferred back to the clean humeral stump. The functional result was similar to that of a standard transhumeral replantation, with 30 degrees to 120 degrees of active range of elbow motion, basic grip pattern, and S3 sensibility.

  18. Patterns of activity expressed by juvenile horseshoe crabs.

    PubMed

    Dubofsky, E A; Simpson, S D; Chabot, Christopher C; Watson, Winsor H

    2013-09-01

    Adult American horseshoe crabs, Limulus polyphemus, possess endogenous circadian and circatidal clocks controlling visual sensitivity and locomotion, respectively. The goal of this study was to determine the types of activity rhythms expressed by juvenile horseshoe crabs (n = 24) when exposed to a 14:10 light/dark cycle (LD) for 10 days, followed by 10 days of constant darkness (DD). Horseshoe crab activity was recorded with a digital time-lapse video system that used an infrared-sensitive camera so animals could be monitored at night. In LD, 15 animals expressed daily patterns of activity, 6 displayed a circatidal pattern, and the remaining 3 were arrhythmic. Of the 15 animals with daily patterns of locomotion, 7 had a significant preference (P < 0.05) for diurnal activity and 3 for nocturnal activity; the remainder did not express a significant preference for day or night activity. In DD, 13 horseshoe crabs expressed circatidal rhythms and 8 maintained a pattern of about 24 h. Although these results suggest the presence of a circadian clock influencing circatidal patterns of locomotion, these apparent circadian rhythms may actually represent the expression of just one of the two bouts of activity driven by the putative circalunidian clocks that control their tidal rhythms. Overall, these results indicate that, like adults, juvenile horseshoe crabs express both daily and tidal patterns of activity and that at least one, and maybe both, of these patterns is driven by endogenous clocks.

  19. Stimulation-induced ectopicity and propagation windows in model damaged axons.

    PubMed

    Lachance, Mathieu; Longtin, André; Morris, Catherine E; Yu, Na; Joós, Béla

    2014-12-01

    Neural tissue injuries render voltage-gated Na+ channels (Nav) leaky, thereby altering excitability, disrupting propagation and causing neuropathic pain related ectopic activity. In both recombinant systems and native excitable membranes, membrane damage causes the kinetically-coupled activation and inactivation processes of Nav channels to undergo hyperpolarizing shifts. This damage-intensity dependent change, called coupled left-shift (CLS), yields a persistent or "subthreshold" Nav window conductance. Nodes of Ranvier simulations involving various degrees of mild CLS showed that, as the system's channel/pump fluxes attempt to re-establish ion homeostasis, the CLS elicits hyperexcitability, subthreshold oscillations and neuropathic type action potential (AP) bursts. CLS-induced intermittent propagation failure was studied in simulations of stimulated axons, but pump contributions were ignored, leaving open an important question: does mild-injury (small CLS values, pumps functioning well) render propagation-competent but still quiescent axons vulnerable to further impairments as the system attempts to cope with its normal excitatory inputs? We probe this incipient diffuse axonal injury scenario using a 10-node myelinated axon model. Fully restabilized nodes with mild damage can, we show, become ectopic signal generators ("ectopic nodes") because incoming APs stress Na+ / K+ gradients, thereby altering spike thresholds. Comparable changes could contribute to acquired sodium channelopathies as diverse as epileptic phenomena and to the neuropathic amplification of normally benign sensory inputs. Input spike patterns, we found, propagate with good fidelity through an ectopically firing site only when their frequencies exceed the ectopic frequency. This "propagation window" is a robust phenomenon, occurring despite Gaussian noise, large jitter and the presence of several consecutive ectopic nodes.

  20. Carcinogen-induced trans activation of gene expression.

    PubMed Central

    Kleinberger, T; Flint, Y B; Blank, M; Etkin, S; Lavi, S

    1988-01-01

    We report a new mechanism of carcinogen action by which the expression of several genes was concomitantly enhanced. This mechanism involved the altered activity of cellular factors which modulate the expression of genes under their control. The increased expression was regulated at least in part on the transcriptional level and did not require amplification of the overexpressed genes. This phenomenon was transient; it was apparent as early as 24 h after carcinogen treatment and declined a few days later. Images PMID:2835673

  1. Carcinogen-induced trans activation of gene expression

    SciTech Connect

    Kleinberger, T.; Flint, Y.B.; Blank, M.; Etkin, S.; Lavi, S.

    1988-03-01

    The authors report a new mechanism of carcinogen action by which the expression of several genes was concomitantly enhanced. This mechanism involved the altered activity of cellular factors which modulate the expression of genes under their control. The increased expression was regulated at least in part on the transcriptional level and did not require amplification of the overexpressed genes. This phenomenon was transient; it was apparent as early as 24 h after carcinogen treatment and declined a few days later.

  2. Heparanase expression upregulates platelet adhesion activity and thrombogenicity

    PubMed Central

    Österholm, Cecilia; Zhang, Xiao; Hedin, Ulf; Vlodavsky, Israel; Li, Jin-Ping

    2016-01-01

    Heparanase is an endo-glucuronidase that specifically cleaves heparan sulfate (HS) and heparin polysaccharides. The enzyme is expressed at low levels in normal tissues, but is often upregulated under pathological conditions such as cancer and inflammation. Normal human platelets express exceptionally high levels of heparanase, but the functional consequences of this feature remain unknown. We investigated functional roles of heparanase by comparing the properties of platelets expressing high (Hpa-tg) or low (Ctr) levels of heparanase. Upon activation, Hpa-tg platelets exhibited a much stronger adhesion activity as compared to Ctr platelets, likely contributing to a higher thrombotic activity in a carotid thrombosis model. Furthermore, we found concomitant upregulated expression of both heparanase and CD62P (P-selectin) upon activation of mouse and human platelets. As platelets play important roles in tumor metastasis, these findings indicate contribution of the platelet heparanase to hyper-thrombotic conditions often seen in patients with metastatic cancer. PMID:27129145

  3. Bacterial expression of human kynurenine 3-monooxygenase: solubility, activity, purification.

    PubMed

    Wilson, K; Mole, D J; Binnie, M; Homer, N Z M; Zheng, X; Yard, B A; Iredale, J P; Auer, M; Webster, S P

    2014-03-01

    Kynurenine 3-monooxygenase (KMO) is an enzyme central to the kynurenine pathway of tryptophan metabolism. KMO has been implicated as a therapeutic target in several disease states, including Huntington's disease. Recombinant human KMO protein production is challenging due to the presence of transmembrane domains, which localise KMO to the outer mitochondrial membrane and render KMO insoluble in many in vitro expression systems. Efficient bacterial expression of human KMO would accelerate drug development of KMO inhibitors but until now this has not been achieved. Here we report the first successful bacterial (Escherichia coli) expression of active FLAG™-tagged human KMO enzyme expressed in the soluble fraction and progress towards its purification.

  4. Restricted patterning of vestigial expression in Drosophila wing imaginal discs requires synergistic activation by both Mad and the drifter POU domain transcription factor.

    PubMed

    Certel, K; Hudson, A; Carroll, S B; Johnson, W A

    2000-07-01

    The Drosophila Vestigial protein has been shown to play an essential role in the regulation of cell proliferation and differentiation within the developing wing imaginal disc. Cell-specific expression of vg is controlled by two separate transcriptional enhancers. The boundary enhancer controls expression in cells near the dorsoventral (DV) boundary and is regulated by the Notch signal transduction pathway, while the quadrant enhancer responds to the Decapentaplegic and Wingless morphogen gradients emanating from cells near the anteroposterior (AP) and DV boundaries, respectively. MAD-dependent activation of the vestigial quadrant enhancer results in broad expression throughout the wing pouch but is excluded from cells near the DV boundary. This has previously been thought to be due to direct repression by a signal from the DV boundary; however, we show that this exclusion of quadrant enhancer-dependent expression from the DV boundary is due to the absence of an additional essential activator in those cells. The Drosophila POU domain transcriptional regulator, Drifter, is expressed in all cells within the wing pouch expressing a vgQ-lacZ transgene and is also excluded from the DV boundary. Viable drifter hypomorphic mutations cause defects in cell proliferation and wing vein patterning correlated with decreased quadrant enhancer-dependent expression. Drifter misexpression at the DV boundary using the GAL4/UAS system causes ectopic outgrowths at the distal wing tip due to induction of aberrant Vestigial expression, while a dominant-negative Drifter isoform represses expression of vgQ-lacZ and causes severe notching of the adult wing. In addition, we have identified an essential evolutionarily conserved sequence element bound by the Drifter protein with high affinity and located adjacent to the MAD binding site within the quadrant enhancer. Our results demonstrate that Drifter functions along with MAD as a direct activator of Vestigial expression in the wing pouch.

  5. [Abdominal ectopic pregnancy. A case report and literature review].

    PubMed

    Puch-Ceballos, Eduardo Erik; Vázquez-Castro, Rosbel; Osorio-Pérez, Ana Isabel; Ramos-Ayala, Montserrat; Villarreal-Sosa, Conrado Otoniel; Ruvalcaba-Rivera, Everardo

    2015-07-01

    Abdominal ectopic pregnancy is an extremely rare entity, which represents 1% of all ectopic pregnancies and is associated with high maternal and fetal morbidity and mortality. The maternal mortality risk of an abdominal ectopic pregnancy is seven to eight times greater than the risk of a tubal ectopic pregnancy and is 90 times greater than the risk of intrauterine pregnancy. This is a disease of difficult diagnosis that often takes place late. We report the case of a patient with an abdominal ectopic pregnancy, which was diagnosed by abdominal ultrasound in the second trimester; the patient was suc- cessfully treated with exploratory laparotomy with complete removal of the fetus and placenta. We provide a review of the literature on the risk factors for abdominal ectopic pregnancy, diagnostic tests and therapeutic options.

  6. Neuronal Activity Regulates Hippocampal miRNA Expression

    PubMed Central

    Eacker, Stephen M.; Keuss, Matthew J.; Berezikov, Eugene; Dawson, Valina L.; Dawson, Ted M.

    2011-01-01

    Neuronal activity regulates a broad range of processes in the hippocampus, including the precise regulation of translation. Disruptions in proper translational control in the nervous system are associated with a variety of disorders that fall in the autistic spectrum. MicroRNA (miRNA) represent a relatively recently discovered player in the regulation of translation in the nervous system. We have conducted an in depth analysis of how neuronal activity regulates miRNA expression in the hippocampus. Using deep sequencing we exhaustively identify all miRNAs, including 15 novel miRNAs, expressed in hippocampus of the adult mouse. We identified 119 miRNAs documented in miRBase but less than half of these miRNA were expressed at a level greater than 0.1% of total miRNA. Expression profiling following induction of neuronal activity by electroconvulsive shock demonstrates that most miRNA show a biphasic pattern of expression: rapid induction of specific mature miRNA expression followed by a decline in expression. These results have important implications into how miRNAs influence activity-dependent translational control. PMID:21984899

  7. Ectopic over-expression of BhHsf1, a heat shock factor from the resurrection plant Boea hygrometrica, leads to increased thermotolerance and retarded growth in transgenic Arabidopsis and tobacco.

    PubMed

    Zhu, Yan; Wang, Zhi; Jing, Yanjun; Wang, Lili; Liu, Xia; Liu, Yongxiu; Deng, Xin

    2009-11-01

    Plant heat shock transcription factors (Hsfs) are commonly found to be involved in various stress responses. Several Hsfs displayed dwarf phenotype while conferred stress tolerance when over-expressed. However, the underlying mechanisms were not fully understood. Here we report the cloning and characterization of an Hsf (BhHsf1) from the resurrection plant Boea hygrometrica. Drought, heat and wound can induce BhHsf1 expression. The over-expression of BhHsf1 conferred growth retardation and stress tolerance in both Arabidopsis and tobacco. Evidence was presented to show that the growth retardation of aerial organs in the transgenic plants was resulted from the reduction of cell proliferation. Gene expression profiling using microarray hybridization and pathway analysis showed that Hsps and stress-associated genes were induced whereas the genes related to DNA replication and mitotic cell cycle were down-regulated in BhHsf1 over-expression Arabidopsis, which was in consistence with the observation of the impaired nuclear endoreduplication. Taking together, our results suggest that BhHsf1 may play dual roles in mediating the processes in heat stress tolerance and growth retardation via regulation of target genes related to stress protection and mitotic cell cycle.

  8. Constitutive androstane receptor activation evokes the expression of glycolytic genes.

    PubMed

    Yarushkin, Andrei A; Kazantseva, Yuliya A; Prokopyeva, Elena A; Markova, Diana N; Pustylnyak, Yuliya A; Pustylnyak, Vladimir O

    2016-09-23

    It is well-known that constitutive androstane receptor (CAR) activation by 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP) increases the liver-to-body weight ratio. CAR-mediated liver growth is correlated with increased expression of the pleiotropic transcription factor cMyc, which stimulates cell cycle regulatory genes and drives proliferating cells into S phase. Because glycolysis supports cell proliferation and cMyc is essential for the activation of glycolytic genes, we hypothesized that CAR-mediated up-regulation of cMyc in mouse livers might play a role in inducing the expression of glycolytic genes. The aim of the present study was to examine the effect of long-term CAR activation on glycolytic genes in a mouse model not subjected to metabolic stress. We demonstrated that long-term CAR activation by TCPOBOP increases expression of cMyc, which was correlated with reduced expression of gluconeogenic genes and up-regulation of glucose transporter, glycolytic and mitochondrial pyruvate metabolising genes. These changes in gene expression after TCPOBOP treatment were strongly correlated with changes in levels of glycolytic intermediates in mouse livers. Moreover, we demonstrated a significant positive regulatory effect of TCPOBOP-activated CAR on both mRNA and protein levels of Pkm2, a master regulator of glucose metabolism and cell proliferation. Thus, our findings provide evidence to support the conclusion that CAR activation initiates a transcriptional program that facilitates the coordinated metabolic activities required for cell proliferation.

  9. [Ectopic parathyroid glands. Imaging methods and surgical access].

    PubMed

    Fialová, M; Adámková, J; Adámek, S; Libánský, P; Kubinyi, J

    2014-08-01

    We discuss the benefits of imaging methods in localizing ectopic parathyroid glands in patients with primary hyperparathyroidism. The ectopic localizations are discussed within the context of the orthotopic norm. In the sample of 123 patients, a 23% rate of ectopic parathyroid glands was detected. Three selected case studies are presented, supporting the benefit of SPECT/CT imaging in terms of surgical access strategy selection.

  10. Molar tubal ectopic pregnancy: Report of two cases.

    PubMed

    Mbarki, Chaouki; Jerbi, Emna; Hsayaoui, Najeh; Zouari, Fatma; Ben Brahim, Ehsen; Oueslati, Hedhili

    2015-06-01

    Ectopic molar pregnancy is a rare occurrence and consequently not often considered as a diagnostic possibility. We report two cases of molar hydatidiform tubal pregnancy. Diagnosis of ectopic pregnancy was confirmed on clinical biological and sonographic investigations. Diagnosis of molar pregnancy was done on histopathology. The clinical course was favorable for both patients. Although rare, molar changes can occur at any site of an ectopic pregnancy. Clinical diagnosis of a molar pregnancy is difficult but histopathology is the gold standard for diagnosis.

  11. Recurrent Ectopic Pregnancy in the Tubal Remnant after Salpingectomy

    PubMed Central

    Samiei-Sarir, Bahareh; Diehm, Christopher

    2013-01-01

    We present two cases of ectopic pregnancy located within the remnant tube following ipsilateral salpingectomy. This particular pathology is rare and yet has significant consequences for the patient, with mortality rates 10–15 times higher than other ectopic pregnancies. It demonstrates that salpingectomy does not exclude ectopic pregnancy on the ipsilateral side. We suggest careful clinical consideration and bring attention to the current surgical technique. PMID:24151570

  12. Active AU Based Patch Weighting for Facial Expression Recognition

    PubMed Central

    Xie, Weicheng; Shen, Linlin; Yang, Meng; Lai, Zhihui

    2017-01-01

    Facial expression has many applications in human-computer interaction. Although feature extraction and selection have been well studied, the specificity of each expression variation is not fully explored in state-of-the-art works. In this work, the problem of multiclass expression recognition is converted into triplet-wise expression recognition. For each expression triplet, a new feature optimization model based on action unit (AU) weighting and patch weight optimization is proposed to represent the specificity of the expression triplet. The sparse representation-based approach is then proposed to detect the active AUs of the testing sample for better generalization. The algorithm achieved competitive accuracies of 89.67% and 94.09% for the Jaffe and Cohn–Kanade (CK+) databases, respectively. Better cross-database performance has also been observed. PMID:28146094

  13. Ectopic histone H3S10 phosphorylation causes chromatin structure remodeling in Drosophila.

    PubMed

    Deng, Huai; Bao, Xiaomin; Cai, Weili; Blacketer, Melissa J; Belmont, Andrew S; Girton, Jack; Johansen, Jørgen; Johansen, Kristen M

    2008-02-01

    Histones are subject to numerous post-translational modifications that correlate with the state of higher-order chromatin structure and gene expression. However, it is not clear whether changes in these epigenetic marks are causative regulatory factors in chromatin structure changes or whether they play a mainly reinforcing or maintenance role. In Drosophila phosphorylation of histone H3S10 in euchromatic chromatin regions by the JIL-1 tandem kinase has been implicated in counteracting heterochromatization and gene silencing. Here we show, using a LacI-tethering system, that JIL-1 mediated ectopic histone H3S10 phosphorylation is sufficient to induce a change in higher-order chromatin structure from a condensed heterochromatin-like state to a more open euchromatic state. This effect was absent when a ;kinase dead' LacI-JIL-1 construct without histone H3S10 phosphorylation activity was expressed. Instead, the 'kinase dead' construct had a dominant-negative effect, leading to a disruption of chromatin structure that was associated with a global repression of histone H3S10 phosphorylation levels. These findings provide direct evidence that the epigenetic histone tail modification of H3S10 phosphorylation at interphase can function as a causative regulator of higher-order chromatin structure in Drosophila in vivo.

  14. Radiologic features of primary intracranial ectopic germinomas

    PubMed Central

    Wei, Xin-Hua; Shen, Hui-Cong; Tang, Shou-Xian; Gao, Cui-Hua; Ren, Ji-Liang; Ai, Lin; Dai, Jian-Ping

    2016-01-01

    Abstract Rationale: Germinomas are sensitive to radiation therapy and chemotherapy; therefore, correct imaging diagnosis is crucial for them. However, the imaging findings of germinomas originating from off-midline regions displayed different patterns from those originating from midline areas. Patient concerns: The objective of this study is to describe the radiologic features of primary ectopic germinoma. We reviewed the MR and CT findings of 12 patients with histologically proven off-midline ectopic germinomas with off-midline locations. Interventions: All of these patients underwent conventional MR images and 3 of them underwent diffusion images. Additional CT images were available in 3 patients. Analysis was focused on the shape and entity of tumors in images, signs of hemiatrophy, and the involvement of fibers in diffusion images. Outcomes: Well-defined (8/12) and ill-defined margin masses (4/12) were identified according to the shape of the mass. Multicystic masses were seen in 11 of the 12 patients. The solid component of the tumors had a high density (3/3) with calcifications (2/3) on CT images, iso- to hypointensity in T2WI (11/12) and restricted diffusion on apparent diffusion coefficient (ADC) maps (3/3). Hemiatrophy was observed in 5 cases and progressive hemiatrophy was observed in 1 case. Other signs included mild peritumoral edema (10/12), and hydrocephalus (7/12). Additionally, infiltration of the corticospinal tract (CST) was identified on diffusion tensor imaging (DTI) (2/2). Lessons: The results indicate that multicysitic entities and hypointensities in solid components on T2WI and hemiatrophy are the imaging features of ectopic germinomas. DTI has potential for assessing CST involvement. PMID:28033250

  15. Submandibular ectopic thyroid with normally located thyroid gland.

    PubMed

    Yılmaz, Mahmut Sinan; Aytürk, Semra; Güven, Mehmet; Dilek, Fatma Hüsniye

    2014-01-01

    Ectopic thyroid is a rare developmental anomaly of the thyroid gland which is defined as the presence of thyroid tissue at a site other than the pretracheal area. Nearly 1 to 3% of all ectopic thyroids are located in the lateral neck. Simultaneous submandibular ectopic thyroid tissue presenting with a functional orthotopic thyroid gland is extremely rare. In this article, we report a 37-year-old female case admitted to our clinic with a complaint of swollen neck in whom ultrasonography revealed submandibular ectopic thyroid tissue presenting with an orthotopic thyroid gland.

  16. Ruptured ectopic pregnancy with a negative urine pregnancy test.

    PubMed

    Hughes, Mallory; Lupo, Andrew; Browning, Adrianne

    2017-01-01

    Ectopic pregnancy is commonly seen as a differential diagnosis of first-trimester vaginal bleeding. Often the diagnosis is made based on a combination of exam findings, transvaginal ultrasound, and a positive pregnancy test. Our case describes a patient with a history of ectopic pregnancy treated with methotrexate and serial human chorionic gonadotropin measurements that were decreasing appropriately. At the time of evaluation, her urine pregnancy test was negative; however, she was confirmed to have a ruptured tubal ectopic pregnancy. This case highlights the variable presentation of ectopic pregnancies and the importance of combining exam findings with ultrasound and laboratory results.

  17. Ruptured ectopic pregnancy with a negative urine pregnancy test

    PubMed Central

    Hughes, Mallory; Lupo, Andrew

    2017-01-01

    Ectopic pregnancy is commonly seen as a differential diagnosis of first-trimester vaginal bleeding. Often the diagnosis is made based on a combination of exam findings, transvaginal ultrasound, and a positive pregnancy test. Our case describes a patient with a history of ectopic pregnancy treated with methotrexate and serial human chorionic gonadotropin measurements that were decreasing appropriately. At the time of evaluation, her urine pregnancy test was negative; however, she was confirmed to have a ruptured tubal ectopic pregnancy. This case highlights the variable presentation of ectopic pregnancies and the importance of combining exam findings with ultrasound and laboratory results. PMID:28127150

  18. Ectopic expression of a phytochrome B gene from Chinese cabbage (Brassica rapa L. ssp. pekinensis) in Arabidopsis thaliana promotes seedling de-etiolation, dwarfing in mature plants, and delayed flowering.

    PubMed

    Song, Mei-Fang; Zhang, Shu; Hou, Pei; Shang, Hong-Zhong; Gu, Hai-Ke; Li, Jing-Juan; Xiao, Yang; Guo, Lin; Su, Liang; Gao, Jian-Wei; Yang, Jian-Ping

    2015-04-01

    Phytochrome B (phyB) is an essential red light receptor that predominantly mediates seedling de-etiolation, shade-avoidance response, and flowering time. In this study, we isolate a full-length cDNA of PHYB, designated BrPHYB, from Chinese cabbage (Brassica rapa L. ssp. pekinensis), and we find that BrphyB protein has high amino acid sequence similarity and the closest evolutionary relationship to Arabidopsis thaliana phyB (i.e., AtphyB). Quantitative reverse transcription (RT)-PCR results indicate that the BrPHYB gene is ubiquitously expressed in different tissues under all light conditions. Constitutive expression of the BrPHYB gene in A. thaliana significantly enhances seedling de-etiolation under red- and white-light conditions, and causes dwarf stature in mature plants. Unexpectedly, overexpression of BrPHYB in transgenic A. thaliana resulted in reduced expression of gibberellins biosynthesis genes and delayed flowering under short-day conditions, whereas AtPHYB overexpression caused enhanced expression of FLOWERING LOCUS T and earlier flowering. Our results suggest that BrphyB might play an important role in regulating the development of Chinese cabbage. BrphyB and AtphyB have conserved functions during de-etiolation and vegetative plant growth and divergent functions in the regulation of flowering time.

  19. Altered glutamyl-aminopeptidase activity and expression in renal neoplasms

    PubMed Central

    2014-01-01

    Background Advances in the knowledge of renal neoplasms have demonstrated the implication of several proteases in their genesis, growth and dissemination. Glutamyl-aminopeptidase (GAP) (EC. 3.4.11.7) is a zinc metallopeptidase with angiotensinase activity highly expressed in kidney tissues and its expression and activity have been associated wtih tumour development. Methods In this prospective study, GAP spectrofluorometric activity and immunohistochemical expression were analysed in clear-cell (CCRCC), papillary (PRCC) and chromophobe (ChRCC) renal cell carcinomas, and in renal oncocytoma (RO). Data obtained in tumour tissue were compared with those from the surrounding uninvolved kidney tissue. In CCRCC, classic pathological parameters such as grade, stage and tumour size were stratified following GAP data and analyzed for 5-year survival. Results GAP activity in both the membrane-bound and soluble fractions was sharply decreased and its immunohistochemical expression showed mild staining in the four histological types of renal tumours. Soluble and membrane-bound GAP activities correlated with tumour grade and size in CCRCCs. Conclusions This study suggests a role for GAP in the neoplastic development of renal tumours and provides additional data for considering the activity and expression of this enzyme of interest in the diagnosis and prognosis of renal neoplasms. PMID:24885240

  20. Induced ectopic expression of HigB toxin in Mycobacterium tuberculosis results in growth inhibition, reduced abundance of a subset of mRNAs and cleavage of tmRNA

    PubMed Central

    Schuessler, Dorothée L; Cortes, Teresa; Fivian-Hughes, Amanda S; Lougheed, Kathryn E A; Harvey, Evelyn; Buxton, Roger S; Davis, Elaine O; Young, Douglas B

    2013-01-01

    In Mycobacterium tuberculosis, the genes Rv1954A–Rv1957 form an operon that includes Rv1955 and Rv1956 which encode the HigB toxin and the HigA antitoxin respectively. We are interested in the role and regulation of this operon, since toxin–antitoxin systems have been suggested to play a part in the formation of persister cells in mycobacteria. To investigate the function of the higBA locus, effects of toxin expression on mycobacterial growth and transcript levels were assessed in M. tuberculosis H37Rv wild type and in an operon deletion background. We show that expression of HigB toxin in the absence of HigA antitoxin arrests growth and causes cell death in M. tuberculosis. We demonstrate HigB expression to reduce the abundance of IdeR and Zur regulated mRNAs and to cleave tmRNA in M. tuberculosis, Escherichia coli and Mycobacterium smegmatis. This study provides the first identification of possible target transcripts of HigB in M. tuberculosis. PMID:23927792

  1. Protein inhibitor of activated STAT3 inhibits adipogenic gene expression

    SciTech Connect

    Deng Jianbei; Hua Kunjie; Caveney, Erica J.; Takahashi, Nobuyuki; Harp, Joyce B. . E-mail: jharp@unc.edu

    2006-01-20

    Protein inhibitor of activated STAT3 (PIAS3), a cytokine-induced repressor of signal transducer and activator of transcription 3 (STAT3) and a modulator of a broad array of nuclear proteins, is expressed in white adipose tissue, but its role in adipogenesis is not known. Here, we determined that PIAS3 was constitutively expressed in 3T3-L1 cells at all stages of adipogenesis. However, it translocated from the nucleus to the cytoplasm 4 days after induction of differentiation by isobutylmethylxanthine, dexamethasone, and insulin (MDI). In ob/ob mice, PIAS3 expression was increased in white adipose tissue depots compared to lean mice and was found in the cytoplasm of adipocytes. Overexpression of PIAS3 in differentiating preadipocytes, which localized primarily to the nucleus, inhibited mRNA level gene expression of adipogenic transcription factors C/EBP{alpha} and PPAR{gamma}, as well as their downstream target genes aP2 and adiponectin. PIAS3 also inhibited C/EBP{alpha} promoter activation mediated specifically by insulin, but not dexamethasone or isobutylmethylxanthine. Taken together, these data suggest that PIAS3 may play an inhibitory role in adipogenesis by modulating insulin-activated transcriptional activation events. Increased PIAS3 expression in adipose tissue may play a role in the metabolic disturbances of obesity.

  2. Expression, purification and characterization of inactive and active forms of ERK2 from insect expression system.

    PubMed

    Yan, Kelly; Merritt, Hanne; Crawford, Kenneth; Pardee, Gwynn; Cheng, Jan Marie; Widger, Stephania; Hekmat-Nejad, Mohammad; Zaror, Isabel; Sim, Janet

    2015-06-01

    Extracellular signal-regulated kinase 2 (ERK2) is a serine/threonine protein kinase involved in many cellular programs, such as cell proliferation, differentiation, motility and programed cell-death. It is therefore considered an important target in the treatment of cancer. In an effort to support biochemical screening and small molecule drug discovery, we established a robust system to generate both inactive and active forms of ERK2 using insect expression system. We report here, for the first time, that inactive ERK2 can be expressed and purified with 100% homogeneity in the unphosphorylated form using insect system. This resulted in a significant 20-fold yield improvement compared to that previously reported using bacterial expression system. We also report a newly developed system to generate active ERK2 in insect cells through in vivo co-expression with a constitutively active MEK1 (S218D S222D). Isolated active ERK2 was confirmed to be doubly phosphorylated at the correct sites, T185 and Y187, in the activation loop of ERK2. Both ERK2 forms, inactive and active, were well characterized by biochemical activity assay for their kinase function. Inactive and active ERK2 were the two key reagents that enabled successful high through-put biochemical assay screen and structural drug discovery studies.

  3. Ectopic hippocampal neurogenesis in adolescent male rats following alcohol dependence.

    PubMed

    McClain, Justin A; Morris, Stephanie A; Marshall, S Alexander; Nixon, Kimberly

    2014-07-01

    The adolescent hippocampus is highly vulnerable to alcohol-induced damage, which could contribute to their increased susceptibility to alcohol use disorder. Altered adult hippocampal neurogenesis represents one potential mechanism by which alcohol (ethanol) affects hippocampal function. Based on the vulnerability of the adolescent hippocampus to alcohol-induced damage, and prior reports of long-term alcohol-induced effects on adult neurogenesis, we predicted adverse effects on adult neurogenesis in the adolescent brain following abstinence from alcohol dependence. Thus, we examined neurogenesis in adolescent male rats during abstinence following a 4-day binge model of alcohol dependence. Bromodeoxyuridine and Ki67 immunohistochemistry revealed a 2.2-fold increase in subgranular zone cell proliferation after 7 days of abstinence. Increased proliferation was followed by a 75% increase in doublecortin expression and a 56% increase in surviving bromodeoxyuridine-labeled cells 14 and 35 days post-ethanol exposure, respectively. The majority of newborn cells in ethanol and control groups co-localized with NeuN, indicating a neuronal phenotype and therefore a 1.6-fold increase in hippocampal neurogenesis during abstinence. Although these results mirror the magnitude of reactive neurogenesis described in adult rat studies, ectopic bromodeoxyuridine and doublecortin positive cells were detected in the molecular layer and hilus of adolescent rats displaying severe withdrawal symptoms, an effect that has not been described in adults. The presence of ectopic neuroblasts suggests that a potential defect exists in the functional incorporation of new neurons into the existing hippocampal circuitry for a subset of rats. Age-related differences in functional incorporation could contribute to the increased vulnerability of the adolescent hippocampus to ethanol.

  4. Ectopic pregnancy treatment by combination therapy

    PubMed Central

    Chudecka-Głaz, Anita; Kuźniak, Sławomir; Menkiszak, Janusz

    2016-01-01

    Abstract Detectability of early stages of ectopic pregnancies has increased due to improvements in ultrasonographic and biochemical techniques. Since the patients’ future procreative plans must be taken into consideration when commencing treatment, the goal of this work was to compare the effects of treatment methods and their impact on fertility. The study included 91 patients treated surgically for ectopic pregnancy. The choice of treatment depended on patients’ general condition, ultrasonographic evaluation and serum level of beta-hCG. A combination of laparoscopic and conservative systemic treatment was applied in 70% of cases. More rapid beta-hCG reduction was noted when laparoscopy and intra-oviductal injection of hyperosmolar glucose or methotrexate (MTX) were combined with intramuscular administration of MTX at a dose of 50 mg/m2. Follow-up examination of 66 patients revealed that the greatest number of spontaneous pregnancies (48%) resulted after this combination therapy. We conclude that this combination treatment is safe and provides satisfactory results in terms of future fertility. PMID:28352846

  5. Laboratory models for studying ectopic pregnancy

    PubMed Central

    Brown, Jeremy K; Horne, Andrew W

    2011-01-01

    Purpose of Review Understanding of the aetiology of tubal ectopic pregnancy (tEP) remains incomplete. We aim to summarize the latest advances in laboratory models of tEP that we believe will, ultimately, contribute to improving the diagnosis and management of the condition. Recent Findings Progress in proteome pre-fractionation and multidimensional protein identification technology has proved particularly effective in identifying novel biomarkers of tEP. These, and related global proteomic and genomic approaches, have as yet to be fully exploited in this context but do have substantial potential to inform future hypothesis driven studies. The majority of data generated since 2009 to explain the aetiology of tEP continues to derive from descriptive human ex-vivo studies. In-vitro models of Fallopian tube ciliary and smooth muscle function have improved to a limited degree, on the back of continuing advances in imaging and data acquisition. We believe that the recent development of a primary human Fallopian tube epithelium culture system represents the most significant recent advance in laboratory models for studying ectopic pregnancy. There remain no good animal models of tEP. Summary The establishment of a viable animal model of tEP remains the key obstacle to a complete understanding the aetiology of the condition. PMID:21666470

  6. Humanized mice with ectopic artificial liver tissues.

    PubMed

    Chen, Alice A; Thomas, David K; Ong, Luvena L; Schwartz, Robert E; Golub, Todd R; Bhatia, Sangeeta N

    2011-07-19

    "Humanized" mice offer a window into aspects of human physiology that are otherwise inaccessible. The best available methods for liver humanization rely on cell transplantation into immunodeficient mice with liver injury but these methods have not gained widespread use due to the duration and variability of hepatocyte repopulation. In light of the significant progress that has been achieved in clinical cell transplantation through tissue engineering, we sought to develop a humanized mouse model based on the facile and ectopic implantation of a tissue-engineered human liver. These human ectopic artificial livers (HEALs) stabilize the function of cryopreserved primary human hepatocytes through juxtacrine and paracrine signals in polymeric scaffolds. In contrast to current methods, HEALs can be efficiently established in immunocompetent mice with normal liver function. Mice transplanted with HEALs exhibit humanized liver functions persistent for weeks, including synthesis of human proteins, human drug metabolism, drug-drug interaction, and drug-induced liver injury. Here, mice with HEALs are used to predict the disproportionate metabolism and toxicity of "major" human metabolites using multiple routes of administration and monitoring. These advances may enable manufacturing of reproducible in vivo models for diverse drug development and research applications.

  7. Transforming Growth Factor β1 (TGF-β1) Activates Hepcidin mRNA Expression in Hepatocytes.

    PubMed

    Chen, Simeng; Feng, Teng; Vujić Spasić, Maja; Altamura, Sandro; Breitkopf-Heinlein, Katja; Altenöder, Jutta; Weiss, Thomas S; Dooley, Steven; Muckenthaler, Martina U

    2016-06-17

    The hepatic hormone hepcidin is the master regulator of systemic iron homeostasis. Its expression level is adjusted to alterations in iron levels, inflammatory cues, and iron requirements for erythropoiesis. Bone morphogenetic protein 6 (BMP6) contributes to the iron-dependent control of hepcidin. In addition, TGF-β1 may stimulate hepcidin mRNA expression in murine hepatocytes and human leukocytes. However, receptors and downstream signaling proteins involved in TGF-β1-induced hepcidin expression are still unclear. Here we show that TGF-β1 treatment of mouse and human hepatocytes, as well as ectopic expression of TGF-β1 in mice, increases hepcidin mRNA levels. The hepcidin response to TGF-β1 depends on functional TGF-β1 type I receptor (ALK5) and TGF-β1 type II receptor (TβRII) and is mediated by a noncanonical mechanism that involves Smad1/5/8 phosphorylation. Interestingly, increasing availability of canonical Smad2/3 decreases TGF-β1-induced hepcidin regulation, whereas the BMP6-hepcidin signal was enhanced, indicating a signaling component stoichiometry-dependent cross-talk between the two pathways. Although ALK2/3-dependent hepcidin activation by BMP6 can be modulated by each of the three hemochromatosis-associated proteins: HJV (hemojuvelin), HFE (hemochromatosis protein), and TfR2 (transferrin receptor 2), these proteins do not control the ALK5-mediated hepcidin response to TGF-β1. TGF-β1 mRNA levels are increased in mouse models of iron overload, indicating that TGF-β1 may contribute to hepcidin synthesis under these conditions. In conclusion, these data demonstrate that a complex regulatory network involving TGF-β1 and BMP6 may control the sensing of systemic and/or hepatic iron levels.

  8. The pituitary V3 vasopressin receptor and the corticotroph phenotype in ectopic ACTH syndrome.

    PubMed

    de Keyzer, Y; Lenne, F; Auzan, C; Jégou, S; René, P; Vaudry, H; Kuhn, J M; Luton, J P; Clauser, E; Bertagna, X

    1996-03-01

    Ectopic ACTH secretion occurs in highly differentiated and rather indolent tumors like bronchial carcinoids or, in contrast, in various types of aggressive and poorly differentiated neuroendocrine tumors. We explored this phenomenon using the recently cloned human pituitary V3 vasopressin receptor as an alternate molecular marker of the corticotroph phenotype. Expression of V3 receptor, corticotrophin releasing hormone (CRH) receptor, and proopiomelanocortin (POMC) genes was examined in tumors of pituitary and nonpituitary origin. A comparative RT-PCR approach revealed signals for both V3 receptor and CHR receptor mRNAs in 17 of 18 ACTH-secreting pituitary adenomas, and 6 of 6 normal pituitaries; in six growth hormone- or prolactin-secreting adenomas, a very faint V3 receptor signal was observed in three cases, and CRH receptor signal was undetected in all. Six of eight bronchial carcinoids responsible for the ectopic ACTH syndrome had both POMC and V3 receptor signals as high as those in ACTH-secreting pituitary adenomas; in contrast, no POMC signal and only a very faint V3 receptor signal were detected in six of eight nonsecreting bronchial carcinoids. Northern blot analysis showed V3 receptor mRNA of identical size in ACTH-secreting bronchial carcinoids and pituitary tumors. Other types of nonpituitary tumors responsible for ectopic ACTH syndrome presented much lower levels of both POMC and V3 receptor gene expression than those found in ACTH-secreting bronchial carcinoids. In contrast with the V3 receptor, CRH receptor mRNA was detected in the majority of neuroendocrine tumors irrespective of their POMC status. These results show that expression of the V3 receptor gene participates in the corticotroph phenotype. Its striking association with ACTH-secreting bronchial carcinoids defines a subset of nonpituitary tumors in which ectopic POMC gene expression is but one aspect of a wider process of corticotroph cell differentiation, and opens new possibilities of

  9. Cloning and Expression of Yak Active Chymosin in Pichia pastoris

    PubMed Central

    Luo, Fan; Jiang, Wei Hua; Yang, Yuan Xiao; Li, Jiang; Jiang, Ming Feng

    2016-01-01

    Rennet, a complex of enzymes found in the stomachs of ruminants, is an important component for cheese production. In our study, we described that yak chymosin gene recombinant Pichia pastoris strain could serve as a novel source for rennet production. Yaks total RNA was extracted from the abomasum of an unweaned yak. The yak preprochymosin, prochymosin, and chymosin genes from total RNA were isolated using gene specific primers based on cattle chymosin gene sequence respectively and analyzed their expression pattern byreal time-polymerase chain reaction. The result showed that the chymosin gene expression level of the sucking yaks was 11.45 times higher than one of adult yaks and yak chymosin belongs to Bovidae family in phylogenetic analysis. To express each, the preprochymosin, prochymosin, and chymosin genes were ligated into the expression vector pPICZαA, respectively, and were expressed in Pichia pastoris X33. The results showed that all the recombinant clones of P. pastoris containing the preprochymosin, prochymosin or chymosin genes could produce the active form of recombinant chymosin into the culture supernatant. Heterologous expressed prochymosin (14.55 Soxhlet unit/mL) had the highest enzyme activity of the three expressed chymosin enzymes. Therefore, we suggest that the yak chymosin gene recombinant Pichia pastoris strain could provide an alternative source of rennet production. PMID:27004812

  10. Expression of soluble and active interferon consensus in SUMO fusion expression system in E. coli.

    PubMed

    Peciak, Karolina; Tommasi, Rita; Choi, Ji-won; Brocchini, Steve; Laurine, Emmanuelle

    2014-07-01

    Protein production can be improved if methods for soluble protein expression are developed. Interferon consensus (IFN-con) is used to treat hepatitis C. IFN-con has superior activity compared to other clinically used interferon α subtypes. However IFN-con is a challenging protein to produce in a soluble form using an Escherichia coli expression system. Here we describe the expression of soluble and active recombinant IFN-con in E. coli. The IFN-con gene sequence was optimised for expression in E. coli, which was then cloned into the Champion™ pET SUMO expression vector downstream of the SUMO fusion protein and under strong T7lac promoter. The SUMO-IFN-con fusion protein was efficiently expressed using the SHuffle™ E. coli strain and existed in soluble form as 86-88% of the total IFN-con. After removal of the SUMO fusion partner, approximately 50mg of recombinant IFN-con of at least 98% purity (by RP-HPLC) was obtained from a 1L fermentation culture. Using an A549/EMCV antiviral assay, the specific activity of the recombinant IFN-con was determined to be 960×10(6) IU/mg as calculated to NIBSC standard for IFN-con (3×10(5)pfu/mL virus titre). Comparison of the antiviral activity of the produced IFN-con to IFN α-2a showed that IFN-con displays 2.8 times greater activity, which is in good agreement with what has been reported in the literature for pure protein. IFN-con expression in a soluble form from E. coli allowed us to use a simple, two-step purification process to yield highly pure and active IFN-con which is more efficient than obtaining IFN-con from inclusion bodies.

  11. Inferior ectopic pupil and typical ocular coloboma in RCS rats.

    PubMed

    Tsuji, Naho; Ozaki, Kiyokazu; Narama, Isao; Matsuura, Tetsuro

    2011-08-01

    Ocular coloboma is sometimes accompanied by corectopia in humans and therefore ectopic pupil may indicate ocular coloboma in experimental animals. The RCS strain of rats has a low incidence of microphthalmia. We found that inferior ectopic pupil is associated exclusively with small-sized eyes in this strain. The objective of the current study was to evaluate whether inferior ectopic pupil is associated with iridal coloboma and other types of ocular coloboma in RCS rats. Both eyes of RCS rats were examined clinically, and those with inferior ectopic pupils underwent morphologic and morphometric examinations. In a prenatal study, coronal serial sections of eyeballs from fetuses at gestational day 16.5 were examined by using light microscopy. Ectopic pupils in RCS rats were found exclusively in an inferior position, where the iris was shortened. Fundic examination revealed severe chorioretinal coloboma in all cases of inferior ectopic pupil. The morphologic characteristics closely resembled those of chorioretinal coloboma in humans. Histopathologic examination of primordia showed incomplete closure of the optic fissure in 4 eyeballs of RCS fetuses. Neither F(1) rats nor N(2) (progeny of RCS × BN matings) displayed any ocular anomalies, including ectopic pupils. The RCS strain is a suitable model for human ocular coloboma, and inferior ectopic pupil appears to be a strong indicator of ocular coloboma.

  12. [Ectopic pancreas mimicking advanced gastric malignancy--case report].

    PubMed

    Zawada, Iwona; Lewosiuk, Agnieszka; Hnatyszyn, Krzysztof; Patalan, Michał; Woyke, Stanisław; Kostyrka, Roman; Marlicz, Krzysztof; Starzyńska, Teresa

    2012-04-01

    Ectopic pancreas is the most common type of ectopic tissue in gastrointestinal tract. It is typically asymptomatic, presenting as a small submucosal lesion in prepyloric region of stomach. The diagnosis is usually incidental, during gastroscopy. The patient with symptomatic heterotropic pancreas, mimicking gastric malignancy was described.

  13. Insertion sequence inversions mediated by ectopic recombination between terminal inverted repeats.

    PubMed

    Ling, Alison; Cordaux, Richard

    2010-12-20

    Transposable elements are widely distributed and diverse in both eukaryotes and prokaryotes, as exemplified by DNA transposons. As a result, they represent a considerable source of genomic variation, for example through ectopic (i.e. non-allelic homologous) recombination events between transposable element copies, resulting in genomic rearrangements. Ectopic recombination may also take place between homologous sequences located within transposable element sequences. DNA transposons are typically bounded by terminal inverted repeats (TIRs). Ectopic recombination between TIRs is expected to result in DNA transposon inversions. However, such inversions have barely been documented. In this study, we report natural inversions of the most common prokaryotic DNA transposons: insertion sequences (IS). We identified natural TIR-TIR recombination-mediated inversions in 9% of IS insertion loci investigated in Wolbachia bacteria, which suggests that recombination between IS TIRs may be a quite common, albeit largely overlooked, source of genomic diversity in bacteria. We suggest that inversions may impede IS survival and proliferation in the host genome by altering transpositional activity. They may also alter genomic instability by modulating the outcome of ectopic recombination events between IS copies in various orientations. This study represents the first report of TIR-TIR recombination within bacterial IS elements and it thereby uncovers a novel mechanism of structural variation for this class of prokaryotic transposable elements.

  14. GILT expression in B cells diminishes cathepsin S steady-state protein expression and activity

    PubMed Central

    Phipps-Yonas, Hannah; Semik, Vikki; Hastings, Karen Taraszka

    2013-01-01

    MHC class II-restricted Ag processing requires protein degradation in the endocytic pathway for the activation of CD4+ T cells. Gamma-interferon-inducible lysosomal thiol reductase (GILT) facilitates Ag processing by reducing protein disulfide bonds in this compartment. Lysosomal cysteine protease cathepsin S (CatS) contains disulfide bonds and mediates essential steps in MHC class II-restricted processing, including proteolysis of large polypeptides and cleavage of the invariant chain. We sought to determine whether GILT’s reductase activity regulates CatS expression and function. Confocal microscopy confirmed that GILT and CatS colocalized within lysosomes of B cells. GILT expression posttranscriptionally decreased the steady-state protein expression of CatS in primary B cells and B-cell lines. GILT did not substantially alter the expression of other lysosomal proteins, including H2-M, H2-O, or CatL. GILT’s reductase active site was necessary for diminished CatS protein levels, and GILT expression decreased the half-life of CatS, suggesting that GILT-mediated reduction of protein disulfide bonds enhances CatS degradation. GILT expression decreased the proteolysis of a CatS selective substrate. This study illustrates a physiologic mechanism that regulates CatS and has implications for fine tuning MHC class II-restricted Ag processing and for the development of CatS inhibitors, which are under investigation for the treatment of autoimmune disease. PMID:23012103

  15. Medullary thyroid carcinoma with ectopic adrenocorticotropic hormone syndrome.

    PubMed

    Choi, Hong Seok; Kim, Min Joo; Moon, Chae Ho; Yoon, Jong Ho; Ku, Ha Ra; Kang, Geon Wook; Na, Im Il; Lee, Seung-Sook; Lee, Byung-Chul; Park, Young Joo; Kim, Hong Il; Ku, Yun Hyi

    2014-03-01

    Ectopic adrenocorticotropic hormone (ACTH) syndrome is caused most frequently by a bronchial carcinoid tumor or by small cell lung cancer. Medullary thyroid carcinoma (MTC) is a rare etiology of ectopic ACTH syndrome. We describe a case of Cushing syndrome due to ectopic ACTH production from MTC in a 48-year-old male. He was diagnosed with MTC 14 years ago and underwent total thyroidectomy, cervical lymph node dissection and a series of metastasectomies. MTC was confirmed by the pathological examination of the thyroid and metastatic mediastinal lymph node tissues. Two years after his last surgery, he developed Cushingoid features, such as moon face and central obesity, accompanied by uncontrolled hypertension and new-onset diabetes. The laboratory results were compatible with ectopic ACTH syndrome. A bilateral adrenalectomy improved the clinical and laboratory findings that were associated with Cushing syndrome. This is the first confirmed case of ectopic ACTH syndrome caused by MTC in Korea.

  16. A pragmatic and evidence-based management of ectopic pregnancy.

    PubMed

    Oron, Galia; Tulandi, Togas

    2013-01-01

    The incidence of ectopic pregnancy is approximately 2% of all pregnancies, and it remains the leading cause of death in early pregnancy. Over 95% of ectopic pregnancies are tubal pregnancies, and the remainders are nontubal pregnancies. The highest risk factor for ectopic pregnancy is a previous tubal pregnancy followed by previous tubal surgery, tubal sterilization, tubal pathology, and current intrauterine device use. The apparent increase in the incidence of nontubal ectopic pregnancy including heterotopic pregnancy may be attributed to the increasing number of pregnancies because of in vitro fertilization treatment. In most cases, an ectopic pregnancy can be treated medically with a single dose of methotrexate. Surgical treatment is still needed in women who are hemodynamically unstable and in those who do not fulfill the criteria for methotrexate treatment. Usually surgical treatment can be performed by laparoscopy and in some cases by hysteroscopy. Laparotomy is rarely needed even in women with intraperitoneal bleeding.

  17. Laparoscope resection of ectopic corticosteroid-secreting adrenal adenoma.

    PubMed

    Wang, Xian-Ling; Dou, Jing-Tao; Gao, Jiang-Ping; Zhong, Wen-Wen; Jin, Du; Hui, Lüzhao; Lu, Ju-Ming; Mu, Yi-Ming

    2012-01-01

    Tumors originating from ectopic adrenal tissue are relatively rare. In this article, we describe a case with Cushing's syndrome caused by an ectopic adrenal adenoma. A 38 year-old male patient presenting with cushingoid appearance for 2 years was diagnosed to have ACTH-independent Cushing's syndrome based on endocrinological evaluation. Mutiple radiological examinations detected bilateral adrenal atrophy. When the images were investigated in a more expanded scope, a 3.0×3.5×5.3 cm mass was detected in the anterior of left renal hilum and left renal vein. The mass was successfully resected with intraoperative endoscopy and pathological evaluation revealed an ectopic adrenal tumor. It is suggested that when the endocrinlogically confirmed adrenal neoplasm could not be well and definitely localized, the possibility of ectopic adrenal should be presumed and further radiography examinations should extend to the field where ectopic adrenal usually presents.

  18. Dual Ectopic Thyroid with Normally Located Thyroid: A Case Report

    PubMed Central

    Kumar Choudhury, Bipul; Kaimal Saikia, Uma; Sarma, Dipti; Saikia, Mihir; Dutta Choudhury, Sarojini; Barua, Santanu; Dewri, Swapna

    2011-01-01

    Dual ectopic thyroid is a rare presentation of thyroid ectopia. Only a few cases have been reported in the world literature. Dual ectopic thyroid in the presence of a normally located thyroid is even rarer. We report a case of dual ectopic thyroid in the lingual and submandibular areas in a seventeen-year-old female with hypoplastic thyroid gland in its normal location. The patient presented with a midline swelling at the base of tongue with dysphagia. Thyroid function test revealed primary hypothyroidism. Ultrasonography of the neck showed hypoplastic thyroid in its normal location. A thyroid scan with Technetium-99 m pertechnate showed two intensely hyperfunctioning foci of ectopic thyroid tissue at a higher level in the midline consistent with dual ectopic thyroid, one at the base of tongue and the other in submental region. No uptake was seen in the normal bed. PMID:21765986

  19. Arabidopsis homeodomain-leucine zipper IV proteins promote stomatal development and ectopically induce stomata beyond the epidermis.

    PubMed

    Peterson, Kylee M; Shyu, Christine; Burr, Christian A; Horst, Robin J; Kanaoka, Masahiro M; Omae, Minami; Sato, Yutaka; Torii, Keiko U

    2013-05-01

    The shoot epidermis of land plants serves as a crucial interface between plants and the atmosphere: pavement cells protect plants from desiccation and other environmental stresses, while stomata facilitate gas exchange and transpiration. Advances have been made in our understanding of stomatal patterning and differentiation, and a set of 'master regulatory' transcription factors of stomatal development have been identified. However, they are limited to specifying stomatal differentiation within the epidermis. Here, we report the identification of an Arabidopsis homeodomain-leucine zipper IV (HD-ZIP IV) protein, HOMEODOMAIN GLABROUS2 (HDG2), as a key epidermal component promoting stomatal differentiation. HDG2 is highly enriched in meristemoids, which are transient-amplifying populations of stomatal-cell lineages. Ectopic expression of HDG2 confers differentiation of stomata in internal mesophyll tissues and occasional multiple epidermal layers. Conversely, a loss-of-function hdg2 mutation delays stomatal differentiation and, rarely but consistently, results in aberrant stomata. A closely related HD-ZIP IV gene, Arabidopsis thaliana MERISTEM LAYER1 (AtML1), shares overlapping function with HDG2: AtML1 overexpression also triggers ectopic stomatal differentiation in the mesophyll layer and atml1 mutation enhances the stomatal differentiation defects of hdg2. Consistently, HDG2 and AtML1 bind the same DNA elements, and activate transcription in yeast. Furthermore, HDG2 transactivates expression of genes that regulate stomatal development in planta. Our study highlights the similarities and uniqueness of these two HD-ZIP IV genes in the specification of protodermal identity and stomatal differentiation beyond predetermined tissue layers.

  20. Role of an expansin-like molecule in Dictyostelium morphogenesis and regulation of its gene expression by the signal transducer and activator of transcription protein Dd-STATa.

    PubMed

    Ogasawara, Shun; Shimada, Nao; Kawata, Takefumi

    2009-02-01

    Expansins are proteins involved in plant morphogenesis, exerting their effects on cellulose to extend cell walls. Dictyostelium is an organism that possesses expansin-like molecules, but their functions are not known. In this study, we analyzed the expL7 (expansin-like 7) gene, which has been identified as a putative target of Dd-STATa, a Dictyostelium homolog of the metazoan signal transducer and activator of transcription (STAT) proteins. Promoter fragments of the expL7 were fused to a lacZ reporter and the expression patterns determined. As expected from the behavior of the endogenous expL7 gene, the expL7/lacZ fusion gene was downregulated in Dd-STATa null slugs. In the parental strain, the expL7 promoter was activated in the anterior tip region. Mutational analysis of the promoter identified a sequence that was necessary for expression in tip cells. In addition, an activator sequence for pstAB cells was identified. These sequences act in combination with the repressor region to prevent ectopic expL7 expression in the prespore and prestalk regions of the slug and culminant. Although the expL7 null mutant showed no phenotypic change, the expL7 overexpressor showed aberrant stalk formation. These results indicate that the expansin-like molecule is important for morphogenesis in Dictyostelium.

  1. Expression of protooncogenes during lymphocyte activation by growth factors.

    PubMed

    Bulanova, E G; Budagyan, V M; Yarilin, A A; Mazurenko, N N

    1997-09-01

    Effects of growth factors of non-immune origin including somatotropin (ST) and platelet-derived growth factor (PDGF) on the expression of the proteins encoded by c-fos, c-myc, c-fun, and c-ets family protooncogenes were studied for the first time. The dynamics of the oncoprotein expression in activated CD(3+)-lymphocytes was investigated by immunoblotting. The accumulation of the Fos and Myc proteins was enhanced in T-lymphocytes treated with ST, PDGF, or phytohemagglutinin; the accumulation was maximum at 30-60 min and decreased in 2 h; the data indicate that the oncoproteins participate in the early lymphocyte activation by various growth factors. The Jun protein appears only in 3 h after the onset of lymphocyte activation; this suggests independent participation of Fos in the early stages of lymphocyte activation prior to the appearance of Jun, preceding the joint action of Fos and Jun within the AP-1 transcription complex. The products of the c-ets family are differentially activated by the studied growth factors. Resting lymphocytes actively accumulate the Ets-1 protein; ST and PDGF activation decreases Ets-1 expression in 2 h. The Ets-2 protein is not detected in resting cells and PDGF-activated lymphocytes, whereas lymphocyte activation by ST is associated with accumulation of Ets-2. The data suggest that the product of the c-ets-1 gene is more important in the regulation of resting cells and the product of the c-ets-2 gene is important during activation of lymphocytes by ST. The results indicate that activation of lymphocytes with growth factors of non-immune origin is mediated by several signal transduction pathways.

  2. Lotus hairy roots expressing inducible arginine decarboxylase activity.

    PubMed

    Chiesa, María A; Ruiz, Oscar A; Sánchez, Diego H

    2004-05-01

    Biotechnological uses of plant cell-tissue culture usually rely on constitutive transgene expression. However, such expression of transgenes may not always be desirable. In those cases, the use of an inducible promoter could be an alternative approach. To test this hypothesis, we developed two binary vectors harboring a stress-inducible promoter from Arabidopsis thaliana, driving the beta-glucuronidase reporter gene and the oat arginine decarboxylase. Transgenic hairy roots of Lotus corniculatus were obtained with osmotic- and cold-inducible beta-glucuronidase and arginine decarboxylase activities. The increase in the activity of the latter was accompanied by a significant rise in total free polyamines level. Through an organogenesis process, we obtained L. corniculatus transgenic plants avoiding deleterious phenotypes frequently associated with the constitutive over-expression of arginine decarboxylation and putrescine accumulation.

  3. Ectopic expression of PgRab7 in rice plants (Oryza sativa L.) results in differential tolerance at the vegetative and seed setting stage during salinity and drought stress.

    PubMed

    Tripathy, Manas Kumar; Tiwari, Budhi Sagar; Reddy, Malireddy K; Deswal, Renu; Sopory, Sudhir K

    2017-01-01

    In this work, we have overexpressed a vesicle trafficking protein, Rab7, from a stress-tolerant plant, Pennisetum glaucum, in a high-yielding but stress-sensitive rice variety Pusa Basmati-1 (PB-1). The transgenic rice plants were tested for tolerance against salinity and drought stress. The transgenic plants showed considerable tolerance at the vegetative stage against both salinity (200 mM NaCl) and drought stress (up to 12 days after withdrawing water). The protection against salt and drought stress may be by regulating Na(+) ion homeostasis, as the transgenic plants showed altered expression of multiple transporter genes, including OsNHX1, OsNHX2, OsSOS1, OsVHA, and OsGLRs. In addition, decreased generation and maintenance of lesser reactive oxygen species (ROS), with maintenance of chloroplast grana and photosynthetic machinery was observed. When evaluated for reproductive growth, 89-96 % of seed setting was maintained in transgenic plants during drought stress; however, under salt stress, a 33-53 % decrease in seed setting was observed. These results indicate that PgRab7 overexpression in rice confers differential tolerance at the seed setting stage during salinity and drought stress and could be a favored target for raising drought-tolerant crops.

  4. The multiple ultrasound patterns of ectopic pregnancy.

    PubMed

    Hourani, Roula; Hachem, Kamal; Haddad-Zebouni, Soha; Mansour, Fersan; Elhage, Abdo; Checrallah, Antoine; Ghossain, Michel A

    2008-01-01

    Ectopic pregnancy (EP) has a variable and misleading clinical presentation contributing to the confusion with medical or other gynecological disorders. The rapid recourse to diagnostic methods, human chorionic gonadotropin (beta-hCG) titers and transvaginal ultrasonography, represents the best approach not only in the early diagnosis but also in the management and monitoring of patients with diagnosed EP. The purpose of this article is to provide a pictorial essay about EP and its multiple ultrasound (US) patterns. We present a large spectrum of EP aspects diagnosed on US and confirmed by pathology. We also review miscellaneous gynecologic diseases that may mimic EP on US. Although endovaginal US combined with quantitative (beta-hCG) analysis is an excellent tool for identifying EP, it may be normal sometimes in early pregnancies. Knowledge of all these patterns is helpful in establishing an early correct diagnosis, therefore leading to elective and conservative management in stable patients and preventing tubal rupture or substantial hemorrhage.

  5. C/EBPa-Mediated Activation of MicroRNAs 34a and 223 Inhibits Lef1 Expression To Achieve Efficient Reprogramming into Macrophages

    PubMed Central

    Rodriguez-Ubreva, Javier; Ciudad, Laura; van Oevelen, Chris; Parra, Maribel; Graf, Thomas

    2014-01-01

    MicroRNAs (miRNAs) exert negative effects on gene expression and influence cell lineage choice during hematopoiesis. C/EBPa-induced pre-B cell-to-macrophage transdifferentiation provides an excellent model to investigate the contribution of miRNAs to hematopoietic cell identity, especially because the two cell types involved fall into separate lymphoid and myeloid branches. In this process, efficient repression of the B cell-specific program is essential to ensure transdifferentation and macrophage function. miRNA profiling revealed that upregulation of miRNAs is highly predominant compared with downregulation and that C/EBPa directly regulates several upregulated miRNAs. We also determined that miRNA 34a (miR-34a) and miR-223 sharply accelerate C/EBPa-mediated transdifferentiation, whereas their depletion delays this process. These two miRNAs affect the transdifferentiation efficiency and activity of macrophages, including their lipopolysaccharide (LPS)-dependent inflammatory response. miR-34a and miR-223 directly target and downregulate the lymphoid transcription factor Lef1, whose ectopic expression delays transdifferentiation to an extent similar to that seen with miR-34a and miR-223 depletion. In addition, ectopic introduction of Lef1 in macrophages causes upregulation of B cell markers, including CD19, Pax5, and Ikzf3. Our report demonstrates the importance of these miRNAs in ensuring the erasure of key B cell transcription factors, such as Lef1, and reinforces the notion of their essential role in fine-tuning the control required for establishing cell identity. PMID:24421386

  6. Linking estrogen receptor β expression with inflammatory bowel disease activity

    PubMed Central

    Pierdominici, Marina; Maselli, Angela; Varano, Barbara; Barbati, Cristiana; Cesaro, Paola; Spada, Cristiano; Zullo, Angelo; Lorenzetti, Roberto; Rosati, Marco; Rainaldi, Gabriella; Limiti, Maria Rosaria; Guidi, Luisa

    2015-01-01

    Crohn disease (CD) and ulcerative colitis (UC) are chronic forms of inflammatory bowel disease (IBD) whose pathogenesis is only poorly understood. Estrogens have a complex role in inflammation and growing evidence suggests that these hormones may impact IBD pathogenesis. Here, we demonstrated a significant reduction (p < 0.05) of estrogen receptor (ER)β expression in peripheral blood T lymphocytes from CD/UC patients with active disease (n = 27) as compared to those in remission (n = 21) and healthy controls (n = 29). Accordingly, in a subgroup of CD/UC patients undergoing to anti-TNF-α therapy and responsive to treatment, ERβ expression was higher (p < 0.01) than that observed in not responsive patients and comparable to that of control subjects. Notably, ERβ expression was markedly decreased in colonic mucosa of CD/UC patients with active disease, reflecting the alterations observed in peripheral blood T cells. ERβ expression inversely correlated with interleukin (IL)-6 serum levels and exogenous exposure of both T lymphocytes and intestinal epithelial cells to this cytokine resulted in ERβ downregulation. These results demonstrate that the ER profile is altered in active IBD patients at both mucosal and systemic levels, at least in part due to IL-6 dysregulation, and highlight the potential exploitation of T cell-associated ERβ as a biomarker of endoscopic disease activity. PMID:26497217

  7. The Saccharomyces cerevisiae RDN1 locus is sequestered from interchromosomal meiotic ectopic recombination in a SIR2-dependent manner.

    PubMed Central

    Davis, E S; Shafer, B K; Strathern, J N

    2000-01-01

    Meiotic ectopic recombination occurs at similar frequencies among many sites in the yeast genome, suggesting that all loci are similarly accessible to homology searching. In contrast, we found that his3 sequences integrated in the RDN1 (rDNA) locus were unusually poor participants in meiotic recombination with his3 sequences at other sites. We show that the low rate of meiotic ectopic recombination resulted from the poor ability of RDN1::his3 to act as a donor sequence. SIR2 partially repressed interchromosomal meiotic ectopic recombination at RDN1, consistent with its role in regulating recombination, gene expression, and retrotransposition within RDN1. We propose that RDN1 is physically sequestered from meiotic homology searching mechanisms. PMID:10880466

  8. Suppression of AKT expression by miR-153 produced anti-tumor activity in lung cancer.

    PubMed

    Yuan, Ye; Du, Weijie; Wang, Ying; Xu, Chaoqian; Wang, Jinghao; Zhang, Yang; Wang, Huimin; Ju, Jiaming; Zhao, Liang; Wang, Zhiguo; Lu, Yanjie; Cai, Benzhi; Pan, Zhenwei

    2015-03-15

    Lung cancer is one of the leading causes of cancer death worldwide. microRNAs have been shown to be a novel class of regulators in lung cancer. Here, we explored the role of miR-153 in the pathogenesis of lung cancer and its therapeutic potential. miR-153 was significantly decreased in lung cancer tissues than the adjacent tissues. The protein and mRNA levels of protein kinase B (AKT), which were shown to promote tumor growth, were both increased in lung cancer tissues than adjacent tissues. Overexpression of miR-153 significantly inhibited AKT protein expression, which were abrogated by co-transfection of AMO-153, the specific inhibitor of miR-153. Luciferase assay showed that transfection of miR-153 markedly suppressed the fluorescent intensity of chimeric vectors carrying the 3'UTR of AKT1, while produced no effect on the mutant construct, indicating that AKT is regulated by miR-153. Overexpression of miR-153 significantly inhibited the proliferation and migration, and promoted apoptosis of cultured lung cancer cells in vitro, and suppressed the growth of xenograft tumors in vivo. Interestingly, lung cancer cells with lower endogenous miR-153 expression are more sensitive to ectopic overexpressed miR-153. The IC50 of miR-153 on lung cancer cells is positive correlated with the endogenous miR-153 level, while negative correlated with AKT level. Knockdown of AKT expression suppressed lung cancer cell proliferation. In summary, miR-153 exerted anti-tumor activity in lung cancer by targeting on AKT. The sensitivity of lung cancer cells to miR-153 is determined by its endogenous miR-153 level.

  9. Nerve injury induces robust allodynia and ectopic discharges in Nav1.3 null mutant mice

    PubMed Central

    Nassar, Mohammed A; Baker, Mark D; Levato, Alessandra; Ingram, Rachel; Mallucci, Giovanna; McMahon, Stephen B; Wood, John N

    2006-01-01

    Changes in sodium channel activity and neuronal hyperexcitability contribute to neuropathic pain, a major clinical problem. There is strong evidence that the re-expression of the embryonic voltage-gated sodium channel subunit Nav1.3 underlies neuronal hyperexcitability and neuropathic pain. Here we show that acute and inflammatory pain behaviour is unchanged in global Nav1.3 mutant mice. Surprisingly, neuropathic pain also developed normally in the Nav1.3 mutant mouse. To rule out any genetic compensation mechanisms that may have masked the phenotype, we investigated neuropathic pain in two conditional Nav1.3 mutant mouse lines. We used Nav1.8-Cre mice to delete Nav1.3 in nociceptors at E14 and NFH-Cre mice to delete Nav1.3 throughout the nervous system postnatally. Again normal levels of neuropathic pain developed after nerve injury in both lines. Furthermore, ectopic discharges from damaged nerves were unaffected by the absence of Nav1.3 in global knock-out mice. Our data demonstrate that Nav1.3 is neither necessary nor sufficient for the development of nerve-injury related pain. PMID:17052333

  10. Drosophila gain-of-function mutant RTK torso triggers ectopic Dpp and STAT signaling.

    PubMed Central

    Li, Jinghong; Li, Willis X

    2003-01-01

    Overactivation of receptor tyrosine kinases (RTKs) has been linked to tumorigenesis. To understand how a hyperactivated RTK functions differently from wild-type RTK, we conducted a genome-wide systematic survey for genes that are required for signaling by a gain-of-function mutant Drosophila RTK Torso (Tor). We screened chromosomal deficiencies for suppression of a gain-of-function mutation tor (tor(GOF)), which led to the identification of 26 genomic regions that, when in half dosage, suppressed the defects caused by tor(GOF). Testing of candidate genes in these regions revealed many genes known to be involved in Tor signaling (such as those encoding the Ras-MAPK cassette, adaptor and structural molecules of RTK signaling, and downstream target genes of Tor), confirming the specificity of this genetic screen. Importantly, this screen also identified components of the TGFbeta (Dpp) and JAK/STAT pathways as being required for Tor(GOF) signaling. Specifically, we found that reducing the dosage of thickveins (tkv), Mothers against dpp (Mad), or STAT92E (aka marelle), respectively, suppressed tor(GOF) phenotypes. Furthermore, we demonstrate that in tor(GOF) embryos, dpp is ectopically expressed and thus may contribute to the patterning defects. These results demonstrate an essential requirement of noncanonical signaling pathways for a persistently activated RTK to cause pathological defects in an organism. PMID:12750336

  11. Ectopic jejunal pacemakers and gastric emptying after Roux gastrectomy: Effect of intestinal pacing

    SciTech Connect

    Karlstrom, L.; Kelly, K.A. )

    1989-11-01

    The aims of this study were to determine whether ectopic pacemakers are present after meals in the Roux limbs of dogs after vagotomy and Roux gastrectomy, whether these pacemakers slow gastric emptying of liquids or solids, and whether abolishing the pacemakers with electric pacing might speed any slow emptying that occurs. In six dogs that underwent vagotomy and Roux gastrectomy and in four dogs that underwent vagotomy and Billroth gastrectomy (controls), myoelectric activity of the Roux limb or duodenum was measured during gastric emptying of a 500 kcal mixed meal of 99mTc-labeled cooked egg and 111In-labeled milk. Roux dogs were tested with and without pacing of the Roux limb. Roux dogs showed ectopic pacemaker in the Roux limb that drove the pacesetter potentials of the limb in a reverse, or orad, direction during 57% of the postprandial recordings. Billroth dogs had no ectopic pacemakers (p less than 0.05). Liquids emptied more slowly in Roux dogs (half-life (t1/2) = 121 +/- 15 minutes) than in Billroth dogs (t1/2 = 43 +/- 9 minutes; p less than 0.05), but solids emptied similarly in both groups of dogs (t1/2 approximately 8 hours). Pacing the Roux limb abolished the ectopic pacemakers, restored the slow emptying of liquids to the more rapid rate found in the Billroth dogs (t1/2: paced Roux, 72 +/- 15 minutes; Billroth, 43 +/- 9 minutes; p greater than 0.05) and did not change emptying of solids. The conclusion was that ectopic pacemakers present in the Roux limb after vagotomy and Roux gastrectomy drove the limb in a reverse direction and slowed emptying of liquids after the operation. The defect was corrected by pacing the Roux limb in a forward direction.

  12. Differences in muscle and adipose tissue gene expression and cardio-metabolic risk factors in the members of physical activity discordant twin pairs.

    PubMed

    Leskinen, Tuija; Rinnankoski-Tuikka, Rita; Rintala, Mirva; Seppänen-Laakso, Tuulikki; Pöllänen, Eija; Alen, Markku; Sipilä, Sarianna; Kaprio, Jaakko; Kovanen, Vuokko; Rahkila, Paavo; Oresic, Matej; Kainulainen, Heikki; Kujala, Urho M

    2010-09-16

    High physical activity/aerobic fitness predicts low morbidity and mortality. Our aim was to identify the most up-regulated gene sets related to long-term physical activity vs. inactivity in skeletal muscle and adipose tissues and to obtain further information about their link with cardio-metabolic risk factors. We studied ten same-sex twin pairs (age range 50-74 years) who had been discordant for leisure-time physical activity for 30 years. The examinations included biopsies from m. vastus lateralis and abdominal subcutaneous adipose tissue. RNA was analyzed with the genome-wide Illumina Human WG-6 v3.0 Expression BeadChip. For pathway analysis we used Gene Set Enrichment Analysis utilizing active vs. inactive co-twin gene expression ratios. Our findings showed that among the physically active members of twin pairs, as compared to their inactive co-twins, gene expression in the muscle tissue samples was chronically up-regulated for the central pathways related to energy metabolism, including oxidative phosphorylation, lipid metabolism and supportive metabolic pathways. Up-regulation of these pathways was associated in particular with aerobic fitness and high HDL cholesterol levels. In fat tissue we found physical activity-associated increases in the expression of polyunsaturated fatty acid metabolism and branched-chain amino acid degradation gene sets both of which associated with decreased 'high-risk' ectopic body fat and plasma glucose levels. Consistent with other findings, plasma lipidomics analysis showed up-regulation of the triacylglycerols containing the polyunsaturated fatty acids. Our findings identified skeletal muscle and fat tissue pathways which are associated with the long-term physical activity and reduced cardio-metabolic disease risk, including increased aerobic fitness. In particular, improved skeletal muscle oxidative energy and lipid metabolism as well as changes in adipocyte function and redistribution of body fat are associated with reduced

  13. Induction of ectopic taste buds by SHH reveals the competency and plasticity of adult lingual epithelium.

    PubMed

    Castillo, David; Seidel, Kerstin; Salcedo, Ernesto; Ahn, Christina; de Sauvage, Frederic J; Klein, Ophir D; Barlow, Linda A

    2014-08-01

    Taste buds are assemblies of elongated epithelial cells, which are innervated by gustatory nerves that transmit taste information to the brain stem. Taste cells are continuously renewed throughout life via proliferation of epithelial progenitors, but the molecular regulation of this process remains unknown. During embryogenesis, sonic hedgehog (SHH) negatively regulates taste bud patterning, such that inhibition of SHH causes the formation of more and larger taste bud primordia, including in regions of the tongue normally devoid of taste buds. Here, using a Cre-lox system to drive constitutive expression of SHH, we identify the effects of SHH on the lingual epithelium of adult mice. We show that misexpression of SHH transforms lingual epithelial cell fate, such that daughter cells of lingual epithelial progenitors form cell type-replete, onion-shaped taste buds, rather than non-taste, pseudostratified epithelium. These SHH-induced ectopic taste buds are found in regions of the adult tongue previously thought incapable of generating taste organs. The ectopic buds are composed of all taste cell types, including support cells and detectors of sweet, bitter, umami, salt and sour, and recapitulate the molecular differentiation process of endogenous taste buds. In contrast to the well-established nerve dependence of endogenous taste buds, however, ectopic taste buds form independently of both gustatory and somatosensory innervation. As innervation is required for SHH expression by endogenous taste buds, our data suggest that SHH can replace the need for innervation to drive the entire program of taste bud differentiation.

  14. Induction of ectopic taste buds by SHH reveals the competency and plasticity of adult lingual epithelium

    PubMed Central

    Castillo, David; Seidel, Kerstin; Salcedo, Ernesto; Ahn, Christina; de Sauvage, Frederic J.; Klein, Ophir D.; Barlow, Linda A.

    2014-01-01

    Taste buds are assemblies of elongated epithelial cells, which are innervated by gustatory nerves that transmit taste information to the brain stem. Taste cells are continuously renewed throughout life via proliferation of epithelial progenitors, but the molecular regulation of this process remains unknown. During embryogenesis, sonic hedgehog (SHH) negatively regulates taste bud patterning, such that inhibition of SHH causes the formation of more and larger taste bud primordia, including in regions of the tongue normally devoid of taste buds. Here, using a Cre-lox system to drive constitutive expression of SHH, we identify the effects of SHH on the lingual epithelium of adult mice. We show that misexpression of SHH transforms lingual epithelial cell fate, such that daughter cells of lingual epithelial progenitors form cell type-replete, onion-shaped taste buds, rather than non-taste, pseudostratified epithelium. These SHH-induced ectopic taste buds are found in regions of the adult tongue previously thought incapable of generating taste organs. The ectopic buds are composed of all taste cell types, including support cells and detectors of sweet, bitter, umami, salt and sour, and recapitulate the molecular differentiation process of endogenous taste buds. In contrast to the well-established nerve dependence of endogenous taste buds, however, ectopic taste buds form independently of both gustatory and somatosensory innervation. As innervation is required for SHH expression by endogenous taste buds, our data suggest that SHH can replace the need for innervation to drive the entire program of taste bud differentiation. PMID:24993944

  15. Prefrontal parvalbumin interneurons shape neuronal activity to drive fear expression.

    PubMed

    Courtin, Julien; Chaudun, Fabrice; Rozeske, Robert R; Karalis, Nikolaos; Gonzalez-Campo, Cecilia; Wurtz, Hélène; Abdi, Azzedine; Baufreton, Jerome; Bienvenu, Thomas C M; Herry, Cyril

    2014-01-02

    Synchronization of spiking activity in neuronal networks is a fundamental process that enables the precise transmission of information to drive behavioural responses. In cortical areas, synchronization of principal-neuron spiking activity is an effective mechanism for information coding that is regulated by GABA (γ-aminobutyric acid)-ergic interneurons through the generation of neuronal oscillations. Although neuronal synchrony has been demonstrated to be crucial for sensory, motor and cognitive processing, it has not been investigated at the level of defined circuits involved in the control of emotional behaviour. Converging evidence indicates that fear behaviour is regulated by the dorsomedial prefrontal cortex (dmPFC). This control over fear behaviour relies on the activation of specific prefrontal projections to the basolateral complex of the amygdala (BLA), a structure that encodes associative fear memories. However, it remains to be established how the precise temporal control of fear behaviour is achieved at the level of prefrontal circuits. Here we use single-unit recordings and optogenetic manipulations in behaving mice to show that fear expression is causally related to the phasic inhibition of prefrontal parvalbumin interneurons (PVINs). Inhibition of PVIN activity disinhibits prefrontal projection neurons and synchronizes their firing by resetting local theta oscillations, leading to fear expression. Our results identify two complementary neuronal mechanisms mediated by PVINs that precisely coordinate and enhance the neuronal activity of prefrontal projection neurons to drive fear expression.

  16. Amphioxus allantoicase: molecular cloning, expression and enzymatic activity.

    PubMed

    Wang, Yongjun; Zhang, Shicui; Liu, Zhenhui; Li, Hongyan; Wang, Lei

    2005-06-01

    Allantoicase, one of the purine metabolism enzymes, is progressively truncated during the chordate evolution, yet it is unknown when its activity became phylogenetically extinct. In this study, a cDNA encoding allantoicase was isolated from the gut cDNA library of amphioxus Branchiostoma belcheri tsingtauense. It is 2441 bp long, and contains an open reading frame encoding a protein of 392 amino acid residues. RT-PCR analysis showed that amphioxus allantoicase was strongly expressed in the hepatic caecum, and weakly expressed in other tissues including hind-gut, gill, muscle, notochord, testis and ovary. The parallel experiment was performed measuring the allantoicase activity in the same tissues revealed that its activity was high in the hepatic caecum, but low or undetectable in other tissues examined. These suggest that allantoicase remains in action in the primitive chordate amphioxus.

  17. LAG3 expression in active Mycobacterium tuberculosis infections.

    PubMed

    Phillips, Bonnie L; Mehra, Smriti; Ahsan, Muhammad H; Selman, Moises; Khader, Shabaana A; Kaushal, Deepak

    2015-03-01

    Mycobacterium tuberculosis (MTB) is a highly successful pathogen because of its ability to persist in human lungs for long periods of time. MTB modulates several aspects of the host immune response. Lymphocyte-activation gene 3 (LAG3) is a protein with a high affinity for the CD4 receptor and is expressed mainly by regulatory T cells with immunomodulatory functions. To understand the function of LAG3 during MTB infection, a nonhuman primate model of tuberculosis, which recapitulates key aspects of natural human infection in rhesus macaques (Macaca mulatta), was used. We show that the expression of LAG3 is highly induced in the lungs and particularly in the granulomatous lesions of macaques experimentally infected with MTB. Furthermore, we show that LAG3 expression is not induced in the lungs and lung granulomas of animals exhibiting latent tuberculosis infection. However, simian immunodeficiency virus-induced reactivation of latent tuberculosis infection results in an increased expression of LAG3 in the lungs. This response is not observed in nonhuman primates infected with non-MTB bacterial pathogens, nor with simian immunodeficiency virus alone. Our data show that LAG3 was expressed primarily on CD4(+) T cells, presumably by regulatory T cells but also by natural killer cells. The expression of LAG3 coincides with high bacterial burdens and changes in the host type 1 helper T-cell response.

  18. Expression and characterization of a Talaromyces marneffei active phospholipase B expressed in a Pichia pastoris expression system

    PubMed Central

    He, Yan; Li, Linghua; Hu, Fengyu; Chen, Wanshan; Lei, Huali; Chen, Xiejie; Cai, Weiping; Tang, Xiaoping

    2016-01-01

    Phospholipase B is a virulence factor for several clinically important pathogenic fungi, including Candida albicans, Cryptococcus neoformans and Aspergillus fumigatus, but its role in the thermally dimorphic fungus Talaromyces marneffei remains unclear. Here, we provide the first report of the expression of a novel phospholipase gene, designated TmPlb1, from T. marneffei in the eukaryotic expression system of Pichia pastoris GS115. Sensitive real-time quantitative reverse-transcription PCR (qRT-PCR) demonstrated that the expression of TmPlb1 increased 1.85-fold in the yeast phase compared with the mycelial phase. TmPlb1 contains an open reading frame (ORF) of 732 bp that encodes a protein of 243 amino acids. The conserved serine, aspartate and histidine catalytic triad and the G-X-S-X-G domain of TmPLB1 provide the structural basis for its molecular activity. The ORF of TmPlb1 was successfully cloned into a pPIC9K vector containing an α-mating factor secretion signal that allowed the secretory expression of TmPLB1 in P. pastoris. The heterologous protein expression began 12 h after methanol induction and peaked at 96 h. Through analysis with SDS–polyacrylamide gel electrophoresis (SDS-PAGE), western blotting and mass spectrometry, we confirmed that TmPLB1 was successfully expressed. Through Ni-affinity chromatography, TmPLB1 was highly purified, and its concentration reached 240.4 mg/L of culture medium. With specific substrates, the phospholipase A1 and phospholipase A2 activities of TmPLB1 were calculated to be 5.96 and 1.59 U/mg, respectively. The high purity and activity of the TmPLB1 obtained here lay a solid foundation for further investigation. PMID:27876784

  19. Variation in stability of endogenous reference genes in fallopian tubes and endometrium from healthy and ectopic pregnant women.

    PubMed

    Gebeh, Alpha K; Marczylo, Emma L; Amoako, Akwasi A; Willets, Jonathon M; Konje, Justin C

    2012-01-01

    RT-qPCR is commonly employed in gene expression studies in ectopic pregnancy. Most use RN18S1, β-actin or GAPDH as internal controls without validation of their suitability as reference genes. A systematic study of the suitability of endogenous reference genes for gene expression studies in ectopic pregnancy is lacking. The aims of this study were therefore to evaluate the stability of 12 reference genes and suggest those that are stable for use as internal control genes in fallopian tubes and endometrium from ectopic pregnancy and healthy non-pregnant controls. Analysis of the results showed that the genes consistently ranked in the top six by geNorm and NormFinder algorithms, were UBC, GAPDH, CYC1 and EIF4A2 (fallopian tubes) and UBC and ATP5B (endometrium). mRNA expression of NAPE-PLD as a test gene of interest varied between the groups depending on which of the 12 reference genes was used as internal controls. This study demonstrates that arbitrary selection of reference genes for normalisation in RT-qPCR studies in ectopic pregnancy without validation, risk producing inaccurate data and should therefore be discouraged.

  20. A missense mutation in the glucosamine-6-phosphate N-acetyltransferase-encoding gene causes temperature-dependent growth defects and ectopic lignin deposition in Arabidopsis.

    PubMed

    Nozaki, Mamoru; Sugiyama, Munetaka; Duan, Jun; Uematsu, Hiroshi; Genda, Tatsuya; Sato, Yasushi

    2012-08-01

    To study the regulatory mechanisms underlying lignin biosynthesis, we isolated and characterized lignescens (lig), a previously undescribed temperature-sensitive mutant of Arabidopsis thaliana that exhibits ectopic lignin deposition and growth defects under high-temperature conditions. The lig mutation was identified as a single base transition in GNA1 encoding glucosamine-6-phosphate N-acetyltransferase (GNA), a critical enzyme of UDP-N-acetylglucosamine (UDP-GlcNAc) biosynthesis. lig harbors a glycine-to-serine substitution at residue 68 (G68S) of GNA1. Enzyme activity assays of the mutant protein (GNA1(G68S)) showed its thermolability relative to the wild-type protein. The lig mutant exposed to the restrictive temperature contained a significantly smaller amount of UDP-GlcNAc than did the wild type. The growth defects and ectopic lignification of lig were suppressed by the addition of UDP-GlcNAc. Since UDP-GlcNAc is an initial sugar donor of N-glycan synthesis and impaired N-glycan synthesis is known to induce the unfolded protein response (UPR), we examined possible relationships between N-glycan synthesis, UPR, and the lig phenotype. N-glycans were reduced and LUMINAL BINDING PROTEIN3, a typical UPR gene, was expressed in lig at the restrictive temperature. Furthermore, treatment with UPR-inducing reagents phenocopied the lig mutant. Our data collectively suggest that impairment of N-glycan synthesis due to a shortage of UDP-GlcNAc leads to ectopic lignin accumulation, mostly through the UPR.

  1. B cell repertoire expansion occurs in meningeal ectopic lymphoid tissue

    PubMed Central

    Lehmann-Horn, Klaus; Wang, Sheng-zhi; Sagan, Sharon A.; Zamvil, Scott S.

    2016-01-01

    Ectopic lymphoid tissues (ELT) can be found in multiple sclerosis (MS) and other organ-specific inflammatory conditions. Whether ELT in the meninges of central nervous system (CNS) autoimmune disease exhibit local germinal center (GC) activity remains unknown. In an experimental autoimmune encephalomyelitis model of CNS autoimmunity, we found activation-induced cytidine deaminase, a GC-defining enzyme, in meningeal ELT (mELT) densely populated by B and T cells. To determine GC activity in mELT, we excised meningeal lymphoid aggregates using laser capture microscopy and evaluated B cell repertoires in mELT and secondary lymphoid organs by next-generation immune repertoire sequencing. We found immunoglobulin heavy chain variable region sequences that were unique to mELT and had accumulated functionally relevant somatic mutations, together indicating localized antigen-driven affinity maturation. Our results suggest that B cells in mELT actively participate in CNS autoimmunity, which may be relevant to mELT in MS and ELT in other chronic inflammatory conditions. PMID:27942581

  2. Prokaryotic expression of a constitutively expressed Tephrosia villosa defensin and its potent antifungal activity.

    PubMed

    Vijayan, S; Guruprasad, Lalitha; Kirti, P B

    2008-10-01

    Plant defensins are small, highly stable, cysteine-rich antimicrobial peptides produced by the plants for inhibiting a broad-spectrum of microbial pathogens. Some of the well-characterized plant defensins exhibit potent antifungal activity on certain pathogenic fungal species only. We characterized a defensin, TvD1 from a weedy leguminous herb, Tephrosia villosa. The open reading frame of the cDNA was 228 bp, which codes for a peptide with 75 amino acids. Expression analyses indicated that this defensin is expressed constitutively in T. villosa with leaf, stem, root, and seed showing almost similar levels of high expression. The recombinant peptide (rTvD1), expressed in the Escherichia coli expression system, exhibited potent in vitro antifungal activity against several filamentous soil-borne fungal pathogens. The purified peptide also showed significant inhibition of root elongation in Arabidopsis seedlings, subsequently affecting the extension of growing root hairs indicating that it has the potential to disturb the plant growth and development.

  3. Fatty acid amide hydrolase ablation promotes ectopic lipid storage and insulin resistance due to centrally mediated hypothyroidism.

    PubMed

    Brown, Whitney H; Gillum, Matthew P; Lee, Hui-Young; Camporez, Joao Paulo G; Zhang, Xian-man; Jeong, Jin Kwon; Alves, Tiago C; Erion, Derek M; Guigni, Blas A; Kahn, Mario; Samuel, Varman T; Cravatt, Benjamin F; Diano, Sabrina; Shulman, Gerald I

    2012-09-11

    Fatty acid amide hydrolase (FAAH) knockout mice are prone to excess energy storage and adiposity, whereas mutations in FAAH are associated with obesity in humans. However, the molecular mechanism by which FAAH affects energy expenditure (EE) remains unknown. Here we show that reduced energy expenditure in FAAH(-/-) mice could be attributed to decreased circulating triiodothyronine and thyroxine concentrations secondary to reduced mRNA expression of both pituitary thyroid-stimulating hormone and hypothalamic thyrotropin-releasing hormone. These reductions in the hypothalamic-pituitary-thyroid axis were associated with activation of hypothalamic peroxisome proliferating-activated receptor γ (PPARγ), and increased hypothalamic deiodinase 2 expression. Infusion of NAEs (anandamide and palmitoylethanolamide) recapitulated increases in PPARγ-mediated decreases in EE. FAAH(-/-) mice were also prone to diet-induced hepatic insulin resistance, which could be attributed to increased hepatic diacylglycerol content and protein kinase Cε activation. Our data indicate that FAAH deletion, and the resulting increases in NAEs, predispose mice to ectopic lipid storage and hepatic insulin resistance by promoting centrally mediated hypothyroidism.

  4. Persistent trophoblastic tissue following salpingostomy for unruptured ectopic pregnancy

    SciTech Connect

    Rivlin, M.E.; Meeks, G.R.; Cowan, B.D.; Bates, G.W.

    1985-02-01

    Radioimmunoassay of beta-hCG was used to diagnose an ectopic pregnancy in a 30 year old patient and the site of pregnancy was determined by ultrasonography. A salpingostomy was performed; the ectopic pregnancy and the residual trophoblastic tissue were removed. Six weeks later a right salpingectomy was performed to remove persistent trophoblastic tissue. Histologic examination of the surgical specimen demonstrated viable chorionic villi. Serial measurements of beta-hCG are recommended following conservative surgery for ectopic gestation to assure the patient and the surgeon that the tube contains no residual products of conception.

  5. Ectopic presacral choroid plexus cyst in a neonate.

    PubMed

    Gross, Eitan; Koplewitz, Benjamin Z; Arbell, Dan; Fellig, Jakob; Udassin, Raphael

    2009-05-01

    An unusual case of a presacral ectopic choroid plexus cyst in a neonate is described. After birth, a soft lump was noticed at the left buttock. Imaging studies including sonography and magnetic resonance imaging demonstrated a presacral cystic lesion extending to the buttocks, composed of several septated cystic masses with no connection to the spinal canal or rectum. After total resection, the tumor was diagnosed as an ectopic choroid plexus cyst. To our knowledge, this is the first case report in the English literature of a presacral ectopic choroid plexus cyst.

  6. Management of Ectopically Erupting Maxillary Incisors: A Case Series

    PubMed Central

    Suresh, Kotumachagi Sangappa; Uma, HL; Nagarathna, J

    2015-01-01

    ABSTRACT Eruption disturbances related to the position include ectopic eruption and transpositions. The occurrence of ectopic eruption is most commonly associated with maxillary incisors. The normal eruption, position and morphology of these teeth are crucial to craniofacial development, facial esthetics as well as phonetics. It is essential that the clinicians have thorough knowledge of the eruption disturbances in order to make an appropriate, as well as timely intervention, as dictated by the complexity of the problem. How to cite this article: Suresh KS, Uma HL, Nagarathna J, Kumar P. Management of Ectopically Erupting Maxillary Incisors: A Case Series. Int J Clin Pediatr Dent 2015;8(3):227-233. PMID:26604543

  7. Management of Ectopically Erupting Maxillary Incisors: A Case Series.

    PubMed

    Suresh, Kotumachagi Sangappa; Uma, H L; Nagarathna, J; Kumar, Pravin

    2015-01-01

    Eruption disturbances related to the position include ectopic eruption and transpositions. The occurrence of ectopic eruption is most commonly associated with maxillary incisors. The normal eruption, position and morphology of these teeth are crucial to craniofacial development, facial esthetics as well as phonetics. It is essential that the clinicians have thorough knowledge of the eruption disturbances in order to make an appropriate, as well as timely intervention, as dictated by the complexity of the problem. How to cite this article: Suresh KS, Uma HL, Nagarathna J, Kumar P. Management of Ectopically Erupting Maxillary Incisors: A Case Series. Int J Clin Pediatr Dent 2015;8(3):227-233.

  8. Papillary Carcinoma in Median Aberrant Thyroid (Ectopic) - Case Report

    PubMed Central

    K, Shashidhar; Deshmane, Vijaya Laxmi; Kumar, Veerendra; Arjunan, Ravi

    2014-01-01

    Median ectopic thyroid may be encountered anywhere from the foramen caecum to the diaphragm. Non lingual median aberrant thyroid (incomplete descent) usually found in the infrahyoid region and malignant transformation in this ectopic thyroid tissue is very rare. We report an extremely rare case of papillary carcinoma in non lingual median aberrant thyroid in a 25-year-old female. The differentiation between a carcinoma arising in the median ectopic thyroid tissue and a metastatic papillary carcinoma from an occult primary in the main thyroid gland is also discussed. PMID:25121039

  9. Diverse intracellular pathogens activate type III interferon expression from peroxisomes.

    PubMed

    Odendall, Charlotte; Dixit, Evelyn; Stavru, Fabrizia; Bierne, Helene; Franz, Kate M; Durbin, Ann Fiegen; Boulant, Steeve; Gehrke, Lee; Cossart, Pascale; Kagan, Jonathan C

    2014-08-01

    Type I interferon responses are considered the primary means by which viral infections are controlled in mammals. Despite this view, several pathogens activate antiviral responses in the absence of type I interferons. The mechanisms controlling type I interferon-independent responses are undefined. We found that RIG-I like receptors (RLRs) induce type III interferon expression in a variety of human cell types, and identified factors that differentially regulate expression of type I and type III interferons. We identified peroxisomes as a primary site of initiation of type III interferon expression, and revealed that the process of intestinal epithelial cell differentiation upregulates peroxisome biogenesis and promotes robust type III interferon responses in human cells. These findings highlight the importance of different intracellular organelles in specific innate immune responses.

  10. Activating frataxin expression by repeat-targeted nucleic acids

    PubMed Central

    Li, Liande; Matsui, Masayuki; Corey, David R.

    2016-01-01

    Friedreich's ataxia is an incurable genetic disorder caused by a mutant expansion of the trinucleotide GAA within an intronic FXN RNA. This expansion leads to reduced expression of frataxin (FXN) protein and evidence suggests that transcriptional repression is caused by an R-loop that forms between the expanded repeat RNA and complementary genomic DNA. Synthetic agents that increase levels of FXN protein might alleviate the disease. We demonstrate that introducing anti-GAA duplex RNAs or single-stranded locked nucleic acids into patient-derived cells increases FXN protein expression to levels similar to analogous wild-type cells. Our data are significant because synthetic nucleic acids that target GAA repeats can be lead compounds for restoring curative FXN levels. More broadly, our results demonstrate that interfering with R-loop formation can trigger gene activation and reveal a new strategy for upregulating gene expression. PMID:26842135

  11. Humanizing the Protease-Activated Receptor (PAR) Expression Profile in Mouse Platelets by Knocking PAR1 into the Par3 Locus Reveals PAR1 Expression Is Not Tolerated in Mouse Platelets

    PubMed Central

    French, Shauna L.; Paramitha, Antonia C.; Moon, Mitchell J.; Dickins, Ross A.; Hamilton, Justin R.

    2016-01-01

    Anti-platelet drugs are the mainstay of pharmacotherapy for heart attack and stroke prevention, yet improvements are continually sought. Thrombin is the most potent activator of platelets and targeting platelet thrombin receptors (protease-activated receptors; PARs) is an emerging anti-thrombotic approach. Humans express two PARs on their platelets–PAR1 and PAR4. The first PAR1 antagonist was recently approved for clinical use and PAR4 antagonists are in early clinical development. However, pre-clinical studies examining platelet PAR function are challenging because the platelets of non-primates do not accurately reflect the PAR expression profile of human platelets. Mice, for example, express Par3 and Par4. To address this limitation, we aimed to develop a genetically modified mouse that would express the same repertoire of platelet PARs as humans. Here, human PAR1 preceded by a lox-stop-lox was knocked into the mouse Par3 locus, and then expressed in a platelet-specific manner (hPAR1-KI mice). Despite correct targeting and the predicted loss of Par3 expression and function in platelets from hPAR1-KI mice, no PAR1 expression or function was detected. Specifically, PAR1 was not detected on the platelet surface nor internally by flow cytometry nor in whole cell lysates by Western blot, while a PAR1-activating peptide failed to induce platelet activation assessed by either aggregation or surface P-selectin expression. Platelets from hPAR1-KI mice did display significantly diminished responsiveness to thrombin stimulation in both assays, consistent with a Par3-/- phenotype. In contrast to the observations in hPAR1-KI mouse platelets, the PAR1 construct used here was successfully expressed in HEK293T cells. Together, these data suggest ectopic PAR1 expression is not tolerated in mouse platelets and indicate a different approach is required to develop a small animal model for the purpose of any future preclinical testing of PAR antagonists as anti-platelet drugs. PMID

  12. Aryl hydrocarbon receptor activity modulates prolactin expression in the pituitary

    PubMed Central

    Moran, Tyler B.; Brannick, Katherine E.; Raetzman, Lori T.

    2012-01-01

    Pituitary tumors account for 15% of intracranial neoplasms, however the extent to which environmental toxicants contribute to the proliferation and hormone expression of pituitary cells is unknown. Aryl-hydrocarbon receptor (AhR) interacting protein (AIP) loss of function mutations cause somatotroph and lactotroph adenomas in humans. AIP sequesters AhR and inhibits its transcriptional function. Because of the link between AIP and pituitary tumors, we hypothesize that exposure to dioxins, potent exogenous ligands for AhR that are persistent in the environment, may predispose to pituitary dysfunction through activation of AhR. In the present study, we examined the effect of AhR activation on proliferation and endogenous pituitary hormone expression in the GH3 rat somato-lactotrope tumor cell line and the effect of loss of AhR action in knockout mice. GH3 cells respond to nM doses of the reversible AhR agonist β-naphthoflavone with a robust induction of Cyp1a1. Although mRNA levels of the anti-proliferative signaling cytokine TGFbeta1 are suppressed upon β-naphthoflavone treatment, we did not observe an alteration in cell proliferation. AhR activation with β-naphthoflavone suppresses Ahr expression and impairs expression of prolactin (PRL), but not growth hormone (GH) mRNA in GH3 cells. In mice, loss of Ahr similarly leads to a reduction in Prl mRNA at P3, while Gh is unaffected. Additionally, there is a significant reduction pituitary hormones Lhb and Fshb in the absence of Ahr. Overall, these results demonstrate that AhR is important for pituitary hormone expression and suggests environmental dioxins can exert endocrine disrupting effects at the pituitary. PMID:22975028

  13. Adaptation of muscle gene expression to changes in contractile activity

    NASA Technical Reports Server (NTRS)

    Booth, F. W.; Babij, P.; Thomason, D. B.; Wong, T. S.; Morrison, P. R.

    1987-01-01

    A review of the existing literature regarding the effects of different types of physical activities on the gene expression of adult skeletal muscles leads us to conclude that each type of exercise training program has, as a result, a different phenotype, which means that there are multiple mechanisms, each producing a unique phenotype. A portion of the facts which support this position is presented and interpreted here. [Abstract translated from the original French by NASA].

  14. Ectopically tethered CP190 induces large-scale chromatin decondensation

    NASA Astrophysics Data System (ADS)

    Ahanger, Sajad H.; Günther, Katharina; Weth, Oliver; Bartkuhn, Marek; Bhonde, Ramesh R.; Shouche, Yogesh S.; Renkawitz, Rainer

    2014-01-01

    Insulator mediated alteration in higher-order chromatin and/or nucleosome organization is an important aspect of epigenetic gene regulation. Recent studies have suggested a key role for CP190 in such processes. In this study, we analysed the effects of ectopically tethered insulator factors on chromatin structure and found that CP190 induces large-scale decondensation when targeted to a condensed lacO array in mammalian and Drosophila cells. In contrast, dCTCF alone, is unable to cause such a decondensation, however, when CP190 is present, dCTCF recruits it to the lacO array and mediates chromatin unfolding. The CP190 induced opening of chromatin may not be correlated with transcriptional activation, as binding of CP190 does not enhance luciferase activity in reporter assays. We propose that CP190 may mediate histone modification and chromatin remodelling activity to induce an open chromatin state by its direct recruitment or targeting by a DNA binding factor such as dCTCF.

  15. Expression and activation of proteases in co-cultures.

    PubMed

    Paduch, Roman; Kandefer-Szerszeń, Martyna

    2011-01-01

    The present study concerned the expression and activation of metalloproteinase-2 (MMP-2), metalloproteinase-9 (MMP-9) and the urokinase plasminogen activator/urokinase plasminogen activator receptor (uPA/uPAR) system in co-cultures of human colon carcinoma cell spheroids (HT29, LS180, SW948) with human normal colon epithelium (CCD 841 CoTr), myofibroblasts (CCD-18Co) and endothelial cells (HUVEC). Additionally, the influence of monensin on the production and function of the proteases was tested. Tumor cells expressed small amounts of MMP-2, MMP-9 and uPA. Normal cells generally produced proportionally higher concentrations of these proteases (especially MMP-2, compared with significantly smaller yields of MMP-9 and significantly lower amounts of uPAR than tumors. In co-cultures of tumor spheroids with normal cell monolayers, the concentration of the proteases was equal to the sum of the enzymes produced in monocultures of both types of cells. The highest activity of uPA, measured as the reduction of the chromogenic substrate (S-2444), was detected in supernatants and lysates of endothelial cells. Interestingly, in normal cells, the higher expression of proteases, mainly uPA, measured as the level of protein concentration, was closely linked with their lower activity and inversely, in tumor cells, the low level of the expression of the enzymes correlated with their high enzymatic activity. In zymography analysis, mainly pro-MMPs were detected both in culture supernatants and cell lysates. The highest amounts of active forms of the MMPs were detected in tumor spheroids co-cultured with endothelial cells. Monensin inhibited MMPs and uPA secretion but significantly increased uPAR release, mainly from normal cells. In conclusion, during direct interactions of tumor cells with normal cells, MMPs and the uPA/uPAR system play an important role in the degradation of ECM and tumor development, but as we found, there is a reverse relationship between the concentration and the

  16. Expression of biologically active murine interleukin-18 in Lactococcus lactis.

    PubMed

    Feizollahzadeh, Sadegh; Khanahmad, Hossein; Rahimmanesh, Ilnaz; Ganjalikhani-Hakemi, Mazdak; Andalib, Alireza; Sanei, Mohammad Hossein; Rezaei, Abbas

    2016-11-01

    The food-grade bacterium Lactococcus lactis is increasingly used for heterologous protein expression in therapeutic and industrial applications. The ability of L. lactis to secrete biologically active cytokines may be used for the generation of therapeutic cytokines. Interleukin (IL)-18 enhances the immune response, especially on mucosal surfaces, emphasizing its therapeutic potential. However, it is produced as an inactive precursor and has to be enzymatically cleaved for maturation. We genetically manipulated L. lactis to secrete murine IL-18. The mature murine IL-18 gene was inserted downstream of a nisin promoter in pNZ8149 plasmid and the construct was used to transform L. lactis NZ3900. The transformants were selected on Elliker agar and confirmed by restriction enzyme digestion and sequencing. The expression and secretion of IL-18 protein was verified by SDS-PAGE, western blotting and ELISA. The biological activity of recombinant IL-18 was determined by its ability to induce interferon (IFN)-γ production in L. lactis co-cultured with murine splenic T cells. The amounts of IL-18 in bacterial lysates and supernatants were 3-4 μg mL(-1) and 0.6-0.7 ng mL(-1), respectively. The successfully generated L. lactis strain that expressed biologically active murine IL-18 can be used to evaluate the possible therapeutic effects of IL-18 on mucosal surfaces.

  17. Human airway epithelia express catalytically active NEU3 sialidase.

    PubMed

    Lillehoj, Erik P; Hyun, Sang Won; Feng, Chiguang; Zhang, Lei; Liu, Anguo; Guang, Wei; Nguyen, Chinh; Sun, Wenji; Luzina, Irina G; Webb, Tonya J; Atamas, Sergei P; Passaniti, Antonino; Twaddell, William S; Puché, Adam C; Wang, Lai-Xi; Cross, Alan S; Goldblum, Simeon E

    2014-05-01

    Sialic acids on glycoconjugates play a pivotal role in many biological processes. In the airways, sialylated glycoproteins and glycolipids are strategically positioned on the plasma membranes of epithelia to regulate receptor-ligand, cell-cell, and host-pathogen interactions at the molecular level. We now demonstrate, for the first time, sialidase activity for ganglioside substrates in human airway epithelia. Of the four known mammalian sialidases, NEU3 has a substrate preference for gangliosides and is expressed at mRNA and protein levels at comparable abundance in epithelia derived from human trachea, bronchi, small airways, and alveoli. In small airway and alveolar epithelia, NEU3 protein was immunolocalized to the plasma membrane, cytosolic, and nuclear subcellular fractions. Small interfering RNA-induced silencing of NEU3 expression diminished sialidase activity for a ganglioside substrate by >70%. NEU3 immunostaining of intact human lung tissue could be localized to the superficial epithelia, including the ciliated brush border, as well as to nuclei. However, NEU3 was reduced in subepithelial tissues. These results indicate that human airway epithelia express catalytically active NEU3 sialidase.

  18. Expression and Activity of a Novel Cathelicidin from Domestic Cats

    PubMed Central

    Leonard, Brian C.; Chu, Hiutung; Johns, Jennifer L.; Gallo, Richard L.; Moore, Peter F.; Marks, Stanley L.; Bevins, Charles L.

    2011-01-01

    Cathelicidins are small cationic antimicrobial peptides found in many species including primates, mammals, marsupials, birds and even more primitive vertebrates, such as the hagfish. Some animals encode multiple cathelicidins in their genome, whereas others have only one. This report identifies and characterizes feline cathelicidin (feCath) as the sole cathelicidin in domestic cats (Felis catus). Expression of feCath is predominantly found in the bone marrow, with lower levels of expression in the gastrointestinal tract and skin. By immunocytochemistry, feCath localizes to the cytoplasm of neutrophils in feline peripheral blood. Structurally, the mature feCath sequence is most similar to a subgroup of cathelicidins that form linear α-helices. feCath possesses antimicrobial activity against E. coli D31, Salmonella enterica serovar Typhimurium (IR715), Listeria monocytogenes and Staphylococcus pseudintermedius (clinical isolate) similar to that of the human ortholog, LL-37. In contrast, feCath lacks the DNA binding activity seen with LL-37. Given its similarity in sequence, structure, tissue expression, and antimicrobial activity, the cathelicidin encoded by cats, feCath, belongs to the subgroup of linear cathelicidins found not only in humans, but also non-human primates, dogs, mice, and rats. PMID:21533281

  19. Genetic Characterization of the Homeodomain-Independent Activity of the Drosophila Fushi Tarazu Gene Product

    PubMed Central

    Hyduk, D.; Percival-Smith, A.

    1996-01-01

    The gene product of fushi tarazu (FTZ) has a homeodomain (HD)-independent activity. Ectopic expression of a FTZ protein that lacks half the HD in embryos results in the anti-ftz phenotype. We have characterized this FTZ HD-independent activity further. Ectopic expression of the HD-independent FTZ activity, in the absence of FTZ activity expressed from the endogenous ftz gene, was sufficient to result in the anti-ftz phenotype. Since the anti-ftz phenotype is a first instar larvae composed nearly entirely of FTZ-dependent cuticular structures derived from the even-numbered parasegments, this result suggests that expression of the HD-independent FTZ activity is sufficient to establish FTZ-dependent cuticle. Activation of FTZ-dependent Engrailed (EN) expression and activation of the ftz enhancer were HD-independent. The ftz enhancer element, AE-1, was activated by the HD-independent FTZ activity; however, the ftz enhancer element, AE-BS2CCC, which is the same as AE-1 except for the inactivation of two FTZ HD DNA-binding sites, was not. Activation of the ftz enhancer by ectopic expression of FTZ activity was effective only during gastrulation and germ band extension. In the discussion, we propose an explanation for these results. PMID:8852847

  20. Twin tubal pregnancy: A large unruptured ectopic pregnancy.

    PubMed

    Goswami, Deepti; Agrawal, Nidhi; Arora, Vrinda

    2015-11-01

    Twin ectopic pregnancy is a rare occurrence, with an estimated incidence of 1 in 20 000 spontaneous pregnancies. We describe a case of unilateral twin ectopic pregnancy in which the gravid fallopian tube showed no signs of tubal rupture despite marked tubal distension. A 25-year-old woman presented with clinical features suggestive of large right-sided tubal ectopic pregnancy. Serum β-human chorionic gonadotropin was 10 800 IU/mL. Laparotomy revealed markedly distended right fallopian tube. There was no hemoperitoneum. The tube contained twin gestational sacs. The crown-rump length of the embryos was 2 cm. The ectopic gestation was thus unique, in that despite marked tubal distension, the trophoblastic invasion was not significant to cause tubal rupture. There may be a role for medical management based on individual gestational sac size in selected cases of twin tubal pregnancy in which there is no evidence of hemoperitoneum.

  1. RCP induces Slug expression and cancer cell invasion by stabilizing β1 integrin.

    PubMed

    Hwang, M H; Cho, K H; Jeong, K J; Park, Y-Y; Kim, J M; Yu, S-L; Park, C G; Mills, G B; Lee, H Y

    2017-02-23

    Rab coupling protein (RCP)-induced tumor cell migration has been implicated in tumor pathophysiology and patient outcomes. In the present study, we demonstrate that RCP stabilizes β1 integrin leading to increased β1 integrin levels and activation of a signaling cascade culminating in Slug induction, epithelial-to-mesenchymal transition and increased invasion. Ectopic expression of RCP induced Slug expression. Silencing β1 integrin efficiently inhibited RCP-induced Slug expression and subsequent cancer cell invasion. Conversely, ectopic expression of β1 integrin was sufficient to induce Slug expression. Pharmacological inhibition of integrin linked kinase (ILK), EGFR and NF-κB, as well as transfection of a dominant-negative mutant of Ras (RasN17), significantly inhibited RCP-induced Slug expression and cancer cell invasion. Strikingly, ectopic expression of RCP was sufficient to enhance metastasis of ovarian cancer cells to the lung. Collectively, we demonstrate a mechanism by which RCP promotes cancer cell aggressiveness through sequential β1 integrin stabilization, activation of an ILK/EGFR/Ras/NF-κB signaling cascade and subsequent Slug expression.

  2. A case of an ectopic prolactinoma.

    PubMed

    Simsir, Ilgin Yildirim; Kocabas, Gokcen Unal; Sahin, Serap Baydur; Erdogan, Mehmet; Cetinkalp, Sevki; Saygili, Fusun; Yilmaz, Candeger; Ozgen, Ahmet Gokhan

    2012-02-01

    A 34-year-old female presented to our clinic with a 1.5 year history of secondary amenorrhea and galactorrhea. Prolactin (PRL) level was found to be 151.89 ng/ml. Pituitary imaging was reported to be normal. An examination of the patient revealed that PRL level was still high so the dose of cabergoline was further increased and subsequently, bromocriptine was added to the treatment. There was no reduction in PRL levels in controls. A scanning was performed to look for an ectopic focus. Abdominal computerized tomography revealed a heterogenous mass lesion originating from the uterus. Octreotide scintigraphy was performed and we observed an involvement consistent with the mass in the uterus. The patient underwent abdominal total hysterectomy. PRL dropped to 0.4 ng/ml the next day after the operation. The pathology result was a low-grade malignant mesenchymal tumor. Prolactin was found to be immunohistochemically negative. However, galactorrhea disappeared postoperative and PRL levels are still low. Elevated levels of PRL, resistant to bromocriptine and cabergoline, rapidly returned to normal after hysterectomy, which obviously indicates that hyperprolactinemia was associated with the myoma of the uterus.

  3. Diagnosis and management of intramural ectopic pregnancy.

    PubMed

    Bannon, Kimberly; Fernandez, Carlos; Rojas, David; Levine, Elliot M; Locher, Stephen

    2013-01-01

    Intramural pregnancy, a gestation completely surrounded by the myometrium located within the uterine wall with separation from the uterine cavity, is an extremely unusual form of pregnancy. Complications resulting from intramural pregnancy include inevitable uterine rupture with resultant hemorrhage and possible hysterectomy if diagnosis is not made early and treatment is not initiated. An asymptomatic patient was initially diagnosed with a missed abortion at approximately 6 weeks of gestation after a routine ultrasound. Suction curettage was performed approximately 1 month after the initial diagnosis. The pathology specimen failed to reveal placental villi. Ultimately, the diagnosis of intramural pregnancy was made via ultrasound and a computed tomography scan. She was treated with a single dose of systemically administered methotrexate. Over a period of 4 months, β-human chorionic gonadotropin levels trended downward; however, the intramural pregnancy failed to resolve completely, and a persistent mass remained. The intramural pregnancy was removed using the da Vinci laparoscopic procedure (Intuitive Surgical, Sunnyvale, CA). If the diagnosis of intramural ectopic pregnancy is made sufficiently early, conservative measures can be taken, which can preserve a patient's future fertility.

  4. MAPK Phosphatase AP2C3 Induces Ectopic Proliferation of Epidermal Cells Leading to Stomata Development in Arabidopsis

    PubMed Central

    Kazanaviciute, Vaiva; Magyar, Zoltan; Ayatollahi, Zahra; Unterwurzacher, Verena; Choopayak, Chonnanit; Boniecka, Justyna; Murray, James A. H.; Bogre, Laszlo; Meskiene, Irute

    2010-01-01

    In plant post-embryonic epidermis mitogen-activated protein kinase (MAPK) signaling promotes differentiation of pavement cells and inhibits initiation of stomata. Stomata are cells specialized to modulate gas exchange and water loss. Arabidopsis MAPKs MPK3 and MPK6 are at the core of the signaling cascade; however, it is not well understood how the activity of these pleiotropic MAPKs is constrained spatially so that pavement cell differentiation is promoted only outside the stomata lineage. Here we identified a PP2C-type phosphatase termed AP2C3 (Arabidopsis protein phosphatase 2C) that is expressed distinctively during stomata development as well as interacts and inactivates MPK3, MPK4 and MPK6. AP2C3 co-localizes with MAPKs within the nucleus and this localization depends on its N-terminal extension. We show that other closely related phosphatases AP2C2 and AP2C4 are also MAPK phosphatases acting on MPK6, but have a distinct expression pattern from AP2C3. In accordance with this, only AP2C3 ectopic expression is able to stimulate cell proliferation leading to excess stomata development. This function of AP2C3 relies on the domains required for MAPK docking and intracellular localization. Concomitantly, the constitutive and inducible AP2C3 expression deregulates E2F-RB pathway, promotes the abundance and activity of CDKA, as well as changes of CDKB1;1 forms. We suggest that AP2C3 downregulates the MAPK signaling activity to help maintain the balance between differentiation of stomata and pavement cells. PMID:21203456

  5. [Bilateral tubal ectopic pregnancy in a spontaneous cycle--a case report].

    PubMed

    Kubiaczyk, Filip; Suchocki, Sławomir; Puskarz, Roman; Zaborowski, Adam; Zaranek, Krzysztof

    2014-08-01

    Ectopic pregnancy occurs in 0.5-2% of all pregnancies. A double ectopic pregnancy is very rare and has the incidence of 1 in 725 ectopic pregnancies. We describe a case of a 30-year-old patient with double ectopic pregnancy located in tubas in spontaneous cycle--diagnosed and treated at the Gynecology and Obstetrics Hospital in Walbrzych. The patient had no risk factors for ectopic pregnancy. This case shows that even the diagnosis of an evident ectopic pregnancy cannot exclude the possibility of a double ectopic pregnancy.

  6. Pharmacological and Genetic Modulation of REV-ERB Activity and Expression Affects Orexigenic Gene Expression

    PubMed Central

    Amador, Ariadna; Wang, Yongjun; Banerjee, Subhashis; Kameneka, Theodore M.; Solt, Laura A.; Burris, Thomas P.

    2016-01-01

    The nuclear receptors REV-ERBα and REV-ERBβ are transcription factors that play pivotal roles in the regulation of the circadian rhythm and various metabolic processes. The circadian rhythm is an endogenous mechanism, which generates entrainable biological changes that follow a 24-hour period. It regulates a number of physiological processes, including sleep/wakeful cycles and feeding behaviors. We recently demonstrated that REV-ERB-specific small molecules affect sleep and anxiety. The orexinergic system also plays a significant role in mammalian physiology and behavior, including the regulation of sleep and food intake. Importantly, orexin genes are expressed in a circadian manner. Given these overlaps in function and circadian expression, we wanted to determine whether the REV-ERBs might regulate orexin. We found that acute in vivo modulation of REV-ERB activity, with the REV-ERB-specific synthetic ligand SR9009, affects the circadian expression of orexinergic genes in mice. Long term dosing with SR9009 also suppresses orexinergic gene expression in mice. Finally, REV-ERBβ-deficient mice present with increased orexinergic transcripts. These data suggest that the REV-ERBs may be involved in the repression of orexinergic gene expression. PMID:26963516

  7. A rare case of ectopic thoracic kidney and spleen.

    PubMed

    Krishnan, Babu; Lakshminarayanan, P; Sudhakar, M K; Vishwanathan, K N

    2011-07-01

    A 23-year-old male presented with fever of 5 days duration. His peripheral smear was positive for Plasmodium vivax. He was treated for malaria and responded. During investigation with USG it was found that he had absent left kidney. An abdominal contrast enhanced CT scan revealed ectopic kidney and spleen in the left hemithorax. This was a rare case of ectopic thoracic kidney and spleen.

  8. COX-2 gene promoter DNA methylation status in eutopic and ectopic endometrium of Egyptian women with endometriosis.

    PubMed

    Zidan, Haidy E; Rezk, Noha A; Alnemr, Amr Abd Almohsen; Abd El Ghany, Amany M

    2015-11-01

    The pathophysiology of COX-2 expression in endometriosis is a matter of debate. The aim was to investigate the role of DNA methylation of the NF-IL6 site within the promoter of COX-2 gene in the pathogenesis of endometriosis. The endometrial tissues (ectopic and eutopic) were collected from 60 women with endometriosis and 30 women without endometriosis (control group). The methylation status of COX-2 was examined by methylation-specific PCR. Quantitative real-time PCR (RT-PCR) was performed to measure COX-2 mRNA levels in endometrial tissues. We found significantly higher levels of COX-2 in ectopic endometriotic tissue compared with eutopic tissue. Also, we found that the frequencies of methylation status of the NF-IL6 site within the COX-2 promoter in the eutopic and ectopic endometrial tissues of endometriosis groups were significantly decreased in comparison to controls (P=0.002, P=0.000 respectively). Our study demonstrated that DNA hypomethylation of the NF-IL6 site within the promoter of COX-2 gene could be a key mechanism for its elevated expression in the eutopic and ectopic tissues of endometriosis.

  9. Ectopic Granule Cells of the Rat Dentate Gyrus

    PubMed Central

    Scharfman, Helen; Goodman, Jeffrey; McCloskey, Daniel

    2007-01-01

    Granule cells of the mammalian dentate gyrus normally form a discrete layer, and virtually all granule cells migrate to this location. Exceptional granule cells that are positioned incorrectly, in ‘ectopic’ locations, are rare. Although the characteristics of such ectopic granule cells appear similar in many respects to granule cells located in the granule cell layer, their rare occurrence has limited a full evaluation of their structure and function. More information about ectopic granule cells has been obtained by studying those that develop after experimental manipulations that increase their number. For example, after severe seizures, the number of ectopic granule cells located in the hilus increases dramatically. These experimentally induced ectopic granule cells may not be equivalent to normal ectopic granule cells necessarily, but the vastly increased numbers have allowed much more information to be obtained. Remarkably, the granule cells that are positioned ectopically develop intrinsic properties and an axonal projection that are similar to granule cells that are located normally, i.e., in the granule cell layer. However, dendritic structure and synaptic structure/function appear to differ. These studies have provided new insight into a rare type of granule cell in the dentate gyrus, and the plastic characteristics of dentate granule cells that appear to depend on the location of the cell body. PMID:17148946

  10. CRISPR-mediated Activation of Latent HIV-1 Expression.

    PubMed

    Limsirichai, Prajit; Gaj, Thomas; Schaffer, David V

    2016-03-01

    Complete eradication of HIV-1 infection is impeded by the existence of cells that harbor chromosomally integrated but transcriptionally inactive provirus. These cells can persist for years without producing viral progeny, rendering them refractory to immune surveillance and antiretroviral therapy and providing a permanent reservoir for the stochastic reactivation and reseeding of HIV-1. Strategies for purging this latent reservoir are thus needed to eradicate infection. Here, we show that engineered transcriptional activation systems based on CRISPR/Cas9 can be harnessed to activate viral gene expression in cell line models of HIV-1 latency. We further demonstrate that complementing Cas9 activators with latency-reversing compounds can enhance latent HIV-1 transcription and that epigenome modulation using CRISPR-based acetyltransferases can also promote viral gene activation. Collectively, these results demonstrate that CRISPR systems are potentially effective tools for inducing latent HIV-1 expression and that their use, in combination with antiretroviral therapy, could lead to improved therapies for HIV-1 infection.

  11. CRISPR-mediated Activation of Latent HIV-1 Expression

    PubMed Central

    Limsirichai, Prajit; Gaj, Thomas; Schaffer, David V

    2016-01-01

    Complete eradication of HIV-1 infection is impeded by the existence of cells that harbor chromosomally integrated but transcriptionally inactive provirus. These cells can persist for years without producing viral progeny, rendering them refractory to immune surveillance and antiretroviral therapy and providing a permanent reservoir for the stochastic reactivation and reseeding of HIV-1. Strategies for purging this latent reservoir are thus needed to eradicate infection. Here, we show that engineered transcriptional activation systems based on CRISPR/Cas9 can be harnessed to activate viral gene expression in cell line models of HIV-1 latency. We further demonstrate that complementing Cas9 activators with latency-reversing compounds can enhance latent HIV-1 transcription and that epigenome modulation using CRISPR-based acetyltransferases can also promote viral gene activation. Collectively, these results demonstrate that CRISPR systems are potentially effective tools for inducing latent HIV-1 expression and that their use, in combination with antiretroviral therapy, could lead to improved therapies for HIV-1 infection. PMID:26607397

  12. Ectopic splenic autotransplantation following traumatic injury: A case report

    PubMed Central

    ZHANG, JINHE; YIN, JILIN; WANG, XINLU; LING, YINGCHEN; QUAN, JIANGTAO

    2015-01-01

    A 41-year-old male patient was admitted to the General Hospital of Guangzhou Military Command due to upper abdominal pain persisting for 12 h. Computed tomography (CT) and positron emission tomography/CT scans revealed multiple soft-tissue shadows in the abdominal cavity, peritoneum and Glisson's capsule, but the metabolic activity was at normal levels. A small area of low-density shadows near the tail of the pancreas and multiple shadows of enlarged lymph nodes were identified around the porta hepatis and the pancreas, with a mildly increased metabolic activity. On the basis of the CT images the patient was diagnosed with pancreatitis. Radionuclide imaging showed the absence of the spleen from its normal position (following splenectomy), but abnormal phagocytosis of multiple red blood cells was observed in the abdomen, which was diagnosed as ectopic splenic autotransplantation (ESAT). The patient subsequently recovered well following symptomatic treatment. ESAT in trauma patients requires urgent surgery in order to remove the damaged spleen and artificially cultivate partial splenic tissue. PMID:26640582

  13. Rare Presentation of Chorioadenoma Destruens as Acute Haemoperitoneum Mimicking Ruptured Ectopic Pregnancy.

    PubMed

    Sinha, M; Kaur, R; Gupta, R; Rani, R; Aggarwal, A

    2014-01-01

    Gestational trophoblastic neoplasms (GTN) are proliferative degenerative disorders of placental elements and include complete or partial mole (90%), invasivemole (5-8%), choriocarcinoma (1-2%) and placental site tumor (1-2%). Chorioadenoma destruens is a trophoblastic tumor, characterized by myometrial invasion through direct extension or via venous channels. We present a case of invasive mole eroding uterus and uterine vasculature, causing sudden rupture of uterus with massive haemoperitoneum mimicking ectopic pregnancy. A 20 year old G1P0 at 6 weeks gestation presented in Casualty of Kasturba Hospital complaining of severe acute onset lower abdominal pain for one hour. Clinical examination revealed shock. Sonography suggested ectopic pregnancy and immediate exploratory laparotomy was decided. On laparotomy, 2000cc of haemoperitoneum was noted. Grape like vesicles protruding through fundal perforation with profuse active bleeding was seen. Bleeding persisted despite evacuation. Step wise uterine devascularisation failed to achieve haemostasis. Total abdominal hysterectomy was performed as a life saving measure.

  14. EBNA2 and activated Notch induce expression of BATF.

    PubMed

    Johansen, Lisa M; Deppmann, Christopher D; Erickson, Kimberly D; Coffin, William F; Thornton, Tina M; Humphrey, Sean E; Martin, Jennifer M; Taparowsky, Elizabeth J

    2003-05-01

    The immortalization of human B lymphocytes by Epstein-Barr virus (EBV) requires the virus-encoded transactivator EBNA2 and the products of both viral and cellular genes which serve as EBNA2 targets. In this study, we identified BATF as a cellular gene that is up-regulated dramatically within 24 h following the infection of established and primary human B cells with EBV. The transactivation of BATF is mediated by EBNA2 in a B-cell-specific manner and is duplicated in non-EBV-infected B cells by the expression of mammalian Notch proteins. In contrast to other target genes activated by EBNA2, the BATF gene encodes a member of the AP-1 family of transcription factors that functions as a negative regulator of AP-1 activity and as an antagonist of cell growth. A potential role for BATF in promoting EBV latency is supported by studies in which BATF was shown to negatively impact the expression of a BZLF1 reporter gene and to reduce the frequency of lytic replication in latently infected cells. The identification of BATF as a cellular target of EBV provides important new information on how programs of viral and cellular gene expression may be coordinated to promote viral latency and control lytic-cycle entry.

  15. LYAR promotes colorectal cancer cell mobility by activating galectin-1 expression.

    PubMed

    Wu, Yupeng; Liu, Ming; Li, Zhuchen; Wu, Xiao-Bin; Wang, Ying; Wang, Yadong; Nie, Min; Huang, Feifei; Ju, Junyi; Ma, Chi; Tan, Renxiang; Zen, Ke; Zhang, Chen-Yu; Fu, Keqin; Chen, Yu-Gen; Wang, Ming-Rong; Zhao, Quan

    2015-10-20

    Colorectal cancer (CRC) is one of the leading causes of cancer-related death worldwide. However, the molecular mechanisms of CRC pathogenesis are not fully understood. In this study, we report the characterization of LYAR (Ly-1 antibody reactive clone) as a key regulator of the migration and invasion of human CRC cells. Immunohistochemistry analysis demonstrated that LYAR is expressed at a higher level in metastatic CRC tissues. We found that LYAR promoted the migratory and invasive capabilities of CRC cells. Gene expression profile analysis of CRC cells showed that LGALS1, which encodes the galectin-1 protein, was a potential target of LYAR. The ChIP assay and gene reporter assays indicated that LYAR directly bound to the LGALS1 promoter. The ectopic expression of galectin-1 partially restored the mobile potential of LYAR knocked-down cells, which suggests that galectin-1 contributed to the LYAR-promoted cell migration and invasion of CRC cells. Thus, this study revealed a novel mechanism by which the transcription factor LYAR may promote tumor cell migration and invasion by upregulating galectin-1 gene expression in CRC.

  16. Immunohistology of ectopic secondary lymph follicles in subcutaneous nodules from patients with hyperreactive onchocerciasis (sowda)

    PubMed Central

    Tenner-Racz, Klara; Korten, Simone; Hoerauf, Achim; Büttner, Dietrich W.

    2010-01-01

    Ectopic secondary lymph follicles emerge in patients with autoimmune or infectious diseases, e.g. in the synovium in rheumatoid arthritis or the skin in Borrelia burgdorferi infection, but ectopic localisations in the skin are rarely described for helminth infections. We investigated the cellular composition of secondary lymph follicles in subcutaneous nodules from eight patients with hyperreactive onchocerciasis (synonymous “localised” form or sowda) using immunohistology. CD3- and CD45RO-positive T cells and CD20-positive B cells were present in the mantle zone. The germinal centre was characterised by many B cells and CD35-positive follicular dendritic cells, which formed a network of attached IgE- and CD23-positive cells with the low-affinity IgE (epsilon) receptor. Few of the B cells were labelled for IgG1, IgG2 and IgG4, whereas in other zones of the nodule IgG1 was expressed by plasma cells and IgG1-coated dead microfilariae. B cells and few macrophages expressed the MHC class II molecule HLA-DR. Mature CD68-positive tingible body macrophages with phagocytosed leukocytes and CD57-positive lymphocytes occurred in the germinal centre. Macrophages in the germinal centre only weakly expressed alpha1-antichymotrypsin in contrast to macrophages in other zones of the onchocercoma. Furthermore, the multifunctional cytokine TGF-beta was only weakly expressed by macrophages and lymphocytes in the secondary follicles. Only few tryptase-positive mast cells, calprotectin-positive young macrophages, eosinophils and neutrophils occurred in the secondary follicles, although these cells were abundant in the onchocercomas. In conclusion, the ectopic secondary lymph follicles in onchocercomas and lymph nodes from hyperreactive onchocerciasis patients are equally composed. Electronic supplementary material The online version of this article (doi:10.1007/s00436-010-1912-0) contains supplementary material, which is available to authorized users. PMID:20524133

  17. Constitutive Smad signaling and Smad-dependent collagen gene expression in mouse embryonic fibroblasts lacking peroxisome proliferator-activated receptor-{gamma}

    SciTech Connect

    Ghosh, Asish K Wei, Jun; Wu, Minghua; Varga, John

    2008-09-19

    Transforming growth factor-{beta} (TGF-{beta}), a potent inducer of collagen synthesis, is implicated in pathological fibrosis. Peroxisome proliferator-activated receptor-{gamma} (PPAR-{gamma}) is a nuclear hormone receptor that regulates adipogenesis and numerous other biological processes. Here, we demonstrate that collagen gene expression was markedly elevated in mouse embryonic fibroblasts (MEFs) lacking PPAR-{gamma} compared to heterozygous control MEFs. Treatment with the PPAR-{gamma} ligand 15d-PGJ{sub 2} failed to down-regulate collagen gene expression in PPAR-{gamma} null MEFs, whereas reconstitution of these cells with ectopic PPAR-{gamma} resulted in their normalization. Compared to control MEFs, PPAR-{gamma} null MEFs displayed elevated levels of the Type I TGF-{beta} receptor (T{beta}RI), and secreted more TGF-{beta}1 into the media. Furthermore, PPAR-{gamma} null MEFs showed constitutive phosphorylation of cellular Smad2 and Smad3, even in the absence of exogenous TGF-{beta}, which was abrogated by the ALK5 inhibitor SB431542. Constitutive Smad2/3 phosphorylation in PPAR-{gamma} null MEFs was associated with Smad3 binding to its cognate DNA recognition sequences, and interaction with coactivator p300 previously implicated in TGF-{beta} responses. Taken together, these results indicate that loss of PPAR-{gamma} in MEFs is associated with upregulation of collagen synthesis, and activation of intracellular Smad signal transduction, due, at least in part, to autocrine TGF-{beta} stimulation.

  18. Osteoprotegerin expression and sensitivity in otosclerosis with different histological activity.

    PubMed

    Karosi, Tamás; Csomor, Péter; Szalmás, Anita; Kónya, József; Petkó, Mihály; Sziklai, István

    2011-03-01

    Otosclerosis is a complex bone dystrophy of the human otic capsule leading to conductive and sensorineural hearing loss. Since otosclerosis may, at least in part, be considered as an autoimmune-inflammatory disease, disturbed balance of TNF-alpha and osteoprotegerin (OPG) expression has been implicated in the pathological bone remodeling. It has been supposed that active otosclerosis is characterized by decreased or missing local OPG production with invariable OPG sensitivity of the otosclerotic foci. Ankylotic stapes footplates (n = 41) removed by stapedectomy were processed to histological examination, OPG-specific RT-PCR, tissue culturing and alkaline-phosphatase (AP) activity assessment, respectively. OPG concentration of serum specimens (n = 41) was measured by ELISA. Cortical bone fragments harvested from the external ear canal were used as negative controls of otosclerosis. Among 41 ankylotic stapes footplates, 22 active and 19 inactive otosclerosis cases were histologically diagnosed. OPG expression was significantly lower (p < 0.001) in active otosclerosis compared to inactive cases. Osteoclast cultures originated from active otosclerotic foci showed a considerable susceptibility against external OPG dosage, which resulted in a significant decrease of AP activity (p < 0.001). In contrast, OPG serum levels were in the normal range (5-100 ng/ml) indicating a non-systemic bone resorption. In conclusion, secondary decreased local OPG production might play an important role in the pathogenesis of otosclerotic bone remodeling disorder. As to previous and current results, decreased OPG sensitivity of lesion-forming cells should be excluded. These observations may indicate the potential role of recombinant OPG treatment in early stages of otosclerosis.

  19. Urokinase type plasminogen activator receptor expression in colorectal neoplasms

    PubMed Central

    Suzuki, S; Hayashi, Y; Wang, Y; Nakamura, T; Morita, Y; Kawasaki, K; Ohta, K; Aoyama, N; Kim, S; Itoh, H; Kuroda, Y; Doe, W

    1998-01-01

    Background—The urokinase type plasminogen activator receptor (uPAR) may play a critical role in cancer invasion and metastasis. 
Aims—To study the involvement of uPAR in colorectal carcinogenesis. 
Methods—The cellular expression and localisation of uPAR were investigated in colorectal adenomas and invasive carcinomas by in situ hybridisation, immunohistochemistry, and northern and western blot analyses. 
Results—uPAR mRNA expression was found mainly in the cytoplasm of dysplastic epithelial cells of 30% of adenomas with mild (19%), moderate (21%), and severe (47%) dysplasia, and in that of carcinomatous cells of 85% of invasive carcinomas: Dukes' stages A (72%), B (93%), and C (91%). Some stromal cells in the adjacent neoplastic epithelium were faintly positive. Immunoreactivity for uPAR was detected in dysplastic epithelial cells of 14% of adenomas and in carcinomatous cells of 49% of invasive carcinomas. uPAR mRNA and protein concentrations were significantly higher in severe than in mild or moderate dysplasia (p<0.05); they were notably higher in Dukes' stage A than in severe dysplasia (p<0.05), and significantly higher in Dukes' stage B than in stage A (p<0.05), but those in stage B were not different from those in stage C or in metastatic colorectal carcinomas of the liver. 
Conclusions—Colorectal adenoma uPAR, expressed essentially in dysplastic epithelial cells, was upregulated with increasing severity of atypia, and increased notably during the critical transition from severe dysplasic adenoma to invasive carcinoma. These findings implicate uPAR expression in the invasive and metastatic processes of colorectal cancer. 

 Keywords: urokinase type plasminogen activator receptor; colorectal adenoma; colorectal cancer; adenoma-carcinoma sequence PMID:9824607

  20. Endometrial cysteine-rich secretory protein 3 is inhibited by human chorionic gonadotrophin, and is increased in the decidua of tubal ectopic pregnancy

    PubMed Central

    Horne, A.W.; Duncan, W.C.; King, A.E.; Burgess, S.; Lourenco, P.C.; Cornes, P.; Ghazal, P.; Williams, A.R.; Udby, L.; Critchley, H.O.D.

    2009-01-01

    Ectopic pregnancy (EP) remains a considerable cause of morbidity and occasional mortality. Currently, there is no reliable test to differentiate ectopic from intrauterine gestation. We have previously used array technology to demonstrate that differences in gene expression in decidualized endometrium from women with ectopic and intrauterine gestations could be used to identify candidate diagnostic biomarkers for EP. The aim of this study was to further investigate the decidual gene with the highest fold increase in EP, cysteine-rich secretory protein-3 (CRISP-3). Decidualized endometrium from gestation-matched women undergoing surgical termination of pregnancy (n = 8), evacuation of uterus for miscarriage (n = 6) and surgery for EP (n = 11) was subjected to quantitative RT–PCR, morphological assessment, immunohistochemistry and western blot analysis. Sera were analysed for progesterone and human chorionic gonadotrophin (hCG) levels. Immortalized endometrial epithelial cells were cultured with physiological concentrations of hCG. CRISP-3 mRNA and protein expression were greater in endometrium from ectopic when compared with intrauterine pregnancies (P < 0.05). CRISP-3 protein was localized to epithelium and granulocytes of endometrium. CRISP-3 serum concentrations were not different in women with ectopic compared with intrauterine pregnancies. CRISP-3 expression in endometrium was not related to the degree of decidualization or to serum progesterone levels. Endometrial CRISP-3 expression was inversely proportional to serum hCG concentrations (P < 0.001). Stimulation of endometrial epithelial cells with hCG in vitro caused a reduction in CRISP-3 expression (P < 0.01). The measurement of CRISP-3 in endometrium could provide an additional tool in the diagnosis of failing early pregnancy of unknown location. The absence of a local reduction in expression of CRISP-3 in decidualized endometrium of women with EP may be due to reduced exposure to hCG due to the ectopic

  1. Endometrial cysteine-rich secretory protein 3 is inhibited by human chorionic gonadotrophin, and is increased in the decidua of tubal ectopic pregnancy.

    PubMed

    Horne, A W; Duncan, W C; King, A E; Burgess, S; Lourenco, P C; Cornes, P; Ghazal, P; Williams, A R; Udby, L; Critchley, H O D

    2009-05-01

    Ectopic pregnancy (EP) remains a considerable cause of morbidity and occasional mortality. Currently, there is no reliable test to differentiate ectopic from intrauterine gestation. We have previously used array technology to demonstrate that differences in gene expression in decidualized endometrium from women with ectopic and intrauterine gestations could be used to identify candidate diagnostic biomarkers for EP. The aim of this study was to further investigate the decidual gene with the highest fold increase in EP, cysteine-rich secretory protein-3 (CRISP-3). Decidualized endometrium from gestation-matched women undergoing surgical termination of pregnancy (n = 8), evacuation of uterus for miscarriage (n = 6) and surgery for EP (n = 11) was subjected to quantitative RT-PCR, morphological assessment, immunohistochemistry and western blot analysis. Sera were analysed for progesterone and human chorionic gonadotrophin (hCG) levels. Immortalized endometrial epithelial cells were cultured with physiological concentrations of hCG. CRISP-3 mRNA and protein expression were greater in endometrium from ectopic when compared with intrauterine pregnancies (P < 0.05). CRISP-3 protein was localized to epithelium and granulocytes of endometrium. CRISP-3 serum concentrations were not different in women with ectopic compared with intrauterine pregnancies. CRISP-3 expression in endometrium was not related to the degree of decidualization or to serum progesterone levels. Endometrial CRISP-3 expression was inversely proportional to serum hCG concentrations (P < 0.001). Stimulation of endometrial epithelial cells with hCG in vitro caused a reduction in CRISP-3 expression (P < 0.01). The measurement of CRISP-3 in endometrium could provide an additional tool in the diagnosis of failing early pregnancy of unknown location. The absence of a local reduction in expression of CRISP-3 in decidualized endometrium of women with EP may be due to reduced exposure to hCG due to the ectopic

  2. Oxalomalate affects the inducible nitric oxide synthase expression and activity.

    PubMed

    Irace, Carlo; Esposito, Giuseppe; Maffettone, Carmen; Rossi, Antonietta; Festa, Michela; Iuvone, Teresa; Santamaria, Rita; Sautebin, Lidia; Carnuccio, Rosa; Colonna, Alfredo

    2007-03-13

    Inducible nitric oxide synthase (iNOS) is an homodimeric enzyme which produces large amounts of nitric oxide (NO) in response to inflammatory stimuli. Several factors affect the synthesis and catalytic activity of iNOS. Particularly, dimerization of NOS monomers is promoted by heme, whereas an intracellular depletion of heme and/or L-arginine considerably decreases NOS resistance to proteolysis. In this study, we found that oxalomalate (OMA, oxalomalic acid, alpha-hydroxy-beta-oxalosuccinic acid), an inhibitor of both aconitase and NADP-dependent isocitrate dehydrogenase, inhibited nitrite production and iNOS protein expression in lipopolysaccharide (LPS)-activated J774 macrophages, without affecting iNOS mRNA content. Furthermore, injection of OMA precursors to LPS-stimulated rats also decreased nitrite production and iNOS expression in isolated peritoneal macrophages. Interestingly, alpha-ketoglutarate or succinyl-CoA administration reversed OMA effect on NO production, thus correlating NO biosynthesis with the anabolic capacity of Krebs cycle. When protein synthesis was blocked by cycloheximide in LPS-activated J774 cells treated with OMA, iNOS protein levels, evaluated by Western blot analysis and (35)S-metabolic labelling, were decreased, suggesting that OMA reduces iNOS biosynthesis and induces an increase in the degradation rate of iNOS protein. Moreover, we showed that OMA inhibits the activity of the iNOS from lung of LPS-treated rats by enzymatic assay. Our results, demonstrating that OMA acts regulating synthesis, catalytic activity and degradation of iNOS, suggest that this compound might have a potential role in reducing the NO overproduction occurring in some pathological conditions.

  3. Gamma-band activity reflects attentional guidance by facial expression.

    PubMed

    Müsch, Kathrin; Siegel, Markus; Engel, Andreas K; Schneider, Till R

    2017-02-01

    Facial expressions attract attention due to their motivational significance. Previous work focused on attentional biases towards threat-related, fearful faces, although healthy participants tend to avoid mild threat. Growing evidence suggests that neuronal gamma (>30Hz) and alpha-band activity (8-12Hz) play an important role in attentional selection, but it is unknown if such oscillatory activity is involved in the guidance of attention through facial expressions. Thus, in this magnetoencephalography (MEG) study we investigated whether attention is shifted towards or away from fearful faces and characterized the underlying neuronal activity in these frequency ranges in forty-four healthy volunteers. We employed a covert spatial attention task using neutral and fearful faces as task-irrelevant distractors and emotionally neutral Gabor patches as targets. Participants had to indicate the tilt direction of the target. Analysis of the neuronal data was restricted to the responses to target Gabor patches. We performed statistical analysis at the sensor level and used subsequent source reconstruction to localize the observed effects. Spatially selective attention effects in the alpha and gamma band were revealed in parieto-occipital regions. We observed an attentional cost of processing the face distractors, as reflected in lower task performance on targets with short stimulus onset asynchrony (SOA <150ms) between faces and targets. On the neuronal level, attentional orienting to face distractors led to enhanced gamma band activity in bilateral occipital and parietal regions, when fearful faces were presented in the same hemifield as targets, but only in short SOA trials. Our findings provide evidence that both top-down and bottom-up attentional biases are reflected in parieto-occipital gamma-band activity.

  4. IKBKE is over-expressed in glioma and contributes to resistance of glioma cells to apoptosis via activating NF-κB.

    PubMed

    Guan, Hongyu; Zhang, Heng; Cai, Junchao; Wu, Jueheng; Yuan, Jie; Li, Jun; Huang, Zhengsong; Li, Mengfeng

    2011-02-01

    IκB kinase-ε (IKBKE), a member of the IκB kinase (IKK) family, has been identified as an oncogenic protein and found to be up-regulated in breast cancer, ovarian cancer and prostate cancer. Nonetheless, the expression status and functional significance of IKBKE in human glioma remain unexplored. For the first time, we have demonstrated that mRNA and protein levels of IKBKE were robustly up-regulated in glioma cell lines and human primary glioma tissues. Immunohistochemistry analysis revealed that 53.5% (38/71) paraffin-embedded archived glioma specimens exhibited high levels of IKBKE expression. Intriguingly, there was no significant difference in IKBKE expression among different grades of glioma. To understand the biological function of IKBKE in the development and progression of human glioma, glioma cells lines ectopically over-expressing IKBKE were established and tested for their responsiveness to apoptotic inducers. Our data showed that IKBKE over-expression inhibited cell apoptosis induced by UV irradiation or adriamycin and, in contrast, shRNAi-mediated suppression of IKBKE increased the sensitivity of glioma cells to the apoptotic inducers. Importantly, we found that up-regulated IKBKE could induce the expression of Bcl-2 through activating NF-κB signalling, and that, specifically, we identified IκB as a critical component for this signalling cascade. The current study suggests that up-regulation of IKBKE may represent an important molecular hallmark that is biologically and clinically relevant to the development and progression, as well as the chemo- and radio-resistance, of the disease.

  5. Cell surface expression of biologically active influenza C virus HEF glycoprotein expressed from cDNA.

    PubMed

    Pekosz, A; Lamb, R A

    1999-10-01

    The hemagglutinin, esterase, and fusion (HEF) glycoprotein of influenza C virus possesses receptor binding, receptor destroying, and membrane fusion activities. The HEF cDNAs from influenza C/Ann Arbor/1/50 (HEF-AA) and influenza C/Taylor/1223/47 (HEF-Tay) viruses were cloned and expressed, and transport of HEF to the cell surface was monitored by susceptibility to cleavage by exogenous trypsin, indirect immunofluorescence microscopy, and flow cytometry. Previously it has been found in studies with the C/Johannesburg/1/66 strain of influenza C virus (HEF-JHB) that transport of HEF to the cell surface is severely inhibited, and it is thought that the short cytoplasmic tail, Arg-Thr-Lys, is involved in blocking HEF cell surface expression (F. Oeffner, H.-D. Klenk, and G. Herrler, J. Gen. Virol. 80:363-369, 1999). As the cytoplasmic tail amino acid sequences of HEF-AA and HEF-Tay are identical to that of HEF-JHB, the data indicate that cell surface expression of HEF-AA and HEF-Tay is not inhibited by this amino acid sequence. Furthermore, the abundant cell surface transport of HEF-AA and HEF-Tay indicates that their cell surface expression does not require coexpression of another viral protein. The HEF-AA and HEF-Tay HEF glycoproteins bound human erythrocytes, promoted membrane fusion in a low-pH and trypsin-dependent manner, and displayed esterase activity, indicating that the HEF glycoprotein alone mediates all three known functions at the cell surface.

  6. Altered Activity and Expression of Cytosolic Peptidases in Colorectal Cancer

    PubMed Central

    Perez, Itxaro; Blanco, Lorena; Sanz, Begoña; Errarte, Peio; Ariz, Usue; Beitia, Maider; Fernández, Ainhoa; Loizate, Alberto; Candenas, M Luz; Pinto, Francisco M; Gil, Javier; López, José I.; Larrinaga, Gorka

    2015-01-01

    Background and Objective: The role of peptidases in carcinogenic processes and their potential usefulness as tumor markers in colorectal cancer (CRC) have been classically attributed to cell-surface enzymes. The objective of the present study was to analyze the activity and mRNA expression of three cytosolic peptidases in the CRC and to correlate the obtained results with classic histopathological parameters for tumor prognosis and survival. Methods: The activity and mRNA levels of puromycin-sensitive aminopeptidase (PSA), aminopeptidase B (APB) and pyroglutamyl-peptidase I (PGI) were measured by fluorimetric and quantitative RT-PCR methods in colorectal mucosa and tumor tissues and plasma samples from CRC patients (n=81). Results: 1) PSA and APB activity was higher in adenomas and carcinomas than in the uninvolved mucosa. 2) mRNA levels of PSA and PGI was lower in tumors. 3) PGI activity in CRC tissue correlated negatively with histological grade, tumor size and 5-year overall suvival of CRC patients. 4) Higher plasmatic APB activity was independently associated with better 5-year overall survival. Conclusions: Data suggest that cytosolic peptidases may be involved in colorectal carcinogenesis and point to the determination of this enzymes as a valuable method in the determination of CRC prognosis. PMID:26078706

  7. Cardiac afferent activity modulates the expression of racial stereotypes

    PubMed Central

    Azevedo, Ruben T.; Garfinkel, Sarah N.; Critchley, Hugo D.; Tsakiris, Manos

    2017-01-01

    Negative racial stereotypes tend to associate Black people with threat. This often leads to the misidentification of harmless objects as weapons held by a Black individual. Yet, little is known about how bodily states impact the expression of racial stereotyping. By tapping into the phasic activation of arterial baroreceptors, known to be associated with changes in the neural processing of fearful stimuli, we show activation of race-threat stereotypes synchronized with the cardiovascular cycle. Across two established tasks, stimuli depicting Black or White individuals were presented to coincide with either the cardiac systole or diastole. Results show increased race-driven misidentification of weapons during systole, when baroreceptor afferent firing is maximal, relative to diastole. Importantly, a third study examining the positive Black-athletic stereotypical association fails to demonstrate similar modulations by cardiac cycle. We identify a body–brain interaction wherein interoceptive cues can modulate threat appraisal and racially biased behaviour in context-dependent ways. PMID:28094772

  8. Giant hepatic hemangioma and cross-fused ectopic kidney in a spaceflight participant.

    PubMed

    Jennings, Richard T; Garriott, Owen K; Bogomolov, Valery V; Pochuev, Vladimir I; Morgun, Valery V; Garriott, Richard A

    2010-02-01

    Commercial spaceflight participants are typically older than traditional astronauts and often have medical conditions that make medical certification for flight difficult. This case report considers a 43-yr-old spaceflight participant who planned a short-duration Soyuz flight to the International Space Station (ISS). While he participated in many hazardous activities such as parachuting, hang gliding, scuba diving, Antarctic and jungle exploration, and deep sea submersible operations, he knew that several of his medical conditions precluded serving as a career astronaut. At the time of his initial spaceflight prescreen examination, he was known to have previous bilateral photorefractive keratectomy (PRK) for myopia and a cross-fused left ectopic kidney that would be disqualifying for a career astronaut. During the evaluation for the left single cross-fused ectopic kidney, a giant hepatic hemangioma was also discovered. In order to medically qualify for flight, the giant hepatic hemangioma was surgically removed. This case summary investigat*es the implications of a single cross-fused left ectopic kidney and the decision process and treatment implications for spaceflight medical certification in an individual with an asymptomatic giant hepatic hemangioma.

  9. Brain Activity while Reading Sentences with Kanji Characters Expressing Emotions

    NASA Astrophysics Data System (ADS)

    Yuasa, Masahide; Saito, Keiichi; Mukawa, Naoki

    In this paper, we describe the brain activity associated with kanji characters expressing emotion, which are places at the end of a sentence. Japanese people use a special kanji character in brackets at the end of sentences in text messages such as those sent through e-mail and messenger tools. Such kanji characters plays a role to expresses the sender's emotion (such as fun, laughter, sadness, tears), like emoticons. It is a very simple and effective way to convey the senders' emotions and his/her thoughts to the receiver. In this research, we investigate the effects of emotional kanji characters by using an fMRI study. The experimental results show that both the right and left inferior frontal gyrus, which have been implicated on verbal and nonverbal information, were activated. We found that we detect a sentence with an emotional kanji character as the verbal and nonverval information, and a sentence with emotional kanji characters enrich communication between the sender and the reciever.

  10. Expression and phosphorylation of the AS160_v2 splice variant supports GLUT4 activation and the Warburg effect in multiple myeloma

    PubMed Central

    2013-01-01

    Background Multiple myeloma (MM) is a fatal plasma cell malignancy exhibiting enhanced glucose consumption associated with an aerobic glycolytic phenotype (i.e., the Warburg effect). We have previously demonstrated that myeloma cells exhibit constitutive plasma membrane (PM) localization of GLUT4, consistent with the dependence of MM cells on this transporter for maintenance of glucose consumption rates, proliferative capacity, and viability. The purpose of this study was to investigate the molecular basis of constitutive GLUT4 plasma membrane localization in MM cells. Findings We have elucidated a novel mechanism through which myeloma cells achieve constitutive GLUT4 activation involving elevated expression of the Rab-GTPase activating protein AS160_v2 splice variant to promote the Warburg effect. AS160_v2-positive MM cell lines display constitutive Thr642 phosphorylation, known to be required for inactivation of AS160 Rab-GAP activity. Importantly, we show that enforced expression of AS160_v2 is required for GLUT4 PM translocation and activation in these select MM lines. Furthermore, we demonstrate that ectopic expression of a full-length, phospho-deficient AS160 mutant is sufficient to impair constitutive GLUT4 cell surface residence, which is characteristic of MM cells. Conclusions This is the first study to tie AS160 de-regulation to increased glucose consumption rates and the Warburg effect in cancer. Future studies investigating connections between the insulin/IGF-1/AS160_v2/GLUT4 axis and FDG-PET positivity in myeloma patients are warranted and could provide rationale for therapeutically targeting this pathway in MM patients with advanced disease. PMID:24280290

  11. Ectopic parathyroid glands and their anatomical, clinical and surgical implications.

    PubMed

    Noussios, G; Anagnostis, P; Natsis, K

    2012-11-01

    Ectopic parathyroid glands result from aberrant migration during early stages of development and lack of successful identification may lead to lack of success in parathyroid surgery. They constitute a common etiology of persistent or recurrent hyperparathyroidism, when they are missed at initial diagnosis. Their prevalence is about 2-43% in anatomical series and up to 16% and 14% in patients with primary and secondary hyperparathyroidism, respectively. Ectopic inferior parathyroids are most frequently found in the anterior mediastinum, in association with the thymus or the thyroid gland, while the most common position for ectopic superior parathyroids is the tracheoesophageal groove and retroesophageal region. Neck ultrasound and 99mTc Sestamibi scan are first-line imaging modalities, although with low sensitivity and specificity. However, their combination with modern techniques, such as single photon emission computed tomography (SPECT) alone or in combination with CT (SPECT/CT) increases their diagnostic accuracy. Fine needle-aspiration cytology of a lesion suspicious for parathyroid tissue and measurement of parathyroid hormone (PTH) in the aspired material further assist to the successful preoperative localization of ectopic glands. Common sites for surgical investigation are the upper thyroid pole and the upper vascular thyroid stalk behind the hypopharynx and cervical esophagus for the superior parathyroids, and the carotid artery bifurcation and the thymic tongue, for the inferior parathyroids. Radioguided minimally invasive parathyroidectomy after successful localization, assisted by rapid PTH measurement postoperatively, significantly improves surgical outcomes in patients with ectopic parathyroid adenomas.

  12. Hydatidiform mole presentation as a tubal ectopic pregnancy.

    PubMed

    Nakeer, Tabassum; Shahid, Muhammad; Ansari, Muhammad Asad; Nakeer, Rooham

    2014-05-01

    Presentation of hydatidiform mole as tubal ectopic pregnancy is very rare. These patients usually present with ectopic pregnancy and are later diagnosed with hydatidiform mole on the basis of histological examination following surgery. We present the case of a 32-year-old female who presented with abdominal pain and vaginal bleed since 2 days of presentation. She was vitally stable. There was mild tenderness in hypogastrium and left iliac fossa. Pelvic examination showed mild bleeding per vaginum and fullness in both fornices. The patient was suspected of having an ectopic pregnancy. Ultrasonography of pelvis revealed fluid in cul-de-sac and a sac like mass of 1.8 cm attached to the left ovary. On laparotomy, there was a left sided tubal ectopic pregnancy and subsequently left salpingectomy was done. Histopathology of the tissue sample showed features of partial hydatidiform mole. Ectopic pregnancy can present as hydatidiform mole in rare cases for which histological examination of the tissue is required to establish the diagnosis.

  13. Upper gastrointestinal ectopic variceal bleeding treated with various endoscopic modalities

    PubMed Central

    Park, Sang Woo; Cho, Eunae; Jun, Chung Hwan; Choi, Sung Kyu; Kim, Hyun Soo; Park, Chang Hwan; Rew, Jong Sun; Cho, Sung Bum; Kim, Hee Joon; Han, Mingui; Cho, Kyu Man

    2017-01-01

    Abstract Rationale: Ectopic variceal bleeding is a rare (2–5%) but fatal gastrointestinal bleed in patients with portal hypertension. Patients with ectopic variceal bleeding manifest melena, hematochezia, or hematemesis, which require urgent managements. Definitive therapeutic modalities of ectopic varices are not yet standardized because of low incidence. Various therapeutic modalities have been applied on the basis of the experiences of experts or availability of facilities, with varying results. Patient concerns: We have encountered eight cases of gastrointestinal ectopic variceal bleeding in five patients in the last five years. Diagnoses: All patients were diagnosed with liver cirrhosis presenting melena or hematemesis. Interventions: All patients were treated with various endoscopic modalities (endoscopic variceal obturation [EVO] with cyanoacrylate in five cases, endoscopic variceal band ligation (EVL) in two cases, hemoclipping in one case). Outcomes: Satisfactory hemostasis was achieved without radiologic interventions in all cases. EVO and EVL each caused one case of portal biliopathy, and EVL induced ulcer bleeding in one case. Lessons: EVO generally accomplished better results of variceal obturations than EVL or hemoclipping, without serious adverse events. EVO may be an effective modality for control of ectopic variceal bleeding without radiologic intervention or surgery. PMID:28072750

  14. Heterologous expression of an active chitin synthase from Rhizopus oryzae.

    PubMed

    Salgado-Lugo, Holjes; Sánchez-Arreguín, Alejandro; Ruiz-Herrera, José

    2016-12-01

    Chitin synthases are highly important enzymes in nature, where they synthesize structural components in species belonging to different eukaryotic kingdoms, including kingdom Fungi. Unfortunately, their structure and the molecular mechanism of synthesis of their microfibrilar product remain largely unknown, probably because no fungal active chitin synthases have been isolated, possibly due to their extreme hydrophobicity. In this study we have turned to the heterologous expression of the transcript from a small chitin synthase of Rhizopus oryzae (RO3G_00942, Chs1) in Escherichia coli. The enzyme was active, but accumulated mostly in inclusion bodies. High concentrations of arginine or urea solubilized the enzyme, but their dilution led to its denaturation and precipitation. Nevertheless, use of urea permitted the purification of small amounts of the enzyme. The properties of Chs1 (Km, optimum temperature and pH, effect of GlcNAc) were abnormal, probably because it lacks the hydrophobic transmembrane regions characteristic of chitin synthases. The product of the enzyme showed that, contrasting with chitin made by membrane-bound Chs's and chitosomes, was only partially in the form of short microfibrils of low crystallinity. This approach may lead to future developments to obtain active chitin synthases that permit understanding their molecular mechanism of activity, and microfibril assembly.

  15. Thiazolidinediones repress ob gene expression in rodents via activation of peroxisome proliferator-activated receptor gamma.

    PubMed Central

    De Vos, P; Lefebvre, A M; Miller, S G; Guerre-Millo, M; Wong, K; Saladin, R; Hamann, L G; Staels, B; Briggs, M R; Auwerx, J

    1996-01-01

    The ob gene product, leptin, is a signaling factor regulating body weight and energy balance. ob gene expression in rodents is increased in obesity and is regulated by feeding patterns and hormones, such as insulin and glucocorticoids. In humans with gross obesity, ob mRNA levels are higher, but other modulators of human ob expression are unknown. In view of the importance of peroxisome proliferator-activated receptor gamma (PPARgamma) in adipocyte differentiation, we analyzed whether ob gene expression is subject to regulation by factors activating PPARs. Treatment of rats with the PPARalpha activator fenofibrate did not change adipose tissue and body weight and had no significant effect on ob mRNA levels. However, administration of the thiazolidinedione BRL49653, a PPARgamma ligand, increased food intake and adipose tissue weight while reducing ob mRNA levels in rats in a dose-dependent manner. The inhibitory action of the thiazolidinedione BRL49653 on ob mRNA levels was also observed in vitro. Thiazolidinediones reduced the expression of the human ob promoter in primary adipocytes, however, in undifferentiated 3T3-L1 preadipocytes lacking endogenous PPARgamma, cotransfection of PPARgamma was required to observe the decrease. In conclusion, these data suggest that PPARgamma activators reduce ob mRNA levels through an effect of PPARgamma on the ob promoter. PMID:8770873

  16. CD8 T Cells Are Required for the Formation of Ectopic Germinal Centers in Rheumatoid Synovitis

    PubMed Central

    Kang, Young Mo; Zhang, Xiaoyu; Wagner, Ulf G.; Yang, Hongyu; Beckenbaugh, Robert D.; Kurtin, Paul J.; Goronzy, Jörg J.; Weyand, Cornelia M.

    2002-01-01

    The assembly of inflammatory lesions in rheumatoid arthritis is highly regulated and typically leads to the formation of lymphoid follicles with germinal center (GC) reactions. We used microdissection of such extranodal follicles to analyze the colonizing T cells. Although the repertoire of follicular T cells was diverse, a subset of T cell receptor (TCR) sequences was detected in multiple independent follicles and not in interfollicular zones, suggesting recognition of a common antigen. Unexpectedly, the majority of shared TCR sequences were from CD8 T cells that were highly enriched in the synovium and present in low numbers in the periphery. To examine their role in extranodal GC reactions, CD8 T cells were depleted in human synovium-SCID mouse chimeras. Depletion of synovial CD8 T cells caused disintegration of the GC-containing follicles. In the absence of CD8 T cells, follicular dendritic cells disappeared, production of lymphotoxin-α1β2 markedly decreased, and immunoglobulin (Ig) secretion ceased. Immunohistochemical studies demonstrated that these CD8 T cells accumulated at the edge of the mantle zone. Besides their unique localization, they were characterized by the production of interferon (IFN)-γ, lack of the pore-forming enzyme perforin, and expression of CD40 ligand. Perifollicular IFN-γ+ CD8 T cells were rare in secondary lymphoid tissues but accounted for the majority of IFN-γ+ cells in synovial infiltrates. We propose that CD8+ T cells regulate the structural integrity and functional activity of GCs in ectopic lymphoid follicles. PMID:12021312

  17. Transgenic chickens expressing human urokinase-type plasminogen activator.

    PubMed

    Lee, Sung Ho; Gupta, Mukesh Kumar; Ho, Young Tae; Kim, Teoan; Lee, Hoon Taek

    2013-09-01

    Urokinase-type plasminogen activator is a serine protease that is clinically used in humans for the treatment of thrombolytic disorders and vascular diseases such as acute ischemic stroke and acute peripheral arterial occlusion. This study explored the feasibility of using chickens as a bioreactor for producing human urokinase-type plasminogen activator (huPA). Recombinant huPA gene, under the control of a ubiquitous Rous sarcoma virus promoter, was injected into the subgerminal cavity of freshly laid chicken eggs at stage X using the replication-defective Moloney murine leukemia virus (MoMLV)-based retrovirus vectors encapsidated with VSV-G (vesicular stomatitis virus G) glycoprotein. A total of 38 chicks, out of 573 virus-injected eggs, hatched and contained the huPA gene in their various body parts. The mRNA transcript of the huPA gene was present in various organs, including blood and egg, and was germ-line transmitted to the next generation. The level of active huPA protein was 16-fold higher in the blood of the transgenic chicken than in the nontransgenic chicken (P < 0.05). The expression of huPA protein in eggs increased from 7.82 IU/egg in the G0 generation to 17.02 IU/egg in the G1 generation. However, huPA-expressing embryos had reduced survival and hatchability at d 18 and 21 of incubation, respectively, and the blood clotting time was significantly higher in transgenic chickens than their nontransgenic counterparts (P < 0.05). Furthermore, adult transgenic rooster showed reduced (P < 0.05) fertility, as revealed by reduced volume of semen ejaculate, sperm concentration, and sperm viability. Taken together, our data suggest that huPA transgenic chickens could be successfully produced by the retroviral vector system. Transgenic chickens, expressing the huPA under the control of a ubiquitous promoter, may not only be used as a bioreactor for pharming of the huPA drug but also be useful for studying huPA-induced bleeding and other disorders.

  18. [Ectopic pregnancy: Its current interest in Primary Health Care].

    PubMed

    López-Luque, P R; Bergal-Mateo, G J; López-Olivares, M C

    2014-01-01

    An ectopic pregnancy is the implantation and development of the ovum fertilized outside the endometrial cavity. Its incidence has increased in the last 30 years, and although its morbimortality has decreased, it is still the first cause of mortality in the first trimester of the pregnancy. Early suspicion is important, particularly in women of fertile age and with risk factors indicative of an extrauterine gestation. The symptomatology is usually amenorrhea, abdominal pain, metrorrhagia, general pregnancy symptoms, and even syncope and shock. The diagnosis of ectopic pregnancy is based on the clinical information, analytical results on mother blood and urine, ultrasound examination, transvaginal culdocentesis, laparoscopic or laparotomic inspection, and a histological study. The treatment can be surgical (salpingostomy or salpingectomy), medical (methotrexate) or expectant, depending on the factors of the ectopic pregnancy: early diagnosis, presence of acute complications, clinical condition of the patient, etc.

  19. Pancreatic solitary fibrous tumor causing ectopic adrenocorticotropic hormone syndrome.

    PubMed

    Murakami, Keigo; Nakamura, Yasuhiro; Felizola, Saulo J A; Morimoto, Ryo; Satoh, Fumitoshi; Takanami, Kentaro; Katakami, Hideki; Hirota, Seiichi; Takeda, Yoshiyu; Meguro-Horike, Makiko; Horike, Shin-Ichi; Unno, Michiaki; Sasano, Hironobu

    2016-11-15

    Solitary fibrous tumors occasionally present with hypoglycemia because of the excessive release of insulin-like growth factor II. We report the first case of pancreatic solitary fibrous tumor causing ectopic adrenocorticotropic hormone syndrome. An 82-year-old Japanese man presented with lower limb edema, uncontrolled hypertension, hypokalemia, and baseline hypercortisolism. Distal pancreatectomy was performed after the clinical diagnosis of a neuroendocrine tumor with ectopic secretion of adrenocorticotropic hormone. On histological examination, the tumor showed spindle cells in a fascicular arrangement. The diagnosis of the solitary fibrous tumor was confirmed by the identification of the NAB2-STAT6 fusion gene and positive immuno-histochemical staining for STAT6 and CD34. Using quantitative real-time polymerase chain reaction, mRNA that encoded proopiomelanocortin, precursor of adrenocorticotropic hormone, was detected. Proopiomelanocortin production through the demethylation of the promoter region Domain IV was detected. Pancreatic solitary fibrous tumors represent a new cause of ectopic adrenocorticotropic hormone syndrome.

  20. Polyphenols from Chilean Propolis and Pinocembrin Reduce MMP-9 Gene Expression and Activity in Activated Macrophages

    PubMed Central

    Saavedra, Nicolás; Cuevas, Alejandro; Cavalcante, Marcela F.; Dörr, Felipe A.; Saavedra, Kathleen; Zambrano, Tomás; Abdalla, Dulcineia S. P.; Salazar, Luis A.

    2016-01-01

    Polyphenols from diverse sources have shown anti-inflammatory activity. In the context of atherosclerosis, macrophages play important roles including matrix metalloproteinases synthesis involved in degradation of matrix extracellular components affecting the atherosclerotic plaque stability. We prepared a propolis extract and pinocembrin in ethanol solution. Propolis extract was chemically characterized using LC-MS. The effect of treatments on gene expression and proteolytic activity was measured in vitro using murine macrophages activated with LPS. Cellular toxicity associated with both treatments and the vehicle was determined using MTT and apoptosis/necrosis detection assays. MMP-9 gene expression and proteolytic activity were measured using qPCR and zymography, respectively. Thirty-two compounds were identified in the propolis extract, including pinocembrin among its major components. Treatment with either ethanolic extract of propolis or pinocembrin inhibits MMP-9 gene expression in a dose-dependent manner. Similarly, an inhibitory effect was observed in proteolytic activity. However, the effect showed by ethanolic extract of propolis was higher than the effect of pinocembrin, suggesting that MMP-9 inhibition results from a joint contribution between the components of the extract. These data suggest a potential role of polyphenols from Chilean propolis in the control of extracellular matrix degradation in atherosclerotic plaques. PMID:27119082

  1. Diosgenin does not express estrogenic activity: a uterotrophic assay.

    PubMed

    Medigović, Ivana; Ristić, Nataša; Živanović, Jasmina; Šošić-Jurjević, Branka; Filipović, Branko; Milošević, Verica; Nestorović, Nataša

    2014-04-01

    This study assessed the effects of diosgenin on estrogenic activity using a uterotrophic assay. Immature female rats received diosgenin orally at doses of 200, 100, or 20 mg/kg body mass; and 17α ethynylestradiol at doses of 1 or 0.3 μg/kg, daily, for 3 consecutive days from day 19 to day 21. Controls were distributed among 2 groups: an intact control group and a vehicle control group. Animals were sacrificed 24 h after the last application of diosgenin, estradiol, or vehicle (22nd day of life). Uterine wet weight, stereological and histomorphometrical changes, immunohistochemical expression of estrogen receptor alpha (ERα), progesterone receptor (PR), and the expression of lactoferrin (LF) were examined. Diosgenin did not affect the uterine wet weight, epithelium height, volume densities of endometrium, endometrial epithelia, number of endometrial glands, or histological appearance of vaginal epithelia. ERα, PR, and LF immunostaining intensity were not altered in the animals that received diosgenin. High-potency reference ER agonist 17α-ethynylestradiol induced a significant increase in all of the measured parameters, and as expected, decreased ERα immunostaining intensity. Based on these data, it can be concluded that diosgenin, at doses of 20-200 mg/kg, did not act as an estrogen agonist in the immature rat uterotrophic assay.

  2. Delayed puberty due to pituitary stalk dysgenesis and ectopic neurohypophysis.

    PubMed

    Yoo, Hye Jin; Choi, Kyung Mook; Ryu, Ohk Hyun; Suh, Sang Il; Kim, Nan Hee; Baik, Sei Hyun; Choi, Dong Seop

    2006-03-01

    Hypopituitarism is not a common cause of delayed puberty. A 22 year old man was referred to our clinic because of the absence of the development of secondary sexual characteristics. The patient had no complaints of physical discomfort. Random serum testosterone and luteinizing hormone level were obtained and found to be low. The combined pituitary function stimulation test revealed a partial hypopituitarism. A pituitary magnetic resonance imaging (MRI) was obtained and showed decreased pituitary stalk enhancement and ectopic neurohypophysis. Therefore, we conclude that the delayed puberty was a result of hypopituitarism due to pituitary stalk dysgenesis and ectopic neurohypophysis. The patient was started on hormone replacement therapy and gradually developed secondary sexual characteristics.

  3. Heterotopic pregnancy: discovery of ectopic pregnancy after elective abortion.

    PubMed

    DeFrancesch, F; DiLeo, L; Martinez, J

    1999-03-01

    We report a case of combined intrauterine and tubal pregnancy in a 23-year-old woman. The patient came to the emergency department complaining of lower abdominal pain after having had an elective abortion 2 weeks earlier. Her physician had done pelvic ultrasonography, noting an intrauterine pregnancy before the abortion. Our working diagnosis in the emergency department was retained products of conception versus postabortion endometritis. Pelvic ultrasonography in the emergency department revealed an ectopic pregnancy without evidence of retained products of conception, and the patient had a right salpingotomy with removal of the ectopic fetus without complications.

  4. Pulmonary agenesis associated with dextrocardia, sternal defects, and ectopic kidney.

    PubMed

    Eroglu, Atilla; Alper, Fatih; Turkyilmaz, Atila; Karaoglanoglu, Nurettin; Okur, Adnan

    2005-12-01

    Pulmonary agenesis is a rare embryological defect, usually unilateral, and is associated with skeletal, cardiovascular, and other anomalies. A 22-month-old girl was admitted to our clinic with recurrent pulmonary infections. A chest X-ray and multidetector computed tomography (MDCT) revealed pulmonary agenesis with dextrocardia, sternal defects, and a right pelvic ectopic kidney. We report on the first known case of right-sided pulmonary agenesis combined with isolated dextrocardia, sternal defects, and an ectopic kidney. The importance of MDCT for the diagnosis of pulmonary agenesis and associated other anomalies is demonstrated.

  5. Expression of Active Human Tissue-Type Plasminogen Activator in Escherichia coli

    PubMed Central

    Qiu, Ji; Swartz, James R.; Georgiou, George

    1998-01-01

    The formation of native disulfide bonds in complex eukaryotic proteins expressed in Escherichia coli is extremely inefficient. Tissue plasminogen activator (tPA) is a very important thrombolytic agent with 17 disulfides, and despite numerous attempts, its expression in an active form in bacteria has not been reported. To achieve the production of active tPA in E. coli, we have investigated the effect of cooverexpressing native (DsbA and DsbC) or heterologous (rat and yeast protein disulfide isomerases) cysteine oxidoreductases in the bacterial periplasm. Coexpression of DsbC, an enzyme which catalyzes disulfide bond isomerization in the periplasm, was found to dramatically increase the formation of active tPA both in shake flasks and in fermentors. The active protein was purified with an overall yield of 25% by using three affinity steps with, in sequence, lysine-Sepharose, immobilized Erythrina caffra inhibitor, and Zn-Sepharose resins. After purification, approximately 180 μg of tPA with a specific activity nearly identical to that of the authentic protein can be obtained per liter of culture in a high-cell-density fermentation. Thus, heterologous proteins as complex as tPA may be produced in an active form in bacteria in amounts suitable for structure-function studies. In addition, these results suggest the feasibility of commercial production of extremely complex proteins in E. coli without the need for in vitro refolding. PMID:9835579

  6. Ectopic norrin induces growth of ocular capillaries and restores normal retinal angiogenesis in Norrie disease mutant mice.

    PubMed

    Ohlmann, Andreas; Scholz, Michael; Goldwich, Andreas; Chauhan, Bharesh K; Hudl, Kristiane; Ohlmann, Anne V; Zrenner, Eberhart; Berger, Wolfgang; Cvekl, Ales; Seeliger, Mathias W; Tamm, Ernst R

    2005-02-16

    Norrie disease is an X-linked retinal dysplasia that presents with congenital blindness, sensorineural deafness, and mental retardation. Norrin, the protein product of the Norrie disease gene (NDP), is a secreted protein of unknown biochemical function. Norrie disease (Ndp(y/-)) mutant mice that are deficient in norrin develop blindness, show a distinct failure in retinal angiogenesis, and completely lack the deep capillary layers of the retina. We show here that the transgenic expression of ectopic norrin under control of a lens-specific promoter restores the formation of a normal retinal vascular network in Ndp(y/-) mutant mice. The improvement in structure correlates with restoration of neuronal function in the retina. In addition, lenses of transgenic mice with ectopic expression of norrin show significantly more capillaries in the hyaloid vasculature that surrounds the lens during development. In vitro, lenses of transgenic mice in coculture with microvascular endothelial cells induce proliferation of the cells. Transgenic mice with ectopic expression of norrin show more bromodeoxyuridine-labeled retinal progenitor cells at embryonic day 14.5 and thicker retinas at postnatal life than wild-type littermates, indicating a putative direct neurotrophic effect of norrin. These data provide direct evidence that norrin induces growth of ocular capillaries and that pharmacologic modulation of norrin might be used for treatment of the vascular abnormalities associated with Norrie disease or other vascular disorders of the retina.

  7. The efficacy of the systemic methotrexate treatment in caesarean scar ectopic pregnancy: A quantitative review of English literature.

    PubMed

    Bodur, S; Özdamar, Ö; Kılıç, S; Gün, I

    2015-04-01

    To determine the efficacy and safety of primary medical treatment with systemic methotrexate (MTX) in caesarean scar ectopic pregnancy, we conducted a Medline/PubMed search on the relevant English literature from January 1978 to January 2012. The search yielded 27 publications of 40 cases of caesarean scar ectopic pregnancy. The literature search showed a very liberal use of systemic MTX treatment with unfavourable outcomes, although the major determinant of the clinical efficacy was found in here to be β-hCG level together with embryonic cardiac activity (ECA) status. A caesarean scar ectopic pregnancy presented with a serum β-hCG concentration of ≤ 12,000 mIU/ml (odds ratio, OR 5.68, 95% confidence interval, CI, 1.37-23.48) and absence of ECA (OR 4.80, 95% CI, 1.14-20.08) was found to be associated with higher efficacy rate of primary systemic MTX treatment. Administration of primary systemic MTX treatment was found to be ideal for a caesarean scar ectopic pregnancy presented before 8 weeks' gestation, with a β-hCG concentration of ≤ 12,000 mIU/ml together with an absent ECA (OR 14.52, 95% CI, 2.36-89.09).

  8. Active Learning of Regular Expressions for Entity Extraction.

    PubMed

    Bartoli, Alberto; De Lorenzo, Andrea; Medvet, Eric; Tarlao, Fabiano

    2017-03-24

    We consider the automatic synthesis of an entity extractor, in the form of a regular expression, from examples of the desired extractions in an unstructured text stream. This is a long-standing problem for which many different approaches have been proposed, which all require the preliminary construction of a large dataset fully annotated by the user. In this paper, we propose an active learning approach aimed at minimizing the user annotation effort: the user annotates only one desired extraction and then merely answers extraction queries generated by the system. During the learning process, the system digs into the input text for selecting the most appropriate extraction query to be submitted to the user in order to improve the current extractor. We construct candidate solutions with genetic programming (GP) and select queries with a form of querying-by-committee, i.e., based on a measure of disagreement within the best candidate solutions. All the components of our system are carefully tailored to the peculiarities of active learning with GP and of entity extraction from unstructured text. We evaluate our proposal in depth, on a number of challenging datasets and based on a realistic estimate of the user effort involved in answering each single query. The results demonstrate high accuracy with significant savings in terms of computational effort, annotated characters, and execution time over a state-of-the-art baseline.

  9. Cloning, expression and biological activity of equine interleukin (IL)-5.

    PubMed

    Cunningham, F M; Vandergrifft, E; Bailey, S R; Sepulveda, M F; Goode, N T; Horohov, D W

    2003-09-15

    The cytokine, interleukin (IL)-5 stimulates eosinophil differentiation, activation and survival and can prime these cells, increasing the response to other mediators. In view of its many effects on eosinophils, IL-5 has been implicated in the pathogenesis of allergic disease in man. Here we report the cloning of equine IL-5 and expression of the recombinant protein by transfection of Chinese hamster ovary (CHO) cells. The cloned cDNA sequence consisted of 405 nucleotides and encoded a protein of 135 amino acids. There is >85% identity with feline, bovine, ovine, canine, and human IL-5 sequences at the nucleotide and protein level. Supernatants containing equine IL-5 were also examined for biological activity. CHO supernatant containing equine recombinant (eqr) IL-5, like the human ortholog (hrIL-5), induced concentration dependent equine eosinophil adherence to autologous serum-coated plastic (9.7+/-1.5% with a 1:100 dilution of eqrIL-5 and 9.1+/-1.6% adherence with 1 nM hrIL-5; n = 4). The eqr protein also caused concentration dependent superoxide production (11.9+/-2.4 nmol (reduced cytochrome (cyt) C)/10(6) cells at a 1:50 dilution, n = 4). In contrast, hrIL-5 only caused significant superoxide production when diluted in conditioned CHO medium, an effect that was inhibited by the anti-human mAb, TRFK5 (4.4+/-0.3 versus 0.3+/-0.4 nmol/10(6) cells for 0.5 nM hrIL-5 in the presence of the isotype matched IgG1 control (10 microM) and TRFK5 (10 microM), respectively). TRFK5 also significantly inhibited hrIL-5 induced adherence at concentrations of 0.3 microg/ml and above but had no significant inhibitory effect on either superoxide or adherence caused by eqrIL-5. These results demonstrate that equine IL-5 expressed by CHO cells stimulates equine eosinophils, suggesting that this cytokine could play a role in eosinophil recruitment and activation in equine allergic disease. The anti-human and murine moAb TRFK5 does not appear to recognise the equine protein.

  10. Transcriptome analysis of ectopic chloroplast development in green curd cauliflower (Brassica oleracea L. var. botrytis)

    PubMed Central

    2011-01-01

    Background Chloroplasts are the green plastids where photosynthesis takes place. The biogenesis of chloroplasts requires the coordinate expression of both nuclear and chloroplast genes and is regulated by developmental and environmental signals. Despite extensive studies of this process, the genetic basis and the regulatory control of chloroplast biogenesis and development remain to be elucidated. Results Green cauliflower mutant causes ectopic development of chloroplasts in the curd tissue of the plant, turning the otherwise white curd green. To investigate the transcriptional control of chloroplast development, we compared gene expression between green and white curds using the RNA-seq approach. Deep sequencing produced over 15 million reads with lengths of 86 base pairs from each cDNA library. A total of 7,155 genes were found to exhibit at least 3-fold changes in expression between green and white curds. These included light-regulated genes, genes encoding chloroplast constituents, and genes involved in chlorophyll biosynthesis. Moreover, we discovered that the cauliflower ELONGATED HYPOCOTYL5 (BoHY5) was expressed higher in green curds than white curds and that 2616 HY5-targeted genes, including 1600 up-regulated genes and 1016 down-regulated genes, were differently expressed in green in comparison to white curd tissue. All these 1600 up-regulated genes were HY5-targeted genes in the light. Conclusions The genome-wide profiling of gene expression by RNA-seq in green curds led to the identification of large numbers of genes associated with chloroplast development, and suggested the role of regulatory genes in the high hierarchy of light signaling pathways in mediating the ectopic chloroplast development in the green curd cauliflower mutant. PMID:22112144

  11. Clones of ectopic stem cells in the regeneration of muscle defects in vivo.

    PubMed

    Yang, Rujing; Chen, Mo; Lee, Chang Hun; Yoon, Richard; Lal, Shan; Mao, Jeremy J

    2010-10-20

    Little is known about whether clones of ectopic, non-muscle stem cells contribute to muscle regeneration. Stem/progenitor cells that are isolated for experimental research or therapeutics are typically heterogeneous. Non-myogenic lineages in a heterogeneous population conceptually may compromise tissue repair. In this study, we discovered that clones of mononucleated stem cells of human tooth pulp fused into multinucleated myotubes that robustly expressed myosin heavy chain in vitro with or without co-culture with mouse skeletal myoblasts (C2C12 cells). Cloned cells were sustainably Oct4+, Nanog+ and Stro1+. The fusion indices of myogenic clones were approximately 16-17 folds greater than their parent, heterogeneous stem cells. Upon infusion into cardio-toxin induced tibialis anterior muscle defects, undifferentiated clonal progenies not only engrafted and colonized host muscle, but also expressed human dystrophin and myosin heavy chain more efficaciously than their parent heterogeneous stem cell populations. Strikingly, clonal progenies yielded ∼9 times more human myosin heavy chain mRNA in regenerating muscles than those infused with their parent, heterogeneous stem cells. The number of human dystrophin positive cells in regenerating muscles infused with clonal progenies was more than ∼3 times greater than muscles infused with heterogeneous stem cells from which clonal progenies were derived. These findings suggest the therapeutic potential of ectopic myogenic clones in muscle regeneration.

  12. Restriction of Ectopic Eye Formation by Drosophila Teashirt and Tiptop to the Developing Antenna

    PubMed Central

    Datta, Rhea R.; Lurye, Jessica M.; Kumar, Justin P.

    2009-01-01

    Summary In Drosophila, the retinal determination network comprises a set of nuclear factors whose loss-of-function phenotypes often include the complete or near total elimination of the developing eye. These genes also share the ability of being able to induce ectopic eye formation when forcibly expressed in non-retinal tissues such as the antennae, legs, halteres, wings and genitals. However, it appears that the ability to redirect and transform tissue fates is limited; not all tissues and cell populations can be forced into adopting an eye fate. In this report we demonstrate that ectopic<