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Sample records for mta1 gene expression

  1. MTA3 regulates CGB5 and Snail genes in trophoblast

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Ying [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, Grand Rapids, MI 49503 (United States); Miyazaki, Jun [Department of Obstetrics and Gynecology, Fujita Health University School of Medicine, Fujita Health University, Toyoake (Japan); Division of Molecular Genetics, Institute for Comprehensive Medical Science, Fujita Health University, Toyoake (Japan); Nishizawa, Haruki [Department of Obstetrics and Gynecology, Fujita Health University School of Medicine, Fujita Health University, Toyoake (Japan); Kurahashi, Hiroki [Division of Molecular Genetics, Institute for Comprehensive Medical Science, Fujita Health University, Toyoake (Japan); Leach, Richard, E-mail: Richard.Leach@hc.msu.edu [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, Grand Rapids, MI 49503 (United States); Department of Obstetrics, Gynecology and Women’s Health, Spectrum Health Medical Group, Grand Rapids, MI 49503 (United States); Wang, Kai, E-mail: Kai.Wang@hc.msu.edu [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, Grand Rapids, MI 49503 (United States)

    2013-04-19

    Highlights: •Impaired MTA3, raised CGB5 and Snail expression are associated with preeclampsia. •Knock-down of MTA3 causes up-regulation of CGB5 and Snail genes in BeWo cells. •MTA3 occupies CGB5 and Snail gene promoters in BeWo cells. -- Abstract: Secreted by the placental trophoblast, human chorionic gonadotropin (hCG) is an important hormone during pregnancy and is required for the maintenance of pregnancy. Previous studies have shown that dys-regulation of hCG expression is associated with preeclampsia. However, the exact relationship between altered hCG levels and development of preeclampsia is unknown. Metastasis associated protein 3 (MTA3), a chromatin remodeling protein, is abundantly expressed in the placental trophoblasts, but its function is unknown. In breast cancer, MTA3 has been shown to repress the expression of Snail and cell migration. However, whether MTA3 acts similarly in the trophoblast has not been investigated. In the present study, we examined the role of MTA3 in regulating the hCG β-subunit gene (gene name: CGB5) and Snail expression in the trophoblast cell line, BeWo, as well as its relevance to the high hCG expression levels seen in preeclampsia. First, we investigated MTA3 expression in preeclamptic placenta as compared to normal control placenta via gene expression microarray and qRT-PCR and found that MTA3 was significantly down-regulated, whereas both CGB5 and Snail were up-regulated in preeclamptic placenta. Secondly, we knocked down MTA3 gene in trophoblast cell line BeWo and found Snail and hCG were both up-regulated, suggesting that MTA3 represses Snail and hCG gene expression in trophoblasts. Next, we cloned the CGB5 and Snail promoters into the pGL3-basic vector individually and found that silencing of MTA3 by siRNA resulted in an increase of both CGB5 and Snail promoter activities. To confirm that this MTA3 inhibition is a direct effect, we performed a chromatin immune-precipitation (ChIP) assay and found that MTA3

  2. MTA3 regulates CGB5 and Snail genes in trophoblast

    International Nuclear Information System (INIS)

    Chen, Ying; Miyazaki, Jun; Nishizawa, Haruki; Kurahashi, Hiroki; Leach, Richard; Wang, Kai

    2013-01-01

    Highlights: •Impaired MTA3, raised CGB5 and Snail expression are associated with preeclampsia. •Knock-down of MTA3 causes up-regulation of CGB5 and Snail genes in BeWo cells. •MTA3 occupies CGB5 and Snail gene promoters in BeWo cells. -- Abstract: Secreted by the placental trophoblast, human chorionic gonadotropin (hCG) is an important hormone during pregnancy and is required for the maintenance of pregnancy. Previous studies have shown that dys-regulation of hCG expression is associated with preeclampsia. However, the exact relationship between altered hCG levels and development of preeclampsia is unknown. Metastasis associated protein 3 (MTA3), a chromatin remodeling protein, is abundantly expressed in the placental trophoblasts, but its function is unknown. In breast cancer, MTA3 has been shown to repress the expression of Snail and cell migration. However, whether MTA3 acts similarly in the trophoblast has not been investigated. In the present study, we examined the role of MTA3 in regulating the hCG β-subunit gene (gene name: CGB5) and Snail expression in the trophoblast cell line, BeWo, as well as its relevance to the high hCG expression levels seen in preeclampsia. First, we investigated MTA3 expression in preeclamptic placenta as compared to normal control placenta via gene expression microarray and qRT-PCR and found that MTA3 was significantly down-regulated, whereas both CGB5 and Snail were up-regulated in preeclamptic placenta. Secondly, we knocked down MTA3 gene in trophoblast cell line BeWo and found Snail and hCG were both up-regulated, suggesting that MTA3 represses Snail and hCG gene expression in trophoblasts. Next, we cloned the CGB5 and Snail promoters into the pGL3-basic vector individually and found that silencing of MTA3 by siRNA resulted in an increase of both CGB5 and Snail promoter activities. To confirm that this MTA3 inhibition is a direct effect, we performed a chromatin immune-precipitation (ChIP) assay and found that MTA3

  3. MTA1 regulation of ERβ pathway in salivary gland carcinoma cells

    International Nuclear Information System (INIS)

    Ohshiro, Kazufumi; Kumar, Rakesh

    2015-01-01

    Abstracts: Although Metastatic-tumor antigen 1 (MTA1) is differentially expressed in metastatic cancer and coregulates the status and activity of nuclear receptors, its role upon estrogen receptor β (ERβ) – a potent tumor suppressor, remains poorly understood. Here we investigated whether MTA1 regulates the expression and functions of ERβ, an ER isoform predominantly expressed in salivary gland cancer cells. We found that the depletion of the endogenous MTA1 in the HSG and HSY salivary duct carcinoma cell lines enhances the expression of ERβ while MTA1 overexpression augmented the expression of ERβ in salivary duct carcinoma cells. Furthermore, MTA1 knockdown inhibited the proliferations and invasion of HSG and HSY cells. The noted ERβ downregulation by MTA1 overexpression involves the process of proteasomal degradation, as a proteasome inhibitor could block it. In addition, both MTA1 knockdown and ERβ overexpression attenuated the cell migration and inhibited the ERK1/2 signaling in the both cell lines. These findings imply that MTA1 dysregulation in a subset of salivary gland cancer might promote aggressive phenotypes by compromising the tumor suppressor activity of ERβ, and hence, MTA1-ERβ axis might serve a new therapeutic target for the salivary gland cancer. - Highlights: • MTA1 silencing upregulates ERβ expression in salivary gland carcinoma cells. • MTA1 overexpression downregulates ERβ expression via proteasomal degradation. • Upregulation of ERβ expression inhibits cell migration and ERK signaling. • MTA1 knockdown inhibits cell proliferation and invasion

  4. MTA family of coregulators in nuclear receptor biology and pathology

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    Manavathi, Bramanandam; Singh, Kamini; Kumar, Rakesh

    2007-01-01

    Nuclear receptors (NRs) rely on coregulators (coactivators and corepressors) to modulate the transcription of target genes. By interacting with nucleosome remodeling complexes, NR coactivators potentiate transcription, whereas corepressors inhibit transcription of the target genes. Metastasis-associated proteins (MTA) represent an emerging family of novel NR coregulators. In general, MTA family members form independent nucleosome remodeling and deacetylation (NuRD) complexes and repress the transcription of different genes by recruiting histone deacetylases onto their target genes. However, MTA1 also acts as a coactivator in a promoter-context dependent manner. Recent findings that repression of estrogen receptor transactivation functions by MTA1, MTA1s, and MTA2 and regulation of MTA3 by estrogen signaling have indicated the significance of these proteins in NR signaling. Here, we highlight the action of MTA proteins on NR signaling and their roles in pathophysiological conditions. PMID:18174918

  5. Curcumin increases the sensitivity of Paclitaxel-resistant NSCLC cells to Paclitaxel through microRNA-30c-mediated MTA1 reduction.

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    Lu, Yimin; Wang, Jun; Liu, Lei; Yu, Lequn; Zhao, Nian; Zhou, Xingju; Lu, Xudong

    2017-04-01

    Non-small-cell lung cancer is one of the most lethal cancers in the worldwide. Although Paclitaxel-based combinational therapies have long been used as a standard treatment in aggressive non-small-cell lung cancers, Paclitaxel resistance emerges as a major clinical problem. It has been demonstrated that Curcumin from Curcuma longa as a traditional Chinese medicine can inhibit cancer cell proliferation. However, the role of Curcumin in Paclitaxel-resistant non-small-cell lung cancer cells is not clear. In this study, we investigated the effect of Curcumin on the Paclitaxel-resistant non-small-cell lung cancer cells and found that Curcumin treatment markedly increased the sensitivity of Paclitaxel-resistant non-small-cell lung cancer cells to Paclitaxel. Mechanically, the study revealed that Curcumin could reduce the expression of metastasis-associated gene 1 (MTA1) gene through upregulation of microRNA-30c in Paclitaxel-resistant non-small-cell lung cancer cells. During the course, MTA1 reduction sensitized Paclitaxel-resistant non-small-cell lung cancer cells and enhanced the effect of Paclitaxel. Taken together, our studies indicate that Curcumin increases the sensitivity of Paclitaxel-resistant non-small-cell lung cancer cells to Paclitaxel through microRNA-30c-mediated MTA1 reduction. Curcumin might be a potential adjuvant for non-small-cell lung cancer patients during Paclitaxel treatment.

  6. Transcriptome and proteomic analyses reveal multiple differences associated with chloroplast development in the spaceflight-induced wheat albino mutant mta.

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    Kui Shi

    Full Text Available Chloroplast development is an integral part of plant survival and growth, and occurs in parallel with chlorophyll biosynthesis. However, little is known about the mechanisms underlying chloroplast development in hexaploid wheat. Here, we obtained a spaceflight-induced wheat albino mutant mta. Chloroplast ultra-structural observation showed that chloroplasts of mta exhibit abnormal morphology and distribution compared to wild type. Photosynthetic pigments content was also significantly decreased in mta. Transcriptome and chloroplast proteome profiling of mta and wild type were done to identify differentially expressed genes (DEGs and proteins (DEPs, respectively. In total 4,588 DEGs including 1,980 up- and 2,608 down-regulated, and 48 chloroplast DEPs including 15 up- and 33 down-regulated were identified in mta. Classification of DEGs revealed that most were involved in chloroplast development, chlorophyll biosynthesis, or photosynthesis. Besides, transcription factors such as PIF3, GLK and MYB which might participate in those pathways were also identified. The correlation analysis between DEGs and DEPs revealed that the transcript-to-protein in abundance was functioned into photosynthesis and chloroplast relevant groups. Real time qPCR analysis validated that the expression level of genes encoding photosynthetic proteins was significantly decreased in mta. Together, our results suggest that the molecular mechanism for albino leaf color formation in mta is a thoroughly regulated and complicated process. The combined analysis of transcriptome and proteome afford comprehensive information for further research on chloroplast development mechanism in wheat. And spaceflight provides a potential means for mutagenesis in crop breeding.

  7. Bioactivity, physical and chemical properties of MTA mixed with propylene glycol

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    Vaishali Prakash NATU

    2015-08-01

    Full Text Available AbstractObjective To investigate the physical (setting time, hardness, flowability, microstructure and chemical (pH change, calcium release, crystallinity properties and the biological outcomes (cell survival and differentiation of mineral trioxide aggregate (MTA mixed using different proportions of propylene glycol (PG and water.Material and Methods White MTA was mixed with different water/PG ratios (100/0, 80/20 and 50/50. Composition (XRD, microstructure (SEM, setting time (ASTM C266-13, flowability (ANSI/ADA 57-2000, Knoop hardness (100 g/10 s and chemical characteristics (pH change and Ca2+ release for 7 days were evaluated. Cell proliferation, osteo/odontoblastic gene expression and mineralization induced by MTA mixed with PG were evaluated. MTA discs (5 mm in diameter, 2 mm thick were prepared and soaked in culture medium for 7 days. Next, the discs were removed and the medium used to culture dental pulp stem cells (DPSC for 28 days. Cells survival was evaluated using MTS assay (24, 72 and 120 h and differentiation with RT-PCR (ALP, OCN, Runx2, DSPP and MEPE and alizarin red staining (7 and 14 days. Data were analysed using one-way ANOVA and Tukey’s post-hoc analysis (a=0.05.Results The addition of PG significantly increased setting time, flowability and Ca2+ release, but it compromised the hardness of the material. SEM showed that 50/50 group resulted porous material after setting due to the incomplete setting reaction, as shown by XRD analysis. The addition of PG (80/20 and 50/50 was not capable to improve cell proliferation or to enhance gene expression, and mineralized deposition of DPSC after 7 and 14 days as compared to the 100/0.Conclusion Except for flowability, the addition of PG did not promote further improvements on the chemical and physical properties evaluated, and it was not capable of enhancing the bioactivity of the MTA.

  8. Upregulation of metastasis-associated gene 2 promotes cell proliferation and invasion in nasopharyngeal carcinoma

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    Wu MH

    2016-03-01

    Full Text Available Minhua Wu,1,2,* Xiaoxia Ye,2,* Xubin Deng,3,* Yanxia Wu,4 Xiaofang Li,4 Lin Zhang11Department of Histology and Embryology, Southern Medical University, Guangzhou, 2Department of Histology and Embryology, Guangdong Medical University, Zhanjiang, 3Affiliated Cancer Hospital of Guangzhou Medical University, Cancer Center of Guangzhou Medical University, Guangzhou, 4Pathological Diagnosis and Research Center, Affiliated Hospital of Guangdong Medical University, Zhanjiang, People’s Republic of China*These authors contributed equally to this workAims: Metastasis-associated gene 2 (MTA2 is reported to play an important role in tumor progression, but little is known about the role of MTA2 in nasopharyngeal carcinoma (NPC. The aim of the study was to explore the expression and function of MTA2 in NPC.Methods: Expression of MTA2 in NPC tissues and cell lines was detected by immunohistochemistry and Western blotting. Relationship between MTA2 expression and clinicopathological features was analyzed. Stable MTA2-overexpressing and MTA2-siliencing NPC cells were established by transfection with plasmids encoding MTA2 cDNA and lentivirus-mediated short hairpin RNA, respectively. Cell viability was determined by Cell Counting Kit-8 and colony formation assay. Cell migration ability was evaluated by wound healing and transwell invasion assay. The impact of MTA2 knockdown on growth and metastasis of CNE2 cells in vivo was determined by nude mouse xenograft models. Expression of several Akt pathway proteins was detected by Western blotting.Results: MTA2 was upregulated in NPC tissues and three NPC cell lines detected (CNE1, CNE2, and HNE1. MTA2 expression was related to clinical stage and lymph node metastasis of patients with NPC. MTA2 upregulation promoted proliferation and invasion of CNE1 cells, while MTA2 depletion had opposite effects on CNE2 cells. Moreover, MTA2 depletion suppressed growth and metastasis of CNE2 cells in vivo. MTA2 overexpression

  9. A novel, highly conserved metallothionein family in basidiomycete fungi and characterization of two representative SlMTa and SlMTb genes in the ectomycorrhizal fungus Suillus luteus.

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    Nguyen, Hoai; Rineau, François; Vangronsveld, Jaco; Cuypers, Ann; Colpaert, Jan V; Ruytinx, Joske

    2017-07-01

    The basidiomycete Suillus luteus is an important member of the ectomycorrhizal community that thrives in heavy metal polluted soils covered with pioneer pine forests. This study aimed to identify potential heavy metal chelators in S. luteus. Two metallothionein (MT) coding genes, SlMTa and SlMTb, were identified. When heterologously expressed in yeast, both SlMTa and SlMTb can rescue the Cu sensitive mutant from Cu toxicity. In S. luteus, transcription of both SlMTa and SlMTb is induced by Cu but not Cd or Zn. Several putative Cu-sensing and metal-response elements are present in the promoter sequences. These results indicate that SlMTa and SlMTb function as Cu-thioneins. Homologs of the S. luteus MTs are present in 49 species belonging to 10 different orders of the subphylum Agaricomycotina and are remarkably conserved. The length of the proteins, number and distribution of cysteine residues indicate a novel family of fungal MTs. The ubiquitous and highly conserved features of these MTs suggest that they are important for basic cellular functions in species in the subphylum Agaricomycotina. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  10. Novel nanohybrid MTA compositions

    DEFF Research Database (Denmark)

    Lovschall, Henrik; Ranjkesh, Bahram

    Original Mineral Trioxide Aggregate (MTA) has been recommended for various uses in endodontics. MTA is a calcia, silica, alumina containing hydraulic cement with long setting time, calcium hydroxide release, and high pH. It possesses antibacterial and sealing properties. Interestingly, recent...... studies describe hydroxyapatite crystals form over MTA when it comes in contact with simulated physiological tissue fluid. On the basis of available evidence it appears that MTA efficiently develop superior marginal adaptation in humid environments. Observations suggest MTA is suitable material for pulp...... modified calcia-silica-alumina-sulphate composition with soluble fluorides, radiocontrast material, and nanosilicate. iMTA thereby demonstrate a remarkably high compressive strength in comparison to several commonly used dental cements. Reduction of iMTA setting time to 2-10 min with either flowable...

  11. Staining Potential of Neo MTA Plus, MTA Plus, and Biodentine Used for Pulpotomy Procedures.

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    Camilleri, Josette

    2015-07-01

    Mineral trioxide aggregate (MTA) used for pulpotomy procedures in immature permanent teeth can reduce treatment to 1 session as opposed to classic calcium hydroxide therapy, which necessitates multiple appointments. The main disadvantage of MTA use is crown discoloration after treatment. The aim of this study was to characterize 3 materials that are used for pulpotomy procedures in immature permanent teeth and assess their color stability in the presence of sodium hypochlorite solution. Hydrated Neo MTA Plus (Avalon Biomed Inc, Bradenton, FL), MTA Plus (Avalon Biomed Inc), and Biodentine (Septodont, Saint-Maur-des-Fossés, France) were characterized after immersion in Hank's balanced salt solution for 1 day and 28 days using a combination of scanning electron microscopy, energy-dispersive spectroscopy, and X-ray diffraction analysis. The color stability of the 3 materials in contact with water or sodium hypochlorite was evaluated by photography, spectrophotometry, and X-ray diffraction analysis. All the materials hydrated and produced calcium hydroxide as a by-product of hydration at early age. All materials interacted with synthetic tissue fluid, forming a calcium phosphate phase. MTA Plus exhibited discoloration in contact with sodium hypochlorite. All the materials tested are suitable to be used in the treatment of immature teeth because they all produced calcium hydroxide, which is necessary to induce dentin bridge formation and continued root formation. Neo MTA Plus and Biodentine are suitable alternatives to MTA, and they do not exhibit discoloration. Copyright © 2015 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  12. Cytotoxicity and Initial Biocompatibility of Endodontic Biomaterials (MTA and Biodentine™) Used as Root-End Filling Materials.

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    Escobar-García, Diana María; Aguirre-López, Eva; Méndez-González, Verónica; Pozos-Guillén, Amaury

    2016-01-01

    Objective. The aim of this study was to evaluate the cytotoxicity and cellular adhesion of Mineral Trioxide Aggregate (MTA) and Biodentine (BD) on periodontal ligament fibroblasts (PDL). Methods. PDL cells were obtained from nonerupted third molars and cultured; MTS cellular profusion test was carried out in two groups: MTA and BD, with respective controls at different time periods. Also, the LIVE/DEAD assay was performed at 24 h. For evaluation of cellular adhesion, immunocytochemistry was conducted to discern the expression of Integrin β1 and Vinculin at 12 h and 24 h. Statistical analysis was performed by the Kruskal-Wallis and Mann-Whitney U tests. Results. MTA and BD exhibited living cells up to 7 days. More expressions of Integrin β1 and Vinculin were demonstrated in the control group, followed by BD and MTA, which also showed cellular loss and morphological changes. There was a significant difference in the experimental groups cultured for 5 and 7 days compared with the control, but there was no significant statistical difference between both cements. Conclusions. Neither material was cytotoxic during the time evaluated. There was an increase of cell adhesion through the expression of focal contacts observed in the case of BD, followed by MTA, but not significantly.

  13. Effect of ProRoot MTA, Portland cement, and amalgam on the expression of fibronectin, collagen I, and TGFβ by human periodontal ligament fibroblasts in vitro.

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    Fayazi, Sara; Ostad, Seyed Nasser; Razmi, Hasan

    2011-01-01

    Today many materials have been introduced for root-end filling materials. One of them is mineral trioxide aggregate (MTA) that is mentioned as a gold standard. The purpose of this in vitro study was to evaluate the reaction of human periodontal ligament fibroblasts to the root-end filling materials, such as ProRoot MTA, Portland cement, and amalgam. Eight impacted teeth were extracted in aseptic condition. The tissues around the roots were used to obtain fibroblast cells. After cell proliferation, they were cultured in the chamber slides and the extracts of the materials were added to the wells. Immunocytochemical method for measuring the expression of Fibronectin, collagen I and transforming growth factor beta (TGF®) was performed by Olysia Bioreport Imaging Software. The results were analyzed by SPSS 13.0 and Tukey post hoc test with PPortland cement group showed the most expression of collagen significantly and after 1 week, Portland cement and MTA groups had the most expression of collagen but there was no significant difference between these 2 groups. After 1 week, the Portland cement group demonstrated a higher amount of TGF® and fibronectin. The results suggest that Portland cement can be used as a less expensive root filling material with low toxicity. It has better effects than amalgam on the fibroblasts.

  14. Physicochemical properties of calcium silicate-based formulations MTA Repair HP and MTA Vitalcem.

    Science.gov (United States)

    Guimarães, Bruno Martini; Prati, Carlo; Duarte, Marco Antonio Hungaro; Bramante, Clovis Monteiro; Gandolfi, Maria Giovanna

    2018-04-05

    This study aimed to analyze the following physicochemical properties: radiopacity, final setting time, calcium release, pH change, solubility, water sorption, porosity, surface morphology, and apatite-forming ability of two calcium silicate-based materials. We tested MTA Repair HP and MTA Vitalcem in comparison with conventional MTA, analyzing radiopacity and final setting time. Water absorption, interconnected pores and apparent porosity were measured after 24-h immersion in deionized water at 37°C. Calcium and pH were tested up to 28 d in deionized water. We analyzed data using two-way ANOVA with Student-Newman-Keuls tests (pcalcium release at 28 d (pcalcium phosphate on their surface after 28 d in HBSS. MTA Repair HP and MTA Vitalcem had extended alkalinizing activity and calcium release that favored calcium phosphate nucleation. The presence of the plasticizer in MTA HP might increase its solubility and porosity. The radiopacifier calcium tungstate can be used to replace bismuth oxide.

  15. Characterization of a new sealer MTA/HAp; Caracterizacao de um novo cimento endodontico MTA/HAp

    Energy Technology Data Exchange (ETDEWEB)

    Barros, C.M.B.; Oliveira, S.V.; Costa, A.C.F.M. [Universidade Federal de Campina Grande (UFCG), PB (Brazil); Dantas, D.C.R.E.; Fontes, L.B.C. [Universidade Estadual da Paraiba (UEPB), PB (Brazil); Viana, K.M.S., E-mail: kalineviana@ect.ufrn.br [Universidade Federal do Rio Grande do Norte (UFRN), Natal, RN (Brazil)

    2012-07-01

    Filling materials (cement-based calcium hydroxide, amalgam, glass ionomer and IRM) are often used as sealers. However, none of them meets the desirable characteristics of a suitable sealing material. This study aims to characterize the new sealer, MTA/HAp from the incorporation of 5% by weight of synthetic hydroxyapatite to the MTA and to investigate the influence of the addition of HAp on the characteristic structural, chemical and consistency of the MTA. For this, we used MTA Angelus ® and HAp synthesized in the laboratory. We used the ratio by mass 95%MTA/5%HAp, where the hydroxyapatite added the characteristic of the MTA osteoconduction. The tests were performed: XRD, XRF and consistency. The results showed that both the new cement had the same crystalline phases as the same chemical composition of its precursors, yet having suitable consistency (plastic mass) of a sealer as well as the pure MTA. (author)

  16. In vitro cytotoxicity of white MTA, MTA Fillapex® and Portland cement on human periodontal ligament fibroblasts.

    Science.gov (United States)

    Yoshino, Patrícia; Nishiyama, Celso Kenji; Modena, Karin Cristina da Silva; Santos, Carlos Ferreira; Sipert, Carla Renata

    2013-01-01

    The aim of this study was to compare the in vitro cytotoxicity of white mineral trioxide aggregate (MTA), MTA Fillapex® and Portland cement (PC) on human cultured periodontal ligament fibroblasts. Periodontal ligament fibroblast culture was established and the cells were used for cytotoxic tests after the fourth passage. Cell density was set at 1.25 X10 4 cells/well in 96-well plates. Endodontic material extracts were prepared by placing sealer/cement specimens (5x3mm) in 1mL of culture medium for 72 h. The extracts were then serially two-fold diluted and inserted into the cell-seeded wells for 24, 48 and 72 h. MTT assay was employed for analysis of cell viability. Cell supernatants were tested for nitric oxide using the Griess reagent system. MTA presented cytotoxic effect in undiluted extracts at 24 and 72 h. MTA Fillapex® presented the highest cytotoxic levels with important cell viability reduction for pure extracts and at ½ and ¼ dilutions. In this study, PC did not induce alterations in fibroblast viability. Nitric oxide was detected in extract-treated cell supernatants and also in the extracts only, suggesting presence of nitrite in the soluble content of the tested materials. In the present study, MTA Fillapex displayed the highest cytotoxic effect on periodontal ligament fibroblasts followed by white MTA and PC.

  17. Chemistry and Bioactivity of NeoMTA Plus™ versus MTA Angelus® Root Repair Materials

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    Sawsan T. Abu Zeid

    2017-01-01

    Full Text Available Objectives. To analyse the chemistry and bioactivity of NeoMTA Plus in comparison with the conventional root repair materials. Method and Materials. Unhydrated and hydrated (initial and final sets materials were analysed by Fourier transform infrared (FTIR spectroscopy and X-ray diffraction (XRD. For bioactivity study, small holes of dentin discs were filled with either materials, immersed in PBS for 15 days, and analysed with FTIR and scanning electron microscope with energy dispersive X-ray (SEM/EDX. The calculation of crystallinity and carbonate/phosphate (CO3/PO4 ratio of surface precipitates (from FTIR and calcium/phosphate (Ca/P ratio (from EDX was statistically analysed using t-test or ANOVA, respectively, at 0.05 significance. Results. Both materials are tricalcium silicate-based that finally react to be calcium silicate hydrate. NeoMTA Plus has relatively high aluminium and sulfur content, with tantalum oxide as an opacifier instead of zirconium oxide in MTA Angelus. NeoMTA Plus showed better apatite formation, higher crystallinity and Ca/P but lower CO3/PO4 ratio than MTA Angelus. SEM showed globular structure with a small particle size in NeoMTA Plus while spherical structure with large particle size in MTA Angelus. Conclusion. Due to fast setting, higher crystallinity, and better bioactivity of NeoMTA Plus, it can be used as a pulp and root repair material.

  18. Evaluation of bacterial leakage of four root- end filling materials: Gray Pro Root MTA, White Pro Root MTA, Root MTA and Portland Cement (type I

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    Zarabian M.

    2005-07-01

    Full Text Available Background and Aim: Today several materials have been used for root- end filling in endodontic surgery. Optimal properties of Pro Root MTA in in-vitro and in-vivo studies has been proven. On the other hand, based on some studies, Root MTA (Iranian Pro Root MTA and Portland cement are similar to Pro Root MTA in physical and biologic properties. The aim of this study was to evaluate bacterial leakage (amount and mean leakage time of four root- end filling materials. Materials and Methods: In this experimental in-vitro study, seventy six extracted single- rooted human teeth were randomly divided into six groups for root-end filling with gray Pro Root MTA, white Pro Root MTA, Root MTA (Iranian Pro Root MTA, Portland Cement (type I and positive and negative control groups. Root canals were instrumented using the step- back technique. Root- end filling materials were placed in 3mm ultra sonic retro preparations. Samples and microleakage model system were sterilized in autoclave. The apical 3-4 mm of the roots were immersed in phenol red with 3% lactose broth culture medium. The coronal access of each specimen was inoculated every 24h with a suspension of Streptococcus sanguis (ATCC 10556. Culture media were observed every 24h for colour change indicating bacterial contamination for 60 days. Statistical analysis was performed using log- rank test with P<0.05 as the limit of significance. Results: At the end of study 50%, 56.25%, 56.25% and 50% of specimens filled with Gray Pro Root MTA, White Pro Root MTA. Root MTA and Portland Cement (type I had evidence of leakage respectively. The mean leakage time was 37.19±6.29, 36.44±5.81, 37.69±5.97 and 34.81±6.67 days respectively. Statistical analysis of data showed no significant difference among the leakage (amount and mean leakage time of the four tested root- end filling materials (P=0.9958. Conclusion: Based on the results of this study, there were no significant differences in leakage among the four

  19. RNAi-Based Identification of Gene-Specific Nuclear Cofactor Networks Regulating Interleukin-1 Target Genes

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    Johanna Meier-Soelch

    2018-04-01

    Full Text Available The potent proinflammatory cytokine interleukin (IL-1 triggers gene expression through the NF-κB signaling pathway. Here, we investigated the cofactor requirements of strongly regulated IL-1 target genes whose expression is impaired in p65 NF-κB-deficient murine embryonic fibroblasts. By two independent small-hairpin (shRNA screens, we examined 170 genes annotated to encode nuclear cofactors for their role in Cxcl2 mRNA expression and identified 22 factors that modulated basal or IL-1-inducible Cxcl2 levels. The functions of 16 of these factors were validated for Cxcl2 and further analyzed for their role in regulation of 10 additional IL-1 target genes by RT-qPCR. These data reveal that each inducible gene has its own (quantitative requirement of cofactors to maintain basal levels and to respond to IL-1. Twelve factors (Epc1, H2afz, Kdm2b, Kdm6a, Mbd3, Mta2, Phf21a, Ruvbl1, Sin3b, Suv420h1, Taf1, and Ube3a have not been previously implicated in inflammatory cytokine functions. Bioinformatics analysis indicates that they are components of complex nuclear protein networks that regulate chromatin functions and gene transcription. Collectively, these data suggest that downstream from the essential NF-κB signal each cytokine-inducible target gene has further subtle requirements for individual sets of nuclear cofactors that shape its transcriptional activation profile.

  20. Characterization of a new sealer MTA/HAp

    International Nuclear Information System (INIS)

    Barros, C.M.B.; Oliveira, S.V.; Costa, A.C.F.M.; Dantas, D.C.R.E.; Fontes, L.B.C.; Viana, K.M.S.

    2012-01-01

    Filling materials (cement-based calcium hydroxide, amalgam, glass ionomer and IRM) are often used as sealers. However, none of them meets the desirable characteristics of a suitable sealing material. This study aims to characterize the new sealer, MTA/HAp from the incorporation of 5% by weight of synthetic hydroxyapatite to the MTA and to investigate the influence of the addition of HAp on the characteristic structural, chemical and consistency of the MTA. For this, we used MTA Angelus ® and HAp synthesized in the laboratory. We used the ratio by mass 95%MTA/5%HAp, where the hydroxyapatite added the characteristic of the MTA osteoconduction. The tests were performed: XRD, XRF and consistency. The results showed that both the new cement had the same crystalline phases as the same chemical composition of its precursors, yet having suitable consistency (plastic mass) of a sealer as well as the pure MTA. (author)

  1. Interaction of dental pulp stem cells with Biodentine and MTA after exposure to different environments

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    Anastasia Agrafioti

    Full Text Available ABSTRACT Objective: The aim of the present study was to evaluate and compare the cytotoxic effects of Biodentine and MTA on dental pulp stem cells (DPSCs and to assess cell viability and adherence after material exposure to an acidic environment. Material and Methods: DPSCs were cultured either alone or in contact with either: Biodentine; MTA set for 1 hour; or MTA set for 24 hours. After 4 and 7 days, cell viability was measured using the MTT assay. Biodentine and MTA were also prepared and packed into standardized bovine dentin disks and divided into three groups according to the storage media (n=6/group: freshly mixed materials without storage medium (Group A; materials stored in saline (Group B; materials stored in citric acid buffered at pH 5.4 (Group C. After 24 hours, DPSCs were introduced in the wells and cell adherence, viability, and cellular morphology were observed via confocal microscopy after three days of culture. Cell viability was analyzed using repeated-measures analysis of variance test with Tukey's post hoc tests (α=0.05. Results: Biodentine expressed significantly higher cell viability compared with all other groups after 4 days, with no differences after 7 days. Notably, cell viability was significantly greater in 24-hour set MTA compared with 1-hour set MTA and control groups after 7 days. Material exposure to an acidic environment showed an increase in cell adherence and viability in both groups. Conclusions: Biodentine induced a significantly accelerated cell proliferation compared with MTA. Setting of these materials in the presence of citric acid enhanced DPSC viability and adherence.

  2. Insight into the architecture of the NuRD complex: structure of the RbAp48-MTA1 subcomplex.

    Science.gov (United States)

    Alqarni, Saad S M; Murthy, Andal; Zhang, Wei; Przewloka, Marcin R; Silva, Ana P G; Watson, Aleksandra A; Lejon, Sara; Pei, Xue Y; Smits, Arne H; Kloet, Susan L; Wang, Hongxin; Shepherd, Nicholas E; Stokes, Philippa H; Blobel, Gerd A; Vermeulen, Michiel; Glover, David M; Mackay, Joel P; Laue, Ernest D

    2014-08-08

    The nucleosome remodeling and deacetylase (NuRD) complex is a widely conserved transcriptional co-regulator that harbors both nucleosome remodeling and histone deacetylase activities. It plays a critical role in the early stages of ES cell differentiation and the reprogramming of somatic to induced pluripotent stem cells. Abnormalities in several NuRD proteins are associated with cancer and aging. We have investigated the architecture of NuRD by determining the structure of a subcomplex comprising RbAp48 and MTA1. Surprisingly, RbAp48 recognizes MTA1 using the same site that it uses to bind histone H4, showing that assembly into NuRD modulates RbAp46/48 interactions with histones. Taken together with other results, our data show that the MTA proteins act as scaffolds for NuRD complex assembly. We further show that the RbAp48-MTA1 interaction is essential for the in vivo integration of RbAp46/48 into the NuRD complex. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. A Histologic Evaluation on Tissue Reaction to Three Implanted Materials (MTA, Root MTA and Portland Cement Type I in the Mandible of Cats

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    F. Sasani

    2004-09-01

    Full Text Available Statement of Problem: Nowadays Mineral Trioxide aggregate (MTA is widely used for root end fillings, pulp capping, perforation repair and other endodontic treatments.Investigations have shown similar physical and chemical properties for Portland cement and Root MTA with those described for MTA.Purpose: The aim of this in vitro study was to evaluate the tissue reaction to implanted MTA, Portland cement and Root MTA in the mandible of cats.Materials and Methods: Under asepsis condition and general anesthesia, a mucoperiosteal flap, following the application of local anesthesia, was elevated to expose mandibular symphysis. Two small holes in both sides of mandible were drilled. MTA, Portland cement and Root MTA were mixed according to the manufacturers, recommendation and placed in bony cavities. In positive control group, the test material was Zinc oxide powder plus tricresoformalin. In negative control group, the bony cavities were left untreated. After 3,6 and 12 weeks, the animals were sacrificed and the mandibular sections were prepared for histologic examination under light microscope. The presence and thickness of inflammation, presence of fibrosis capsule, the severity of fibrosis and bone formation were investigated. The data were submitted to Exact Fisher test, chi square test and Kruskal-Wallis test for statistical analysis.Results: No statistically significant differences were found in the degree of inflammation,presence of fibrotic capsule, severity of fibrosis and inflammation thickness between Root MTA, Portland cement and MTA (P>0.05. There was no statistical difference in boneformation between MTA and Portland cement (P>0.05. However, bone formation was not found in any of the Root MTA specimens and the observed tissue was exclusively of fibrosis type.Conclusion: The physical and histological results observed with MTA are similar to those of Root MTA and Portland cement. Additionally, all of these three materials are biocompatible

  4. A comparative histologic study on furcal perforation repair with Root MTA and Pro Root MTA in fully developed teeth in dog

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    Rahimi S.

    2005-07-01

    Full Text Available Background and Aim: The goal of endodontics is to seal the root canal system from the orifice to apical constriction completely and tridimensionally.Hence perforations during root canal therapy, because of caries or resorptions must be sealed and obturated with ideal materials. The aim of this study was to histologically compare two kinds of mineral trioxide aggregate Root MTA and Pro Root MTA for furcal perforation repair in developed teeth in dog. Materials and Methods: In this experimental study, thirty teeth consisting of second, third and fourth mandibular premolars of five German shepherd dogs were selected. Twenty-four teeth were randomly divided into four experimental groups (6 teeth each. One pair of Root MTA and Pro Root MTA groups studied in one month and the other in three months intervals. Positive and negative control groups was each contained three teeth. In positive control group, perforations were not treated and negative control group contained intact teeth. In experimental groups perforations repaired after one week exposure to oral cavity with Root MTA or Pro Root MTA. After time intervals animals were subjected to vital perfusion and 6 m histologic sections were prepared. Inflammation and hard tissue formation were ranked by Cox criteria. Data were analysed using Mann-Whitney and Chi-Square statistical tests with P0.05. Conclusion: Mineral Trioxide Aggregate is an adequate material for furcal perforation repair in dog’s teeth. Root MTA could be a good substitute for Pro Root MTA considering the lower cost and similar characteristics.

  5. Analysis of marginal seal of ProRoot MTA, MTA Angelus biodentine, and glass ionomer cement as root-end filling materials: An in vitro study

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    Sakshi Malhotra

    2015-01-01

    Conclusion: Microleakage was present in all the samples. Least amount of apical dye microleakage was seen in biodentine with mean value of 0.16 mm followed by ProRoot MTA 0.68 mm, MTA Angelus 0.74 mm, and GIC 1.53 mm. The best sealing ability was seen in biodentine, and this difference was statistically significant.

  6. Presence of arsenic in different types of MTA and white and gray Portland cement.

    Science.gov (United States)

    Monteiro Bramante, Clóvis; Demarchi, Ana Claudia Cardoso Oliveira; de Moraes, Ivaldo Gomes; Bernadineli, Norberti; Garcia, Roberto Brandão; Spångberg, Lars S W; Duarte, Marco Antonio Hungaro

    2008-12-01

    The presence of arsenic in various types of mineral trioxide aggregate (MTA) and Portland cements were evaluated to verify if they comply with the ISO-recommended limit for water-based cements of 2 mg arsenic/kg material. An amount of 5 mL of hydrochloric acid was added to 2 g each of MTA and Portland cement to be analyzed. After 15 minutes, the material was filtered and the volume of supernatant was diluted with reagent-grade water up to 40 mL. Atomic absorption spectrophotometry readings were performed in triplicate. The following mean values were obtained: CPM (Egeo, Buenos Aires, Argentina) 11.06 mg/kg; CPM sealer (Egeo) 10.30 mg/kg; MTA-Obtura (Angelus, Londrina, PR, Brazil) 0.39 mg/kg; Experimental MTA: 10.30 mg/kg; White MTA-Angelus (Angelus) 1.03 mg/kg; Gray MTA-Angelus (Angelus) 5.91 mg/kg; ProRoot-MTA (Dentsply/Tulsa Dental Specialties, Tulsa, OK) 5.25 mg/kg; Gray Portland cement (Votorantim Cimentos, Cubatão, SP, Brazil): 34.27 mg/kg; and White Portland cement (Cimento Rio Branco, Rio de Janeiro, RJ, Brazil) 0.52 mg/kg. All tested materials presented arsenic in their composition. The form of arsenic was not analyzed nor the toxicity of the arsenic found. Only MTA-Obtura, White MTA-Angelus, and White Portland cement presented arsenic levels below the limit set in the ISO 9917-1 standard.

  7. The effect of blood and synthetic tissue fluid on the microhardness of ProRoot MTA, OrthoMTA and RetroMTA

    Directory of Open Access Journals (Sweden)

    Noushin Shokouhinejad

    2017-12-01

    Results: Exposure to blood significantly decreased the microhardness of all materials at all three points of 0.5, 2 and 3.5 mm (P<0.001. At level of 0.5 and 2 mm distant from blood, OrthoMTA showed significantly the least microhardness value; however, at the point of 3.5 mm, the microhardness of RetroMTA was higher than the two other materials (P<0.001. After exposure of samples to distilled water or PBS, no significant difference was found between the materials at any levels of 0.5, 2, and 3.5 mm (P<0.01. Conclusion: Blood exposure resulted in the decrease of microhardness of internal part of the materials.

  8. Comparison between visual assessment of MTA and hippocampal volumes in an elderly, non-demented population

    Energy Technology Data Exchange (ETDEWEB)

    Cavallin, Lena; Axelsson, Rimma [CLINTEC, Div. of Medical Imaging and Technology, Karolinska Inst., Karolinska Univ. Hospital, Stockholm (Sweden); Dept. of Radiology, Karolinska Univ. Hospital, Stockholm (Sweden)], e-mail: lena.cavallin@karolinska.se; Bronge, Lena [CLINTEC, Div. of Medical Imaging and Technology, Karolinska Inst., Karolinska Univ. Hospital, Stockholm (Sweden); Aleris Diagnostics, Stockholm (Sweden); Zhang, Yi [NVS, Novum, Karolinska Inst., Karolinska Univ. Hospital, Stockholm (Sweden); Oeksengaard, Anne-Rita [NVS, Novum, Karolinska Inst., Karolinska Univ. Hospital, Stockholm (Sweden); Ulleval Univ. Hospital and Asker and Baerum Hospital, Oslo (Norway); Wahlund, Lars-Olof [NVS, Novum, Karolinska Inst., Karolinska Univ. Hospital, Stockholm (Sweden); Swedish Brain Power, Karolinska Univ. Hospital, Stockholm (Sweden); Fratiglioni, Laura [ARC Karolinska Inst. Stockholm (Sweden)

    2012-06-15

    Background: It is important to have a replicable easy method for monitoring atrophy progression in Alzheimer's disease. Volumetric methods for calculating hippocampal volume are time-consuming and commonly used in research. Visual assessments of medial temporal lobe atrophy (vaMTA) is a rapid method for clinical use. This method has not been tested in a large non-demented population in comparison with volumetry measurements. Since hippocampal volume decreases with time even in normal aging there is also a need to study the normal age differences of medial temporal lobe atrophy. Purpose: To compare visual assessment of medial temporal lobe atrophy (vaMTA) with hippocampal volume in a healthy, non-demented elderly population. To describe normal ageing using vaMTA. Material and Methods: Non-demented individuals aged 60, 66, 72, 78, 81, 84, and {>=}87 years old were recruited from the Swedish National study on Ageing and Care in Kungsholmen (SNAC-K), Sweden. Standard magnetic resonance imaging (MRI) scans, vaMTA, and calculations of hippocampal volumes were performed in 544 subjects. Results: Significant correlation (rs = -0.32, P < 0.001, sin; and rs = -0.26, P < 0.001, dx) was found between hippocampal volume measurements and vaMTA. In normal ageing, almost 95% of {<=}66-year-olds had a medial temporal lobe atrophy (MTA) score {<=}1, with possible scores ranging from 0 to 4. Subjects aged 72, 78, and 81 years scored {<=}2, while the two oldest age groups had scores {<=}3. Conclusion: There was a highly significant correlation between volumetric measurements of the hippocampus and MTA scoring. In normal ageing, there is increasing MTA score. For non-demented elderly individuals {<=}70 years, an MTA score of 0-1 may be considered normal, compared with MTA {<=}2 for 70-80-years and MTA 3 for >80-year-old individuals.

  9. Nidogen-1 regulates laminin-1-dependent mammary-specific gene expression

    Energy Technology Data Exchange (ETDEWEB)

    Pujuguet, Philippe; Simian, Marina; Liaw, Jane; Timpl, Rupert; Werb, Zena; Bissell, Mina J..

    2000-02-01

    Nidogen-1 (entactin) acts as a bridge between the extracellular matrix molecules laminin-1 and type IV collagen, and thus participates in the assembly of basement membranes. To investigate the role of nidogen-1 in regulating cell-type-specific gene expression in mammary epithelium, we designed a culture microecosystem in which each component, including epithelial cells, mesenchymal cells, lactogenic hormones and extracellular matrix, could be controlled. We found that primary and established mesenchymal and myoepithelial cells synthesized and secreted nidogen-1, whereas expression was absent in primary and established epithelial cells. In an epithelial cell line containing mesenchymal cells, nidogen-1 was produced by the mesenchymal cells but deposited between the epithelial cells. In this mixed culture, mammary epithelial cells express b-casein in the presence of lactogenic hormones. Addition of either laminin-1 plus nidogen-1, or laminin-1 alone to mammary epithelial cells induced b- casein production. We asked whether recombinant nidogen-1 alone could signal directly for b-casein. Nidogen-1 did not induce b-casein synthesis in epithelial cells, but it augmented the inductive capacity of laminin-1. These data suggest that nidogen-1 can cooperate with laminin-1 to regulate b-casein expression. Addition of full length nidogen-1 to the mixed cultures had no effect on b-casein gene expression; however, a nidogen-1 fragment containing the laminin-1 binding domain, but lacking the type IV collagen-binding domain, had a dominant negative effect on b-casein expression. These data point to a physiological role for nidogen-1 in the basement membrane-induced gene expression by epithelial cells.

  10. Cryogenics for the MuCool Test Area (MTA)

    International Nuclear Information System (INIS)

    Darve, Christine; Norris, Barry; Pei, Liujin

    2006-01-01

    MuCool Test Area (MTA) is a complex of buildings at Fermi National Accelerator Laboratory, which are dedicated to operate components of a cooling cell to be used for Muon Collider and Neutrino Factory R and D. The long-term goal of this facility is to test ionization cooling principles by operating a 25-liter liquid hydrogen (LH2) absorber embedded in a 5 Tesla superconducting solenoid magnet. The MTA solenoid magnet will be used with RF cavities exposed to a high intensity beam. Cryogens used at the MTA include LHe, LN2 and LH2. The latter dictates stringent system design for hazardous locations. The cryogenic plant is a modified Tevatron refrigerator based on the Claude cycle. The implementation of an in-house refrigerator system and two 300 kilowatt screw compressors is under development. The helium refrigeration capacity is 500 W at 14 K. In addition the MTA solenoid magnet will be batch-filled with LHe every 2 days using the same cryo-plant. This paper reviews cryogenic systems used to support the Muon Collider and Neutrino Factory R and D programs and emphasizes the feasibility of handling cryogenic equipment at MTA in a safe manner

  11. Photo-Induced depopulation of the 180mTa isomer

    Science.gov (United States)

    Bhike, Megha; Krishichayan, Fnu; Tornow, W.

    2015-10-01

    The 180mTa nucleus is the rarest isotope in the universe, existing only in an isomeric state at 77.2 keV (Jπ = 9-) with half-life of greater than 7.1 ×1015 years. The stellar production of this high-spin isomer has been a challenging astrophysical problem. Cross-section measurements for the depopulation of the 180mTa isomer with monoenergetic photon beams of energies 2.5 and 3.1 MeV have been carried out at the HI γS facility. The activated Ta foils of natural abundance and containing 14.4 mg of 180mTa were γ-ray counted at TUNL's low background facility using a 13% planar HPGe detector. A 8'' ×12'' NaI detector in combination with the standard HI γS scintillator paddle system was employed for absolute photon-flux determination. Preliminary results will be discussed, and measurements at lower energies are planned. This work was supported by the U.S. DOE under Grant NO. DE-FG02-97ER41033.

  12. Iodine-131 dose dependent gene expression in thyroid cancers and corresponding normal tissues following the Chernobyl accident.

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    Michael Abend

    Full Text Available The strong and consistent relationship between irradiation at a young age and subsequent thyroid cancer provides an excellent model for studying radiation carcinogenesis in humans. We thus evaluated differential gene expression in thyroid tissue in relation to iodine-131 (I-131 doses received from the Chernobyl accident. Sixty three of 104 papillary thyroid cancers diagnosed between 1998 and 2008 in the Ukrainian-American cohort with individual I-131 thyroid dose estimates had paired RNA specimens from fresh frozen tumor (T and normal (N tissue provided by the Chernobyl Tissue Bank and satisfied quality control criteria. We first hybridized 32 randomly allocated RNA specimen pairs (T/N on 64 whole genome microarrays (Agilent, 4×44 K. Associations of differential gene expression (log(2(T/N with dose were assessed using Kruskall-Wallis and trend tests in linear mixed regression models. While none of the genes withstood correction for the false discovery rate, we selected 75 genes with a priori evidence or P kruskall/P trend <0.0005 for validation by qRT-PCR on the remaining 31 RNA specimen pairs (T/N. The qRT-PCR data were analyzed using linear mixed regression models that included radiation dose as a categorical or ordinal variable. Eleven of 75 qRT-PCR assayed genes (ACVR2A, AJAP1, CA12, CDK12, FAM38A, GALNT7, LMO3, MTA1, SLC19A1, SLC43A3, ZNF493 were confirmed to have a statistically significant differential dose-expression relationship. Our study is among the first to provide direct human data on long term differential gene expression in relation to individual I-131 doses and to identify a set of genes potentially important in radiation carcinogenesis.

  13. Appendix 1:Upregulated genes in gene expression profile (P<0.05 ...

    Indian Academy of Sciences (India)

    lazi

    Appendix 1: Upregulated genes in gene expression profile«P2). Probe_s. Gene_Symbol pvalues foldchange. Probe_S. et_ID. Gene_Symbol pvalues foldchange. et_ID. 1370355. 1393751. Scd1. 1.35E-04. 25.77. Loc1009122508.06E-03. 2.55. -at at. 1398250. 1370870. Acot1. 2.43E-02. 12.18. Me1.

  14. Histologic evaluation of pulpal response to MTA and capsaicin in cats

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    Zahrasadat Madani

    2012-09-01

    Full Text Available Introduction: In direct pup capping, the exposed pulp is directly capped with a capping material to provoke a dentinal bridge formation to seal the exposed area.The aim of this study was to evaluate the pulpal response to MTA and capsaicin in cats. Methods: The sample for this experimental study consisted of 24 canines of cats, weighed approximately 3-4kg. After sedation, the teeth were cut and exposed at 3mm above cementoenamel junction, then divided into two groups: 1 direct pulp capping with MTA 2 direct pulp capping with Capsaicin. The cavities were filled with glass ionomer cement (Fuji IX. The cats were sacrificed in the first, second, and fourth weeks 4 canine teeth were extracted for the purpose of histologic analysis. 6Serial sections were cut parallel to the longitudinal axis of the canines. The sections were stained with hematoxylin & eosin, and were observed and analyzed using a light microscope. Results: In both groups treated with MTA and capsaicin, the inflammation decreased during weeks1, 2, and 4. Less inflammation was seen in MTA group but there was no statistical difference between the two groups (p=0.22. In the group treated with capsaicin, necrosis was observed in every 12 samples, but in the other group treated with MTA, only 1 pulp necrosis was seen in the first week (p=0.000. There was no significant difference in dentinal barrier formation, inflammatory response, and soft tissue changes between the two groups. Conclusions: The results of this study showed that capsaicin can decrease the severity of inflammation, but it is ineffective in dentinal barrier formation.

  15. Comparison of Nasal Epithelial Smoking-Induced Gene Expression on Affymetrix Exon 1.0 and Gene 1.0 ST Arrays

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    Xiaoling Zhang

    2013-01-01

    Full Text Available We have previously defined the impact of tobacco smoking on nasal epithelium gene expression using Affymetrix Exon 1.0 ST arrays. In this paper, we compared the performance of the Affymetrix GeneChip Human Gene 1.0 ST array with the Human Exon 1.0 ST array for detecting nasal smoking-related gene expression changes. RNA collected from the nasal epithelium of five current smokers and five never smokers was hybridized to both arrays. While the intersample correlation within each array platform was relatively higher in the Gene array than that in the Exon array, the majority of the genes most changed by smoking were tightly correlated between platforms. Although neither array dataset was powered to detect differentially expressed genes (DEGs at a false discovery rate (FDR <0.05, we identified more DEGs than expected by chance using the Gene ST array. These findings suggest that while both platforms show a high degree of correlation for detecting smoking-induced differential gene expression changes, the Gene ST array may be a more cost-effective platform in a clinical setting for gene-level genomewide expression profiling and an effective tool for exploring the host response to cigarette smoking and other inhaled toxins.

  16. Psychotic Symptoms in Attention-Deficit/Hyperactivity Disorder: An Analysis of the MTA Database.

    Science.gov (United States)

    Vitiello, Benedetto; Perez Algorta, Guillermo; Arnold, L Eugene; Howard, Andrea L; Stehli, Annamarie; Molina, Brooke S G

    2017-04-01

    To assess the prevalence of psychotic symptoms among youths (14-25 years of age) with a childhood diagnosis of attention-deficit/hyperactivity disorder (ADHD) combined type. Participants in the Multimodal Treatment Study of Children with ADHD (MTA) and a local normative comparison group (LNCG) were systematically assessed 6, 8, 10, 12, 14, and 16 years after the original enrollment at a mean age of 8.5 years. Trained research assistants administered a psychosis screener, and positive screens were referred to study clinicians to confirm or exclude psychosis. Possible associations between screening positive and alcohol or substance use were assessed. Data were available from 509 MTA participants (88% of original MTA sample; mean age 25.1 years) and 276 LNCG participants (96% of original sample; mean age 24.6 years) at year 16. Twenty-six MTA participants (5%; 95% CI 3-7) and 11 LNCG participants (4%; 95% CI 2-6) screened positive for at least 1 psychotic symptom (p = .60). Most psychotic symptoms were transient. The prevalence of clinician-confirmed psychotic symptoms was 1.1% (95% CI 0.2-2.1) in the MTA group and 0.7% (0-1.7) in the LNCG (p = .72). Greater cannabis use was reported by those who screened positive (p ADHD increased the risk for psychotic symptoms. In the ADHD and normative comparison groups, more frequent cannabis use was associated with a greater likelihood of experiencing psychotic symptoms, thus supporting the recommendation that youth should not use cannabis. Published by Elsevier Inc.

  17. Genomewide analysis of gene expression associated with Tcof1 in mouse neuroblastoma

    International Nuclear Information System (INIS)

    Mogass, Michael; York, Timothy P.; Li, Lin; Rujirabanjerd, Sinitdhorn; Shiang, Rita

    2004-01-01

    Mutations in the Treacher Collins syndrome gene, TCOF1, cause a disorder of craniofacial development. We manipulated the levels of Tcof1 and its protein treacle in a murine neuroblastoma cell line to identify downstream changes in gene expression using a microarray platform. We identified a set of genes that have similar expression with Tcof1 as well as a set of genes that are negatively correlated with Tcof1 expression. We also showed that the level of Tcof1 and treacle expression is downregulated during differentiation of neuroblastoma cells into neuronal cells. Inhibition of Tcof1 expression by siRNA induced morphological changes in neuroblastoma cells that mimic differentiation. Thus, expression of Tcof1 and treacle synthesis play an important role in the proliferation of neuroblastoma cells and we have identified genes that may be important in this pathway

  18. Gene expression response to EWS–FLI1 in mouse embryonic cartilage

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    Miwa Tanaka

    2014-12-01

    Full Text Available Ewing's sarcoma is a rare bone tumor that affects children and adolescents. We have recently succeeded to induce Ewing's sarcoma-like small round cell tumor in mice by expression of EWS–ETS fusion genes in murine embryonic osteochondrogenic progenitors. The Ewing's sarcoma precursors are enriched in embryonic superficial zone (eSZ cells of long bone. To get insights into the mechanisms of Ewing's sarcoma development, gene expression profiles between EWS–FLI1-sensitive eSZ cells and EWS–FLI1-resistant embryonic growth plate (eGP cells were compared using DNA microarrays. Gene expression of eSZ and eGP cells (total, 30 samples was evaluated with or without EWS–FLI1 expression 0, 8 or 48 h after gene transduction. Our data provide useful information for gene expression responses to fusion oncogenes in human sarcoma.

  19. Differential expression and interaction of host factors augment HIV-1 gene expression in neonatal mononuclear cells

    International Nuclear Information System (INIS)

    Sundaravaradan, Vasudha; Mehta, Roshni; Harris, David T.; Zack, Jerome A.; Ahmad, Nafees

    2010-01-01

    We have previously shown a higher level of HIV-1 replication and gene expression in neonatal (cord) blood mononuclear cells (CBMC) compared with adult blood cells (PBMC), which could be due to differential expression of host factors. We performed the gene expression profile of CBMC and PBMC and found that 8013 genes were expressed at higher levels in CBMC than PBMC and 8028 genes in PBMC than CBMC, including 1181 and 1414 genes upregulated after HIV-1 infection in CBMC and PBMC, respectively. Several transcription factors (NF-κB, E2F, HAT-1, TFIIE, Cdk9, Cyclin T1), signal transducers (STAT3, STAT5A) and cytokines (IL-1β, IL-6, IL-10) were upregulated in CBMC than PBMC, which are known to influence HIV-1 replication. In addition, a repressor of HIV-1 transcription, YY1, was down regulated in CBMC than PBMC and several matrix metalloproteinase (MMP-7, -12, -14) were significantly upregulated in HIV-1 infected CBMC than PBMC. Furthermore, we show that CBMC nuclear extracts interacted with a higher extent to HIV-1 LTR cis-acting sequences, including NF-κB, NFAT, AP1 and NF-IL6 compared with PBMC nuclear extracts and retroviral based short hairpin RNA (shRNA) for STAT3 and IL-6 down regulated their own and HIV-1 gene expression, signifying that these factors influenced differential HIV-1 gene expression in CBMC than PBMC.

  20. MTA and calcium hydroxide for pulp capping MTA e hidróxido de cálcio para proteção pulpar direta

    Directory of Open Access Journals (Sweden)

    Alexandra Mussolino de Queiroz

    2005-06-01

    Full Text Available This study evaluated the biocompatibility of mineral trioxide aggregate (MTA after direct capping of exposed pulp tissue in dog's teeth. Class I cavities were prepared in 26 teeth from 3 adult dogs. MTA was applied over the exposed pulp in 13 teeth and paste of calcium hydroxide plus distilled water (control was applied in the remaining 13 teeth. After 90 days, the animals were killed; the maxilla and mandible were dissected and sectioned to obtain individual roots. The samples were processed histologically. The pulp and periapical response observed with the use of MTA was similar to that of calcium hydroxide paste. In all specimens, there was a dentin bridge obliterating the exposure, an intact odontoblastic layer, no inflammatory cells, normal connective pulp tissue, normal apical and periapical regions and no bone tissue changes. Similar to calcium hydroxide, MTA presented excellent response when used for direct pulp capping.O objetivo deste trabalho foi avaliar a biocompatibilidade do agregado de trióxido mineral (MTA, após proteção pulpar direta em dentes de cães. Foram preparadas cavidades de Classe I, em 26 dentes de 3 cães adultos. O MTA foi aplicado sobre 13 dentes e a pasta de hidróxido de cálcio (grupo controle foi aplicada sobre os 13 dentes remanescentes. Após 90 dias, os animais foram mortos, a maxila e a mandíbula foram dissecadas e os dentes foram seccionados para obtenção de raízes individualizadas. Os espécimes foram processados histologicamente. A resposta do tecido pulpar e periapical foi semelhante para o MTA e o hidróxido de cálcio. Em todos os espécimes havia ponte de dentina obliterando o local da exposição pulpar, camada odontoblástica íntegra, ausência de células inflamatórias, tecido pulpar normal, e ausência de alterações na região periapical e óssea. Da mesma maneira que o hidróxido de cálcio, o MTA apresentou excelente biocompatibilidade quando usado para proteção pulpar direta.

  1. Alu Elements as Novel Regulators of Gene Expression in Type 1 Diabetes Susceptibility Genes?

    Science.gov (United States)

    Kaur, Simranjeet; Pociot, Flemming

    2015-07-13

    Despite numerous studies implicating Alu repeat elements in various diseases, there is sparse information available with respect to the potential functional and biological roles of the repeat elements in Type 1 diabetes (T1D). Therefore, we performed a genome-wide sequence analysis of T1D candidate genes to identify embedded Alu elements within these genes. We observed significant enrichment of Alu elements within the T1D genes (p-value genes harboring Alus revealed significant enrichment for immune-mediated processes (p-value genes harboring inverted Alus (IRAlus) within their 3' untranslated regions (UTRs) that are known to regulate the expression of host mRNAs by generating double stranded RNA duplexes. Our in silico analysis predicted the formation of duplex structures by IRAlus within the 3'UTRs of T1D genes. We propose that IRAlus might be involved in regulating the expression levels of the host T1D genes.

  2. Imaging gene expression in gene therapy

    International Nuclear Information System (INIS)

    Wiebe, Leonard I.

    1997-01-01

    Full text. Gene therapy can be used to introduce new genes, or to supplement the function of indigenous genes. At the present time, however, there is non-invasive test to demonstrate efficacy of the gene transfer and expression processes. It has been postulated that scintigraphic imaging can offer unique information on both the site at which the transferred gene is expressed, and the degree of expression, both of which are critical issue for safety and clinical efficacy. Many current studies are based on 'suicide gene therapy' of cancer. Cells modified to express these genes commit metabolic suicide in the presence of an enzyme encoded by the transferred gene and a specifically-convertible pro drug. Pro drug metabolism can lead to selective metabolic trapping, required for scintigraphy. Herpes simplex virus type-1 thymidine kinase (H S V-1 t k + ) has been use for 'suicide' in vivo tumor gene therapy. It has been proposed that radiolabelled nucleosides can be used as radiopharmaceuticals to detect H S V-1 t k + gene expression where the H S V-1 t k + gene serves a reporter or therapeutic function. Animal gene therapy models have been studied using purine-([ 18 F]F H P G; [ 18 F]-A C V), and pyrimidine- ([ 123 / 131 I]I V R F U; [ 124 / 131I ]) antiviral nucleosides. Principles of gene therapy and gene therapy imaging will be reviewed and experimental data for [ 123 / 131I ]I V R F U imaging with the H S V-1 t k + reporter gene will be presented

  3. Effect of TPA and HTLV-1 Tax on BRCA1 and ERE controlled genes expression.

    Science.gov (United States)

    Jabareen, Azhar; Abu-Jaafar, Aya; Abou-Kandil, Ammar; Huleihel, Mahmoud

    2017-07-18

    Interference with the expression and/or functions of the multifunctional tumor suppressor BRCA1 leads to a high risk of breast and ovarian cancers. BRCA1 expression is usually activated by the estrogen (E2) liganded ERα receptor. Activated ERα is considered as a potent transcription factor which activates various genes expression by 2 pathways. A classical pathway, ERα binds directly to E2-responsive elements (EREs) in the promoters of the responsive genes and a non-classical pathway where ERα indirectly binds with the appropriate gene promoter. In our previous study, HTLV-1Tax was found to strongly inhibit ERα induced BRCA1 expression while stimulating ERα induced ERE dependent genes. TPA is a strong PKC activator which found to induce the expression of HTLV-1. Here we examined the effect of TPA on the expression of BRCA1 and genes controlled by ERE region in MCF-7 cells and on Tax activity on these genes. Our results showed strong stimulatory effect of TPA on both BRCA1 and ERE expression without treatment with E2. Tax did not show any significant effect on these TPA activities. It seems that TPA activation of BRCA1 and ERE expression is dependent on PKC activity but not through the NFκB pathway. However, 53BP1 may be involved in this TPA activity because its overexpression significantly reduced the TPA stimulatory effect on BRCA1 and ERE expression. Additionally, our Chip assay results probably exclude possible involvement of ERα pathway in this TPA activity because TPA did not interfere with the binding of ERα to both BRCA1 promoter and ERE region.

  4. Gerakan Religio-Kultural MTA Dakwah : Mobilisasi dan Tafsir-Tanding

    NARCIS (Netherlands)

    Sunarwoto,

    2012-01-01

    This article investigates the role of Majelis Tafsir Al-Qur’an (MTA), a dakwah movement operating aminly in Central Java. MTA is known as the Muslim purist movement whose mission focuses on the purification of Islam by utilizing the slogan back to the Qur’an and Sunnah. Using the social movement

  5. Involvement of Resveratrol and ω-3 Polyunsaturated Fatty Acids on Sirtuin 1 Gene Expression in THP1 Cells.

    Science.gov (United States)

    Tsuchiya, Takafumi; Endo, Ayano; Tsujikado, Kyoko; Inukai, Toshihiko

    2017-10-01

    Resveratrol, a kind of polyphenol, has the potential to activate the longevity gene in several cells, in the same manner as calorie restriction. We investigated the effect of resveratrol and ω-3-line polyunsaturated fatty acid on surtuin 1 (SIRT1) gene expression in human monocytes (THP1) cells. We examined the gene expression of THP1 cells using real-time polymerase chain reaction and Western blotting analysis. Resveratol, eicosapentaenoic acid (EPA) and docosahexaeanoic acid (DHA) as n-3 polyunsaturated fatty acid were added on THP1 cells. We observed the changes in the SIRT1 gene expression in those cells, under various doses of agents and in time courses. Then, we examined the interaction of glucose and mannitol on those agents׳ effect of the gene expression. The concentration range of glucose and mannitol was from 5-20mM, respectively. The SIRT1 gene expression could be defined in 24 and 48 hours both in real-time polymerase chain reaction analysis and in Western blotting. Resveratrol showed SIRT1 gene expression in a dose-dependent manner in the range of 0-20μM in both analyses. Although EPA at 10μM showed marked increase in SIRT1 gene expression compared to control condition in Western blotting, this phenomenon was not in dose-dependent manner. DHA did not exhibit any augmentation of SIRT1 gene expression in a dose-dependent manner in the range of 0-20μM in both analyses. We refined the dose-dependent inhibition of the SIRT1 gene expression within 20mM glucose medium. Although 20mM did not exhibit any inhibition, 10μM resveratrol induced the gene expression compared to control medium. Both 5 and 15mM mannitol medium did not significantly alter basic gene expression and 10μM resveratrol-induced gene expression. The present results suggest that resveratrol and EPA, but not DHA, markedly activated the SIRT1 gene expression in THP1 cells, and that high glucose medium could inhibit the basic gene expression, but not powerful resveratrol-induced gene

  6. Imaging gene expression in gene therapy

    Energy Technology Data Exchange (ETDEWEB)

    Wiebe, Leonard I. [Alberta Univ., Edmonton (Canada). Noujaim Institute for Pharmaceutical Oncology Research

    1997-12-31

    Full text. Gene therapy can be used to introduce new genes, or to supplement the function of indigenous genes. At the present time, however, there is non-invasive test to demonstrate efficacy of the gene transfer and expression processes. It has been postulated that scintigraphic imaging can offer unique information on both the site at which the transferred gene is expressed, and the degree of expression, both of which are critical issue for safety and clinical efficacy. Many current studies are based on `suicide gene therapy` of cancer. Cells modified to express these genes commit metabolic suicide in the presence of an enzyme encoded by the transferred gene and a specifically-convertible pro drug. Pro drug metabolism can lead to selective metabolic trapping, required for scintigraphy. Herpes simplex virus type-1 thymidine kinase (H S V-1 t k{sup +}) has been use for `suicide` in vivo tumor gene therapy. It has been proposed that radiolabelled nucleosides can be used as radiopharmaceuticals to detect H S V-1 t k{sup +} gene expression where the H S V-1 t k{sup +} gene serves a reporter or therapeutic function. Animal gene therapy models have been studied using purine-([{sup 18} F]F H P G; [{sup 18} F]-A C V), and pyrimidine- ([{sup 123}/{sup 131} I]I V R F U; [{sup 124}/{sup 131I}]) antiviral nucleosides. Principles of gene therapy and gene therapy imaging will be reviewed and experimental data for [{sup 123}/{sup 131I}]I V R F U imaging with the H S V-1 t k{sup +} reporter gene will be presented

  7. Gene Expression of Glucose Transporter 1 (GLUT1), Hexokinase 1 and Hexokinase 2 in Gastroenteropancreatic Neuroendocrine Tumors

    DEFF Research Database (Denmark)

    Binderup, Tina; Knigge, Ulrich; Federspiel, Birgitte Hartnack

    2013-01-01

    -associated genes and to compare this with FDG-PET imaging as well as with the cellular proliferation index in two cancer entities with different malignant potential. Using real-time PCR, gene expression of GLUT1, HK1 and HK2 were studied in 34 neuroendocrine tumors (NETs) in comparison with 14 colorectal...... adenocarcinomas (CRAs). The Ki67 proliferation index and, when available, FDG-PET imaging was compared with gene expression. Overexpression of GLUT1 gene expression was less frequent in NETs (38%) compared to CRAs (86%), P = 0.004. HK1 was overexpressed in 41% and 71% of NETs and CRAs, respectively (P = 0.......111) and HK2 was overexpressed in 50% and 64% of NETs and CRAs, respectively (P = 0.53). There was a significant correlation between the Ki67 proliferation index and GLUT1 gene expression for the NETs (R = 0.34, P = 0.047), but no correlation with the hexokinases. FDG-PET identified foci in significantly...

  8. [An 18 month evaluation of MM-MTA pulpotomy on primary decayed molars].

    Science.gov (United States)

    Abou Chedid, J C; Mchayleh, N; Khalil, I; Melki, B; Hardan, L S

    2015-12-01

    Pulpotomy is the most performed and controversial therapeutic in pediatric dentistry. Formocresol is known to have a toxic effect on living tissues, a mutagenic and carcinogenic potential with a systemic uptake of formocresol via pulpotomized teeth, other alternative products have been investigated. 40 molars were pulpotomized using Micro Mega Mineral Trioxide Aggregate (MM-MTA), which eliminates the need for the use of formocresol. The effects of this material were evaluated both clinically and radiographically. Post-operative control examinations were performed at 1, 6, 12, and 18 months trying to detect spontaneous or stimulated pain, pathological tooth mobility, abscesses or fistulas, internal or external pathological tooth resorption, periapical bone destruction, or canal obliteration. Pain was absent at 18 months post operatively. Thirty six molar treated with the MM-MTA didn't show any mobility or pain, one molar presented a pathological resorption and one molar presented an abscess without a fistula at 12 month. The observations were compared to others related to formocresol, ferric sulfate, MTA, and laser pulpotomies, using the Chi-square test x2. The abundance of positive result strongly demonstrate that the MM-MTA pulpotomy on carious temporary molars is a promising technique.

  9. Radiopacity evaluation of Portland and MTA-based cements by digital radiographic system.

    Science.gov (United States)

    Borges, Alvaro Henrique; Pedro, Fabio Luiz Miranda; Semanoff-Segundo, Alex; Miranda, Carlos Eduardo Saraiva; Pécora, Jesus Djalma; Cruz Filho, Antônio Miranda

    2011-01-01

    The aim of the present study was to evaluate the radiopacity of Portland and MTA-based cements using the Digora TM digital radiographic system. The performed tests followed specification number 57 from the American National Standard Institute/American Dental Association (2000) for endodontic sealing materials. The materials were placed in 5 acrylic plates, especially designed for this experiment, along with a graduated aluminum stepwedge varying from 1 to 10 mm in thickness. The set was radiographed at a 30 cm focus-object distance and with 0.2 s exposure time. After the radiographs were taken, the optical laser readings of radiographs were performed by Digora TM system. Five radiographic density readings were performed for each studied material and for each step of the aluminum scale. White ProRoot MTA (155.99±8.04), gray ProRoot MTA (155.96±16.30) and MTA BIO (143.13±16.94) presented higher radiopacity values (pPortland (119.76±22.34), gray Portland (109.71±4.90) and white structural Portland (99.59±12.88) presented lower radiopacity values (pcements were the only materials presenting radiopacity within the ANSI/ADA specifications.

  10. Prognostic significance of glucose transporter-1 (GLUT1) gene expression in rectal cancer after preoperative chemoradiotherapy

    International Nuclear Information System (INIS)

    Saigusa, Susumu; Toiyama, Yuji; Tanaka, Koji; Okugawa, Yoshinaga; Fujikawa, Hiroyuki; Matsushita, Kohei; Uchida, Keiichi; Inoue, Yasuhiro; Kusunoki, Masato

    2012-01-01

    Most cancer cells exhibit increased glycolysis. The elevated glucose transporter 1 (GLUT1) expression has been reported to be associated with resistance to therapeutic agents and a poor prognosis. We wondered whether GLUT1 expression was associated with the clinical outcome in rectal cancer after preoperative chemoradiotherapy (CRT), and whether glycolysis inhibition could represent a novel anticancer treatment. We obtained total RNA from residual cancer cells using microdissection from a total of 52 rectal cancer specimens from patients who underwent preoperative CRT. We performed transcriptional analyzes, and studied the association of the GLUT1 gene expression levels with the clinical outcomes. In addition, we examined each proliferative response of three selected colorectal cancer cell lines to a glycolysis inhibitor, 3-bromopyruvic acid (3-BrPA), with regard to their expression of the GLUT1 gene. An elevated GLUT1 gene expression was associated with a high postoperative stage, the presence of lymph node metastasis, and distant recurrence. Moreover, elevated GLUT1 gene expression independently predicted both the recurrence-free and overall survival. In the in vitro studies, we observed that 3-BrPA significantly suppressed the proliferation of colon cancer cells with high GLUT1 gene expression, compared with those with low expression. An elevated GLUT1 expression may be a useful predictor of distant recurrence and poor prognosis in rectal cancer patients after preoperative CRT. (author)

  11. Evaluating the role of CRM1-mediated export for adenovirus gene expression

    International Nuclear Information System (INIS)

    Carter, Christoph C.; Izadpanah, Reza; Bridge, Eileen

    2003-01-01

    A complex of the Adenovirus (Ad) early region 1b 55-kDa (E1b-55kDa) and early region 4 ORF6 34-kDa (E4-34kDa) proteins promotes viral late gene expression. E1b-55kDa and E4-34kDa have leucine-rich nuclear export signals (NESs) similar to that of HIV Rev. It was proposed that E1b-55kDa and/or E4-34kDa might promote the export of Ad late mRNA via their Rev-like NESs, and the transport receptor CRM1. We treated infected cells with the cytotoxin leptomycin B to inhibit CRM1-mediated export; treatment initially delays the onset of late gene expression, but this activity completely recovers as the late phase progresses. We find that the E1b-55kDa NES is not required to promote late gene expression. Previous results showed that E4-34kDa-mediated late gene expression does not require an intact NES (J. Virol. 74 (2000), 6684-6688). Our results indicate that these Ad regulatory proteins promote late gene expression without intact NESs or active CRM1

  12. Angiogenic Gene Signature Derived from Subtype Specific Cell Models Segregate Proneural and Mesenchymal Glioblastoma

    Directory of Open Access Journals (Sweden)

    Aman Sharma

    2017-07-01

    Full Text Available Intertumoral molecular heterogeneity in glioblastoma identifies four major subtypes based on expression of molecular markers. Among them, the two clinically interrelated subtypes, proneural and mesenchymal, are the most aggressive with proneural liable for conversion to mesenchymal upon therapy. Using two patient-derived novel primary cell culture models (MTA10 and KW10, we developed a minimal but unique four-gene signature comprising genes vascular endothelial growth factor A (VEGF-A, vascular endothelial growth factor B (VEGF-B and angiopoietin 1 (ANG1, angiopoietin 2 (ANG2 that effectively segregated the proneural (MTA10 and mesenchymal (KW10 glioblastoma subtypes. The cell culture preclassified as mesenchymal showed elevated expression of genes VEGF-A, VEGF-B and ANG1, ANG2 as compared to the other cell culture model that mimicked the proneural subtype. The differentially expressed genes in these two cell culture models were confirmed by us using TCGA and Verhaak databases and we refer to it as a minimal multigene signature (MMS. We validated this MMS on human glioblastoma tissue sections with the use of immunohistochemistry on preclassified (YKL-40 high or mesenchymal glioblastoma and OLIG2 high or proneural glioblastoma tumor samples (n = 30. MMS segregated mesenchymal and proneural subtypes with 83% efficiency using a simple histopathology scoring approach (p = 0.008 for ANG2 and p = 0.01 for ANG1. Furthermore, MMS expression negatively correlated with patient survival. Importantly, MMS staining demonstrated spatiotemporal heterogeneity within each subclass, adding further complexity to subtype identification in glioblastoma. In conclusion, we report a novel and simple sequencing-independent histopathology-based biomarker signature comprising genes VEGF-A, VEGF-B and ANG1, ANG2 for subtyping of proneural and mesenchymal glioblastoma.

  13. Regional gene expression of LOX-1, VCAM-1, and ICAM-1 in aorta of HIV-1 transgenic rats.

    Directory of Open Access Journals (Sweden)

    Anne Mette Fisker Hag

    Full Text Available BACKGROUND: Increased prevalence of atherosclerotic cardiovascular disease in HIV-infected patients has been observed. The cause of this accelerated atherosclerosis is a matter of controversy. As clinical studies are complicated by a multiplicity of risk-factors and a low incidence of hard endpoints, studies in animal models could be attractive alternatives. METHODOLOGY/PRINCIPAL FINDINGS: We evaluated gene expression of lectin-like oxidized-low-density-lipoprotein receptor-1 (LOX-1, vascular cell adhesion molecule-1 (VCAM-1, and intercellular adhesion molecule-1 (ICAM-1 in HIV-1 transgenic (HIV-1Tg rats; these genes are all thought to play important roles in early atherogenesis. Furthermore, the plasma level of sICAM-1 was measured. We found that gene expressions of LOX-1 and VCAM-1 were higher in the aortic arch of HIV-1Tg rats compared to controls. Also, the level of sICAM-1 was elevated in the HIV-1Tg rats compared to controls, but the ICAM-1 gene expression profile did not show any differences between the groups. CONCLUSIONS/SIGNIFICANCE: HIV-1Tg rats have gene expression patterns indicating endothelial dysfunction and accelerated atherosclerosis in aorta, suggesting that HIV-infection per se may cause atherosclerosis. This transgenic rat model may be a very promising model for further studies of the pathophysiology behind HIV-associated cardiovascular disease.

  14. The putative protein methyltransferase LAE1 controls cellulase gene expression in Trichoderma reesei

    Science.gov (United States)

    Seiboth, Bernhard; Karimi, Razieh Aghcheh; Phatale, Pallavi A; Linke, Rita; Hartl, Lukas; Sauer, Dominik G; Smith, Kristina M; Baker, Scott E; Freitag, Michael; Kubicek, Christian P

    2012-01-01

    Summary Trichoderma reesei is an industrial producer of enzymes that degrade lignocellulosic polysaccharides to soluble monomers, which can be fermented to biofuels. Here we show that the expression of genes for lignocellulose degradation are controlled by the orthologous T. reesei protein methyltransferase LAE1. In a lae1 deletion mutant we observed a complete loss of expression of all seven cellulases, auxiliary factors for cellulose degradation, β-glucosidases and xylanases were no longer expressed. Conversely, enhanced expression of lae1 resulted in significantly increased cellulase gene transcription. Lae1-modulated cellulase gene expression was dependent on the function of the general cellulase regulator XYR1, but also xyr1 expression was LAE1-dependent. LAE1 was also essential for conidiation of T. reesei. Chromatin immunoprecipitation followed by high-throughput sequencing (‘ChIP-seq’) showed that lae1 expression was not obviously correlated with H3K4 di- or trimethylation (indicative of active transcription) or H3K9 trimethylation (typical for heterochromatin regions) in CAZyme coding regions, suggesting that LAE1 does not affect CAZyme gene expression by directly modulating H3K4 or H3K9 methylation. Our data demonstrate that the putative protein methyltransferase LAE1 is essential for cellulase gene expression in T. reesei through mechanisms that remain to be identified. PMID:22554051

  15. Sealing ability of mineral trioxide aggregate (MTA) combined with distilled water, chlorhexidine, and doxycycline.

    Science.gov (United States)

    Arruda, Roberta A A; Cunha, Rodrigo S; Miguita, Kenner B; Silveira, Cláudia F M; De Martin, Alexandre S; Pinheiro, Sérgio L; Rocha, Daniel G P; Bueno, Carlos E S

    2012-09-01

    The aim of this study was to evaluate the sealing ability of mineral trioxide aggregate (MTA Bio) combined with different mixing agents (distilled water, chlorhexidine, doxycycline), used as an apical root-end filling material. Forty-two extracted human teeth were divided into three groups (n = 12); six teeth were used as controls. Root-ends were resected at 90 degrees, 3 mm from the apex. Root-end cavities were prepared using ultrasonic tips and filled with MTA Bio plus distilled water, 2% chlorhexidine solution, or 10% doxycycline solution. Apical sealing was assessed by microleakage of 50% silver nitrate solution. Roots were longitudinally sectioned in a buccolingual plane and analyzed using an operating microscope (20× magnification). Depth of dye leakage into the dentinal walls was measured in millimeters. Results were analyzed using ANOVA and Tukey's test (P = 0.05). MTA Bio plus distilled water showed significantly higher mean leakage results (1.06 mm) when compared with MTA Bio plus doxycycline (0.61 mm), and higher, although not significant, results when compared with MTA Bio plus chlorhexidine (0.79 mm). In conclusion, replacing distilled water with two biologically active mixing agents (doxycycline and chlorhexidine) did not alter the sealing properties of MTABio. The antimicrobial properties of these combinations should be further investigated.

  16. SATB1 tethers multiple gene loci to reprogram expression profiledriving breast cancer metastasis

    Energy Technology Data Exchange (ETDEWEB)

    Han, Hye-Jung; Kohwi, Yoshinori; Kohwi-Shigematsu, Terumi

    2006-07-13

    Global changes in gene expression occur during tumor progression, as indicated by expression profiling of metastatic tumors. How this occurs is poorly understood. SATB1 functions as a genome organizer by folding chromatin via tethering multiple genomic loci and recruiting chromatin remodeling enzymes to regulate chromatin structure and expression of a large number of genes. Here we show that SATB1 is expressed at high levels in aggressive breast cancer cells, and is undetectable in non-malignant breast epithelial cells. Importantly, RNAi-mediated removal of SATB1 from highly-aggressive MDA-MB-231 cells altered the expression levels of over 1200 genes, restored breast-like acinar polarity in three-dimensional cultures, and prevented the metastastic phenotype in vivo. Conversely, overexpression of SATB1 in the less-aggressive breast cancer cell line Hs578T altered the gene expression profile and increased metastasis dramatically in vivo. Thus, SATB1 is a global regulator of gene expression in breast cancer cells, directly regulating crucial metastasis-associated genes, including ERRB2 (HER2/NEU), TGF-{beta}1, matrix metalloproteinase 3, and metastasin. The identification of SATB1 as a protein that re-programs chromatin organization and transcription profiles to promote breast cancer metastasis suggests a new model for metastasis and may provide means of therapeutic intervention.

  17. Learning-dependent gene expression of CREB1 isoforms in the molluscan brain

    Directory of Open Access Journals (Sweden)

    Hisayo Sadamoto

    2010-05-01

    Full Text Available Cyclic AMP-responsive element binding protein1 (CREB1 has multiple functions in gene regulation. Various studies have reported that CREB1-dependent gene induction is necessary for memory formation and long-lasting behavioral changes in both vertebrates and invertebrates. In the present study, we characterized Lymnaea CREB1 (LymCREB1 mRNA isoforms of spliced variants in the central nervous system (CNS of the pond snail Lymnaea stagnalis. Among these spliced variants, the three isoforms that code a whole LymCREB1 protein are considered to be the activators for gene regulation. The other four isoforms, which code truncated LymCREB1 proteins with no kinase inducible domain, are the repressors. For a better understanding of the possible roles of different LymCREB1 isoforms, the expression level of these isoform mRNAs was investigated by a real-time quantitative RT-PCR method. Further, we examined the changes in gene expression for all the isoforms in the CNS after conditioned taste aversion (CTA learning or backward conditioning as a control. The results showed that CTA learning increased LymCREB1 gene expression, but it did not change the activator/repressor ratio. Our findings showed that the repressor isoforms, as well as the activator ones, are expressed in large amounts in the CNS, and the gene expression of CREB1 isoforms appeared to be specific for the given stimulus. This was the first quantitative analysis of the expression patterns of CREB1 isoforms at the mRNA level and their association with learning behavior.

  18. Regulation of glutamate transporter 1 (GLT-1) gene expression by cocaine self-administration and withdrawal.

    Science.gov (United States)

    Kim, Ronald; Sepulveda-Orengo, Marian T; Healey, Kati L; Williams, Emily A; Reissner, Kathryn J

    2018-01-01

    Downregulation of the astroglial glutamate transporter GLT-1 is observed in the nucleus accumbens (NAc) following administration of multiple drugs of abuse. The decrease in GLT-1 protein expression following cocaine self-administration is dependent on both the amount of cocaine self-administered and the length of withdrawal, with longer access to cocaine and longer withdrawal periods leading to greater decreases in GLT-1 protein. However, the mechanism(s) by which cocaine downregulates GLT-1 protein remains unknown. We used qRT-PCR to examine gene expression of GLT-1 splice isoforms (GLT-1A, GLT-1B) in the NAc, prelimbic cortex (PL) and basolateral amygdala (BLA) of rats, following two widely used models of cocaine self-administration: short-access (ShA) self-administration, and the long-access (LgA) self-administration/incubation model. While downregulation of GLT-1 protein is observed following ShA cocaine self-administration and extinction, this model did not lead to a change in GLT-1A or GLT-1B gene expression in any brain region examined. Forced abstinence following ShA cocaine self-administration also was without effect. In contrast, LgA cocaine self-administration and prolonged abstinence significantly decreased GLT-1A gene expression in the NAc and BLA, and significantly decreased GLT-1B gene expression in the PL. No change was observed in NAc GLT-1A gene expression one day after LgA cocaine self-administration, indicating withdrawal-induced decreases in GLT-1A mRNA. In addition, LgA cocaine self-administration and withdrawal induced hypermethylation of the GLT-1 gene in the NAc. These results indicate that a decrease in NAc GLT-1 mRNA is only observed after extended access to cocaine combined with protracted abstinence, and that epigenetic mechanisms likely contribute to this effect. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Effect of MTA-based sealer on the healing of periapical lesions.

    Science.gov (United States)

    Gomes-Filho, João Eduardo; Watanabe, Simone; Cintra, Luciano Tavares Angelo; Nery, Mauro Juvenal; Dezan-Júnior, Eloi; Queiroz, India Olinta Azevedo; Lodi, Carolina Simonetti; Basso, Maria Daniela

    2013-01-01

    Some manufacturers have recently added specific components to improve the ease of handling and insertion material properties of MTA in order to create MTA-based sealers. The aim of this study was to evaluate the healing of periapical lesions in canine teeth after a single session of endodontic treatment with MTA Fillapex® compared with Sealapex® or Endo-CPM-Sealer®. Sixty-two root canals were performed on two 1-year-old male dogs. After coronal access and pulp extirpation, the canals were exposed to the oral cavity for 6 months in order to induce periapical lesions. The root canals were prepared, irrigated with a solution of 2.5% sodium hypochlorite and filled with gutta-percha and different sealers, according to the following groups: 1) Sealapex®; 2) Endo-CPM-Sealer®; and 3) MTA Fillapex®. Some teeth with periapical lesions were left untreated for use as positive controls. Healthy teeth were used as negative controls. After 6 months, the animals were sacrificed and serial sections from the roots were prepared for histomorphologic analysis and stained with hematoxylin and eosin and the Brown and Brenn technique. The lesions were scored according to pre-established histomorphologic parameters and the scores statistically analyzed using the Kruskal-Wallis test. All 3 materials produced similar patterns of healing (p>0.05); in particular, persistent inflammation and absence of complete periapical tissue healing were consistently noted. Preparation of the infected root canals followed by filling with the materials studied was insufficient to provide complete healing of the periapical tissues.

  20. Comparative evaluation of antimicrobial action of MTA, calcium hydroxide and Portland cement Avaliação comparativa da ação antimicrobiana do MTA, hidróxido de cálcio e cimento Portland

    Directory of Open Access Journals (Sweden)

    Caroline Sousa Ribeiro

    2006-10-01

    Full Text Available The present study aimed to evaluate and compare the antimicrobial effect of MTA Dentsply, MTA Angelus, Calcium Hydroxide and Portland cement. Four reference bacterial strains were used: Pseudomonas aeruginosa, Escherichia coli, Bacteroides fragilis, and Enterococcus faecalis. Plates containing Mueller-Hinton agar supplemented with 5% sheep blood, hemin, and menadione were inoculated with the bacterial suspensions. Subsequently, wells were prepared and immediately filled with materials and incubated at 37ºC for 48 hours under anaerobic conditions, except P. aeruginosa. The diameters of inhibition zones were measured, and data analyzed using ANOVA and the Tukey test with 1% level of significance. MTA Dentsply, MTA Angelus and Portland cement inhibited the growth of P. aeruginosa. Calcium Hydroxide was effective against P. aeruginosa and B. fragillis. Under anaerobic conditions, which may hamper the formation of reactive oxygen species, the materials failed to inhibit E. faecalis, and E. coli.O objetivo do presente trabalho foi avaliar e comparar o efeito antimicrobiano do MTA Dentsply, MTA Angelus, hidróxido de cálcio e cimento Portland sobre quatro cepas bacterianas: Pseudomonas aeruginosa, Escherichia coli, Bacteroides fragilis, e Enterococcus faecalis. Placas contendo agar Muller-Hinton suplementadas com 5% de sangue de carneiro, hemina e menadiona foram inoculadas com as suspensões bacterianas. Poços foram confeccionados com auxílio de perfuradores e imediatamente preenchidos com os materiais, e incubados a 37ºC por 48 horas em atmosfera de anaerobiose, exceto P. aeruginosa. O diâmetro dos halos de inibição foi medido e os dados analisados usando o teste estatístico ANOVA e o de Tukey com nível de significância de 1%. O MTA Dentsply, MTA Angelus e Cimento Portland inibiram o crescimento da P.aeruginosa. O hidróxido de cálcio foi efetivo contra P. aeruginosa e B. fragillis. Sob atmosfera de anaerobiose, condição que pode

  1. Image of HSV1-TK gene expression with {sup 123}IVDU

    Energy Technology Data Exchange (ETDEWEB)

    Kim, S. Y.; Woo, K. S.; Chung, W. S. [Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2005-07-01

    The liver is an important target organ for gene transfer due to its capacity for synthesizing serum protein and its involvement in numerous genetic diseases. So livertargeted gene transfer is significant tool for expanding the treatment options and gene function studies. Gene transfer methods commonly use recombinant viral vector. However, viral vectors also have various disadvantages for example immune recognition after adenoviral vector delivery and potential viralassociated toxicity including helper virus replication and insertional mutagenesis. In contrast, nonviral vectors such as naked plasmid DNA(pDNA) and cationic liposomal systems exhibit low immunogenicity and repeated administration is possible(Ledley et al.,1992; Nabel et al.,1993). These are attractive vectors for in vivo gene transfer because of their suitable characteristics such as biodegradability, minimal toxicity, nonimmunogenicity, and simplicity of use. But non-viral gene delivery, has problems associated with limited efficiency at gene expression. hydrodynamic-based produce has very high level efficiency of gene extraction in liver or soild tumor. In mice, hydrodynamic-based produce was reported that a high level of transgene expression could be obtained in the liver by intravenous injection of large volume( 8{approx}10% of body weight) and high-speed ( Kobayashi N et al., 2004 ). HSV1-TK is one of the most widely use effect gene systems sued for imaging gene expression, in association with its use as a suicide gene, or as a reporter gene In non-invasive imaging of the HSV1-TK system, many nucleoside derivatives have developed as prodrug for tumor proliferation imaging or as anti-viral drugs. Several 5-substituted uracil nucleoside derivatives have been identified to have high sensitivity and selective accumulation in HSV1-TK expression cell. This producer has been used hydrodynamic-based produce, we investigated to image of herpes simplex virus type 1 thymidine kinase (HSV1-tk) gene with (E

  2. Gene expression profile analysis of Ligon lintless-1 (Li1) mutant reveals important genes and pathways in cotton leaf and fiber development.

    Science.gov (United States)

    Ding, Mingquan; Jiang, Yurong; Cao, Yuefen; Lin, Lifeng; He, Shae; Zhou, Wei; Rong, Junkang

    2014-02-10

    Ligon lintless-1 (Li1) is a monogenic dominant mutant of Gossypium hirsutum (upland cotton) with a phenotype of impaired vegetative growth and short lint fibers. Despite years of research involving genetic mapping and gene expression profile analysis of Li1 mutant ovule tissues, the gene remains uncloned and the underlying pathway of cotton fiber elongation is still unclear. In this study, we report the whole genome-level deep-sequencing analysis of leaf tissues of the Li1 mutant. Differentially expressed genes in leaf tissues of mutant versus wild-type (WT) plants are identified, and the underlying pathways and potential genes that control leaf and fiber development are inferred. The results show that transcription factors AS2, YABBY5, and KANDI-like are significantly differentially expressed in mutant tissues compared with WT ones. Interestingly, several fiber development-related genes are found in the downregulated gene list of the mutant leaf transcriptome. These genes include heat shock protein family, cytoskeleton arrangement, cell wall synthesis, energy, H2O2 metabolism-related genes, and WRKY transcription factors. This finding suggests that the genes are involved in leaf morphology determination and fiber elongation. The expression data are also compared with the previously published microarray data of Li1 ovule tissues. Comparative analysis of the ovule transcriptomes of Li1 and WT reveals that a number of pathways important for fiber elongation are enriched in the downregulated gene list at different fiber development stages (0, 6, 9, 12, 15, 18dpa). Differentially expressed genes identified in both leaf and fiber samples are aligned with cotton whole genome sequences and combined with the genetic fine mapping results to identify a list of candidate genes for Li1. Copyright © 2013 Elsevier B.V. All rights reserved.

  3. Mineral trioxide aggregate induces osteoblastogenesis via Atf6

    Directory of Open Access Journals (Sweden)

    Toyonobu Maeda

    2015-06-01

    Full Text Available Mineral trioxide aggregate (MTA has been recommended for various uses in endodontics. To understand the effects of MTA on alveolar bone, we examined whether MTA induces osteoblastic differentiation using MC3T3-E1 cells. MTA enhanced mineralization concomitant with alkaline phosphatase activity in a dose- and time-dependent manner. MTA increased production of collagens (Type I and Type III and matrix metalloproteinases (MMP-9 and MMP-13, suggesting that MTA affects bone matrix remodeling. MTA also induced Bglap (osteocalcin but not Bmp2 (bone morphogenetic protein-2 mRNA expression. We observed induction of Atf6 (activating transcription factor 6, an endoplasmic reticulum (ER stress response transcription factor mRNA expression and activation of Atf6 by MTA treatment. Forced expression of p50Atf6 (active form of Atf6 markedly enhanced Bglap mRNA expression. Chromatin immunoprecipitation assay was performed to investigate the increase in p50Atf6 binding to the Bglap promoter region by MTA treatment. Furthermore, knockdown of Atf6 gene expression by introduction of Tet-on Atf6 shRNA expression vector abrogated MTA-induced mineralization. These results suggest that MTA induces in vitro osteoblastogenesis through the Atf6–osteocalcin axis as ER stress signaling. Therefore, MTA in endodontic treatment may affect alveolar bone healing in the resorbed region caused by pulpal infection.

  4. Regulation of RNA-dependent RNA polymerase 1 and isochorismate synthase gene expression in Arabidopsis.

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    Lydia J R Hunter

    Full Text Available RNA-dependent RNA polymerases (RDRs function in anti-viral silencing in Arabidopsis thaliana and other plants. Salicylic acid (SA, an important defensive signal, increases RDR1 gene expression, suggesting that RDR1 contributes to SA-induced virus resistance. In Nicotiana attenuata RDR1 also regulates plant-insect interactions and is induced by another important signal, jasmonic acid (JA. Despite its importance in defense RDR1 regulation has not been investigated in detail.In Arabidopsis, SA-induced RDR1 expression was dependent on 'NON-EXPRESSER OF PATHOGENESIS-RELATED GENES 1', indicating regulation involves the same mechanism controlling many other SA- defense-related genes, including pathogenesis-related 1 (PR1. Isochorismate synthase 1 (ICS1 is required for SA biosynthesis. In defensive signal transduction RDR1 lies downstream of ICS1. However, supplying exogenous SA to ics1-mutant plants did not induce RDR1 or PR1 expression to the same extent as seen in wild type plants. Analysing ICS1 gene expression using transgenic plants expressing ICS1 promoter:reporter gene (β-glucuronidase constructs and by measuring steady-state ICS1 transcript levels showed that SA positively regulates ICS1. In contrast, ICS2, which is expressed at lower levels than ICS1, is unaffected by SA. The wound-response hormone JA affects expression of Arabidopsis RDR1 but jasmonate-induced expression is independent of CORONATINE-INSENSITIVE 1, which conditions expression of many other JA-responsive genes. Transiently increased RDR1 expression following tobacco mosaic virus inoculation was due to wounding and was not a direct effect of infection. RDR1 gene expression was induced by ethylene and by abscisic acid (an important regulator of drought resistance. However, rdr1-mutant plants showed normal responses to drought.RDR1 is regulated by a much broader range of phytohormones than previously thought, indicating that it plays roles beyond those already suggested in virus

  5. Clinical Survey of Successfulness of DPC with MTA and Calcium Hydroxide

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    E.Yasini

    2005-02-01

    Full Text Available Statement of Problem: The pulp may be exposed during remove of caries or by traumatic and iatrogenicinjuries. The material used to DPC, must be biocompatible, scalable and prevent bacterial leakage.Furthermore, it is better to stimulate dentinogenesis. Calcium hydroxide is most usual material for DPC. But itcannot provide a suitablcbiologic seal. Mineral irioxide aggregate (MTA is also a suitable material for DPC.Purpose: The aim of this study was to compare the clinical success of DPC done with MTA or calciumhydroxide cement.Materials and Methods: In this study eighteen posterior teeth without previous spontaneous pain thatinvolved mechanicaly exposure was randomly selected and capped with MTA or calcium hydroxide. These teeth were evaluated by clinical vitality test and radiograph after 3 and 12 months, and patients werequestioned about irritative or spontancus pain in this period. Statistical analysis was carried out by Usher exacttest (a-0.05.Results: This study showed that in calcium hydroxid groud three had irreversible pulpitis and two teeth had symptom and signs of reversible pulpitis. While, In MTA group all cases were normaL and no clinical sign was evident.Conclusion: MTA exhibited better results than calcium hydroxide cement for the capping of the pulp in human

  6. Characterization of un-hydrated and hydrated BioAggregate™ and MTA Angelus™.

    Science.gov (United States)

    Camilleri, J; Sorrentino, F; Damidot, D

    2015-04-01

    BioAggregate™ is a novel material introduced for use as a root-end filling material. It is tricalcium silicate-based, free of aluminium and uses tantalum oxide as radiopacifier. BioAggregate contains additives to enhance the material performance. The purpose of this research was to characterize the un-hydrated and hydrated forms of BioAggregate using a combination of techniques, verify whether the additives if present affect the properties of the set material and compare these properties to those of MTA Angelus™. Un-hydrated and hydrated BioAggregate and MTA Angelus were assessed. Un-hydrated cement was tested for chemical composition, specific surface area, mineralogy and kinetics of hydration. The set material was investigated for mineralogy, microstructure and bioactivity. Scanning electron microscopy, X-ray energy dispersive spectroscopic analysis, X-ray fluorescence spectroscopy, X-ray diffraction and isothermal calorimetry were employed. The specific surface area was investigated using a gas adsorption method with nitrogen as the probe. BioAggregate was composed of tricalcium silicate, tantalum oxide, calcium phosphate and silicon dioxide and was free of aluminium. On hydration, the tricalcium silicate produced calcium silicate hydrate and calcium hydroxide. The former was deposited around the cement grains, while the latter reacted with the silicon dioxide to form additional calcium silicate hydrate. This resulted in reduction of calcium hydroxide in the aged cement. MTA Angelus reacted in a similar fashion; however, since it contained no additives, the calcium hydroxide was still present in the aged cement. Bioactivity was demonstrated by deposition of hydroxyapatite. BioAggregate exhibited a high specific surface area. Nevertheless, the reactivity determined by isothermal calorimetry appeared to be slow compared to MTA Angelus. The tantalum oxide as opposed to bismuth oxide was inert, and tantalum was not leached in solution. BioAggregate exhibited

  7. Expression of Xanthophyll Biosynthetic Genes during Light-Dependent Chloroplast Differentiation1

    Science.gov (United States)

    Woitsch, Sonja; Römer, Susanne

    2003-01-01

    In higher plants, etioplast to chloroplast differentiation is characterized by dramatic ultrastructural changes of the plastid and a concomitant increase in chlorophylls and carotenoids. Whereas the formation and function of carotenes and their oxygenated derivatives, the xanthophylls, have been well studied, little is known about the regulation of the genes involved in xanthophyll biosynthesis. Here, we analyze the expression of three xanthophyll biosynthetic genes (i.e. β-carotene hydroxylase [bhy], zeaxanthin epoxidase [zep], and violaxanthin de-epoxidase [vde]) during de-etiolation of seedlings of tobacco (Nicotiana tabacum L. cv Samsun) under different light conditions. White-light illumination caused an increase in the amount of all corresponding mRNAs. The expression profiles of bhy and zep not only resembled each other but were also similar to the pattern of a gene encoding a major light-harvesting protein of photosystem II. This finding indicates a coordinated synthesis during formation of the antenna complex. In contrast, the expression pattern of vde was clearly different. Furthermore, the gene expression of bhy was shown to be modulated after illumination with different white-light intensities. The expression of all xanthophyll biosynthetic genes under examination was up-regulated upon exposure to red, blue, and white light. Gene expression of bhy and vde but not of zep was more pronounced under red-light illumination, pointing at an involvement of the phytochrome system. Expression analysis in the presence of the photosynthetic electron transport inhibitors 3-(3,4-dichlorophenyl)-1,1-dimethyl-urea and 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone indicated a redox control of transcription of two of the xanthophyll biosynthetic genes (bhy and zep). PMID:12857831

  8. Effect of MTA-based sealer on the healing of periapical lesions

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    Joao Eduardo GOMES-FILHO

    2013-06-01

    Full Text Available Some manufacturers have recently added specific components to improve the ease of handling and insertion material properties of MTA in order to create MTA-based sealers. Objective The aim of this study was to evaluate the healing of periapical lesions in canine teeth after a single session of endodontic treatment with MTA Fillapex® compared with Sealapex® or Endo-CPM-Sealer®. Material and Methods Sixty-two root canals were performed on two 1-year-old male dogs. After coronal access and pulp extirpation, the canals were exposed to the oral cavity for 6 months in order to induce periapical lesions. The root canals were prepared, irrigated with a solution of 2.5% sodium hypochlorite and filled with gutta-percha and different sealers, according to the following groups: 1 Sealapex®; 2 Endo-CPM-Sealer®; and 3 MTA Fillapex®. Some teeth with periapical lesions were left untreated for use as positive controls. Healthy teeth were used as negative controls. After 6 months, the animals were sacrificed and serial sections from the roots were prepared for histomorphologic analysis and stained with hematoxylin and eosin and the Brown and Brenn technique. The lesions were scored according to pre-established histomorphologic parameters and the scores statistically analyzed using the Kruskal-Wallis test. Results All 3 materials produced similar patterns of healing (p>0.05; in particular, persistent inflammation and absence of complete periapical tissue healing were consistently noted. Conclusions Preparation of the infected root canals followed by filling with the materials studied was insufficient to provide complete healing of the periapical tissues.

  9. REDD1 induction regulates the skeletal muscle gene expression signature following acute aerobic exercise.

    Science.gov (United States)

    Gordon, Bradley S; Steiner, Jennifer L; Rossetti, Michael L; Qiao, Shuxi; Ellisen, Leif W; Govindarajan, Subramaniam S; Eroshkin, Alexey M; Williamson, David L; Coen, Paul M

    2017-12-01

    The metabolic stress placed on skeletal muscle by aerobic exercise promotes acute and long-term health benefits in part through changes in gene expression. However, the transducers that mediate altered gene expression signatures have not been completely elucidated. Regulated in development and DNA damage 1 (REDD1) is a stress-induced protein whose expression is transiently increased in skeletal muscle following acute aerobic exercise. However, the role of this induction remains unclear. Because REDD1 altered gene expression in other model systems, we sought to determine whether REDD1 induction following acute exercise altered the gene expression signature in muscle. To do this, wild-type and REDD1-null mice were randomized to remain sedentary or undergo a bout of acute treadmill exercise. Exercised mice recovered for 1, 3, or 6 h before euthanization. Acute exercise induced a transient increase in REDD1 protein expression within the plantaris only at 1 h postexercise, and the induction occurred in both cytosolic and nuclear fractions. At this time point, global changes in gene expression were surveyed using microarray. REDD1 induction was required for the exercise-induced change in expression of 24 genes. Validation by RT-PCR confirmed that the exercise-mediated changes in genes related to exercise capacity, muscle protein metabolism, neuromuscular junction remodeling, and Metformin action were negated in REDD1-null mice. Finally, the exercise-mediated induction of REDD1 was partially dependent upon glucocorticoid receptor activation. In all, these data show that REDD1 induction regulates the exercise-mediated change in a distinct set of genes within skeletal muscle. Copyright © 2017 the American Physiological Society.

  10. Alzheimer's Disease Risk Polymorphisms Regulate Gene Expression in the ZCWPW1 and the CELF1 Loci.

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    Celeste M Karch

    Full Text Available Late onset Alzheimer's disease (LOAD is a genetically complex and clinically heterogeneous disease. Recent large-scale genome wide association studies (GWAS have identified more than twenty loci that modify risk for AD. Despite the identification of these loci, little progress has been made in identifying the functional variants that explain the association with AD risk. Thus, we sought to determine whether the novel LOAD GWAS single nucleotide polymorphisms (SNPs alter expression of LOAD GWAS genes and whether expression of these genes is altered in AD brains. The majority of LOAD GWAS SNPs occur in gene dense regions under large linkage disequilibrium (LD blocks, making it unclear which gene(s are modified by the SNP. Thus, we tested for brain expression quantitative trait loci (eQTLs between LOAD GWAS SNPs and SNPs in high LD with the LOAD GWAS SNPs in all of the genes within the GWAS loci. We found a significant eQTL between rs1476679 and PILRB and GATS, which occurs within the ZCWPW1 locus. PILRB and GATS expression levels, within the ZCWPW1 locus, were also associated with AD status. Rs7120548 was associated with MTCH2 expression, which occurs within the CELF1 locus. Additionally, expression of several genes within the CELF1 locus, including MTCH2, were highly correlated with one another and were associated with AD status. We further demonstrate that PILRB, as well as other genes within the GWAS loci, are most highly expressed in microglia. These findings together with the function of PILRB as a DAP12 receptor supports the critical role of microglia and neuroinflammation in AD risk.

  11. Dynamic gene expression analysis in a H1N1 influenza virus mouse pneumonia model.

    Science.gov (United States)

    Bao, Yanyan; Gao, Yingjie; Shi, Yujing; Cui, Xiaolan

    2017-06-01

    H1N1, a major pathogenic subtype of influenza A virus, causes a respiratory infection in humans and livestock that can range from a mild infection to more severe pneumonia associated with acute respiratory distress syndrome. Understanding the dynamic changes in the genome and the related functional changes induced by H1N1 influenza virus infection is essential to elucidating the pathogenesis of this virus and thereby determining strategies to prevent future outbreaks. In this study, we filtered the significantly expressed genes in mouse pneumonia using mRNA microarray analysis. Using STC analysis, seven significant gene clusters were revealed, and using STC-GO analysis, we explored the significant functions of these seven gene clusters. The results revealed GOs related to H1N1 virus-induced inflammatory and immune functions, including innate immune response, inflammatory response, specific immune response, and cellular response to interferon-beta. Furthermore, the dynamic regulation relationships of the key genes in mouse pneumonia were revealed by dynamic gene network analysis, and the most important genes were filtered, including Dhx58, Cxcl10, Cxcl11, Zbp1, Ifit1, Ifih1, Trim25, Mx2, Oas2, Cd274, Irgm1, and Irf7. These results suggested that during mouse pneumonia, changes in the expression of gene clusters and the complex interactions among genes lead to significant changes in function. Dynamic gene expression analysis revealed key genes that performed important functions. These results are a prelude to advancements in mouse H1N1 influenza virus infection biology, as well as the use of mice as a model organism for human H1N1 influenza virus infection studies.

  12. [Eukaryotic expression of Leptospira interrogans lipL32/1-ompL1/1 fusion gene encoding genus-specific protein antigens and the immunoreactivity of expression products].

    Science.gov (United States)

    Yan, Jie; Zhao, Shou-feng; Mao, Ya-fei; Ruan, Ping; Luo, Yi-hui; Li, Shu-ping; Li, Li-wei

    2005-01-01

    To construct the eukaryotic expression system of L.interrogans lipL32/1-ompL1/1 fusion gene and to identify the immunoreactivity of expression products. PCR with linking primer was used to construct the fusion gene lipL32/1-ompL1/1. The P.pastoris eukaryotic expression system of the fusion gene, pPIC9K-lipL32/1-ompL1/1-P. pastorisGS115, was constructed after the fusion gene was cloned and sequenced. Colony with phenotype His(+)Mut(+) was isolated by using MD and MM plates and His(+) Mut(+) transformant with high resistance to G418 was screened out by using YPD plate. Using lysate of His(+) Mut(+) colony with high copies of the target gene digested with yeast lyase as the template and 5'AOX1 and 3'AOX1 as the primers, the target fusion gene in chromosome DNA of the constructed P. pastoris engineering strain was detected by PCR. Methanol in BMMY medium was used to induce the target recombinant protein rLipL32/1-rOmpL1/1 expression. rLipL32/1-rOmpL1/1 in the medium supernatant was extracted by using ammonium sulfate precipitation and Ni-NTA affinity chromatography. Output and immunoreactivity of rLipL32/1-rOmpL1/1 were measured by SDS-PAGE and Western blot methods, respectively. Amplification fragments of the obtained fusion gene lipL32/1-ompL1/1 was 1794 bp in size. The homogeneity of nucleotide and putative amino acid sequences of the fusion gene were as high as 99.94 % and 100 %, respectively, compared with the sequences of original lipL32/1 and ompL1/1 genotypes. The constructed eukaryotic expression system was able to secrete rLipL32/1-rOmpL1/1 with an output of 10 % of the total proteins in the supernatant, which located the expected position after SDS-PAGE. The rabbit anti-rLipL32/1 and anti-rOmpL1/1 sera could combine the expressed rLipL32/1-rOmpL1/1. An eukaryotic expression system with high efficiency in P.pastoris of L.interrogans lipL32/1-ompL1/1 fusion gene was successfully constructed in this study. The expressed fusion protein shows specific

  13. The bromodomain protein LEX-1 acts with TAM-1 to modulate gene expression in C. elegans.

    Science.gov (United States)

    Tseng, Rong-Jeng; Armstrong, Kristin R; Wang, Xiaodong; Chamberlin, Helen M

    2007-11-01

    In many organisms, repetitive DNA serves as a trigger for gene silencing. However, some gene expression is observed from repetitive genomic regions such as heterochromatin, suggesting mechanisms exist to modulate the silencing effects. From a genetic screen in C. elegans, we have identified mutations in two genes important for expression of repetitive sequences: lex-1 and tam-1. Here we show that lex-1 encodes a protein containing an ATPase domain and a bromodomain. LEX-1 is similar to the yeast Yta7 protein, which maintains boundaries between silenced and active chromatin. tam-1 has previously been shown to encode a RING finger/B-box protein that modulates gene expression from repetitive DNA. We find that lex-1, like tam-1, acts as a class B synthetic multivulva (synMuv) gene. However, since lex-1 and tam-1 mutants have normal P granule localization, it suggests they act through a mechanism distinct from other class B synMuvs. We observe intragenic (interallelic) complementation with lex-1 and a genetic interaction between lex-1 and tam-1, data consistent with the idea that the gene products function in the same biological process, perhaps as part of a protein complex. We propose that LEX-1 and TAM-1 function together to influence chromatin structure and to promote expression from repetitive sequences.

  14. Effect of diode laser irradiation on the apical sealing of MTA retrofillings Efeito da irradiação de laser de diodo no selamento apical em retrobturações com MTA

    Directory of Open Access Journals (Sweden)

    Eliana Barbosa de Souza

    2006-09-01

    Full Text Available Apical sealing is essential for the success of paraendodontic surgery, so any procedure that may favor an adequate sealing of the apical remainder should be performed. The purpose of this study was to evaluate the influence of diode laser irradiation on the apical sealing of root-end cavities with MTA retrofillings. Root canals in twenty extracted human teeth were shaped with K-files and filled with gutta-percha. The apexes were cut off and root-end preparations were performed. The roots were divided randomly in 2 groups. Group 1 (ten specimens was retrofilled with MTA. Group 2 was irradiated with diode laser, with 1 W for 20 seconds, on the apical surface and root end cavity before retrofilling with MTA. The specimens had their external surfaces impermeabilized with cyanoacrylate, except for the apical surface, and were then immersed in 1% rhodamine B dye for 72 h and placed in plaster stone. After that, the specimens were submitted to longitudinal abrasion until half of the root remained. The linear dye leakage was observed in these mid-roots between the root canal wall and retrofilling. The linear dye leakage was measured with Image Lab software, and the results were statistically analyzed with Student's t test. There were no statistically significant differences between the two groups (p > 0.05. The diode laser irradiation did not improve the apical sealing of MTA retrofillings under the conditions of this in vitro study.O selamento apical é fundamental para o sucesso da cirurgia parendodôntica. Assim, procedimentos que melhorem o selamento do remanescente apical devem ser utilizados. O objetivo deste estudo foi verificar se a irradiação de laser de diodo poderia aumentar o selamento apical em cavidades retrógradas obturadas com MTA. Foram utilizadas 20 raízes de dentes humanos extraídos que, após preparo com lima tipo K, tiveram seus canais obturados com guta-percha. Os ápices foram cortados e sofreram preparo de cavidades retr

  15. Ascl1 Coordinately Regulates Gene Expression and the Chromatin Landscape during Neurogenesis

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    Alexandre A.S.F. Raposo

    2015-03-01

    Full Text Available The proneural transcription factor Ascl1 coordinates gene expression in both proliferating and differentiating progenitors along the neuronal lineage. Here, we used a cellular model of neurogenesis to investigate how Ascl1 interacts with the chromatin landscape to regulate gene expression when promoting neuronal differentiation. We find that Ascl1 binding occurs mostly at distal enhancers and is associated with activation of gene transcription. Surprisingly, the accessibility of Ascl1 to its binding sites in neural stem/progenitor cells remains largely unchanged throughout their differentiation, as Ascl1 targets regions of both readily accessible and closed chromatin in proliferating cells. Moreover, binding of Ascl1 often precedes an increase in chromatin accessibility and the appearance of new regions of open chromatin, associated with de novo gene expression during differentiation. Our results reveal a function of Ascl1 in promoting chromatin accessibility during neurogenesis, linking the chromatin landscape at Ascl1 target regions with the temporal progression of its transcriptional program.

  16. Gene expression profiles in Parkinson disease prefrontal cortex implicate FOXO1 and genes under its transcriptional regulation.

    Directory of Open Access Journals (Sweden)

    Alexandra Dumitriu

    2012-06-01

    Full Text Available Parkinson disease (PD is a complex neurodegenerative disorder with largely unknown genetic mechanisms. While the degeneration of dopaminergic neurons in PD mainly takes place in the substantia nigra pars compacta (SN region, other brain areas, including the prefrontal cortex, develop Lewy bodies, the neuropathological hallmark of PD. We generated and analyzed expression data from the prefrontal cortex Brodmann Area 9 (BA9 of 27 PD and 26 control samples using the 44K One-Color Agilent 60-mer Whole Human Genome Microarray. All samples were male, without significant Alzheimer disease pathology and with extensive pathological annotation available. 507 of the 39,122 analyzed expression probes were different between PD and control samples at false discovery rate (FDR of 5%. One of the genes with significantly increased expression in PD was the forkhead box O1 (FOXO1 transcription factor. Notably, genes carrying the FoxO1 binding site were significantly enriched in the FDR-significant group of genes (177 genes covered by 189 probes, suggesting a role for FoxO1 upstream of the observed expression changes. Single-nucleotide polymorphisms (SNPs selected from a recent meta-analysis of PD genome-wide association studies (GWAS were successfully genotyped in 50 out of the 53 microarray brains, allowing a targeted expression-SNP (eSNP analysis for 52 SNPs associated with PD affection at genome-wide significance and the 189 probes from FoxO1 regulated genes. A significant association was observed between a SNP in the cyclin G associated kinase (GAK gene and a probe in the spermine oxidase (SMOX gene. Further examination of the FOXO1 region in a meta-analysis of six available GWAS showed two SNPs significantly associated with age at onset of PD. These results implicate FOXO1 as a PD-relevant gene and warrant further functional analyses of its transcriptional regulatory mechanisms.

  17. MTA Computer Based Evaluation System.

    Science.gov (United States)

    Brenner, Lisa P.; And Others

    The MTA PLATO-based evaluation system, which has been implemented by a consortium of schools of medical technology, is designed to be general-purpose, modular, data-driven, and interactive, and to accommodate other national and local item banks. The system provides a comprehensive interactive item-banking system in conjunction with online student…

  18. Over-expression of KdSOC1 gene affected plantlet morphogenesis in Kalanchoe daigremontiana.

    Science.gov (United States)

    Zhu, Chen; Wang, Li; Chen, Jinhua; Liu, Chenglan; Zeng, Huiming; Wang, Huafang

    2017-07-17

    Kalanchoe daigremontiana reproduces asexually by producing plantlets along the leaf margin. The aim of this study was to identify the function of the SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 gene in Kalanchoe daigremontiana (KdSOC1) during plantlet morphogenesis. In this study, KdSOC1 gene expression was detected at stem cell niche during in vitro somatic embryogenesis and plantlet morphogenesis. Disrupting endogenous auxin transportation suppressed the KdSOC1 gene response. Knockdown of the KdSOC1 gene caused a defect in cotyledon formation during the early heart stage of somatic embryogenesis. Over-expression (OE) of the KdSOC1 gene resulted in asymmetric plantlet distribution, a reduced number of plantlets, thicker leaves, and thicker vascular fibers. Higher KdPIN1 gene expression and auxin content were found in OE plant compared to those of wild-type plant leaves, which indicated possible KdSOC1 gene role in affecting auxin distribution and accumulation. KdSOC1 gene OE in DR5-GUS Arabidopsis reporting lines resulted in an abnormal auxin response pattern during different stages of somatic embryogenesis. In summary, the KdSOC1 gene OE might alter auxin distribution and accumulation along leaf margin to initiate plantlet formation and distribution, which is crucial for plasticity during plantlet formation under various environmental conditions.

  19. Similar expression of through-and-through fluid movement along orthograde apical plugs of MTA Bio and white Portland cement.

    Science.gov (United States)

    De-Deus, G; Audi, C; Murad, C; Fidel, S; Fidel, R

    2008-12-01

    To compare the sealing ability of four hydraulic cements when used as an apical plug in teeth with wide-open apices. A sample of 70 maxillary central incisors were divided into four groups (n = 15) and a further 10 teeth served as controls. An artificial open apex was created in the teeth using Gates Glidden drills numbers 6-1 in a crown-down manner until the size 1 bur passed through the foramen. A divergent open apex was prepared to a size of 1.24 mm at the foramen by retrograde apical transportation using a number 8 (0.60) Profile Series 29 0.4 taper instrument inserted to the length of the cutting blade. In G1, the open apices were repaired with WMTA Angelus whilst in G2, G3 and G4 MTA Bio, Pro-Root MTA and Portland cement was employed respectively. Each root was assembled in a hermetic cell to allow the evaluation of fluid filtration. Leakage was measured by the movement of an air bubble travelling within a pipette connected to the teeth. Measurements of the air bubble movement were made after 10 min at a constant pressure of 50 cm H(2)O. The Kruskal-Wallis H-test was applied to the fluid flow data to detect differences between the experimental groups (P 0.05). Fluid movement through teeth with open apices and filled with four hydraulic cements was similar. All cements allowed fluid movement.

  20. The hematopoietic transcription factor PU.1 regulates RANK gene expression in myeloid progenitors

    International Nuclear Information System (INIS)

    Kwon, Oh Hyung; Lee, Chong-Kil; Lee, Young Ik; Paik, Sang-Gi; Lee, Hyun-Jun

    2005-01-01

    Osteoclasts are bone resorbing cells of hematopoietic origin. The hematopoietic transcription factor PU.1 is critical for osteoclastogenesis; however, the molecular mechanisms of PU.1-regulated osteoclastogenesis have not been explored. Here, we present evidence that the receptor activator of nuclear factor κB (RANK) gene that has been shown to be crucial for osteoclastogenesis is a transcriptional target of PU.1. The PU.1 -/- progenitor cells failed to express the RANK gene and reconstitution of PU.1 in these cells induced RANK expression. Treatment of the PU.1 reconstituted cells with M-CSF and RANKL further augmented the RANK gene expression. To explore the regulatory mechanism of the RANK gene expression by PU.1, we have cloned the human RANK promoter. Transient transfection assays have revealed that the 2.2-kb RANK promoter was functional in a monocyte line RAW264.7, whereas co-transfection of PU.1 transactivated the RANK promoter in HeLa cells. Taken together, these results suggest that PU.1 regulates the RANK gene transcription and this may represent one of the key roles of PU.1 in osteoclast differentiation

  1. Enhancement of solubility in Escherichia coli and purification of an aminotransferase from Sphingopyxis sp. MTA144 for deamination of hydrolyzed fumonisin B1

    Directory of Open Access Journals (Sweden)

    Hartinger Doris

    2010-08-01

    Full Text Available Abstract Background Fumonisin B1 is a cancerogenic mycotoxin produced by Fusarium verticillioides and other fungi. Sphingopyxis sp. MTA144 can degrade fumonisin B1, and a key enzyme in the catabolic pathway is an aminotransferase which removes the C2-amino group from hydrolyzed fumonisin B1. In order to study this aminotransferase with respect to a possible future application in enzymatic fumonisin detoxification, we attempted expression of the corresponding fumI gene in E. coli and purification of the enzyme. Since the aminotransferase initially accumulated in inclusion bodies, we compared the effects of induction level, host strain, expression temperature, solubility enhancers and a fusion partner on enzyme solubility and activity. Results When expressed from a T7 promoter at 30°C, the aminotransferase accumulated invariably in inclusion bodies in DE3 lysogens of the E. coli strains BL21, HMS174, Rosetta 2, Origami 2, or Rosetta-gami. Omission of the isopropyl-beta-D-thiogalactopyranoside (IPTG used for induction caused a reduction of expression level, but no enhancement of solubility. Likewise, protein production but not solubility correlated with the IPTG concentration in E. coli Tuner(DE3. Addition of the solubility enhancers betaine and sorbitol or the co-enzyme pyridoxal phosphate showed no effect. Maltose-binding protein, used as an N-terminal fusion partner, promoted solubility at 30°C or less, but not at 37°C. Low enzyme activity and subsequent aggregation in the course of purification and cleavage indicated that the soluble fusion protein contained incorrectly folded aminotransferase. Expression in E. coli ArcticExpress(DE3, which co-expresses two cold-adapted chaperonins, at 11°C finally resulted in production of appreciable amounts of active enzyme. Since His tag-mediated affinity purification from this strain was hindered by co-elution of chaperonin, two steps of chromatography with optimized imidazole concentration in the

  2. Regional gene expression of LOX-1, VCAM-1, and ICAM-1 in aorta of HIV-1 transgenic rats

    DEFF Research Database (Denmark)

    Hag, Anne Mette Fisker; Kristoffersen, Ulrik Sloth; Pedersen, Sune Folke

    2009-01-01

    endpoints, studies in animal models could be attractive alternatives. METHODOLOGY/PRINCIPAL FINDINGS: We evaluated gene expression of lectin-like oxidized-low-density-lipoprotein receptor-1 (LOX-1), vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) in HIV-1...... transgenic (HIV-1Tg) rats; these genes are all thought to play important roles in early atherogenesis. Furthermore, the plasma level of sICAM-1 was measured. We found that gene expressions of LOX-1 and VCAM-1 were higher in the aortic arch of HIV-1Tg rats compared to controls. Also, the level of sICAM-1......-infection per se may cause atherosclerosis. This transgenic rat model may be a very promising model for further studies of the pathophysiology behind HIV-associated cardiovascular disease....

  3. Restricted expression of recombination activating gene (RAG-1) in mouse lymphoid tissues

    International Nuclear Information System (INIS)

    Yamamoto, Akihito; Fujinaga, Hiroyuki; Hamatani, Kiyohiro; Atsuta, Mitsuru.

    1993-03-01

    In an attempt to determine the distribution of recombinase activity in the mouse thymus, spleen, and lymph nodes, we used the in situ hybridization method to examine the expression of the recombination activating genes RAG-1 and RAG-2. Expression of RAG-1 was found in most cortical thymocytes but not in the majority of medullary thymocytes. Although hybridization signals of RAG-2 were not as intense as those of RAG-1, the localization of RAG-2 transcripts was similar to that of RAG-1. In the spleen, expression of RAG-1 was found only in limited cells near the splenic sinus, and the majority of the cells within the follicle were negative for RAG-1 transcript. In nude mice, RAG-1-expressing cells were detected in the same regions, which suggests that in situ hybridization signals of RAG-1 in the spleen are due to the cells of B-cell origin. In the lymph nodes, expression of RAG-1 was found only in the medullary region. Expression of RAG-2 transcript in the spleen and the lymph nodes, if any, was too faint to allow determination of the specific localization. These results suggest that most of the cortical thymocytes and some cells in the spleen are capable of rearranging T-cell receptor genes and immunoglobulin genes, respectively, but the possible involvement of the RAG-1 transcript in RAG-1-positive cells of the spleen and the lymph nodes in functions other than the rearrangement of genes could not be ruled out. (author)

  4. Functional imaging: monitoring heme oxygenase-1 gene expression in vivo

    Science.gov (United States)

    Zhang, Weisheng; Reilly-Contag, Pamela; Stevenson, David K.; Contag, Christopher H.

    1999-07-01

    The regulation of genetic elements can be monitored in living animals using photoproteins as reporters. Heme oxygenase (HO) is the key catabolic enzyme in the heme degradation pathway. Here, HO expression serves as a model for in vivo functional imaging of transcriptional regulation of a clinically relevant gene. HO enzymatic activity is inhibited by heme analogs, metalloporphyrins, but many members of this family of compounds also activate transcription of the HO-1 promoter. The degree of transcriptional activation by twelve metalloporphyrins, differing at the central metal and porphyrin ring substituents, was evaluated in both NIH 3T3 stable lines and transgenic animals containing HO-1 promoter-luciferase gene fusions. In the correlative cell culture assays, the metalloporphyrins increased transcription form the full length HO promoter fusion to varying degrees, but none increased transcription from a truncated HO-1 promoter. These results suggested that one or both of the two distal enhancer elements located at -4 and -10 Kb upstream from transcriptional start are required for HO-1 induction by heme and its analogs. The full-length HO-1-luc fusion was then evaluated as a transgene in mice. It was possible to monitor the effects of the metalloporphyrins, SnMP and ZnPP, in living animals over time. This spatiotemporal analyses of gene expression in vivo implied that alterations in porphyrin ring substituents and the central metal may affect the extent of gene activation. These data further indicate that using photoprotein reporters, subtle differences in gene expression can be monitored in living animals.

  5. USP2 Regulates the Intracellular Localization of PER1 and Circadian Gene Expression

    DEFF Research Database (Denmark)

    Yang, Yaoming; Duguay, David; Fahrenkrug, Jan

    2014-01-01

    . Although Per1 mRNA expression rhythm remained intact in the Usp2 KO MEFs, the expression profiles of other core clock genes were altered. This was also true for the expression of clock-controlled genes (e.g., Dbp, Tef, Hlf, E4bp4). A similar phase advance of PER1 nuclear localization rhythm and alteration...

  6. Domestication-driven Gossypium profilin 1 (GhPRF1) gene transduces early flowering phenotype in tobacco by spatial alteration of apical/floral-meristem related gene expression.

    Science.gov (United States)

    Pandey, Dhananjay K; Chaudhary, Bhupendra

    2016-05-13

    Plant profilin genes encode core cell-wall structural proteins and are evidenced for their up-regulation under cotton domestication. Notwithstanding striking discoveries in the genetics of cell-wall organization in plants, little is explicit about the manner in which profilin-mediated molecular interplay and corresponding networks are altered, especially during cellular signalling of apical meristem determinacy and flower development. Here we show that the ectopic expression of GhPRF1 gene in tobacco resulted in the hyperactivation of apical meristem and early flowering phenotype with increased flower number in comparison to the control plants. Spatial expression alteration in CLV1, a key meristem-determinacy gene, is induced by the GhPRF1 overexpression in a WUS-dependent manner and mediates cell signalling to promote flowering. But no such expression alterations are recorded in the GhPRF1-RNAi lines. The GhPRF1 transduces key positive flowering regulator AP1 gene via coordinated expression of FT4, SOC1, FLC1 and FT1 genes involved in the apical-to-floral meristem signalling cascade which is consistent with our in silico profilin interaction data. Remarkably, these positive and negative flowering regulators are spatially controlled by the Actin-Related Protein (ARP) genes, specifically ARP4 and ARP6 in proximate association with profilins. This study provides a novel and systematic link between GhPRF1 gene expression and the flower primordium initiation via up-regulation of the ARP genes, and an insight into the functional characterization of GhPRF1 gene acting upstream to the flowering mechanism. Also, the transgenic plants expressing GhPRF1 gene show an increase in the plant height, internode length, leaf size and plant vigor. Overexpression of GhPRF1 gene induced early and increased flowering in tobacco with enhanced plant vigor. During apical meristem determinacy and flower development, the GhPRF1 gene directly influences key flowering regulators through ARP-genes

  7. Undifferentiated embryonic cell transcription factor 1 regulates ESC chromatin organization and gene expression

    DEFF Research Database (Denmark)

    Kooistra, Susanne M; van den Boom, Vincent; Thummer, Rajkumar P

    2010-01-01

    Previous reports showed that embryonic stem (ES) cells contain hyperdynamic and globally transcribed chromatin-properties that are important for ES cell pluripotency and differentiation. Here, we demonstrate a role for undifferentiated embryonic cell transcription factor 1 (UTF1) in regulating ES...... cell chromatin structure. Using chromatin immunoprecipitation-on-chip analysis, we identified >1,700 UTF1 target genes that significantly overlap with previously identified Nanog, Oct4, Klf-4, c-Myc, and Rex1 targets. Gene expression profiling showed that UTF1 knock down results in increased expression...... of a large set of genes, including a significant number of UTF1 targets. UTF1 knock down (KD) ES cells are, irrespective of the increased expression of several self-renewal genes, Leukemia inhibitory factor (LIF) dependent. However, UTF1 KD ES cells are perturbed in their differentiation in response...

  8. Comparative study of physico-chemical properties of MTA-based and Portland cements.

    Science.gov (United States)

    Borges, Alvaro H; Pedro, Fábio L M; Miranda, Carlos E S; Semenoff-Segundo, Alex; Pécora, Jesus D; Filho, Antônio M Cruz

    2010-01-01

    The purpose of this investigation was to evaluate the physicochemical properties of gray and white structural and nonstructural Portland cement, gray and white ProRoot MTA and MTA BIO. The water/powder ratio, setting time, solubility and pH (hydrogen-ion potential) changes of the materials were evaluated. Tests followed specification #57 from the American National Standard Institute/American Dental Association (2000) for endodontic sealing materials and pH was determined by a digital pH meter. The test results were statistically analyzed by variance analyses for global comparison and by the complementary Tukey's test for pairwise comparisons (5%). Considering the water/powder ratio, no significant difference (p > 0.05) was observed among the cements. MTA BIO (33.10 +/- 2.30) had the lowest setting time (p Portland cement (2.55 +/- 0.08) had the highest solubility (p 0.05) was observed among materials when considering pH evaluation. The pH levels were highly alkaline immediately after immersion in solution, remaining stable throughout the test period. The authors conclude that the cements had similar water/powder proportions. MTA BIO had the shortest setting time and gray ProRoot MTA had the lowest solubility. All cements had similar behavior in the pH analysis.

  9. Expression of the tumor suppressor genes NF2, 4.1B, and TSLC1 in canine meningiomas.

    Science.gov (United States)

    Dickinson, P J; Surace, E I; Cambell, M; Higgins, R J; Leutenegger, C M; Bollen, A W; LeCouteur, R A; Gutmann, D H

    2009-09-01

    Meningiomas are common primary brain tumors in dogs; however, little is known about the molecular genetic mechanisms involved in their tumorigenesis. Several tumor suppressor genes have been implicated in meningioma pathogenesis in humans, including the neurofibromatosis 2 (NF2), protein 4.1B (4.1 B), and tumor suppressor in lung cancer-1 (TSLC1) genes. We investigated the expression of these tumor suppressor genes in a series of spontaneous canine meningiomas using quantitative real-time reverse transcription polymerase chain reaction (RT-PCR) (NF2; n = 25) and western blotting (NF2/merlin, 4.1B, TSLC1; n = 30). Decreased expression of 4.1B and TSLC1 expression on western blotting was seen in 6/30 (20%) and in 15/30 (50%) tumors, respectively, with 18/30 (60%) of meningiomas having decreased or absent expression of one or both proteins. NF2 gene expression assessed by western blotting and RT-PCR varied considerably between individual tumors. Complete loss of NF2 protein on western blotting was not seen, unlike 4.1B and TSLC1. Incidence of TSLC1 abnormalities was similar to that seen in human meningiomas, while perturbation of NF2 and 4.1B appeared to be less common than reported for human tumors. No association was observed between tumor grade, subtype, or location and tumor suppressor gene expression based on western blot or RT-PCR. These results suggest that loss of these tumor suppressor genes is a frequent occurrence in canine meningiomas and may be an early event in tumorigenesis in some cases. In addition, it is likely that other, as yet unidentified, genes play an important role in canine meningioma formation and growth.

  10. Identification of Phosphoglycerate Kinase 1 (PGK1 as a reference gene for quantitative gene expression measurements in human blood RNA

    Directory of Open Access Journals (Sweden)

    Unger Elizabeth R

    2011-09-01

    Full Text Available Abstract Background Blood is a convenient sample and increasingly used for quantitative gene expression measurements with a variety of diseases including chronic fatigue syndrome (CFS. Quantitative gene expression measurements require normalization of target genes to reference genes that are stable and independent from variables being tested in the experiment. Because there are no genes that are useful for all situations, reference gene selection is an essential step to any quantitative reverse transcription-PCR protocol. Many publications have described appropriate genes for a wide variety of tissues and experimental conditions, however, reference genes that may be suitable for the analysis of CFS, or human blood RNA derived from whole blood as well as isolated peripheral blood mononuclear cells (PBMCs, have not been described. Findings Literature review and analyses of our unpublished microarray data were used to narrow down the pool of candidate reference genes to six. We assayed whole blood RNA from Tempus tubes and cell preparation tube (CPT-collected PBMC RNA from 46 subjects, and used the geNorm and NormFinder algorithms to select the most stable reference genes. Phosphoglycerate kinase 1 (PGK1 was one of the optimal normalization genes for both whole blood and PBMC RNA, however, additional genes differed for the two sample types; Ribosomal protein large, P0 (RPLP0 for PBMC RNA and Peptidylprolyl isomerase B (PPIB for whole blood RNA. We also show that the use of a single reference gene is sufficient for normalization when the most stable candidates are used. Conclusions We have identified PGK1 as a stable reference gene for use with whole blood RNA and RNA derived from PBMC. When stable genes are selected it is possible to use a single gene for normalization rather than two or three. Optimal normalization will improve the ability of results from PBMC RNA to be compared with those from whole blood RNA and potentially allows comparison of

  11. Differential gene expression and Hog1 interaction with osmoresponsive genes in the extremely halotolerant black yeast Hortaea werneckii

    Directory of Open Access Journals (Sweden)

    Plemenitaš Ana

    2007-08-01

    Full Text Available Abstract Background Fluctuations in external salinity force eukaryotic cells to respond by changes in the gene expression of proteins acting in protective biochemical processes, thus counteracting the changing osmotic pressure. The high-osmolarity glycerol (HOG signaling pathway is essential for the efficient up-regulation of the osmoresponsive genes. In this study, the differential gene expression of the extremely halotolerant black yeast Hortaea werneckii was explored. Furthermore, the interaction of mitogen-activated protein kinase HwHog1 and RNA polymerase II with the chromatin in cells adapted to an extremely hypersaline environment was analyzed. Results A cDNA subtraction library was constructed for H. werneckii, adapted to moderate salinity or an extremely hypersaline environment of 4.5 M NaCl. An uncommon osmoresponsive set of 95 differentially expressed genes was identified. The majority of these had not previously been connected with the adaptation of salt-sensitive S. cerevisiae to hypersaline conditions. The transcriptional response in hypersaline-adapted and hypersaline-stressed cells showed that only a subset of the identified genes responded to acute salt-stress, whereas all were differentially expressed in adapted cells. Interaction with HwHog1 was shown for 36 of the 95 differentially expressed genes. The majority of the identified osmoresponsive and HwHog1-dependent genes in H. werneckii have not been previously reported as Hog1-dependent genes in the salt-sensitive S. cerevisiae. The study further demonstrated the co-occupancy of HwHog1 and RNA polymerase II on the chromatin of 17 up-regulated and 2 down-regulated genes in 4.5 M NaCl-adapted H. werneckii cells. Conclusion Extremely halotolerant H. werneckii represents a suitable and highly relevant organism to study cellular responses to environmental salinity. In comparison with the salt-sensitive S. cerevisiae, this yeast shows a different set of genes being expressed at

  12. Photobiomodulation changes type 1 collagen gene expression by pulp fibroblasts

    Science.gov (United States)

    Lourenço Ribeiro Vitor, Luciana; Tavares Oliveira Prado, Mariel; Lourenço Neto, Natalino; Cardoso de Oliveira, Rodrigo; Ferreira Santos, Carlos; Moreira Machado, Maria Aparecida Andrade; Marchini Oliveira, Thais

    2018-06-01

    This study aimed to evaluate type 1 collagen (COL1) gene expression by human pulp fibroblasts from primary teeth (HPF) after the variation of photobiomodulation (PBM) parameters. HPF were obtained from a biorepository, used at 4th passage, and irradiated (InGaAlP—660 nm) varying the power and application time according to the following groups: G1: 1.2 J cm‑2–05 mW–10 s G2: 2.5 J cm‑2–05 mW–20 s G3: 3.7 J cm‑2–05 mW–30 s G4: 5.0 J cm‑2–05 mW–40 s G5: 6.2 J cm‑2–05 mW–50 s G6: 2.5 J cm‑2–10 mW–10 s G7: 3.7 J cm‑2–15 mW–10 s G8: 5.0 J cm‑2–20 mW–10 s G9: 6.2 J cm‑2–25 mW–10 s. The control group (G10) was not irradiated and maintained with DMEM  +  10% SFB. RT-PCR was used to evaluate COL1 gene expression at 6, 12, and 24 h after irradiation. Intra- and intergroup comparisons were performed by two-way ANOVA followed by Tukey test (p  differences among periods (p  differences (p  >  0.05). The energy densities from 2.5 to 5 J cm‑2, regardless of the variation in PBM parameters, biomodulated the COL1 gene expression. At the energy density of 6.2 J cm‑2, longer application time and smaller power changed the pattern of COL1 gene expression by pulp fibroblasts from human primary teeth.

  13. Regulation of human heme oxygenase-1 gene expression under thermal stress.

    Science.gov (United States)

    Okinaga, S; Takahashi, K; Takeda, K; Yoshizawa, M; Fujita, H; Sasaki, H; Shibahara, S

    1996-06-15

    Heme oxygenase-1 is an essential enzyme in heme catabolism, and its human gene promoter contains a putative heat shock element (HHO-HSE). This study was designed to analyze the regulation of human heme oxygenase-1 gene expression under thermal stress. The amounts of heme oxygenase-1 protein were not increased by heat shock (incubation at 42 degrees C) in human alveolar macrophages and in a human erythroblastic cell line, YN-1-0-A, whereas heat shock protein 70 (HSP70) was noticeably induced. However, heat shock factor does bind in vitro to HHO-HSE and the synthetic HHO-HSE by itself is sufficient to confer the increase in the transient expression of a reporter gene upon heat shock. The deletion of the sequence, located downstream from HHO-HSE, resulted in the activation of a reporter gene by heat shock. These results suggest that HHO-HSE is potentially functional but is repressed in vivo. Interestingly, heat shock abolished the remarkable increase in the levels of heme oxygenase-1 mRNA in YN-1-0-A cells treated with hemin or cadmium, in which HSP70 mRNA was noticeably induced. Furthermore, transient expression assays showed that heat shock inhibits the cadmium-mediated activation of the heme oxygenase-1 promoter, whereas the HSP70 gene promoter was activated upon heat shock. Such regulation of heme oxygenase-1 under thermal stress may be of physiologic significance in erythroid cells.

  14. Impact of blood collection and processing on peripheral blood gene expression profiling in type 1 diabetes.

    Science.gov (United States)

    Yip, Linda; Fuhlbrigge, Rebecca; Atkinson, Mark A; Fathman, C Garrison

    2017-08-18

    The natural history of type 1 diabetes (T1D) is challenging to investigate, especially as pre-diabetic individuals are difficult to identify. Numerous T1D consortia have been established to collect whole blood for gene expression analysis from individuals with or at risk to develop T1D. However, with no universally accepted protocol for their collection, differences in sample processing may lead to variances in the results. Here, we examined whether the choice of blood collection tube and RNA extraction kit leads to differences in the expression of genes that are changed during the progression of T1D, and if these differences could be minimized by measuring gene expression directly from the lysate of whole blood. Microarray analysis showed that the expression of 901 genes is highly influenced by sample processing using the PAXgene versus the Tempus system. These included a significant number of lymphocyte-specific genes and genes whose expression has been reported to differ in the peripheral blood of at-risk and T1D patients compared to controls. We showed that artificial changes in gene expression occur when control and T1D samples were processed differently. The sample processing-dependent differences in gene expression were largely due to loss of transcripts during the RNA extraction step using the PAXgene system. The majority of differences were not observed when gene expression was measured in whole blood lysates prepared from blood collected in PAXgene and Tempus tubes. We showed that the gene expression profile of samples processed using the Tempus system is more accurate than that of samples processed using the PAXgene system. Variation in sample processing can result in misleading changes in gene expression. However, these differences can be minimized by measuring gene expression directly in whole blood lysates.

  15. Portland cement versus MTA as a root-end filling material. A pilot study.

    Science.gov (United States)

    da Silva, Sérgio Ribeiro; da Silva Neto, José Dias; Veiga, Daniela Francescato; Schnaider, Taylor Brandão; Ferreira, Lydia Masako

    2015-02-01

    To assess periradicular lesions clinically and by computed tomography (CT) after endodontic surgery using either Portland cement or mineral trioxide aggregate (MTA) as a root-end filling material. Three patients diagnosed with periradicular lesions by cone-beam CT underwent endodontic surgery with root-end filling. Patient A was treated with MTA as the root-end filling material, patient B was treated with Portland cement and patient C had two teeth treated, one with MTA and the other with Portland cement. Six months after surgery, the patients were assessed clinically and by CT scan and the obtained results were compared. Periradicular tissue regeneration was observed in all cases, with no significant differences in bone formation when comparing the use of MTA and Portland cement as root-end filling materials. Both mineral trioxide aggregate and Portland cement were successful in the treatment of periradicular lesions.

  16. Influence of phototherapies on the outcome of complete tibial fractures grafted or not with MTA: Raman spectroscopic study on rabbits

    Science.gov (United States)

    Pinheiro, Antônio L. B.; Soares, Luiz G. P.; da Silva, Aline C. P.; Santos, Nicole R. S.; da Silva, Anna Paula L. T.; Neves, Bruno Luiz R. C.; Soares, Amanda P.; Silveira, Landulfo

    2018-02-01

    The aim of the present study was to assess, by means of Raman spectroscopy, the repair of complete surgical tibial fractures fixed with wire osteosynthesis or miniplates treated or not with infrared laser (λ780 nm) or infrared LED (λ850 +/- 10 nm) lights, 142.8 J/cm2 per treatment, associated or not to the use of mineral trioxide aggregate (MTA) cement. Surgical fractures were created on 36 rabbits and fixed with WO or miniplates and some groups were grafted with MTA. Irradiated groups received lights at every other day for 15 days and sacrifice occurred after 30 days. The results showed that only irradiation with either laser or LED influenced the peaks of phosphate ( 960 cm-1) and carbonated ( 1,070 cm-1) hydroxyapatite. Collagen peak (1,450 cm-1) was influenced by both the use of MTA and irradiation with either laser or LED. It is concluded that the use of either laser or LED phototherapy associated to MTA cement was efficacious on improving the repair of complete tibial fractures treated with wire osteosynthesis or miniplates.

  17. Differential expression of homeobox-containing genes Msx-1 and Msx-2 and homeoprotein Msx-2 expression during chick craniofacial development.

    Science.gov (United States)

    Nishikawa, K; Nakanishi, T; Aoki, C; Hattori, T; Takahashi, K; Taniguchi, S

    1994-03-01

    The expression pattern of chick Msx-1 and Msx-2 homeobox genes in craniofacial primordia was examined by in situ hybridization using cRNA probes. Both genes were expressed in the distal region of the facial primordia, where the distribution of Msx-2 expression was restricted distally within the Msx-1 expression domain. On the contrary, Msx-2 expression in the lateral choroid plexus and cranial skull was broader and more intensive than Msx-1 expression. Our findings suggest that these two genes cooperate to play differential roles in craniofacial development. Msx-2 protein was detected immunohistochemically, and its localization essentially corresponded to the mRNA expression pattern, substantiating the involvement of Msx-2 protein as a transcriptional regulator in developing limb and face.

  18. G-protein inwardly rectifying potassium channel 1 (GIRK 1) gene expression correlates with tumor progression in non-small cell lung cancer

    International Nuclear Information System (INIS)

    Takanami, Iwao; Inoue, Yoshimasa; Gika, Masatoshi

    2004-01-01

    G-protein inwardly rectifying potassium channel 1 (GIRK1) is thought to play a role in cell proliferation in cancer, and GIRK1 gene expression level may define a more aggressive phenotype. We detected GIRK1 expression in tissue specimens from patients with non-small cell lung cancers (NSCLCs) and assessed their clinical characteristics. Using reverse transcription-polymerase chain reaction (RT-PCR) analyses, we quantified the expression of GIRK1 in 72 patients with NSCLCs to investigate the relationship between GIRK1 expression and clinicopathologic factors and prognosis. In 72 NSCLC patients, 50 (69%) samples were evaluated as having high GIRK1 gene expression, and 22 (31%) were evaluated as having low GIRK1 gene expression. GIRK1 gene expression was significantly associated with lymph node metastasis, stage (p = 0.0194 for lymph node metastasis; p = 0.0207 for stage). The overall and stage I survival rates for patients with high GIRK1 gene expressed tumors was significantly worse than for those individuals whose tumors had low GIRK1 expression (p = 0.0004 for the overall group; p = 0.0376 for stage I). These data indicate that GIRK1 may contribute to tumor progression and GIRK1 gene expression can serve as a useful prognostic marker in the overall and stage I NSCLCs

  19. The effect of MTA application on the affected dentine remineralization after partial caries excavation (in vivo)

    Science.gov (United States)

    Pratiwi, A. R.; Meidyawati, R.; Djauharie, N.

    2017-08-01

    On deep carious lesions, only thin dentine remains, causing a high risk of pulp exposure during the removal of all infected dentine. A minimally invasive technique is required, such as a partial caries excavation method in the infected dentine tissue and the use of bioactive material that can promote (Mineral Trioxide Aggregate) MTA remineralization. To compare the remineralization of deep carious lesion-affected dentine with the removal of some and all the infected dentine after the application of MTA. Subjects were divided into two groups: group I had only some parts of the infected dentine removed before MTA application, while group II had all the infected dentine removed before MTA application. Each group was measured on the pixel grey value before the treatment and again four weeks after the MTA application, and then the results were compared. Furthermore, the enhancement of both groups’ grey values were compared. Remineralization occurred in both groups after the MTA application. There was no significant difference in the remineralization level of the affected dentine in both groups I and II four weeks after the MTA application. Remineralization occurred in the affected dentine in both groups, either by removing only some parts or all the infected dentine in the deep carious lesion.

  20. HAP1 gene expression is associated with radiosensitivity in breast cancer cells

    International Nuclear Information System (INIS)

    Wu, Jing; Zhang, Jun-ying; Yin, Li; Wu, Jian-zhong; Guo, Wen-jie; Wu, Jian-feng; Chen, Meng; Xia, You-you; Tang, Jin-hai; Ma, Yong-chao; He, Xia

    2015-01-01

    Highlights: • Overexpression of HAP1 gene promotes apoptosis in MCF-7 cells after irradiation. • HAP1 reduces tumor volume in nude mice xenograft models after irradiation. • HAP1 increases radiosensitivity of breast cancer cells in vitro and vivo. - Abstract: Objectives: The purpose of this study was to investigate the relationship between huntingtin-associated protein1 (HAP1) gene and radiation therapy of breast cancer cells. Methods: HAP1 gene was transfected into breast cancer MCF-7 cells, which was confirmed by quantitative reverse transcription-polymerase chain reaction analysis (qRT-PCR) and Western blot in vitro. The changes of cell radiosensitivity were assessed by colony formation assay. Apoptosis were examined by flow cytometry. The expressions of two radiation-induced genes were evaluated by Western blot. Tumor growth was investigated in nude mice xenograft models in vivo. Results: Our data showed that HAP1 gene expression was significantly increased in HAP1-transfected MCF-7 cells in comparison with the parental cells or negative control cells. The survival rate in MCF-7/HAP1 cells was significantly decreased after irradiation (0, 2, 4, 6, 8 Gy), compared to cells in MCF-7 and MCF-7/Pb groups in vitro. HAP1 gene increased apoptosis in MCF-7 cells after irradiation. Additionally, the tumor volume and weight in MCF-7/HAP1 + RT group were observably lower than in MCF-7/HAP1 group and MCF-7/Pb + RT group. Conclusion: The present study indicated that HAP1 gene expression was related to the radiosensitivity of breast cancer cells and may play an important role in the regulation of cellular radiosensitivity

  1. Interactions between Bmp-4 and Msx-1 act to restrict gene expression to odontogenic mesenchyme.

    Science.gov (United States)

    Tucker, A S; Al Khamis, A; Sharpe, P T

    1998-08-01

    Tooth development is regulated by a reciprocal series of epithelial-mesenchymal interactions. Bmp4 has been identified as a candidate signalling molecule in these interactions, initially as an epithelial signal and then later at the bud stage as a mesenchymal signal (Vainio et al. [1993] Cell 75:45-58). A target gene for Bmp4 signalling is the homeobox gene Msx-1, identified by the ability of recombinant Bmp4 protein to induce expression in mesenchyme. There is, however, no evidence that Bmp4 is the endogenous inducer of Msx-1 expression. Msx-1 and Bmp-4 show dynamic, interactive patterns of expression in oral epithelium and ectomesenchyme during the early stages of tooth development. In this study, we compare the temporal and spatial expression of these two genes to determine whether the changing expression patterns of these genes are consistent with interactions between the two molecules. We show that changes in Bmp-4 expression precede changes in Msx-1 expression. At embryonic day (E)10.5-E11.0, expression patterns are consistent with BMP4 from the epithelium, inducing or maintaining Msx-1 in underlying mesenchyme. At E11.5, Bmp-4 expression shifts from epithelium to mesenchyme and is rapidly followed by localised up-regulation of Msx-1 expression at the sites of Bmp-4 expression. Using cultured explants of developing mandibles, we confirm that exogenous BMP4 is capable of replacing the endogenous source in epithelium and inducing Msx-1 gene expression in mesenchyme. By using noggin, a BMP inhibitor, we show that endogenous Msx-1 expression can be inhibited at E10.5 and E11.5, providing the first evidence that endogenous Bmp-4 from the epithelium is responsible for regulating the early spatial expression of Msx-1. We also show that the mesenchymal shift in Bmp-4 is responsible for up-regulating Msx-1 specifically at the sites of future tooth formation. Thus, we establish that a reciprocal series of interactions act to restrict expression of both genes to future

  2. Rce1, a novel transcriptional repressor, regulates cellulase gene expression by antagonizing the transactivator Xyr1 in Trichoderma reesei.

    Science.gov (United States)

    Cao, Yanli; Zheng, Fanglin; Wang, Lei; Zhao, Guolei; Chen, Guanjun; Zhang, Weixin; Liu, Weifeng

    2017-07-01

    Cellulase gene expression in the model cellulolytic fungus Trichoderma reesei is supposed to be controlled by an intricate regulatory network involving multiple transcription factors. Here, we identified a novel transcriptional repressor of cellulase gene expression, Rce1. Disruption of the rce1 gene not only facilitated the induced expression of cellulase genes but also led to a significant delay in terminating the induction process. However, Rce1 did not participate in Cre1-mediated catabolite repression. Electrophoretic mobility shift (EMSA) and DNase I footprinting assays in combination with chromatin immunoprecipitation (ChIP) demonstrated that Rce1 could bind directly to a cbh1 (cellobiohydrolase 1-encoding) gene promoter region containing a cluster of Xyr1 binding sites. Furthermore, competitive binding assays revealed that Rce1 antagonized Xyr1 from binding to the cbh1 promoter. These results indicate that intricate interactions exist between a variety of transcription factors to ensure tight and energy-efficient regulation of cellulase gene expression in T. reesei. This study also provides important clues regarding increased cellulase production in T. reesei. © 2017 John Wiley & Sons Ltd.

  3. Evolution and Expression Patterns of CYC/TB1 Genes in Anacyclus: Phylogenetic Insights for Floral Symmetry Genes in Asteraceae

    Science.gov (United States)

    Bello, María A.; Cubas, Pilar; Álvarez, Inés; Sanjuanbenito, Guillermo; Fuertes-Aguilar, Javier

    2017-01-01

    Homologs of the CYC/TB1 gene family have been independently recruited many times across the eudicots to control aspects of floral symmetry The family Asteraceae exhibits the largest known diversification in this gene paralog family accompanied by a parallel morphological floral richness in its specialized head-like inflorescence. In Asteraceae, whether or not CYC/TB1 gene floral symmetry function is preserved along organismic and gene lineages is unknown. In this study, we used phylogenetic, structural and expression analyses focused on the highly derived genus Anacyclus (tribe Anthemidae) to address this question. Phylogenetic reconstruction recovered eight main gene lineages present in Asteraceae: two from CYC1, four from CYC2 and two from CYC3-like genes. The species phylogeny was recovered in most of the gene lineages, allowing the delimitation of orthologous sets of CYC/TB1 genes in Asteraceae. Quantitative real-time PCR analysis indicated that in Anacyclus three of the four isolated CYC2 genes are more highly expressed in ray flowers. The expression of the four AcCYC2 genes overlaps in several organs including the ligule of ray flowers, as well as in anthers and ovules throughout development. PMID:28487706

  4. Differentially expressed genes in embryonic cardiac tissues of mice lacking Folr1 gene activity

    Directory of Open Access Journals (Sweden)

    Schwartz Robert J

    2007-11-01

    Full Text Available Abstract Background Heart anomalies are the most frequently observed among all human congenital defects. As with the situation for neural tube defects (NTDs, it has been demonstrated that women who use multivitamins containing folic acid peri-conceptionally have a reduced risk for delivering offspring with conotruncal heart defects 123. Cellular folate transport is mediated by a receptor or binding protein and by an anionic transporter protein system. Defective function of the Folr1 (also known as Folbp1; homologue of human FRα gene in mice results in inadequate transport, accumulation, or metabolism of folate during cardiovascular morphogenesis. Results We have observed cardiovascular abnormalities including outflow tract and aortic arch arterial defects in genetically compromised Folr1 knockout mice. In order to investigate the molecular mechanisms underlying the failure to complete development of outflow tract and aortic arch arteries in the Folr1 knockout mouse model, we examined tissue-specific gene expression difference between Folr1 nullizygous embryos and morphologically normal heterozygous embryos during early cardiac development (14-somite stage, heart tube looping (28-somite stage, and outflow track septation (38-somite stage. Microarray analysis was performed as a primary screening, followed by investigation using quantitative real-time PCR assays. Gene ontology analysis highlighted the following ontology groups: cell migration, cell motility and localization of cells, structural constituent of cytoskeleton, cell-cell adhesion, oxidoreductase, protein folding and mRNA processing. This study provided preliminary data and suggested potential candidate genes for further description and investigation. Conclusion The results suggested that Folr1 gene ablation and abnormal folate homeostasis altered gene expression in developing heart and conotruncal tissues. These changes affected normal cytoskeleton structures, cell migration and

  5. IGF-1 modulates gene expression of proteins involved in inflammation, cytoskeleton, and liver architecture.

    Science.gov (United States)

    Lara-Diaz, V J; Castilla-Cortazar, I; Martín-Estal, I; García-Magariño, M; Aguirre, G A; Puche, J E; de la Garza, R G; Morales, L A; Muñoz, U

    2017-05-01

    Even though the liver synthesizes most of circulating IGF-1, it lacks its receptor under physiological conditions. However, according to previous studies, a damaged liver expresses the receptor. For this reason, herein, we examine hepatic histology and expression of genes encoding proteins of the cytoskeleton, extracellular matrix, and cell-cell molecules and inflammation-related proteins. A partial IGF-1 deficiency murine model was used to investigate IGF-1's effects on liver by comparing wild-type controls, heterozygous igf1 +/- , and heterozygous mice treated with IGF-1 for 10 days. Histology, microarray for mRNA gene expression, RT-qPCR, and lipid peroxidation were assessed. Microarray analyses revealed significant underexpression of igf1 in heterozygous mice compared to control mice, restoring normal liver expression after treatment, which then normalized its circulating levels. IGF-1 receptor mRNA was overexpressed in Hz mice liver, while treated mice displayed a similar expression to that of the controls. Heterozygous mice showed overexpression of several genes encoding proteins related to inflammatory and acute-phase proteins and underexpression or overexpression of genes which coded for extracellular matrix, cytoskeleton, and cell junction components. Histology revealed an altered hepatic architecture. In addition, liver oxidative damage was found increased in the heterozygous group. The mere IGF-1 partial deficiency is associated with relevant alterations of the hepatic architecture and expression of genes involved in cytoskeleton, hepatocyte polarity, cell junctions, and extracellular matrix proteins. Moreover, it induces hepatic expression of the IGF-1 receptor and elevated acute-phase and inflammation mediators, which all resulted in liver oxidative damage.

  6. ELFN1-AS1: A Novel Primate Gene with Possible MicroRNA Function Expressed Predominantly in Human Tumors

    Directory of Open Access Journals (Sweden)

    Dmitrii E. Polev

    2014-01-01

    Full Text Available Human gene LOC100505644 uncharacterized LOC100505644 [Homo sapiens] (Entrez Gene ID 100505644 is abundantly expressed in tumors but weakly expressed in few normal tissues. Till now the function of this gene remains unknown. Here we identified the chromosomal borders of the transcribed region and the major splice form of the LOC100505644-specific transcript. We characterised the major regulatory motifs of the gene and its splice sites. Analysis of the secondary structure of the major transcript variant revealed a hairpin-like structure characteristic for precursor microRNAs. Comparative genomic analysis of the locus showed that it originated in primates de novo. Taken together, our data indicate that human gene LOC100505644 encodes some non-protein coding RNA, likely a microRNA. It was assigned a gene symbol ELFN1-AS1 (ELFN1 antisense RNA 1 (non-protein coding. This gene combines features of evolutionary novelty and predominant expression in tumors.

  7. Alteration in gene expression profile and oncogenicity of esophageal squamous cell carcinoma by RIZ1 upregulation.

    Science.gov (United States)

    Dong, Shang-Wen; Li, Dong; Xu, Cong; Sun, Pei; Wang, Yuan-Guo; Zhang, Peng

    2013-10-07

    To investigate the effect of retinoblastoma protein-interacting zinc finger gene 1 (RIZ1) upregulation in gene expression profile and oncogenicity of human esophageal squamous cell carcinoma (ESCC) cell line TE13. TE13 cells were transfected with pcDNA3.1(+)/RIZ1 and pcDNA3.1(+). Changes in gene expression profile were screened and the microarray results were confirmed by reverse transcription-polymerase chain reaction (RT-PCR). Nude mice were inoculated with TE13 cells to establish ESCC xenografts. After two weeks, the inoculated mice were randomly divided into three groups. Tumors were injected with normal saline, transfection reagent pcDNA3.1(+) and transfection reagent pcDNA3.1(+)/RIZ1, respectively. Tumor development was quantified, and changes in gene expression of RIZ1 transfected tumors were detected by RT-PCR and Western blotting. DNA microarray data showed that RIZ1 transfection induced widespread changes in gene expression profile of cell line TE13, with 960 genes upregulated and 1163 downregulated. Treatment of tumor xenografts with RIZ1 recombinant plasmid significantly inhibited tumor growth, decreased tumor size, and increased expression of RIZ1 mRNA compared to control groups. The changes in gene expression profile were also observed in vivo after RIZ1 transfection. Most of the differentially expressed genes were associated with cell development, supervision of viral replication, lymphocyte costimulatory and immune system development in esophageal cells. RIZ1 gene may be involved in multiple cancer pathways, such as cytokine receptor interaction and transforming growth factor beta signaling. The development and progression of esophageal cancer are related to the inactivation of RIZ1. Virus infection may also be an important factor.

  8. Isolation of MA-ACS Gene Family and Expression Study of MA-ACS1 Gene in Musa acuminata Cultivar Pisang Ambon Lumut

    Directory of Open Access Journals (Sweden)

    LISTYA UTAMI KARMAWAN

    2009-03-01

    Full Text Available Musa acuminata cultivar pisang ambon lumut is a native climacteric fruit from Indonesia. Climacteric fruit ripening process is triggered by the gaseous plant hormone ethylene. The rate limiting enzyme involved in ethylene biosynthesis is ACC synthase (ACS which is encoded by ACS gene family. The objective of this study is to identify MA-ACS gene family in M. acuminata cultivar pisang ambon lumut and to study the MA-ACS1 gene expression. The result showed that there were nine M. acuminata ACS gene family members called MA-ACS1–9. Two of them (MA-ACS1 and MA-ACS2 were assessed using reverse transcriptase PCR (RT-PCR for gene expression study and it was only MA-ACS1 correlated with fruit ripening. The MA-ACS1 gene fragment has been successfully isolated and characterized and it has three introns, four exons, and one stop codon. It also shows highest homology with MACS1 gene from M. acuminata cultivar Hsian Jien Chiao (GenBank accession number AF056164. Expression analysis of MA-ACS1 using quantitative PCR (qPCR showed that MA-ACS1 gene expression increased significantly in the third day, reached maximum at the fifth day, and then decreased in the seventh day after harvesting. The qPCR expression analysis result correlated with the result of physical analysis during fruit ripening.

  9. Effect of MTA and Portland Cement on Fracture Resistance of Dentin

    Directory of Open Access Journals (Sweden)

    Maryam Forghani

    2013-06-01

    Full Text Available Background and aims. It is important to evaluate the effects of endodontic materials on tooth structures to avoid endodontic treatment failure. The aim of the present study was to investigate the effect of mineral trioxide aggregates (MTA and Portland cement (PC on fracture resistance of dentin. Materials and methods. Thirty-six freshly extracted human single-rooted premolar teeth were selected. The crowns were removed and the roots were randomly divided into two experimental groups and one control group. The root samples were longitudinally divided into two halves and a dentin bar (2×2×10 mm was cut from each root section for short-term (2 weeks and long-term (12 weeks evaluations. The root sections in the experimental groups were exposed to MTA or PC, while keeping the control group specimens in physiologic saline. The fracture resistance of each specimen was measured using an Instron testing machine. The results were statistically analyzed using ANOVA, a post hoc Tukey test and paired ttest at 5% significance level. Results. The fracture resistance of MTA-treated specimens significantly increased between 2 and 12 weeks (P0.05. Conclusion. The results showed that MTA increased the fracture resistance of root dentin, while PC had no significant effect on dentin fracture resistance.

  10. [Construction and application of prokaryotic expression system of Leptospira interrogans lipL32/1-lipL41/1 fusion gene].

    Science.gov (United States)

    Luo, Dong-jiao; Yan, Jie; Mao, Ya-fei; Li, Shu-ping; Luo, Yi-hui; Li, Li-wei

    2005-01-01

    To construct lipL32/1-lipL41/1 fusion gene and its prokaryotic expression system and to determine frequencies of carrying and expression of lipL32 and lipL41 genes in L.interrogans wild strains and specific antibody levels in sera from leptospirosis patients. lipL32/1-lipL41/1 fusion gene was constructed using linking primer PCR method and the prokaryotic expression system of the fusion gene done with routine techniques. SDS-PAGE was used to examine expression of the target recombinant protein rLipL32/1-rLipL41/1. Immunogenicity of rLipL32/1-rLipL41/1 was identified by Western blot. PCR and MAT were performed to detect carrying and expression of lipL32 and lipL41 genes in 97 wild L.interrogans strains. Antibodies against products of lipL32 and lipL41 genes in serum samples from 228 leptospirosis patients were detected by ELISA method. The homogeneity of nucleotide and putative amino acid sequence of lipL32/1-lipL41/1 fusion gene were 99.9 % and 99.8 % in comparison with the reported sequences. Expression output of the target recombinant protein rLipL32/1-rLipL41/1, mainly present in inclusion body, accounted for 10 % of the total bacterial proteins. Both the rabbit antisera against rLipL32/1 and rLipL41/1 could combine to rLipL32/1-rLipL41/1. 97.9 % and 87.6 % of the L.interrogans wild strains had lipL32 and lipL41 genes, respectively. 95.9 % and 84.5 % of the wild strains were positive for MAT with titers of 1:4 - 1:128 using rabbit anti-rLipL32s or anti-rLipL41s sera, respectively. 94.7 % - 97.4 % of the patients'serum samples were positive for rLipL32s antibodies, while 78.5 % - 84.6 % of them were rLipL41s antibodies detectable. lipL32/1-jlipL41/1 fusion gene and its prokaryotic expression system were successfully constructed. The expressed fusion protein had qualified immunogenicity. Both the lipL32 and lipL41 genes are extensively carried and frequently expressed by different serogroups of L.interrogans, and their expression products exhibit cross-antigenicity.

  11. Elicitor and fusarium-induced expression of NPR-1 like genes in banana

    CSIR Research Space (South Africa)

    Endah, R

    2008-11-01

    Full Text Available NPR1 is an essential positive regulator of salicylic acid-induced PR gene expression and systemic acquired resistance. Two novel full-length NPR1-like genes; MNPR1A and MNPR1B, were isolated by application of the PCR and RACE techniques. The two...

  12. Tumor-produced, active Interleukin-1 β regulates gene expression in carcinoma-associated fibroblasts

    International Nuclear Information System (INIS)

    Dudas, Jozsef; Fullar, Alexandra; Bitsche, Mario; Schartinger, Volker; Kovalszky, Ilona; Sprinzl, Georg Mathias; Riechelmann, Herbert

    2011-01-01

    Recently we described a co-culture model of periodontal ligament (PDL) fibroblasts and SCC-25 lingual squamous carcinoma cells, which resulted in conversion of normal fibroblasts into carcinoma-associated fibroblasts (CAFs), and in epithelial-mesenchymal transition (EMT) of SCC-25 cells. We have found a constitutive high interleukin-1β (IL1-β) expression in SCC-25 cells in normal and in co-cultured conditions. In our hypothesis a constitutive IL1expression in SCC-25 regulates gene expression in fibroblasts during co-culture. Co-cultures were performed between PDL fibroblasts and SCC-25 cells with and without dexamethasone (DEX) treatment; IL1-β processing was investigated in SCC-25 cells, tumor cells and PDL fibroblasts were treated with IL1-β. IL1-β signaling was investigated by western blot and immunocytochemistry. IL1-β-regulated genes were analyzed by real-time qPCR. SCC-25 cells produced 16 kD active IL1-β, its receptor was upregulated in PDL fibroblasts during co-culture, which induced phosphorylation of interleukin-1 receptor-associated kinase-1 (IRAK-1), and nuclear translocalization of NFκBα. Several genes, including interferon regulatory factor 1 (IRF1) interleukin-6 (IL-6) and prostaglandin-endoperoxide synthase 2 (COX-2) were induced in CAFs during co-culture. The most enhanced induction was found for IL-6 and COX-2. Treatment of PDL fibroblasts with IL1-β reproduced a time- and dose-dependent upregulation of IL1-receptor, IL-6 and COX-2. A further proof was achieved by DEX inhibition for IL1-β-stimulated IL-6 and COX-2 gene expression. Constitutive expression of IL1-β in the tumor cells leads to IL1-β-stimulated gene expression changes in tumor-associated fibroblasts, which are involved in tumor progression. -- Graphical abstract: SCC-25 cells produce active, processed IL1-β. PDL fibroblasts possess receptor for IL1-β, and its expression is increased 4.56-times in the presence of SCC-25 tumor cells. IL1-β receptor expression in

  13. Tumor-produced, active Interleukin-1 {beta} regulates gene expression in carcinoma-associated fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Dudas, Jozsef, E-mail: Jozsef.Dudas@i-med.ac.at [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria); Fullar, Alexandra, E-mail: fullarsz@gmail.com [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria); 1st Institute of Pathology and Experimental Cancer Research, Semmelweis University, Ulloei ut 26, H-1085 Budapest (Hungary); Bitsche, Mario, E-mail: Mario.Bitsche@i-med.ac.at [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria); Schartinger, Volker, E-mail: Volker.Schartinger@i-med.ac.at [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria); Kovalszky, Ilona, E-mail: koval@korb1.sote.hu [1st Institute of Pathology and Experimental Cancer Research, Semmelweis University, Ulloei ut 26, H-1085 Budapest (Hungary); Sprinzl, Georg Mathias, E-mail: Georg.Sprinzl@i-med.ac.at [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria); Riechelmann, Herbert, E-mail: Herbert.Riechelmann@i-med.ac.at [Department of Otorhinolaryngology, Medical University Innsbruck, Anichstrasse 35, A-6020 Innsbruck (Austria)

    2011-09-10

    Recently we described a co-culture model of periodontal ligament (PDL) fibroblasts and SCC-25 lingual squamous carcinoma cells, which resulted in conversion of normal fibroblasts into carcinoma-associated fibroblasts (CAFs), and in epithelial-mesenchymal transition (EMT) of SCC-25 cells. We have found a constitutive high interleukin-1{beta} (IL1-{beta}) expression in SCC-25 cells in normal and in co-cultured conditions. In our hypothesis a constitutive IL1-{beta} expression in SCC-25 regulates gene expression in fibroblasts during co-culture. Co-cultures were performed between PDL fibroblasts and SCC-25 cells with and without dexamethasone (DEX) treatment; IL1-{beta} processing was investigated in SCC-25 cells, tumor cells and PDL fibroblasts were treated with IL1-{beta}. IL1-{beta} signaling was investigated by western blot and immunocytochemistry. IL1-{beta}-regulated genes were analyzed by real-time qPCR. SCC-25 cells produced 16 kD active IL1-{beta}, its receptor was upregulated in PDL fibroblasts during co-culture, which induced phosphorylation of interleukin-1 receptor-associated kinase-1 (IRAK-1), and nuclear translocalization of NF{kappa}B{alpha}. Several genes, including interferon regulatory factor 1 (IRF1) interleukin-6 (IL-6) and prostaglandin-endoperoxide synthase 2 (COX-2) were induced in CAFs during co-culture. The most enhanced induction was found for IL-6 and COX-2. Treatment of PDL fibroblasts with IL1-{beta} reproduced a time- and dose-dependent upregulation of IL1-receptor, IL-6 and COX-2. A further proof was achieved by DEX inhibition for IL1-{beta}-stimulated IL-6 and COX-2 gene expression. Constitutive expression of IL1-{beta} in the tumor cells leads to IL1-{beta}-stimulated gene expression changes in tumor-associated fibroblasts, which are involved in tumor progression. -- Graphical abstract: SCC-25 cells produce active, processed IL1-{beta}. PDL fibroblasts possess receptor for IL1-{beta}, and its expression is increased 4.56-times in the

  14. Histological and histomorphometrical evaluation of furcation perforations filled with MTA, CPM and ZOE.

    Science.gov (United States)

    da Silva, G F; Guerreiro-Tanomaru, J M; Sasso-Cerri, E; Tanomaru-Filho, M; Cerri, P S

    2011-02-01

    To evaluate the biological response of the periodontium adjacent to furcation perforations in rat molars filled with Endo-CPM-Sealer (CPM), MTA-Angelus (MTA) or zinc oxide-eugenol cement (ZOE). The pulp chamber floors of maxillary right first molar teeth were perforated and sealed with CPM, mineral trioxide aggregate (MTA) or ZOE; the left first molars, without any treatment, were used as controls (CG). After 7, 15, 30 and 60 days, fragments of maxilla were fixed, decalcified and embedded in paraffin. Sections were stained with H&E, Masson's trichrome and submitted to tartrate-resistant acid phosphatase (TRAP) reaction, used as an osteoclast marker. The width of the periodontal space, the numerical density of inflammatory cells and the number of TRAP-positive osteoclasts in the bone surface were measured, and statistical analyses were performed using analysis of variance and Tukey test (P ≤ 0.05). In all experimental groups, the greatest number of inflammatory cells was observed at 7 days, especially in the ZOE group. In this group, the intense inflammatory process was related to a significant increase (P ≤ 0.05) in the number of osteoclasts and, thereby, in an increase in the width of the periodontal space. At 60 days, no significant differences in osteoclast numbers amongst CPM, MTA and CG groups occurred; the periodontal space was also significantly reduced in the experimental groups in comparison with the initial periods. However, in the ZOE group, the periodontal space was significantly larger (P ≤ 0.05) in comparison with MTA-based materials. The periodontium adjacent to perforations filled with MTA and CPM exhibited clear evidence of re-establishment and thus better biocompatibility than ZOE. © 2010 International Endodontic Journal.

  15. Metastatic tumor antigen 3 is a direct corepressor of the Wnt4 pathway

    OpenAIRE

    Zhang, Hao; Singh, Rajesh R.; Talukder, Amjad H.; Kumar, Rakesh

    2006-01-01

    Here we show that expression of MTA3 inhibits ductal branching in virgin and pregnant murine transgenic mammary glands. MTA3 also suppresses the Wnt4 pathway and, thus, these findings parallel phenotypic changes in Wnt4-null mice. MTA3 represses Wnt4 transcription and Wnt4 secretion, inhibiting Wnt-target genes in mammary epithelial cells. Accordingly, knockdown of endogenous MTA3 stimulates Wnt4 expression and Wnt cellular targets. The MTA3–NuRD (nucleosome remodeling and deacetylase) comple...

  16. Identification of Achaete-scute complex-like 1 (ASCL1) target genes and evaluation of DKK1 and TPH1 expression in pancreatic endocrine tumours

    International Nuclear Information System (INIS)

    Johansson, Térèse A; Westin, Gunnar; Skogseid, Britt

    2009-01-01

    ASCL1 role in pancreatic endocrine tumourigenesis has not been established. Recently it was suggested that ASCL1 negatively controls expression of the Wnt signalling antagonist DKK1. Notch signalling regulates expression of TPH1, the rate limiting enzyme in the biosyntesis of serotonin. Understanding the development and proliferation of pancreatic endocrine tumours (PETs) is essential for the development of new therapies. ASCL1 target genes in the pancreatic endocrine tumour cell line BON1 were identified by RNA interference and microarray expression analysis. Protein expressions of selected target genes in PETs were evaluated by immunohistochemistry. 158 annotated ASCL1 target genes were identified in BON1 cells, among them DKK1 and TPH1 that were negatively regulated by ASCL1. An inverse relation of ASCL1 to DKK1 protein expression was observed for 15 out of 22 tumours (68%). Nine tumours displayed low ASCL1/high DKK1 and six tumours high ASCL1/low DKK1 expression. Remaining PETs showed high ASCL1/high DKK1 (n = 4) or low ASCL1/low DKK1 (n = 3) expression. Nine of twelve analysed PETs (75%) showed TPH1 expression with no relation to ASCL1. A number of genes with potential importance for PET tumourigenesis have been identified. ASCL1 negatively regulated the Wnt signalling antagonist DKK1, and TPH1 expression in BON1 cells. In concordance with these findings DKK1 showed an inverse relation to ASCL1 expression in a subset of PETs, which may affect growth control by the Wnt signalling pathway

  17. The ASK1 gene regulates B function gene expression in cooperation with UFO and LEAFY in Arabidopsis.

    Science.gov (United States)

    Zhao, D; Yu, Q; Chen, M; Ma, H

    2001-07-01

    The Arabidopsis floral regulatory genes APETALA3 (AP3) and PISTILLATA (PI) are required for the B function according to the ABC model for floral organ identity. AP3 and PI expression are positively regulated by the LEAFY (LFY) and UNUSUAL FLORAL ORGANS (UFO) genes. UFO encodes an F-box protein, and we have shown previously that UFO genetically interacts with the ASK1 gene encoding a SKP1 homologue; both the F-box containing protein and SKP1 are subunits of ubiquitin ligases. We show here that the ask1-1 mutation can enhance the floral phenotypes of weak lfy and ap3 mutants; therefore, like UFO, ASK1 also interacts with LFY and AP3 genetically. Furthermore, our results from RNA in situ hybridizations indicate that ASK1 regulates early AP3 and PI expression. These results support the idea that UFO and ASK1 together positively regulate AP3 and PI expression. We propose that the UFO and ASK1 proteins are components of a ubiquitin ligase that mediates the proteolysis of a repressor of AP3 and PI expression. Our genetic studies also indicate that ASK1 and UFO play a role in regulating the number of floral organ primordia, and we discuss possible mechanisms for such a regulation.

  18. Dietary Lecithin Decreases Skeletal Muscle COL1A1 and COL3A1 Gene Expression in Finisher Gilts

    Directory of Open Access Journals (Sweden)

    Henny Akit

    2016-06-01

    Full Text Available The purpose of this study was to investigate the effect of dietary lecithin on skeletal muscle gene expression of collagen precursors and enzymes involved in collagen synthesis and degradation. Finisher gilts with an average start weight of 55.9 ± 2.22 kg were fed diets containing either 0, 4, 20 or 80 g/kg soybean lecithin prior to harvest for six weeks and the rectus abdominis muscle gene expression profile was analyzed by quantitative real-time PCR. Lecithin treatment down-regulated Type I (α1 procollagen (COL1A1 and Type III (α1 procollagen (COL3A1 mRNA expression ( p < 0.05, respectively, indicating a decrease in the precursors for collagen synthesis. The α-subunit of prolyl 4-hydroxylase (P4H mRNA expression also tended to be down-regulated ( p = 0.056, indicating a decrease in collagen synthesis. Decreased matrix metalloproteinase-1 (MMP-1 mRNA expression may reflect a positive regulatory response to the reduced collagen synthesis in muscle from the pigs fed lecithin ( p = 0.035. Lecithin had no effect on tissue inhibitor metalloproteinase-1 (TIMP-1, matrix metalloproteinase-13 (MMP-13 and lysyl oxidase mRNA expression. In conclusion, lecithin down-regulated COL1A1 and COL3A1 as well as tended to down-regulate α-subunit P4H expression. However, determination of muscle collagen content and solubility are required to support the gene functions.

  19. Overexpression of transcription factor Sp1 leads to gene expression perturbations and cell cycle inhibition.

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    Emmanuelle Deniaud

    Full Text Available BACKGROUND: The ubiquitous transcription factor Sp1 regulates the expression of a vast number of genes involved in many cellular functions ranging from differentiation to proliferation and apoptosis. Sp1 expression levels show a dramatic increase during transformation and this could play a critical role for tumour development or maintenance. Although Sp1 deregulation might be beneficial for tumour cells, its overexpression induces apoptosis of untransformed cells. Here we further characterised the functional and transcriptional responses of untransformed cells following Sp1 overexpression. METHODOLOGY AND PRINCIPAL FINDINGS: We made use of wild-type and DNA-binding-deficient Sp1 to demonstrate that the induction of apoptosis by Sp1 is dependent on its capacity to bind DNA. Genome-wide expression profiling identified genes involved in cancer, cell death and cell cycle as being enriched among differentially expressed genes following Sp1 overexpression. In silico search to determine the presence of Sp1 binding sites in the promoter region of modulated genes was conducted. Genes that contained Sp1 binding sites in their promoters were enriched among down-regulated genes. The endogenous sp1 gene is one of the most down-regulated suggesting a negative feedback loop induced by overexpressed Sp1. In contrast, genes containing Sp1 binding sites in their promoters were not enriched among up-regulated genes. These results suggest that the transcriptional response involves both direct Sp1-driven transcription and indirect mechanisms. Finally, we show that Sp1 overexpression led to a modified expression of G1/S transition regulatory genes such as the down-regulation of cyclin D2 and the up-regulation of cyclin G2 and cdkn2c/p18 expression. The biological significance of these modifications was confirmed by showing that the cells accumulated in the G1 phase of the cell cycle before the onset of apoptosis. CONCLUSION: This study shows that the binding to DNA

  20. Effects of LED or laser phototherapy on bone defects grafted with MTA and irradiated with laser or LED light: a comparative Raman spectroscopic study

    Science.gov (United States)

    Pinheiro, Antonio L. B.; Soares, Luiz G. P.; Barbosa, Artur Felipe S.; Silveira, Landulfo, Jr.

    2012-03-01

    We studied peaks of calcium hydroxyapatite - CHA on defects grafted with MTA, treated or not with Light Emitting Diode - LED or IR Laser. 54 rats were divided in 6 groups each subdivided into 3 subgroups (15,21,30d). LED (λ850 +/- 10nm) or IR Laser (λ850 nm) was applied over (LED) or in 4 points around the defect at 48 h intervals for 15 days. Raman readings were taken at the surface of the defect. The smaller overall intensity of the peak was found in Group MTA + Laser (1510.2 +/- 274.1) and the highest on Group LED (2322 +/- 715). There were no statistically significant differences between non-irradiated subjects on regards the CHA peaks. On the other hand, there were statistically significant differences between the Group Clot and LED, Clot and Laser, and Clot and MTA + Laser (p =0.01, p = 0.02, p = 0.003). There were no significant differences between Group MTA and MTA + LED (p=0.2) but significant differences were seen between Groups MTA and MTA + Laser (p=0.01). Significant differences were also observed between Groups LED and Laser (p <0.001) and between Groups MTA + LED and MTA + Laser (p=0.009). MTA, due to its characteristics, seemed to be directly affected by the light. However, the use of either phototherapy positively affected bone healing similarly as observed on different studies using other biomaterials. The overall analysis of our results indicated that the use of either light source resulted in a better, more advanced, and of quality bone repair.

  1. IL-1 receptor antagonism and muscle gene expression in patients with type 2 diabetes

    DEFF Research Database (Denmark)

    Berchtold, L. A.; Larsen, C. M.; Vaag, A.

    2009-01-01

    ). To investigate the effects of IL-1Ra in insulin-sensitive tissue, gene expression levels in skeletal muscle from type 2 diabetic patients treated with IL-1Ra were analysed. Methods. Gene expression profiles in vastus lateralis muscle biopsies from five obese patients (BMI>27) were determined before and after 13......RT-PCR, were significantly altered when comparing the number of transcripts before and after treatment for each individual. Conclusion. Treatment with IL-1Ra did not significantly affect gene expression levels in skeletal muscle in this limited and selected sample of obese patients with type 2 diabetes. Larger...

  2. Global loss of bmal1 expression alters adipose tissue hormones, gene expression and glucose metabolism.

    Directory of Open Access Journals (Sweden)

    David John Kennaway

    Full Text Available The close relationship between circadian rhythm disruption and poor metabolic status is becoming increasingly evident, but role of adipokines is poorly understood. Here we investigated adipocyte function and the metabolic status of mice with a global loss of the core clock gene Bmal1 fed either a normal or a high fat diet (22% by weight. Bmal1 null mice aged 2 months were killed across 24 hours and plasma adiponectin and leptin, and adipose tissue expression of Adipoq, Lep, Retn and Nampt mRNA measured. Glucose, insulin and pyruvate tolerance tests were conducted and the expression of liver glycolytic and gluconeogenic enzyme mRNA determined. Bmal1 null mice displayed a pattern of increased plasma adiponectin and plasma leptin concentrations on both control and high fat diets. Bmal1 null male and female mice displayed increased adiposity (1.8 fold and 2.3 fold respectively on the normal diet, but the high fat diet did not exaggerate these differences. Despite normal glucose and insulin tolerance, Bmal1 null mice had increased production of glucose from pyruvate, implying increased liver gluconeogenesis. The Bmal1 null mice had arrhythmic clock gene expression in epigonadal fat and liver, and loss of rhythmic transcription of a range of metabolic genes. Furthermore, the expression of epigonadal fat Adipoq, Retn, Nampt, AdipoR1 and AdipoR2 and liver Pfkfb3 mRNA were down-regulated. These results show for the first time that global loss of Bmal1, and the consequent arrhythmicity, results in compensatory changes in adipokines involved in the cellular control of glucose metabolism.

  3. Cholesterol Transporters ABCA1 and ABCG1 Gene Expression in Peripheral Blood Mononuclear Cells in Patients with Metabolic Syndrome

    Directory of Open Access Journals (Sweden)

    Zahra Tavoosi

    2015-01-01

    Full Text Available ABCA1 and ABCG1 genes encode the cholesterol transporter proteins that play a key role in cholesterol and phospholipids homeostasis. This study was aimed at evaluating and comparing ABCA1 and ABCG1 genes expression in metabolic syndrome patients and healthy individuals. This case-control study was performed on 36 patients with metabolic syndrome and the same number of healthy individuals in Hamadan (west of Iran during 2013-2014. Total RNA was extracted from mononuclear cells and purified using RNeasy Mini Kit column. The expression of ABCA1 and ABCG1 genes was performed by qRT-PCR. Lipid profile and fasting blood glucose were measured using colorimetric procedures. ABCG1 expression in metabolic syndrome patients was significantly lower (about 75% compared to that of control group, while for ABCA1 expression, there was no significant difference between the two studied groups. Comparison of other parameters such as HDL-C, FBS, BMI, waist circumference, and systolic and diastolic blood pressure between metabolic syndrome patients and healthy individuals showed significant differences (P<0.05. Decrease in ABCG1 expression in metabolic syndrome patients compared to healthy individuals suggests that hyperglycemia, related metabolites, and hyperlipidemia over the transporter capacity resulted in decreased expression of ABCG1. Absence of a significant change in ABCA1 gene expression between two groups can indicate a different regulation mechanism for ABCA1 expression.

  4. Periradicular Tissue Responses to Biologically Active Molecules or MTA When Applied in Furcal Perforation of Dogs' Teeth

    Directory of Open Access Journals (Sweden)

    Anna Zairi

    2012-01-01

    Full Text Available The aim of this study was the comparative evaluation of inflammatory reactions and tissue responses to four growth factors, or mineral trioxide aggregate (MTA, or a zinc-oxide-eugenol-based cement (IRM as controls, when used for the repair of furcal perforations in dogs’ teeth. Results showed significantly higher inflammatory cell response in the transforming growth factorβ1 (TGFβ1 and zinc-oxide-eugenol-based cement (IRM groups and higher rates of epithelial proliferation in the TGFβ1, basic fibroblast growth factor (bFGF, and insulin growth factor-I (IGF-I groups compared to the MTA. Significantly higher rates of bone formation were found in the control groups compared to the osteogenic protein-1 (OP-1. Significantly higher rates of cementum formation were observed in the IGF-I and bFGF groups compared to the IRM. None of the biologically active molecules can be suggested for repairing furcal perforations, despite the fact that growth factors exerted a clear stimulatory effect on cementum formation and inhibited collagen capsule formation. MTA exhibited better results than the growth factors.

  5. KAP1 regulates type I interferon/STAT1-mediated IRF-1 gene expression

    International Nuclear Information System (INIS)

    Kamitani, Shinya; Ohbayashi, Norihiko; Ikeda, Osamu; Togi, Sumihito; Muromoto, Ryuta; Sekine, Yuichi; Ohta, Kazuhide; Ishiyama, Hironobu; Matsuda, Tadashi

    2008-01-01

    Signal transducers and activators of transcription (STATs) mediate cell proliferation, differentiation, and survival in immune responses, hematopoiesis, neurogenesis, and other biological processes. Recently, we showed that KAP1 is a novel STAT-binding partner that regulates STAT3-mediated transactivation. KAP1 is a universal co-repressor protein for the KRAB zinc finger protein superfamily of transcriptional repressors. In this study, we found KAP1-dependent repression of interferon (IFN)/STAT1-mediated signaling. We also demonstrated that endogenous KAP1 associates with endogenous STAT1 in vivo. Importantly, a small-interfering RNA-mediated reduction in KAP1 expression enhanced IFN-induced STAT1-dependent IRF-1 gene expression. These results indicate that KAP1 may act as an endogenous regulator of the IFN/STAT1 signaling pathway

  6. In planta gene expression analysis of Xanthomonas oryzae pathovar oryzae, African strain MAI1

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    Verdier Valérie

    2010-06-01

    Full Text Available Abstract Background Bacterial leaf blight causes significant yield losses in rice crops throughout Asia and Africa. Although both the Asian and African strains of the pathogen, Xanthomonas oryzae pv. oryzae (Xoo, induce similar symptoms, they are nevertheless genetically different, with the African strains being more closely related to the Asian X. oryzae pv. oryzicola (Xoc. Results Changes in gene expression of the African Xoo strain MAI1 in the susceptible rice cultivar Nipponbare were profiled, using an SSH Xoo DNA microarray. Microarray hybridization was performed comparing bacteria recovered from plant tissues at 1, 3, and 6 days after inoculation (dai with bacteria grown in vitro. A total of 710 bacterial genes were found to be differentially expressed, with 407 up-regulated and 303 down-regulated. Expression profiling indicated that less than 20% of the 710 bacterial transcripts were induced in the first 24 h after inoculation, whereas 63% were differentially expressed at 6 dai. The 710 differentially expressed genes were one-end sequenced. 535 sequences were obtained from which 147 non-redundant sequences were identified. Differentially expressed genes were related to metabolism, secretion and transport, pathogen adherence to plant tissues, plant cell-wall degradation, IS elements, and virulence. In addition, various other genes encoding proteins with unknown function or showing no similarity to other proteins were also induced. The Xoo MAI1 non-redundant set of sequences was compared against several X. oryzae genomes, revealing a specific group of genes that was present only in MAI1. Numerous IS elements were also found to be differentially expressed. Quantitative real-time PCR confirmed 86% of the identified profile on a set of 14 genes selected according to the microarray analysis. Conclusions This is the first report to compare the expression of Xoo genes in planta across different time points during infection. This work shows that

  7. Gene expression and gene therapy imaging

    International Nuclear Information System (INIS)

    Rome, Claire; Couillaud, Franck; Moonen, Chrit T.W.

    2007-01-01

    The fast growing field of molecular imaging has achieved major advances in imaging gene expression, an important element of gene therapy. Gene expression imaging is based on specific probes or contrast agents that allow either direct or indirect spatio-temporal evaluation of gene expression. Direct evaluation is possible with, for example, contrast agents that bind directly to a specific target (e.g., receptor). Indirect evaluation may be achieved by using specific substrate probes for a target enzyme. The use of marker genes, also called reporter genes, is an essential element of MI approaches for gene expression in gene therapy. The marker gene may not have a therapeutic role itself, but by coupling the marker gene to a therapeutic gene, expression of the marker gene reports on the expression of the therapeutic gene. Nuclear medicine and optical approaches are highly sensitive (detection of probes in the picomolar range), whereas MRI and ultrasound imaging are less sensitive and require amplification techniques and/or accumulation of contrast agents in enlarged contrast particles. Recently developed MI techniques are particularly relevant for gene therapy. Amongst these are the possibility to track gene therapy vectors such as stem cells, and the techniques that allow spatiotemporal control of gene expression by non-invasive heating (with MRI guided focused ultrasound) and the use of temperature sensitive promoters. (orig.)

  8. Evaluation of physico-chemical properties of Portland cements and MTA

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    Jorge Luis Gonçalves

    2010-09-01

    Full Text Available The purpose of this study was to evaluate the hydrogenionic potential and electrical conductivity of Portland cements and MTA, as well as the amount of arsenic and calcium released from these materials. In Teflon molds, samples of each material were agitated and added to plastic flasks containing distilled water for 3, 24, 72 and 168 h. The results were analyzed with a Kruskal-Wallis non-parametric test for global comparisons and a Dunn-Tukey test for pairwise comparisons. The results revealed no significant differences in the pH of the materials (p > 0.05. The electrical conductivity of the cements were not statistically different (p > 0.05. White non-structural cement and MTA BIO released the largest amount of calcium ions into solution (p 0.05. The results indicated that the physico-chemical properties of Portland cements and MTA were similar. Furthermore, all materials produced an alkaline environment and can be considered safe for clinical use because arsenic was not released. The electrical conductivity and the amount of calcium ions released into solution increased over time.

  9. Evaluation of physico-chemical properties of Portland cements and MTA.

    Science.gov (United States)

    Gonçalves, Jorge Luis; Viapiana, Raqueli; Miranda, Carlos Eduardo Saraiva; Borges, Alvaro Henrique; Cruz Filho, Antônio Miranda da

    2010-01-01

    The purpose of this study was to evaluate the hydrogenionic potential and electrical conductivity of Portland cements and MTA, as well as the amount of arsenic and calcium released from these materials. In Teflon molds, samples of each material were agitated and added to plastic flasks containing distilled water for 3, 24, 72 and 168 h. The results were analyzed with a Kruskal-Wallis non-parametric test for global comparisons and a Dunn-Tukey test for pairwise comparisons. The results revealed no significant differences in the pH of the materials (p > 0.05). The electrical conductivity of the cements were not statistically different (p > 0.05). White non-structural cement and MTA BIO released the largest amount of calcium ions into solution (p 0.05). The results indicated that the physico-chemical properties of Portland cements and MTA were similar. Furthermore, all materials produced an alkaline environment and can be considered safe for clinical use because arsenic was not released. The electrical conductivity and the amount of calcium ions released into solution increased over time.

  10. Cloning, expression and characterization of COI1 gene (AsCOI1 from Aquilaria sinensis (Lour. Gilg

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    Yongcui Liao

    2015-09-01

    Full Text Available Aquilaria sinensis, a kind of typically wounding-induced medicinal plant with a great economical value, is widely used in the production of traditional Chinese medicine, perfume and incense. Coronatine-insensitive protein 1 (COI1 acts as a receptor in jasmonate (JA signaling pathway, and regulates the expression of JA-responsive genes in plant defense. However, little is known about the COI1 gene in A. sinensis. Here, based on the transcriptome data, a full-length cDNA sequence of COI1 (termed as AsCOI1 was firstly cloned by RT–PCR and rapid-amplification of cDNA ends (RACE strategies. AsCOI1 is 2330 bp in length (GenBank accession No. KM189194, and contains a complete open frame (ORF of 1839 bp. The deduced protein was composed of 612 amino acids, with a predicted molecular weight of 68.93 kDa and an isoelectric point of 6.56, and was predicted to possess F-box and LRRs domains. Combining bioinformatics prediction with subcellular localization experiment analysis, AsCOI1 was appeared to locate in nucleus. AsCOI1 gene was highly expressed in roots and stems, the major organs of agarwood formation. Methyl jasmonate (MeJA, mechanical wounding and heat stress could significantly induce the expression level of AsCOI1 gene. AsCOI1 is an early wound-responsive gene, and it likely plays some role in agarwood formation.

  11. Ribavirin restores ESR1 gene expression and tamoxifen sensitivity in ESR1 negative breast cancer cell lines

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    Sappok Anne

    2011-12-01

    Full Text Available Abstract Tumor growth is estrogen independent in approximately one-third of all breast cancers, which makes these patients unresponsive to hormonal treatment. This unresponsiveness to hormonal treatment may be explained through the absence of the estrogen receptor alpha (ESR1. The ESR1 gene re-expression through epigenetic modulators such as DNA methyltransferase inhibitors and/or histone deacetylase inhibitors restores tamoxifen sensitivity in ESR1 negative breast cancer cell lines and opens new treatment horizons in patients who were previously associated with a poor prognosis. In the study presented herein, we tested the ability of ribavirin, which shares some structural similarities with the DNA-methyltransferase inhibitor 5-azacytidine and which is widely known as an anti-viral agent in the treatment of hepatitis C, to restore ESR1 gene re-expression in ESR1 negative breast cancer cell lines. In our study we identified ribavirin to restore ESR1 gene re-expression alone and even more in combination with suberoylanilide hydroxamic acid (SAHA - up to 276 fold induction. Ribavirin and analogs could pave the way to novel translational research projects that aim to restore ESR1 gene re-expression and thus the susceptibility to tamoxifen-based endocrine treatment strategies.

  12. Pulpotomy of human primary molars with MTA and Portland cement: a randomised controlled trial.

    Science.gov (United States)

    Sakai, Vivien Thiemy; Moretti, A B S; Oliveira, T M; Fornetti, A P C; Santos, C F; Machado, M A A M; Abdo, R C C

    2009-08-08

    This study compared the clinical and radiographic effectiveness of mineral trioxide aggregate (MTA) and Portland cement (PC) as pulp dressing agents in carious primary teeth. Thirty carious primary mandibular molars of children aged 5-9 years old were randomly assigned to MTA or PC groups, and treated by a conventional pulpotomy technique. The teeth were restored with resin modified glass ionomer cement. Clinical and radiographic successes and failures were recorded at 6, 12, 18 and 24-month follow-up. All pulpotomised teeth were clinically and radiographically successful at all follow-up appointments. Six out of 15 teeth in the PC group and five out of 14 teeth in the MTA group exfoliated throughout the follow-up period. No statistically significant difference regarding dentine bridge formation was found between both groups throughout the follow-up period. As far as pulp canal obliteration is concerned, a statistically significant difference was detected at 6-month follow-up (p <0.05), since the beginning of mineralised material deposition could be radiographically detected in 100% and 57.14% of the teeth treated with PC and MTA, respectively. PC may serve as an effective and less expensive MTA substitute in primary molar pulpotomies. Further studies and longer follow-up assessments are needed.

  13. Global analysis of gene expression in the developing brain of Gtf2ird1 knockout mice.

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    Jennifer O'Leary

    Full Text Available Williams-Beuren Syndrome (WBS is a neurodevelopmental disorder caused by a hemizygous deletion of a 1.5 Mb region on chromosome 7q11.23 encompassing 26 genes. One of these genes, GTF2IRD1, codes for a putative transcription factor that is expressed throughout the brain during development. Genotype-phenotype studies in patients with atypical deletions of 7q11.23 implicate this gene in the neurological features of WBS, and Gtf2ird1 knockout mice show reduced innate fear and increased sociability, consistent with features of WBS. Multiple studies have identified in vitro target genes of GTF2IRD1, but we sought to identify in vivo targets in the mouse brain.We performed the first in vivo microarray screen for transcriptional targets of Gtf2ird1 in brain tissue from Gtf2ird1 knockout and wildtype mice at embryonic day 15.5 and at birth. Changes in gene expression in the mutant mice were moderate (0.5 to 2.5 fold and of candidate genes with altered expression verified using real-time PCR, most were located on chromosome 5, within 10 Mb of Gtf2ird1. siRNA knock-down of Gtf2ird1 in two mouse neuronal cell lines failed to identify changes in expression of any of the genes identified from the microarray and subsequent analysis showed that differences in expression of genes on chromosome 5 were the result of retention of that chromosome region from the targeted embryonic stem cell line, and so were dependent upon strain rather than Gtf2ird1 genotype. In addition, specific analysis of genes previously identified as direct in vitro targets of GTF2IRD1 failed to show altered expression.We have been unable to identify any in vivo neuronal targets of GTF2IRD1 through genome-wide expression analysis, despite widespread and robust expression of this protein in the developing rodent brain.

  14. Two potential hookworm DAF-16 target genes, SNR-3 and LPP-1: gene structure, expression profile, and implications of a cis-regulatory element in the regulation of gene expression.

    Science.gov (United States)

    Gao, Xin; Goggin, Kevin; Dowling, Camille; Qian, Jason; Hawdon, John M

    2015-01-08

    Hookworms infect nearly 700 million people, causing anemia and developmental stunting in heavy infections. Little is known about the genomic structure or gene regulation in hookworms, although recent publication of draft genome assemblies has allowed the first investigations of these topics to be undertaken. The transcription factor DAF-16 mediates multiple developmental pathways in the free living nematode Caenorhabditis elegans, and is involved in the recovery from the developmentally arrested L3 in hookworms. Identification of downstream targets of DAF-16 will provide a better understanding of the molecular mechanism of hookworm infection. Genomic Fragment 2.23 containing a DAF-16 binding element (DBE) was used to identify overlapping complementary expressed sequence tags (ESTs). These sequences were used to search a draft assembly of the Ancylostoma caninum genome, and identified two neighboring genes, snr-3 and lpp-1, in a tail-to-tail orientation. Expression patterns of both genes during parasitic development were determined by qRT-PCR. DAF-16 dependent cis-regulatory activity of fragment 2.23 was investigated using an in vitro reporter system. The snr-3 gene spans approximately 5.6 kb in the genome and contains 3 exons and 2 introns, and contains the DBE in its 3' untranslated region. Downstream from snr-3 in a tail-to-tail arrangement is the gene lpp-1. The lpp-1 gene spans more than 6 kb and contains 10 exons and 9 introns. The A. caninum genome contains 2 apparent splice variants, but there are 7 splice variants in the A. ceylanicum genome. While the gene order is similar, the gene structures of the hookworm genes differ from their C. elegans orthologs. Both genes show peak expression in the late L4 stage. Using a cell culture based expression system, fragment 2.23 was found to have both DAF-16-dependent promoter and enhancer activity that required an intact DBE. Two putative DAF-16 targets were identified by genome wide screening for DAF-16 binding

  15. Night-time restricted feeding normalises clock genes and Pai-1 gene expression in the db/db mouse liver.

    Science.gov (United States)

    Kudo, T; Akiyama, M; Kuriyama, K; Sudo, M; Moriya, T; Shibata, S

    2004-08-01

    An increase in PAI-1 activity is thought to be a key factor underlying myocardial infarction. Mouse Pai-1 (mPai-1) activity shows a daily rhythm in vivo, and its transcription seems to be controlled not only by clock genes but also by humoral factors such as insulin and triglycerides. Thus, we investigated daily clock genes and mPai-1 mRNA expression in the liver of db/db mice exhibiting high levels of glucose, insulin and triglycerides. Locomotor activity was measured using an infrared detection system. RT-PCR or in situ hybridisation methods were applied to measure gene expression. Humoral factors were measured using measurement kits. The db/ db mice showed attenuated locomotor activity rhythms. The rhythmic expression of mPer2 mRNA was severely diminished and the phase of mBmal1 oscillation was advanced in the db/db mouse liver, whereas mPai-1 mRNA was highly and constitutively expressed. Night-time restricted feeding led to a recovery not only from the diminished locomotor activity, but also from the diminished Per2 and advanced mBmal1 mRNA rhythms. Expression of mPai-1 mRNA in db/db mice was reduced to levels far below normal. Pioglitazone treatment slightly normalised glucose and insulin levels, with a slight reduction in mPai-1 gene expression. We demonstrated that Type 2 diabetes impairs the oscillation of the peripheral oscillator. Night-time restricted feeding rather than pioglitazone injection led to a recovery from the diminished locomotor activity, and altered oscillation of the peripheral clock and mPai-1 mRNA rhythm. Thus, we conclude that scheduled restricted food intake may be a useful form of treatment for diabetes.

  16. The expression and activity of thioredoxin reductase 1 splice variants v1 and v2 regulate the expression of genes associated with differentiation and adhesion

    Science.gov (United States)

    Nalvarte, Ivan; Damdimopoulos, Anastasios E.; Rüegg, Joëlle; Spyrou, Giannis

    2015-01-01

    The mammalian redox-active selenoprotein thioredoxin reductase (TrxR1) is a main player in redox homoeostasis. It transfers electrons from NADPH to a large variety of substrates, particularly to those containing redox-active cysteines. Previously, we reported that the classical form of cytosolic TrxR1 (TXNRD1_v1), when overexpressed in human embryonic kidney cells (HEK-293), prompted the cells to undergo differentiation [Nalvarte et al. (2004) J. Biol. Chem. 279, 54510–54517]. In the present study, we show that several genes associated with differentiation and adhesion are differentially expressed in HEK-293 cells stably overexpressing TXNRD1_v1 compared with cells expressing its splice variant TXNRD1_v2. Overexpression of these two splice forms resulted in distinctive effects on various aspects of cellular functions including gene regulation patterns, alteration of growth rate, migration and morphology and susceptibility to selenium-induced toxicity. Furthermore, differentiation of the neuroblastoma cell line SH-SY5Y induced by all-trans retinoic acid (ATRA) increased both TXNRD1_v1 and TXNRD1_v2 expressions along with several of the identified genes associated with differentiation and adhesion. Selenium supplementation in the SH-SY5Y cells also induced a differentiated morphology and changed expression of the adhesion protein fibronectin 1 and the differentiation marker cadherin 11, as well as different temporal expression of the studied TXNRD1 variants. These data suggest that both TXNRD1_v1 and TXNRD1_v2 have distinct roles in differentiation, possibly by altering the expression of the genes associated with differentiation, and further emphasize the importance in distinguishing each unique action of different TrxR1 splice forms, especially when studying the gene silencing or knockout of TrxR1. PMID:26464515

  17. Highly expressed genes within hippocampal sector CA1: implications for the physiology of memory

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    Michael A. Meyer

    2014-06-01

    Full Text Available As the CA1 sector has been implicated to play a key role in memory formation, a dedicated search for highly expressed genes within this region was made from an on-line atlas of gene expression within the mouse brain (GENSAT. From a data base of 1013 genes, 16 were identified that had selective localization of gene expression within the CA1 region, and included Angpt2, ARHGEF6, CCK, Cntnap1, DRD3, EMP1, Epha2, Itm2b, Lrrtm2, Mdk, PNMT, Ppm1e, Ppp2r2d, RASGRP1, Slitrk5, and Sstr4. Of the 16 identified, the most selective and intense localization for both adult and post-natal day 7 was noted for ARHGEF6, which is known to be linked to non-syndromic mental retardation, and has also been localized to dendritic spines. Further research on the role played by ARHGEF6 in memory formation is strongly advocated.

  18. Highly Expressed Genes within Hippocampal Sector CA1: Implications for the Physiology of Memory.

    Science.gov (United States)

    Meyer, Michael A

    2014-04-22

    As the CA1 sector has been implicated to play a key role in memory formation, a dedicated search for highly expressed genes within this region was made from an on-line atlas of gene expression within the mouse brain (GENSAT). From a data base of 1013 genes, 16 were identified that had selective localization of gene expression within the CA1 region, and included Angpt2, ARHGEF6, CCK, Cntnap1, DRD3, EMP1, Epha2, Itm2b, Lrrtm2, Mdk, PNMT, Ppm1e, Ppp2r2d, RASGRP1, Slitrk5, and Sstr4. Of the 16 identified, the most selective and intense localization for both adult and post-natal day 7 was noted for ARHGEF6, which is known to be linked to non-syndromic mental retardation, and has also been localized to dendritic spines. Further research on the role played by ARHGEF6 in memory formation is strongly advocated.

  19. The cauliflower Orange gene enhances petiole elongation by suppressing expression of eukaryotic release factor 1.

    Science.gov (United States)

    Zhou, Xiangjun; Sun, Tian-Hu; Wang, Ning; Ling, Hong-Qing; Lu, Shan; Li, Li

    2011-04-01

    The cauliflower (Brassica oleracea var. botrytis) Orange (Or) gene affects plant growth and development in addition to conferring β-carotene accumulation. This study was undertaken to investigate the molecular basis for the effects of the Or gene mutation in on plant growth. The OR protein was found to interact with cauliflower and Arabidopsis eukaryotic release factor 1-2 (eRF1-2), a member of the eRF1 family, by yeast two-hybrid analysis and by bimolecular fluorescence complementation (BiFC) assay. Concomitantly, the Or mutant showed reduced expression of the BoeRF1 family genes. Transgenic cauliflower plants with suppressed expression of BoeRF1-2 and BoeRF1-3 were generated by RNA interference. Like the Or mutant, the BoeRF1 RNAi lines showed increased elongation of the leaf petiole. This long-petiole phenotype was largely caused by enhanced cell elongation, which resulted from increased cell length and elevated expression of genes involved in cell-wall loosening. These findings demonstrate that the cauliflower Or gene controls petiole elongation by suppressing the expression of eRF1 genes, and provide new insights into the molecular mechanism of leaf petiole regulation. © 2010 The Authors. New Phytologist © 2010 New Phytologist Trust.

  20. Regulation of a transcription factor network by Cdk1 coordinates late cell cycle gene expression.

    Science.gov (United States)

    Landry, Benjamin D; Mapa, Claudine E; Arsenault, Heather E; Poti, Kristin E; Benanti, Jennifer A

    2014-05-02

    To maintain genome stability, regulators of chromosome segregation must be expressed in coordination with mitotic events. Expression of these late cell cycle genes is regulated by cyclin-dependent kinase (Cdk1), which phosphorylates a network of conserved transcription factors (TFs). However, the effects of Cdk1 phosphorylation on many key TFs are not known. We find that elimination of Cdk1-mediated phosphorylation of four S-phase TFs decreases expression of many late cell cycle genes, delays mitotic progression, and reduces fitness in budding yeast. Blocking phosphorylation impairs degradation of all four TFs. Consequently, phosphorylation-deficient mutants of the repressors Yox1 and Yhp1 exhibit increased promoter occupancy and decreased expression of their target genes. Interestingly, although phosphorylation of the transcriptional activator Hcm1 on its N-terminus promotes its degradation, phosphorylation on its C-terminus is required for its activity, indicating that Cdk1 both activates and inhibits a single TF. We conclude that Cdk1 promotes gene expression by both activating transcriptional activators and inactivating transcriptional repressors. Furthermore, our data suggest that coordinated regulation of the TF network by Cdk1 is necessary for faithful cell division.

  1. Characterization of differential gene expression in adrenocortical tumors harboring beta-catenin (CTNNB1) mutations.

    Science.gov (United States)

    Durand, Julien; Lampron, Antoine; Mazzuco, Tania L; Chapman, Audrey; Bourdeau, Isabelle

    2011-07-01

    Mutations of β-catenin gene (CTNNB1) are frequent in adrenocortical adenomas (AA) and adrenocortical carcinomas (ACC). However, the target genes of β-catenin have not yet been identified in adrenocortical tumors. Our objective was to identify genes deregulated in adrenocortical tumors harboring CTNNB1 genetic alterations and nuclear accumulation of β-catenin. Microarray analysis identified a dataset of genes that were differently expressed between AA with CTNNB1 mutations and wild-type (WT) tumors. Within this dataset, the expression profiles of five genes were validated by real time-PCR (RT-PCR) in a cohort of 34 adrenocortical tissues (six AA and one ACC with CTNNB1 mutations, 13 AA and four ACC with WT CTNNB1, and 10 normal adrenal glands) and two human ACC cell lines. We then studied the effects of suppressing β-catenin transcriptional activity with the T-cell factor/β-catenin inhibitors PKF115-584 and PNU74654 on gene expression in H295R and SW13 cells. RT-PCR analysis confirmed the overexpression of ISM1, RALBP1, and PDE2A and the down-regulation of PHYHIP in five of six AA harboring CTNNB1 mutations compared with WT AA (n = 13) and normal adrenal glands (n = 10). RALBP1 and PDE2A overexpression was also confirmed at the protein level by Western blotting analysis in mutated tumors. ENC1 was specifically overexpressed in three of three AA harboring CTNNB1 point mutations. mRNA expression and protein levels of RALBP1, PDE2A, and ENC1 were decreased in a dose-dependent manner in H295R cells after treatment with PKF115-584 or PNU74654. This study identified candidate genes deregulated in CTNNB1-mutated adrenocortical tumors that may lead to a better understanding of the role of the Wnt-β-catenin pathway in adrenocortical tumorigenesis.

  2. Selection and validation of reference genes for quantitative gene expression studies in Erythroxylum coca [v1; ref status: indexed, http://f1000r.es/y1

    Directory of Open Access Journals (Sweden)

    Teresa Docimo

    2013-02-01

    Full Text Available Real-time quantitative PCR is a powerful technique for the investigation of comparative gene expression, but its accuracy and reliability depend on the reference genes used as internal standards. Only genes that show a high level of expression stability are suitable for use as reference genes, and these must be identified on a case-by-case basis. Erythroxylum coca produces and accumulates high amounts of the pharmacologically active tropane alkaloid cocaine (especially in the leaves, and is an emerging model for the investigation of tropane alkaloid biosynthesis. The identification of stable internal reference genes for this species is important for its development as a model species, and would enable comparative analysis of candidate biosynthetic genes in the different tissues of the coca plant. In this study, we evaluated the expression stability of nine candidate reference genes in E. coca (Ec6409, Ec10131, Ec11142, Actin, APT2, EF1α, TPB1, Pex4, Pp2aa3. The expression of these genes was measured in seven tissues (flowers, stems, roots and four developmental leaf stages and the stability of expression was assessed using three algorithms (geNorm, NormFinder and BestKeeper. From our results we conclude that Ec10131 and TPB1 are the most appropriate internal reference genes in leaves (where the majority of cocaine is produced, while Ec10131 and Ec6409 are the most suitable internal reference genes across all of the tissues tested.

  3. Analysis of metal contents in Portland Type V and MTA-based cements.

    Science.gov (United States)

    Dorileo, Maura Cristiane Gonçales Orçati; Bandeca, Matheus Coelho; Pedro, Fábio Luis Miranda; Volpato, Luiz Evaristo Ricci; Guedes, Orlando Aguirre; Dalla Villa, Ricardo; Tonetto, Mateus Rodrigues; Borges, Alvaro Henrique

    2014-01-01

    The aim of this study was to determine, by Atomic Absorption Spectrometry (AAS), the concentration levels of 11 metals in Type V gray and structural white PC, ProRoot MTA, and MTA Bio. Samples, containing one gram of each tested cement, were prepared and transferred to a 100 mL Teflon tube with a mixture of 7.0 mL of nitric acid and 21 mL of hydrochloric acid. After the reaction, the mixture was filtered and then volumed to 50 mL of distilled water. For each metal, specific patterns were determined from universal standards. Arsenic quantification was performed by hydride generator. The analysis was performed five times and the data were statistically analyzed at 5% level of significance. Only the cadmium presented concentration levels of values lower than the quantification limit of the device. The AAS analysis showed increased levels of calcium, nickel, and zinc in structural white PC. Type V PC presented the greatest concentration levels of arsenic, chromium, copper, iron, lead, and manganese (P cements, and the lowest concentration levels were observed in Portland cements, while the highest were observed in ProRoot MTA. Both PC and MTA-based cements showed evidence of metals inclusion.

  4. Effect of MLH1 -93G>A on gene expression in patients with colorectal cancer.

    Science.gov (United States)

    Funck, Alexandre; Santos, Juliana C; Silva-Fernandes, Isabelle J L; Rabenhorst, Silvia H B; Martinez, Carlos A R; Ribeiro, Marcelo L

    2014-09-01

    The DNA repair machinery plays a key role in maintaining genomic stability by preventing the emergence of mutations. Furthermore, the -93G>A polymorphism in the MLH1 gene has been associated with an increased risk of developing colorectal cancer. Therefore, the aim of this study was to examine the expression pattern and effect of this polymorphism in normal and tumour samples from patients with colorectal cancer. The MLH1 -93G>A (rs1800734) polymorphism was detected by PCR-RFLP in 49 cases of colorectal cancer. MLH1 expression was investigated using real-time quantitative PCR. The results indicate a significant decrease in MLH1 expression in tumour samples compared to their normal counterparts. The MLH1 gene was also significantly repressed in samples from patients who had some degree of tumour invasion into other organs. Similarly, those patients who were in a more advanced tumour stage (TNM III and IV) exhibited a significant reduction in MLH1 gene expression. Finally, the mutant genotype AA of MLH1 was associated with a significant decrease in the expression of this gene. This finding suggests that this polymorphism could increase the risk of developing colorectal cancer by a defective mismatch repair system, particularly through the loss of MLH1 expression in an allele-specific manner.

  5. Changes in oil content of transgenic soybeans expressing the yeast SLC1 gene.

    Science.gov (United States)

    Rao, Suryadevara S; Hildebrand, David

    2009-10-01

    The wild type (Wt) and mutant form of yeast (sphingolipid compensation) genes, SLC1 and SLC1-1, have been shown to have lysophosphatidic acid acyltransferase (LPAT) activities (Nageic et al. in J Biol Chem 269:22156-22163, 1993). Expression of these LPAT genes was reported to increase oil content in transgenic Arabidopsis and Brassica napus. It is of interest to determine if the TAG content increase would also be seen in soybeans. Therefore, the wild type SLC1 was expressed in soybean somatic embryos under the control of seed specific phaseolin promoter. Some transgenic somatic embryos and in both T2 and T3 transgenic seeds showed higher oil contents. Compared to controls, the average increase in triglyceride values went up by 1.5% in transgenic somatic embryos. A maximum of 3.2% increase in seed oil content was observed in a T3 line. Expression of the yeast Wt LPAT gene did not alter the fatty acid composition of the seed oil.

  6. Transcriptional coactivator NT-PGC-1α promotes gluconeogenic gene expression and enhances hepatic gluconeogenesis.

    Science.gov (United States)

    Chang, Ji Suk; Jun, Hee-Jin; Park, Minsung

    2016-10-01

    The transcriptional coactivator PGC-1α plays a central role in hepatic gluconeogenesis. We previously reported that alternative splicing of the PGC-1α gene produces an additional transcript encoding the truncated protein NT-PGC-1α NT-PGC-1α is co-expressed with PGC-1α and highly induced by fasting in the liver. NT-PGC-1α regulates tissue-specific metabolism, but its role in the liver has not been investigated. Thus, the objective of this study was to determine the role of hepatic NT-PGC-1α in the regulation of gluconeogenesis. Adenovirus-mediated expression of NT-PGC-1α in primary hepatocytes strongly stimulated the expression of key gluconeogenic enzyme genes (PEPCK and G6Pase), leading to increased glucose production. To further understand NT-PGC-1α function in hepatic gluconeogenesis in vivo, we took advantage of a previously reported FL-PGC-1α -/- mouse line that lacks full-length PGC-1α (FL-PGC-1α) but retains a slightly shorter and functionally equivalent form of NT-PGC-1α (NT-PGC-1α 254 ). In FL-PGC-1α -/- mice, NT-PGC-1α 254 was induced by fasting in the liver and recruited to the promoters of PEPCK and G6Pase genes. The enrichment of NT-PGC-1α 254 at the promoters was closely associated with fasting-induced increase in PEPCK and G6Pase gene expression and efficient production of glucose from pyruvate during a pyruvate tolerance test in FL-PGC-1α -/- mice. Moreover, FL-PGC-1α -/- primary hepatocytes showed a significant increase in gluconeogenic gene expression and glucose production after treatment with dexamethasone and forskolin, suggesting that NT-PGC-1α 254 is sufficient to stimulate the gluconeogenic program in the absence of FL-PGC-1α Collectively, our findings highlight the role of hepatic NT-PGC-1α in stimulating gluconeogenic gene expression and glucose production. © 2016 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

  7. Neurospora crassa glucose - repressible gene -1(Grg-1) promoter controls the expression of neurospora tyrosinase gene in a clock-controlled manner

    International Nuclear Information System (INIS)

    Tarawneh, A. K

    1997-01-01

    In this study sphareroplastes of white Neurospora crassa mutant auxotroph for aromatic am no acids a rom 9 q a-2 inv, was transformed by the pKF-Tyr7-wt DNA construct. This construct contains the promoter of neurospora crassa glucose-repressible gene-1 (G rg-1) usp stream of Neurospora tyrosinase gene. The co transformation of this mutant with pKF-Tyr-7-wt cincture's and the pKAL-1, a plasmid which contains the Neurospora q a-2+ gene transform it to photophor. The transform ant contains the tyrosinase gene which catalyzes the unique step in the synthesis of the black pigment melanin. The activity of the tyrosinase in this transform ant was followed by measuring the absorbance of the dark coloured pigment at 332 nm. The maximum of the tyrosinase activity was shown at 16.36 and 56 hours after the shift of the transformed mycelia from constant light (L L) to constant dark (Dd). The rate of the enzyme activity was changed according to ci radian cycle of 20 hours. This G rg 1/tyrosinase construct provides a good system to study to study the temporal control of gene expression and the interaction between the different environmental c uses that affects gene expression. (author). 20 refs., 4 figs

  8. Production of transgenic pigs over-expressing the antiviral gene Mx1

    Directory of Open Access Journals (Sweden)

    Quanmei Yan

    2014-01-01

    Full Text Available The myxovirus resistance gene (Mx1 has a broad spectrum of antiviral activities. It is therefore an interesting candidate gene to improve disease resistance in farm animals. In this study, we report the use of somatic cell nuclear transfer (SCNT to produce transgenic pigs over-expressing the Mx1 gene. These transgenic pigs express approximately 15–25 times more Mx1 mRNA than non-transgenic pigs, and the protein level of Mx1 was also markedly enhanced. We challenged fibroblast cells isolated from the ear skin of transgenic and control pigs with influenza A virus and classical swine fever virus (CFSV. Indirect immunofluorescence assay (IFA revealed a profound decrease of influenza A proliferation in Mx1 transgenic cells. Growth kinetics showed an approximately 10-fold reduction of viral copies in the transgenic cells compared to non-transgenic controls. Additionally, we found that the Mx1 transgenic cells were more resistant to CSFV infection in comparison to non-transgenic cells. These results demonstrate that the Mx1 transgene can protect against viral infection in cells of transgenic pigs and indicate that the Mx1 transgene can be harnessed to develop disease-resistant pigs.

  9. Mucosal CCR1 gene expression as a marker of molecular activity in Crohn's disease: preliminary data.

    Science.gov (United States)

    Dobre, Maria; Mănuc, Teodora Ecaterina; Milanesi, Elena; Pleşea, Iancu Emil; Ţieranu, Eugen Nicolae; Popa, Caterina; Mănuc, Mircea; Preda, Carmen Monica; Ţieranu, Ioana; Diculescu, Mihai Mircea; Ionescu, Elena Mirela; Becheanu, Gabriel

    2017-01-01

    A series of mechanisms of immune response, inflammation and apoptosis have been demonstrated to contribute to the appearance and evolution of Crohn's disease (CD) through the overexpression of several cytokines and chemokines in a susceptible host. The aim of this study was to identify the differences in gene expression profiles analyzing a panel of candidate genes in the mucosa from patients with active CD (CD-A), patients in remission (CD-R), and normal controls. Nine individuals were enrolled in the study: six CD patients (three with active lesions, three with mucosal healing) and three controls without inflammatory bowel disease (IBD) seen on endoscopy. All the individuals underwent mucosal biopsy during colonoscopy. Gene expression levels of 84 genes previously associated with CD were evaluated by polymerase chain reaction (PCR) array. Ten genes out of 84 were found significantly differentially expressed in CD-A (CCL11, CCL25, DEFA5, GCG, IL17A, LCN2, REG1A, STAT3, MUC1, CCR1) and eight genes in CD-R (CASP1, IL23A, STAT1, STAT3, TNF, CCR1, CCL5, and HSP90B1) when compared to controls. A quantitative gene expression analysis revealed that CCR1 gene was more expressed in CD-A than in CD-R. Our data suggest that CCR1 gene may be a putative marker of molecular activity of Crohn's disease. Following these preliminary data, a confirmation in larger cohort studies could represent a useful method in order to identify new therapeutic targets.

  10. Distinct Calcium Signaling Pathways Regulate Calmodulin Gene Expression in Tobacco1

    Science.gov (United States)

    van der Luit, Arnold H.; Olivari, Claudio; Haley, Ann; Knight, Marc R.; Trewavas, Anthony J.

    1999-01-01

    Cold shock and wind stimuli initiate Ca2+ transients in transgenic tobacco (Nicotiana plumbaginifolia) seedlings (named MAQ 2.4) containing cytoplasmic aequorin. To investigate whether these stimuli initiate Ca2+ pathways that are spatially distinct, stress-induced nuclear and cytoplasmic Ca2+ transients and the expression of a stress-induced calmodulin gene were compared. Tobacco seedlings were transformed with a construct that encodes a fusion protein between nucleoplasmin (a major oocyte nuclear protein) and aequorin. Immunocytochemical evidence indicated targeting of the fusion protein to the nucleus in these plants, which were named MAQ 7.11. Comparison between MAQ 7.11 and MAQ 2.4 seedlings confirmed that wind stimuli and cold shock invoke separate Ca2+ signaling pathways. Partial cDNAs encoding two tobacco calmodulin genes, NpCaM-1 and NpCaM-2, were identified and shown to have distinct nucleotide sequences that encode identical polypeptides. Expression of NpCaM-1, but not NpCaM-2, responded to wind and cold shock stimulation. Comparison of the Ca2+ dynamics with NpCaM-1 expression after stimulation suggested that wind-induced NpCaM-1 expression is regulated by a Ca2+ signaling pathway operational predominantly in the nucleus. In contrast, expression of NpCaM-1 in response to cold shock is regulated by a pathway operational predominantly in the cytoplasm. PMID:10557218

  11. Gene expression

    International Nuclear Information System (INIS)

    Hildebrand, C.E.; Crawford, B.D.; Walters, R.A.; Enger, M.D.

    1983-01-01

    We prepared probes for isolating functional pieces of the metallothionein locus. The probes enabled a variety of experiments, eventually revealing two mechanisms for metallothionein gene expression, the order of the DNA coding units at the locus, and the location of the gene site in its chromosome. Once the switch regulating metallothionein synthesis was located, it could be joined by recombinant DNA methods to other, unrelated genes, then reintroduced into cells by gene-transfer techniques. The expression of these recombinant genes could then be induced by exposing the cells to Zn 2+ or Cd 2+ . We would thus take advantage of the clearly defined switching properties of the metallothionein gene to manipulate the expression of other, perhaps normally constitutive, genes. Already, despite an incomplete understanding of how the regulatory switch of the metallothionein locus operates, such experiments have been performed successfully

  12. Digital gene expression analysis of gene expression differences within Brassica diploids and allopolyploids.

    Science.gov (United States)

    Jiang, Jinjin; Wang, Yue; Zhu, Bao; Fang, Tingting; Fang, Yujie; Wang, Youping

    2015-01-27

    Brassica includes many successfully cultivated crop species of polyploid origin, either by ancestral genome triplication or by hybridization between two diploid progenitors, displaying complex repetitive sequences and transposons. The U's triangle, which consists of three diploids and three amphidiploids, is optimal for the analysis of complicated genomes after polyploidization. Next-generation sequencing enables the transcriptome profiling of polyploids on a global scale. We examined the gene expression patterns of three diploids (Brassica rapa, B. nigra, and B. oleracea) and three amphidiploids (B. napus, B. juncea, and B. carinata) via digital gene expression analysis. In total, the libraries generated between 5.7 and 6.1 million raw reads, and the clean tags of each library were mapped to 18547-21995 genes of B. rapa genome. The unambiguous tag-mapped genes in the libraries were compared. Moreover, the majority of differentially expressed genes (DEGs) were explored among diploids as well as between diploids and amphidiploids. Gene ontological analysis was performed to functionally categorize these DEGs into different classes. The Kyoto Encyclopedia of Genes and Genomes analysis was performed to assign these DEGs into approximately 120 pathways, among which the metabolic pathway, biosynthesis of secondary metabolites, and peroxisomal pathway were enriched. The non-additive genes in Brassica amphidiploids were analyzed, and the results indicated that orthologous genes in polyploids are frequently expressed in a non-additive pattern. Methyltransferase genes showed differential expression pattern in Brassica species. Our results provided an understanding of the transcriptome complexity of natural Brassica species. The gene expression changes in diploids and allopolyploids may help elucidate the morphological and physiological differences among Brassica species.

  13. MSX-1 gene expression and regulation in embryonic palatal tissue.

    Science.gov (United States)

    Nugent, P; Greene, R M

    1998-01-01

    The palatal cleft seen in Msx-1 knock-out mice suggests a role for this gene in normal palate development. The cleft is presumed secondary to tooth and jaw malformations, since in situ hybridization suggests that Msx-1 mRNA is not highly expressed in developing palatal tissue. In this study we demonstrate, by Northern blot analysis, the expression of Msx-1, but not Msx-2, in the developing palate and in primary cultures of murine embryonic palate mesenchymal cells. Furthermore, we propose a role for Msx-1 in retinoic acid-induced cleft palate, since retinoic acid inhibits Msx-1 mRNA expression in palate mesenchymal cells. We also demonstrate that transforming growth factor beta inhibits Msx-1 mRNA expression in palate mesenchymal cells, with retinoic acid and transforming growth factor beta acting synergistically when added simultaneously to these cells. These data suggest a mechanistic interaction between retinoic acid, transforming growth factor beta, and Msx-1 in the etiology of retinoic acid-induced cleft palate.

  14. Rat subcutaneous tissue response to MTA Fillapex® and Portland cement.

    Science.gov (United States)

    Marques, Nádia Carolina Teixeira; Lourenço Neto, Natalino; Fernandes, Ana Paula; Rodini, Camila de Oliveira; Duarte, Marco Antônio Hungaro; Oliveira, Thais Marchini

    2013-01-01

    The aim of this study was to evaluate the response of rat subcutaneous tissue to MTA Fillapex® (Angelus), an experimental root canal filling material based on Portland cement and propylene glycol (PCPG), and a zinc oxide, eugenol and iodoform (ZOEI) paste. These materials were placed in polyethylene tubes and implanted into the dorsal connective tissue of Wistar rats for 7 and 15 days. The specimens were stained with hematoxylin and eosin, and evaluated regarding inflammatory reaction parameters by optical microscopy. The intensity of inflammatory response against the sealers was analyzed by two blinded and previously calibrated examiners for all experimental periods (kappa=0.96). The histological evaluation showed that all materials caused a moderate inflammatory reaction at 7 days, which subsided with time. A greater inflammatory reaction was observed at 7 days in the tubes filled with ZOEI paste. Tubes filled with MTA Fillapex presented some giant cells, macrophages and lymphocytes after 7 days. At 15 days, the presence of fibroblasts and collagen fibers was observed indicating normal tissue healing. The tubes filled with PCPG showed similar results to those observed in MTA Fillapex. At 15 days, the inflammatory reaction was almost absent at the tissue, with several collagen fibers indicating normal tissue healing. Data were analyzed by the nonparametric Kruskal-Wallis test (α=0.05). Statistically significant difference (p0.05). MTA Fillapex and Portland cement added with propylene glycol had greater tissue compatibility than the PCPG paste.

  15. Ex vivo assessment of genotoxicity and cytotoxicity in murine fibroblasts exposed to white MTA or white Portland cement with 15% bismuth oxide.

    Science.gov (United States)

    Zeferino, E G; Bueno, C E S; Oyama, L M; Ribeiro, D A

    2010-10-01

    To evaluate whether white mineral trioxide aggregate (MTA) or white Portland cement with 15% bismuth oxide were able to induce genetic damage and cellular death ex vivo. Aliquots of 1 × 10(4) murine fibroblasts were incubated at 37 °C for 3 h with MTA (white) or white Portland cement with 15% bismuth oxide, at final concentrations ranging from 10 to 1000 μg mL(-1) individually. Data of three independent repeats from the comet assay and the trypan blue exclusion test were assessed by the one-way anova followed by Tukey's test. Mineral trioxide aggregate or Portland cement containing bismuth oxide did not produce genotoxic effects with respect to the single-cell gel (comet) assay data for all concentrations evaluated. Furthermore, no cytotoxicity was observed for MTA or Portland cement. White MTA or white Portland cement containing 15% bismuth oxide were not genotoxic and cytotoxic. © 2010 International Endodontic Journal.

  16. Role of resveratrol in FOXO1-mediated gluconeogenic gene expression in the liver

    Energy Technology Data Exchange (ETDEWEB)

    Park, Joo-Man; Kim, Tae-Hyun [Department of Biochemistry and Molecular Biology, Center for Chronic Metabolic Disease Research, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Brain Korea 21 Project for Medical Sciences, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Bae, Jin-Sik; Kim, Mi-Young [Department of Biochemistry and Molecular Biology, Center for Chronic Metabolic Disease Research, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Kim, Kyung-Sup [Department of Biochemistry and Molecular Biology, Center for Chronic Metabolic Disease Research, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Brain Korea 21 Project for Medical Sciences, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); The Institute of Genetic Science, Yonsei University College of Medicine, 134 Shinchon-dong, Seodaemoon-gu, Seoul 120-752 (Korea, Republic of); Ahn, Yong-Ho, E-mail: yha111@yuhs.ac [Department of Biochemistry and Molecular Biology, Center for Chronic Metabolic Disease Research, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Brain Korea 21 Project for Medical Sciences, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of)

    2010-12-17

    Research highlights: {yields} Insulin-suppression of PEPCK and G6Pase gene expression is counteracted by resveratrol. {yields} Resveratrol upregulates hepatic gluconeogenic genes by attenuating insulin signaling and deacetylating FOXO1, which are SIRT1-independent in the cytosol and SIRT1-dependent in the nucleus, respectively. {yields} Resveratrol increases the binding activity of Foxo1 to the IRE of PEPCK and G6Pase. -- Abstract: During a state of fasting, the blood glucose level is maintained by hepatic gluconeogenesis. SIRT1 is an important metabolic regulator during nutrient deprivation and the liver-specific knockdown of SIRT1 resulted in decreased glucose production. We hypothesize that SIRT1 is responsible for the upregulation of insulin-suppressed gluconeogenic genes through the deacetylation of FOXO1. Treatment of primary cultured hepatocytes with resveratrol increased insulin-repressed PEPCK and G6Pase mRNA levels, which depend on SIRT1 activity. We found that the resveratrol treatment resulted in a decrease in the phosphorylation of Akt and FOXO1, which are independent of SIRT1 action. Fluorescence microscopy revealed that resveratrol caused the nuclear localization of FOXO1. In the nucleus, FOXO1 is deacetylated by SIRT1, which might make it more accessible to the IRE of the PEPCK and G6Pase promoter, causing an increase in their gene expression. Our results indicate that resveratrol upregulates the expression of gluconeogenic genes by attenuating insulin signaling and by deacetylating FOXO1, which are SIRT1-independent in the cytosol and SIRT1-dependent in the nucleus, respectively.

  17. Role of resveratrol in FOXO1-mediated gluconeogenic gene expression in the liver

    International Nuclear Information System (INIS)

    Park, Joo-Man; Kim, Tae-Hyun; Bae, Jin-Sik; Kim, Mi-Young; Kim, Kyung-Sup; Ahn, Yong-Ho

    2010-01-01

    Research highlights: → Insulin-suppression of PEPCK and G6Pase gene expression is counteracted by resveratrol. → Resveratrol upregulates hepatic gluconeogenic genes by attenuating insulin signaling and deacetylating FOXO1, which are SIRT1-independent in the cytosol and SIRT1-dependent in the nucleus, respectively. → Resveratrol increases the binding activity of Foxo1 to the IRE of PEPCK and G6Pase. -- Abstract: During a state of fasting, the blood glucose level is maintained by hepatic gluconeogenesis. SIRT1 is an important metabolic regulator during nutrient deprivation and the liver-specific knockdown of SIRT1 resulted in decreased glucose production. We hypothesize that SIRT1 is responsible for the upregulation of insulin-suppressed gluconeogenic genes through the deacetylation of FOXO1. Treatment of primary cultured hepatocytes with resveratrol increased insulin-repressed PEPCK and G6Pase mRNA levels, which depend on SIRT1 activity. We found that the resveratrol treatment resulted in a decrease in the phosphorylation of Akt and FOXO1, which are independent of SIRT1 action. Fluorescence microscopy revealed that resveratrol caused the nuclear localization of FOXO1. In the nucleus, FOXO1 is deacetylated by SIRT1, which might make it more accessible to the IRE of the PEPCK and G6Pase promoter, causing an increase in their gene expression. Our results indicate that resveratrol upregulates the expression of gluconeogenic genes by attenuating insulin signaling and by deacetylating FOXO1, which are SIRT1-independent in the cytosol and SIRT1-dependent in the nucleus, respectively.

  18. Oocyte-specific gene Oog1 suppresses the expression of spermatogenesis-specific genes in oocytes.

    Science.gov (United States)

    Honda, Shinnosuke; Miki, Yuka; Miyamoto, Yuya; Kawahara, Yu; Tsukamoto, Satoshi; Imai, Hiroshi; Minami, Naojiro

    2018-05-03

    Oog1, an oocyte-specific gene that encodes a protein of 425 amino acids, is present in five copies on mouse chromosomes 4 and 12. In mouse oocytes, Oog1 mRNA expression begins at embryonic day 15.5 and almost disappears by the late two-cell stage. Meanwhile, OOG1 protein is detectable in oocytes in ovarian cysts and disappears by the four-cell stage; the protein is transported to the nucleus in late one-cell to early two-cell stage embryos. In this study, we examined the role of Oog1 during oogenesis in mice. Oog1 RNAi-transgenic mice were generated by expressing double-stranded hairpin Oog1 RNA, which is processed into siRNAs targeting Oog1 mRNA. Quantitative RT-PCR revealed that the amount of Oog1 mRNA was dramatically reduced in oocytes obtained from Oog1-knockdown mice, whereas the abundance of spermatogenesis-associated transcripts (Klhl10, Tekt2, Tdrd6, and Tnp2) was increased in Oog1 knockdown ovaries. Tdrd6 is involved in the formation of the chromatoid body, Tnp2 contributes to the formation of sperm heads, Tekt2 is required for the formation of ciliary and flagellar microtubules, and Klhl10 plays a key role in the elongated sperm differentiation. These results indicate that Oog1 down-regulates the expression of spermatogenesis-associated genes in female germ cells, allowing them to develop normally into oocytes.

  19. Imaging Herpes Simplex Virus Type 1 Amplicon Vector–Mediated Gene Expression in Human Glioma Spheroids

    Directory of Open Access Journals (Sweden)

    Christine Kaestle

    2011-05-01

    Full Text Available Vectors derived from herpes simplex virus type 1 (HSV-1 have great potential for transducing therapeutic genes into the central nervous system; however, inefficient distribution of vector particles in vivo may limit their therapeutic potential in patients with gliomas. This study was performed to investigate the extent of HSV-1 amplicon vector–mediated gene expression in a three-dimensional glioma model of multicellular spheroids by imaging highly infectious HSV-1 virions expressing green fluorescent protein (HSV-GFP. After infection or microscopy-guided vector injection of glioma spheroids at various spheroid sizes, injection pressures and injection times, the extent of HSV-1 vector–mediated gene expression was investigated via laser scanning microscopy. Infection of spheroids with HSV-GFP demonstrated a maximal depth of vector-mediated GFP expression at 70 to 80 μm. A > 80% transduction efficiency was reached only in small spheroids with a diameter of 90%. The results demonstrated that vector-mediated gene expression in glioma spheroids was strongly dependent on the mode of vector application—injection pressure and injection time being the most important parameters. The assessment of these vector application parameters in tissue models will contribute to the development of safe and efficient gene therapy protocols for clinical application.

  20. Effects of mineral trioxide aggregate, BiodentineTM and calcium hydroxide on viability, proliferation, migration and differentiation of stem cells from human exfoliated deciduous teeth

    Directory of Open Access Journals (Sweden)

    Leandro Borges Araújo

    2018-02-01

    Full Text Available Abstract Objective: The aim of the study was to evaluate the effects of the capping materials mineral trioxide aggregate (MTA, calcium hydroxide (CH and BiodentineTM (BD on stem cells from human exfoliated deciduous teeth (SHED in vitro. Material and Methods: SHED were cultured for 1 – 7 days in medium conditioned by incubation with MTA, BD or CH (1 mg/mL, and tested for viability (MTT assay and proliferation (SRB assay. Also, the migration of serum-starved SHED towards conditioned media was assayed in companion plates, with 8 μm-pore-sized membranes, for 24 h. Gene expression of dentin matrix protein-1 (DMP-1 was evaluated by reverse-transcription polymerase chain reaction. Regular culture medium with 10% FBS (without conditioning and culture medium supplemented with 20% FBS were used as controls. Results: MTA, CH and BD conditioned media maintained cell viability and allowed continuous SHED proliferation, with CH conditioned medium causing the highest positive effect on proliferation at the end of the treatment period (compared with BD and MTA (p<0.05. In contrast, we observed increased SHED migration towards BD and MTA conditioned media (compared with CH (p<0.05. A greater amount of DMP-1 gene was expressed in MTA group compared with the other groups from day 7 up to day 21. Conclusion: Our results show that the three capping materials are biocompatible, maintain viability and stimulate proliferation, migration and differentiation in a key dental stem cell population.

  1. NBL1 and anillin (ANLN genes over-expression in pancreatic carcinoma.

    Directory of Open Access Journals (Sweden)

    Dariusz Lange

    2009-12-01

    Full Text Available The aim of the study was to analyze the gene expression profile of pancreatic cancer to derive novel molecular markers of this malignancy. The snap-frozen or RNA-later preserved samples of 18 pancreatic adenocarcinomas, 5 chronic pancreatitis cases and 6 specimens of grossly normal pancreas were used for microarray analysis by HG-U133 Plus 2.0 oligonucleotide Affymetrix arrays. Validation was carried out by real-time quantitative PCR (Q-PCR in the set of 66 samples: 31 of pancreatic cancer, 14 of chronic pancreatitis and 21 of macroscopically unchanged pancreas. By Principal Component Analysis of the microarray data we found a very consistent expression pattern of normal samples and a less homogenous one in chronic pancreatitis. By supervised comparison (corrected p-value 0.001 we observed 11094 probesets differentiating between cancer and normal samples, while only seventy six probesets were significant for difference between cancer and chronic pancreatitis. The only gene occurring within the best 10 genes in both comparisons was S100 calcium binding protein P (S100P, already indicated for its utility as pancreatic cancer marker by earlier microarray-based studies. For validation we selected two genes which appeared as valuable candidates for molecular markers of pancreatic cancer: neuroblastoma, suppression of tumorigenicity 1 (NBL1 and anillin (ANLN. By Q-PCR, we confirmed statistically significant differences in these genes with a 9.5 fold-change difference between NBL1 expression in cancer/normal comparison and a relatively modest difference between cancer and pancreatitis. For ANLN even more distinct differences were observed (cancer/normal 19.8-fold, cancer/pancreatitis 4.0-fold. NBL1 and anillin are promising markers for pancreatic carcinoma molecular diagnostics.

  2. GAD1 Gene Expression in Blood of Patients with First-Episode Psychosis.

    Directory of Open Access Journals (Sweden)

    Jie Yin Yee

    Full Text Available γ-Aminobutyric acid (GABA, the primary inhibitory neurotransmitter, has often been studied in relation to its role in the pathophysiology of schizophrenia. GABA is synthesized from glutamate by glutamic acid decarboxylase (GAD, derived from two genes, GAD1 and GAD2. GAD1 is expressed as both GAD67 and GAD25 mRNA transcripts with the former reported to have a lower expression level in schizophrenia compared to healthy controls and latter was reported to be predominantly expressed fetally, suggesting a role in developmental process. In this study, GAD67 and GAD25 mRNA levels were measured by quantitative PCR (qPCR in peripheral blood of subjects with first-episode psychosis (FEP and from healthy controls. We observed low GAD25 and GAD67 gene expression levels in human peripheral blood. There was no difference in GAD25 and GAD67 gene expression level, and GAD25/GAD67 ratio between patients with FEP and healthy controls. PANSS negative symptoms were associated with levels of GAD25 mRNA transcripts in patients with FEP. While the current study provides information on GAD25 and GAD67 mRNA transcript levels in whole blood of FEP patients, further correlation and validation work between brain regions, cerebrospinal fluid and peripheral blood expression profiling are required to provide a better understanding of GAD25 and GAD67.

  3. Composition and microstructure of MTA and Aureoseal Plus: XRF, EDS, XRD and FESEM evaluation.

    Science.gov (United States)

    Cianconi, L; Palopoli, P; Campanella, V; Mancini, M

    2016-12-01

    The aim of this study was to determine the chemical composition and the phases' microstructure of Aureoseal Plus (OGNA, Italy) and ProRoot MTA (Dentsply Tulsa Dental, USA) and to compare their characteristics. Study Design: Comparing Aureoseal Plus and ProRoot MTA microstructure by means of several analyses type. The chemical analysis of the two cements was assessed following the UNI EN ISO 196-2 norm. X-Ray fluorescence (XRF) was used to determine the element composition. The crystalline structure was analysed quantitatively using x-ray diffraction (XRD). Powders morphology was evaluated using a scanning electron microscope (SEM) with backscattering detectors, and a field emission scanning electron microscope (FESEM). Elemental analysis was performed by energy dispersive x-ray analysis (EDS). The semi-quantitative XRF analysis showed the presence of heavy metal oxides in both cements. The XRD spectra of the two cements reported the presence of dicalcium silicate, tricalcium silicate, tricalcium aluminate, tetracalcium aluminoferrite, bismuth oxide and gypsum. SEM analysis showed that ProRoot MTA powder is less coarse and more homogeneous than Aureoseal. Both powders are formed by particles of different shapes: round, prismatic and oblong. The EDS analysis showed that some ProRoot MTA particles, differently from Aureoseal, contain Ca, Si, Al and Fe. Oblong particles in ProRoot and Aureoseal are rich of bismuth. The strong interest in developing new Portland cement-based endodontic sealers will create materials with increased handling characteristics and physicochemical properties. A thorough investigation on two cement powders was carried out by using XRF, XRD, SEM and EDS analysis. To date there was a lack of studies on Aureoseal Plus. This cement is similar in composition to ProRoot MTA. Despite that it has distinctive elements that could improve its characteristics, resulting in a good alternative to MTA.

  4. Neighboring Genes Show Correlated Evolution in Gene Expression

    Science.gov (United States)

    Ghanbarian, Avazeh T.; Hurst, Laurence D.

    2015-01-01

    When considering the evolution of a gene’s expression profile, we commonly assume that this is unaffected by its genomic neighborhood. This is, however, in contrast to what we know about the lack of autonomy between neighboring genes in gene expression profiles in extant taxa. Indeed, in all eukaryotic genomes genes of similar expression-profile tend to cluster, reflecting chromatin level dynamics. Does it follow that if a gene increases expression in a particular lineage then the genomic neighbors will also increase in their expression or is gene expression evolution autonomous? To address this here we consider evolution of human gene expression since the human-chimp common ancestor, allowing for both variation in estimation of current expression level and error in Bayesian estimation of the ancestral state. We find that in all tissues and both sexes, the change in gene expression of a focal gene on average predicts the change in gene expression of neighbors. The effect is highly pronounced in the immediate vicinity (genes increasing their expression in humans tend to avoid nuclear lamina domains and be enriched for the gene activator 5-hydroxymethylcytosine, we conclude that, most probably owing to chromatin level control of gene expression, a change in gene expression of one gene likely affects the expression evolution of neighbors, what we term expression piggybacking, an analog of hitchhiking. PMID:25743543

  5. Lithium ions induce prestalk-associated gene expression and inhibit prespore gene expression in Dictyostelium discoideum

    NARCIS (Netherlands)

    Peters, Dorien J.M.; Lookeren Campagne, Michiel M. van; Haastert, Peter J.M. van; Spek, Wouter; Schaap, Pauline

    1989-01-01

    We investigated the effect of Li+ on two types of cyclic AMP-regulated gene expression and on basal and cyclic AMP-stimulated inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) levels. Li+ effectively inhibits cyclic AMP-induced prespore gene expression, half-maximal inhibition occurring at about 2mM-LiCl.

  6. Regulation of UGT1A1 and HNF1 transcription factor gene expression by DNA methylation in colon cancer cells

    Directory of Open Access Journals (Sweden)

    Harvey Mario

    2010-01-01

    Full Text Available Abstract Background UDP-glucuronosyltransferase 1A1 (UGT1A1 is a pivotal enzyme involved in metabolism of SN-38, the active metabolite of irinotecan commonly used to treat metastatic colorectal cancer. We previously demonstrated aberrant methylation of specific CpG dinucleotides in UGT1A1-negative cells, and revealed that methylation state of the UGT1A1 5'-flanking sequence is negatively correlated with gene transcription. Interestingly, one of these CpG dinucleotides (CpG -4 is found close to a HNF1 response element (HRE, known to be involved in activation of UGT1A1 gene expression, and within an upstream stimulating factor (USF binding site. Results Gel retardation assays revealed that methylation of CpG-4 directly affect the interaction of USF1/2 with its cognate sequence without altering the binding for HNF1-alpha. Luciferase assays sustained a role for USF1/2 and HNF1-alpha in UGT1A1 regulation in colon cancer cells. Based on the differential expression profiles of HNF1A gene in colon cell lines, we also assessed whether methylation affects its expression. In agreement with the presence of CpG islands in the HNF1A promoter, treatments of UGT1A1-negative HCT116 colon cancer cells with a DNA methyltransferase inhibitor restore HNF1A gene expression, as observed for UGT1A1. Conclusions This study reveals that basal UGT1A1 expression in colon cells is positively regulated by HNF1-alpha and USF, and negatively regulated by DNA methylation. Besides, DNA methylation of HNF1A could also play an important role in regulating additional cellular drug metabolism and transporter pathways. This process may contribute to determine local inactivation of drugs such as the anticancer agent SN-38 by glucuronidation and define tumoral response.

  7. Functional network analysis of genes differentially expressed during xylogenesis in soc1ful woody Arabidopsis plants.

    Science.gov (United States)

    Davin, Nicolas; Edger, Patrick P; Hefer, Charles A; Mizrachi, Eshchar; Schuetz, Mathias; Smets, Erik; Myburg, Alexander A; Douglas, Carl J; Schranz, Michael E; Lens, Frederic

    2016-06-01

    Many plant genes are known to be involved in the development of cambium and wood, but how the expression and functional interaction of these genes determine the unique biology of wood remains largely unknown. We used the soc1ful loss of function mutant - the woodiest genotype known in the otherwise herbaceous model plant Arabidopsis - to investigate the expression and interactions of genes involved in secondary growth (wood formation). Detailed anatomical observations of the stem in combination with mRNA sequencing were used to assess transcriptome remodeling during xylogenesis in wild-type and woody soc1ful plants. To interpret the transcriptome changes, we constructed functional gene association networks of differentially expressed genes using the STRING database. This analysis revealed functionally enriched gene association hubs that are differentially expressed in herbaceous and woody tissues. In particular, we observed the differential expression of genes related to mechanical stress and jasmonate biosynthesis/signaling during wood formation in soc1ful plants that may be an effect of greater tension within woody tissues. Our results suggest that habit shifts from herbaceous to woody life forms observed in many angiosperm lineages could have evolved convergently by genetic changes that modulate the gene expression and interaction network, and thereby redeploy the conserved wood developmental program. © 2016 The Authors. The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.

  8. Gene expression in periodontal tissues following treatment

    Directory of Open Access Journals (Sweden)

    Eisenacher Martin

    2008-07-01

    Full Text Available Abstract Background In periodontitis, treatment aimed at controlling the periodontal biofilm infection results in a resolution of the clinical and histological signs of inflammation. Although the cell types found in periodontal tissues following treatment have been well described, information on gene expression is limited to few candidate genes. Therefore, the aim of the study was to determine the expression profiles of immune and inflammatory genes in periodontal tissues from sites with severe chronic periodontitis following periodontal therapy in order to identify genes involved in tissue homeostasis. Gingival biopsies from 12 patients with severe chronic periodontitis were taken six to eight weeks following non-surgical periodontal therapy, and from 11 healthy controls. As internal standard, RNA of an immortalized human keratinocyte line (HaCaT was used. Total RNA was subjected to gene expression profiling using a commercially available microarray system focusing on inflammation-related genes. Post-hoc confirmation of selected genes was done by Realtime-PCR. Results Out of the 136 genes analyzed, the 5% most strongly expressed genes compared to healthy controls were Interleukin-12A (IL-12A, Versican (CSPG-2, Matrixmetalloproteinase-1 (MMP-1, Down syndrome critical region protein-1 (DSCR-1, Macrophage inflammatory protein-2β (Cxcl-3, Inhibitor of apoptosis protein-1 (BIRC-1, Cluster of differentiation antigen 38 (CD38, Regulator of G-protein signalling-1 (RGS-1, and Finkel-Biskis-Jinkins murine osteosarcoma virus oncogene (C-FOS; the 5% least strongly expressed genes were Receptor-interacting Serine/Threonine Kinase-2 (RIP-2, Complement component 3 (C3, Prostaglandin-endoperoxide synthase-2 (COX-2, Interleukin-8 (IL-8, Endothelin-1 (EDN-1, Plasminogen activator inhibitor type-2 (PAI-2, Matrix-metalloproteinase-14 (MMP-14, and Interferon regulating factor-7 (IRF-7. Conclusion Gene expression profiles found in periodontal tissues following

  9. Tissue-specific alternative splicing and expression of ATP1B2 gene ...

    African Journals Online (AJOL)

    After heat-stress, the expression levels of the different transcripts were lower in different tissues; however, the expression of the ATP1B2-complete transcript increased in heart and lung tissues. The results of this research provide some useful information for further studies into the function of the bovine ATP1B2 gene.

  10. Gene Expression Commons: an open platform for absolute gene expression profiling.

    Directory of Open Access Journals (Sweden)

    Jun Seita

    Full Text Available Gene expression profiling using microarrays has been limited to comparisons of gene expression between small numbers of samples within individual experiments. However, the unknown and variable sensitivities of each probeset have rendered the absolute expression of any given gene nearly impossible to estimate. We have overcome this limitation by using a very large number (>10,000 of varied microarray data as a common reference, so that statistical attributes of each probeset, such as the dynamic range and threshold between low and high expression, can be reliably discovered through meta-analysis. This strategy is implemented in a web-based platform named "Gene Expression Commons" (https://gexc.stanford.edu/ which contains data of 39 distinct highly purified mouse hematopoietic stem/progenitor/differentiated cell populations covering almost the entire hematopoietic system. Since the Gene Expression Commons is designed as an open platform, investigators can explore the expression level of any gene, search by expression patterns of interest, submit their own microarray data, and design their own working models representing biological relationship among samples.

  11. The rules of gene expression in plants: Organ identity and gene body methylation are key factors for regulation of gene expression in Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Gutiérrez Rodrigo A

    2008-09-01

    Full Text Available Abstract Background Microarray technology is a widely used approach for monitoring genome-wide gene expression. For Arabidopsis, there are over 1,800 microarray hybridizations representing many different experimental conditions on Affymetrix™ ATH1 gene chips alone. This huge amount of data offers a unique opportunity to infer the principles that govern the regulation of gene expression in plants. Results We used bioinformatics methods to analyze publicly available data obtained using the ATH1 chip from Affymetrix. A total of 1887 ATH1 hybridizations were normalized and filtered to eliminate low-quality hybridizations. We classified and compared control and treatment hybridizations and determined differential gene expression. The largest differences in gene expression were observed when comparing samples obtained from different organs. On average, ten-fold more genes were differentially expressed between organs as compared to any other experimental variable. We defined "gene responsiveness" as the number of comparisons in which a gene changed its expression significantly. We defined genes with the highest and lowest responsiveness levels as hypervariable and housekeeping genes, respectively. Remarkably, housekeeping genes were best distinguished from hypervariable genes by differences in methylation status in their transcribed regions. Moreover, methylation in the transcribed region was inversely correlated (R2 = 0.8 with gene responsiveness on a genome-wide scale. We provide an example of this negative relationship using genes encoding TCA cycle enzymes, by contrasting their regulatory responsiveness to nitrate and methylation status in their transcribed regions. Conclusion Our results indicate that the Arabidopsis transcriptome is largely established during development and is comparatively stable when faced with external perturbations. We suggest a novel functional role for DNA methylation in the transcribed region as a key determinant

  12. Physicochemical Properties of MTA and Portland Cement after Addition of Aloe Vera.

    Science.gov (United States)

    Henrique Borges, Alvaro; Aguirre Guedes, Orlando; Evaristo Ricci Volpato, Luiz; Siebert Filho, Gilberto; Meireles Borba, Alexandre; Zina, Omar; Piva, Evandro; Estrela, Carlos

    2017-01-01

    The aim of this in vitro study was to determine the liquid-powder ratio, setting time, solubility, dimensional change, pH, and radiopacity of white structural and non-structural Portland cement, ProRoot MTA and MTA Bio, associated with a 2% glycolic solution containing Aloe Vera, as vehicle. Five samples of each material were used for each test, according to the American National Standards Institute/American Dental Association (ANSI/ADA) specification No. 57. Statistical analyses were performed using ANOVA and Tukey's test at 5% significance. When sample distribution was not normal, non-parametric analysis of variance and the Kruskal-Wallis test were used ( α =0.05). No statistical differences were found in liquid-powder ratios among the tested materials. ProRoot MTA showed the longest setting time. Dimensional change values were acceptable in all groups. Also, no significant differences were found in pH values and pH was alkaline in all samples throughout the experiment. Mean radiopacity results obtained for white Portland cements did not meet ANSI/ADA requirements, and were significantly lower than those obtained for MTA-based cements. Finally, Portland cements showed significantly higher mean solubility values compared to the other samples. The physicochemical properties of the tested materials in association with Aloe Vera were compatible with ANSI/ADA requirements, except for the white Portland cements, which failed to meet the radiopacity specification.

  13. [Cloning, subcellular localization, and heterologous expression of ApNAC1 gene from Andrographis paniculata].

    Science.gov (United States)

    Wang, Jian; Qi, Meng-Die; Guo, Juan; Shen, Ye; Lin, Hui-Xin; Huang, Lu-Qi

    2017-03-01

    Andrographis paniculata is widely used as medicinal herb in China for a long time and andrographolide is its main medicinal constituent. To investigate the underlying andrographolide biosynthesis mechanisms, RNA-seq for A. paniculata leaves with MeJA treatment was performed. In A. paniculata transcriptomic data, the expression pattern of one member of NAC transcription factor family (ApNAC1) matched with andrographolide accumulation. The coding sequence of ApNAC1 was cloned by RT-PCR, and GenBank accession number was KY196416. The analysis of bioinformatics showed that the gene encodes a peptide of 323 amino acids, with a predicted relative molecular weight of 35.9 kDa and isoelectric point of 6.14. To confirm the subcellular localization, ApNAC1-GFP was transiently expressed in A. paniculata protoplast. The results indicated that ApNAC1 is a nucleus-localized protein. The analysis of real-time quantitative PCR revealed that ApNAC1 gene predominantly expresses in leaves. Compared with control sample, its expression abundance sharply increased with methyl jasmonate treatment. Based on its expression pattern, ApNAC1 gene might involve in andrographolide biosynthesis. ApNAC1 was heterologously expressed in Escherichia coli and recombinant protein was purified by Ni-NTA agarose. Further study will help us to understand the function of ApNAC1 in andrographolide biosynthesis. Copyright© by the Chinese Pharmaceutical Association.

  14. Differentiation-inducing factor-1 suppresses gene expression of cyclin D1 in tumor cells

    International Nuclear Information System (INIS)

    Yasmin, Tania; Takahashi-Yanaga, Fumi; Mori, Jun; Miwa, Yoshikazu; Hirata, Masato; Watanabe, Yutaka; Morimoto, Sachio; Sasaguri, Toshiyuki

    2005-01-01

    To determine the mechanism by which differentiation-inducing factor-1 (DIF-1), a morphogen of Dictyostelium discoideum, inhibits tumor cell proliferation, we examined the effect of DIF-1 on the gene expression of cyclin D1. DIF-1 strongly reduced the expression of cyclin D1 mRNA and correspondingly decreased the amount of β-catenin in HeLa cells and squamous cell carcinoma cells. DIF-1 activated glycogen synthase kinase-3β (GSK-3β) and inhibition of GSK-3β attenuated the DIF-1-induced β-catenin degradation, indicating the involvement of GSK-3β in this effect. Moreover, DIF-1 reduced the activities of T-cell factor (TCF)/lymphoid enhancer factor (LEF) reporter plasmid and a reporter gene driven by the human cyclin D1 promoter. Eliminating the TCF/LEF consensus site from the cyclin D1 promoter diminished the effect of DIF-1. These results suggest that DIF-1 inhibits Wnt/β-catenin signaling, resulting in the suppression of cyclin D1 promoter activity

  15. Selection of housekeeping genes for normalization by real-time RT-PCR: analysis of Or-MYB1 gene expression in Orobanche ramosa development.

    Science.gov (United States)

    González-Verdejo, C I; Die, J V; Nadal, S; Jiménez-Marín, A; Moreno, M T; Román, B

    2008-08-15

    Real-time PCR has become the method of choice for accurate and in-depth expression studies of candidate genes. To avoid bias, real-time PCR is referred to one or several internal control genes that should not fluctuate among treatments. A need for reference genes in the parasitic plant Orobanche ramosa has emerged, and the studies in this area have not yet been evaluated. In this study, the genes 18S rRNA, Or-act1, Or-tub1, and Or-ubq1 were compared in terms of expression stability using the BestKeeper software program. Among the four common endogenous control genes, Or-act1 and Or-ubq1 were the most stable in O. ramosa samples. In parallel, a study was carried out studying the expression of the transcription factor Or-MYB1 that seemed to be implicated during preinfection stages. The normalization strategy presented here is a prerequisite to accurate real-time PCR expression profiling that, among other things, opens up the possibility of studying messenger RNA levels of low-copy-number-like transcription factors.

  16. Comprehensive Gene Expression Analysis of Rice Aleurone Cells: Probing the Existence of an Alternative Gibberellin Receptor1

    Science.gov (United States)

    Yano, Kenji; Aya, Koichiro; Hirano, Ko; Ordonio, Reynante Lacsamana; Ueguchi-Tanaka, Miyako; Matsuoka, Makoto

    2015-01-01

    Current gibberellin (GA) research indicates that GA must be perceived in plant nuclei by its cognate receptor, GIBBERELLIN INSENSITIVE DWARF1 (GID1). Recognition of GA by GID1 relieves the repression mediated by the DELLA protein, a model known as the GID1-DELLA GA perception system. There have been reports of potential GA-binding proteins in the plasma membrane that perceive GA and induce α-amylase expression in cereal aleurone cells, which is mechanistically different from the GID1-DELLA system. Therefore, we examined the expression of the rice (Oryza sativa) α-amylase genes in rice mutants impaired in the GA receptor (gid1) and the DELLA repressor (slender rice1; slr1) and confirmed their lack of response to GA in gid1 mutants and constitutive expression in slr1 mutants. We also examined the expression of GA-regulated genes by genome-wide microarray and quantitative reverse transcription-polymerase chain reaction analyses and confirmed that all GA-regulated genes are modulated by the GID1-DELLA system. Furthermore, we studied the regulatory network involved in GA signaling by using a set of mutants defective in genes involved in GA perception and gene expression, namely gid1, slr1, gid2 (a GA-related F-box protein mutant), and gamyb (a GA-related trans-acting factor mutant). Almost all GA up-regulated genes were regulated by the four named GA-signaling components. On the other hand, GA down-regulated genes showed different expression patterns with respect to GID2 and GAMYB (e.g. a considerable number of genes are not controlled by GAMYB or GID2 and GAMYB). Based on these observations, we present a comprehensive discussion of the intricate network of GA-regulated genes in rice aleurone cells. PMID:25511432

  17. Orthograde retreatment failure with extruded MTA apical plug in a large periradicular lesion followed by surgical intervention: case report.

    Science.gov (United States)

    Brito-Junior, Manoel; Faria-e-Silva, Andre Luis; Quintino, Alex Carvalho; Moreira-Junior, Gil; Geber, Mauro; Camilo, Carla Cristina; Soares, Janir Alves

    2012-01-01

    Absence of periapical healing after orthograde retreatment using an apical plug with mineral trioxide aggregate (MTA) can require surgical intervention. A patient with a root-filled maxillary central incisor with chronic apical periodontitis and sinus tract was referred for endodontic retreatment. Excessive apical enlargement was verified, indicating an MTA apical plug placement; however, an unintentional extrusion of MTA occurred during this step. The root canal was filled with gutta-percha and sealer, and periodic recalls were scheduled. The sinus tract was observed after six months and the lesion remained unaltered, although the extruded MTA had resorbed; therefore, an apicoectomy and retrograde root-end filling with MTA were performed. Absence of sinus tract, normal clinical aspect, and complete repair of the lesion were observed at the two-year follow-up.

  18. Nuclear factor 1 regulates adipose tissue-specific expression in the mouse GLUT4 gene

    International Nuclear Information System (INIS)

    Miura, Shinji; Tsunoda, Nobuyo; Ikeda, Shinobu; Kai, Yuko; Cooke, David W.; Lane, M. Daniel; Ezaki, Osamu

    2004-01-01

    Previous studies demonstrated that an adipose tissue-specific element(s) (ASE) of the murine GLUT4 gene is located between -551 and -506 in the 5'-flanking sequence and that a high-fat responsive element(s) for down-regulation of the GLUT4 gene is located between bases -701 and -552. A binding site for nuclear factor 1 (NF1), that mediates insulin and cAMP-induced repression of GLUT4 in 3T3-L1 adipocytes is located between bases -700 and -688. To examine the role of NF1 in the regulation of GLUT4 gene expression in white adipose tissues (WAT) in vivo, we created two types of transgenic mice harboring mutated either 5' or 3' half-site of NF1-binding sites in GLUT4 minigene constructs. In both cases, the GLUT4 minigene was not expressed in WAT, while expression was maintained in brown adipose tissue, skeletal muscle, and heart. This was an unexpected finding, since a -551 GLUT4 minigene that did not have the NF1-binding site was expressed in WAT. We propose a model that explains the requirement for both the ASE and the NF1-binding site for expression of GLUT4 in WAT

  19. Effects of dietary cadmium exposure on tissue-specific cadmium accumulation, iron status and expression of iron-handling and stress-inducible genes in rainbow trout: Influence of elevated dietary iron

    Energy Technology Data Exchange (ETDEWEB)

    Kwong, Raymond W.M. [Toxicology Centre, University of Saskatchewan, Saskatoon, SK, S7N 5B3 (Canada); Andres, Jose A. [Department of Biology, University of Saskatchewan, Saskatoon, SK, S7N 5E2 (Canada); Niyogi, Som, E-mail: som.niyogi@usask.ca [Department of Biology, University of Saskatchewan, Saskatoon, SK, S7N 5E2 (Canada)

    2011-03-15

    Recent evidences suggest that dietary cadmium (Cd) uptake likely occurs via the dietary iron (Fe) uptake pathway in freshwater fish, at least in part. The present study investigated the interactive effects of dietary Cd and Fe in juvenile rainbow trout (Oncorhynchus mykiss). Fish were treated for four weeks with four different diets: normal Fe, high Fe, normal Fe plus Cd, and high Fe plus Cd. Physiological parameters, tissue-specific Fe and Cd level, plasma Fe status, and tissue-specific mRNA expression of transferrin, metallothioneins (MT-A and MT-B) and heat shock proteins 70 (HSP70a and HSP70b) were analyzed. Exposure to dietary Cd increased Cd burden in the following order: intestine > kidney > stomach > liver > gill > carcass. Interestingly, high dietary Fe reduced Cd accumulation in the stomach and intestine as well as in the wholebody of fish. Dietary Cd increased hepatic transferrin mRNA expression and total Fe binding capacity in the plasma, indicating the effect of Cd on Fe handling in fish. The mRNA expression of MTs and HSP70s was also increased in various tissues following dietary Cd exposure, however the response profile of different MT and HSP70 genes was not consistent among different tissues. In general, MT-A was more responsive to Cd exposure in the intestine and liver, whereas MT-B was more responsive in the kidney. Similarly, HSP70a expression was more sensitive to Cd exposure than HSP70b, particularly in the intestine. Interestingly, high Fe diet suppressed Cd-induced induction of transferrin, MT and HSP70 genes in various tissues. Overall, our study suggests that elevated dietary Fe can reduce Cd accumulation and ameliorate Cd-induced stress responses in freshwater fish.

  20. Effects of dietary cadmium exposure on tissue-specific cadmium accumulation, iron status and expression of iron-handling and stress-inducible genes in rainbow trout: Influence of elevated dietary iron

    International Nuclear Information System (INIS)

    Kwong, Raymond W.M.; Andres, Jose A.; Niyogi, Som

    2011-01-01

    Recent evidences suggest that dietary cadmium (Cd) uptake likely occurs via the dietary iron (Fe) uptake pathway in freshwater fish, at least in part. The present study investigated the interactive effects of dietary Cd and Fe in juvenile rainbow trout (Oncorhynchus mykiss). Fish were treated for four weeks with four different diets: normal Fe, high Fe, normal Fe plus Cd, and high Fe plus Cd. Physiological parameters, tissue-specific Fe and Cd level, plasma Fe status, and tissue-specific mRNA expression of transferrin, metallothioneins (MT-A and MT-B) and heat shock proteins 70 (HSP70a and HSP70b) were analyzed. Exposure to dietary Cd increased Cd burden in the following order: intestine > kidney > stomach > liver > gill > carcass. Interestingly, high dietary Fe reduced Cd accumulation in the stomach and intestine as well as in the wholebody of fish. Dietary Cd increased hepatic transferrin mRNA expression and total Fe binding capacity in the plasma, indicating the effect of Cd on Fe handling in fish. The mRNA expression of MTs and HSP70s was also increased in various tissues following dietary Cd exposure, however the response profile of different MT and HSP70 genes was not consistent among different tissues. In general, MT-A was more responsive to Cd exposure in the intestine and liver, whereas MT-B was more responsive in the kidney. Similarly, HSP70a expression was more sensitive to Cd exposure than HSP70b, particularly in the intestine. Interestingly, high Fe diet suppressed Cd-induced induction of transferrin, MT and HSP70 genes in various tissues. Overall, our study suggests that elevated dietary Fe can reduce Cd accumulation and ameliorate Cd-induced stress responses in freshwater fish.

  1. The Effect of Laminin-1-Doped Nanoroughened Implant Surfaces: Gene Expression and Morphological Evaluation

    Directory of Open Access Journals (Sweden)

    Humberto Osvaldo Schwartz-Filho

    2012-01-01

    Full Text Available Aim. This study aimed to observe the morphological and molecular effect of laminin-1 doping to nanostructured implant surfaces in a rabbit model. Materials and Methods. Nanostructured implants were coated with laminin-1 (test; dilution, 100 μg/mL and inserted into the rabbit tibiae. Noncoated implants were used as controls. After 2 weeks of healing, the implants were removed and subjected to morphological analysis using scanning electron microscopy (SEM and gene expression analysis using the real-time reverse transcriptase-polymerase chain reaction (RT-PCR. Results. SEM revealed bony tissue attachment for both control and test implants. Real-time RT-PCR analysis showed that the expression of osteoblast markers RUNX-2, osteocalcin, alkaline phosphatase, and collagen I was higher (1.62-fold, 1.53-fold, 1.97-fold, and 1.04-fold, resp. for the implants modified by laminin-1 relative to the control. All osteoclast markers investigated in the study presented higher expression on the test implants than controls as follows: tartrate-resistant acid phosphatase (1.67-fold, calcitonin receptor (1.35-fold, and ATPase (1.25-fold. The test implants demonstrated higher expression of inflammatory markers interleukin-10 (1.53-fold and tumour necrosis factor-α (1.61-fold relative to controls. Conclusion. The protein-doped surface showed higher gene expression of typical genes involved in the osseointegration cascade than the control surface.

  2. Expressão dos genes que codificam as proteínas anexina-1 e galectina-1 nos pólipos rinossinusais e sua modulação pelo glicocorticoide Expression of genes that encode the annexin-1 and galectin-1 proteins in nasal polyposis and their modulation by glucocorticoid

    Directory of Open Access Journals (Sweden)

    Atílio Maximino Fernandes

    2010-04-01

    Full Text Available A fisiopatologia da polipose rinossinusal não é totalmente compreendida, apesar de várias hipóteses em relação ao seu processo inflamatório. OBJETIVOS: Estudo prospectivo da expressão dos genes das proteínas, anexina-1 e a galectina-1, que têm ação anti-inflamatória, e sua modulação pelo glicocorticoide. MATERIAL E MÉTODOS: Onze pacientes portadores de polipose rinossinusal tiveram biopsiados seus pólipos em dois momentos: na ausência de glicocorticoide sistêmico, e na sua presença. Nas duas amostras, foi avaliada a expressão desses genes e comparada com a expressão na mucosa nasal normal do meato médio. RESULTADOS: Verificou-se que a média de expressão dos genes que codifica a anexina-1 e galectina-1 estava predominantemente aumentada, independente do uso do glicocorticoide em relação à mucosa nasal controle. Entretanto, nos pólipos sem uso de corticoide, a média de expressão do gene da anexina-1 foi significativamente maior do que nos pólipos que estavam sob uso de glicocorticoide. Com relação à galectina-1 não houve diferença significativa entre as médias de expressão antes e após o uso de glicocorticoide sistêmico. CONCLUSÃO: Os genes apresentaram um aumento da expressão na mucosa nasal polipoide, independente do uso do glicocorticoide, porém a relação destes dois genes das proteínas anti-inflamatórias com o glicocorticoide não ocorreu da mesma maneira.Rhinosinusal polyps physiopathology is not fully understand, despite numerous hypotheses regarding its inflammatory process. AIMS: a prospective study regarding the gene expression of proteins: anexin-1 and galectin-1, which has an anti-inflammatory action and is modulated by steroids. MATERIALS AND METHODS: eleven patients with rhinosinusal polyps suffered a biopsy of their polyps at two moments: in the absence of systemic steroids and during its use. In the two samples we assessed the expression of these genes and compared it to the normal

  3. Cancer-Predicting Gene Expression Changes in Colonic Mucosa of Western Diet Fed Mlh1 +/- Mice

    Science.gov (United States)

    Dermadi Bebek, Denis; Valo, Satu; Reyhani, Nima; Ollila, Saara; Päivärinta, Essi; Peltomäki, Päivi; Mutanen, Marja; Nyström, Minna

    2013-01-01

    Colorectal cancer (CRC) is the second most common cause of cancer-related deaths in the Western world and interactions between genetic and environmental factors, including diet, are suggested to play a critical role in its etiology. We conducted a long-term feeding experiment in the mouse to address gene expression and methylation changes arising in histologically normal colonic mucosa as putative cancer-predisposing events available for early detection. The expression of 94 growth-regulatory genes previously linked to human CRC was studied at two time points (5 weeks and 12 months of age) in the heterozygote Mlh1 +/- mice, an animal model for human Lynch syndrome (LS), and wild type Mlh1 +/+ littermates, fed by either Western-style (WD) or AIN-93G control diet. In mice fed with WD, proximal colon mucosa, the predominant site of cancer formation in LS, exhibited a significant expression decrease in tumor suppressor genes, Dkk1, Hoxd1, Slc5a8, and Socs1, the latter two only in the Mlh1 +/- mice. Reduced mRNA expression was accompanied by increased promoter methylation of the respective genes. The strongest expression decrease (7.3 fold) together with a significant increase in its promoter methylation was seen in Dkk1, an antagonist of the canonical Wnt signaling pathway. Furthermore, the inactivation of Dkk1 seems to predispose to neoplasias in the proximal colon. This and the fact that Mlh1 which showed only modest methylation was still expressed in both Mlh1 +/- and Mlh1 +/+ mice indicate that the expression decreases and the inactivation of Dkk1 in particular is a prominent early marker for colon oncogenesis. PMID:24204690

  4. Cancer-predicting gene expression changes in colonic mucosa of Western diet fed Mlh1+/- mice.

    Directory of Open Access Journals (Sweden)

    Marjaana Pussila

    Full Text Available Colorectal cancer (CRC is the second most common cause of cancer-related deaths in the Western world and interactions between genetic and environmental factors, including diet, are suggested to play a critical role in its etiology. We conducted a long-term feeding experiment in the mouse to address gene expression and methylation changes arising in histologically normal colonic mucosa as putative cancer-predisposing events available for early detection. The expression of 94 growth-regulatory genes previously linked to human CRC was studied at two time points (5 weeks and 12 months of age in the heterozygote Mlh1(+/- mice, an animal model for human Lynch syndrome (LS, and wild type Mlh1(+/+ littermates, fed by either Western-style (WD or AIN-93G control diet. In mice fed with WD, proximal colon mucosa, the predominant site of cancer formation in LS, exhibited a significant expression decrease in tumor suppressor genes, Dkk1, Hoxd1, Slc5a8, and Socs1, the latter two only in the Mlh1(+/- mice. Reduced mRNA expression was accompanied by increased promoter methylation of the respective genes. The strongest expression decrease (7.3 fold together with a significant increase in its promoter methylation was seen in Dkk1, an antagonist of the canonical Wnt signaling pathway. Furthermore, the inactivation of Dkk1 seems to predispose to neoplasias in the proximal colon. This and the fact that Mlh1 which showed only modest methylation was still expressed in both Mlh1(+/- and Mlh1(+/+ mice indicate that the expression decreases and the inactivation of Dkk1 in particular is a prominent early marker for colon oncogenesis.

  5. Stat1-independent regulation of gene expression in response to IFN-γ

    Science.gov (United States)

    Ramana, Chilakamarti V.; Gil, M. Pilar; Han, Yulong; Ransohoff, Richard M.; Schreiber, Robert D.; Stark, George R.

    2001-01-01

    Although Stat1 is essential for cells to respond fully to IFN-γ, there is substantial evidence that, in the absence of Stat1, IFN-γ can still regulate the expression of some genes, induce an antiviral state and affect cell growth. We have now identified many genes that are regulated by IFN-γ in serum-starved Stat1-null mouse fibroblasts. The proteins induced by IFN-γ in Stat1-null cells can account for the substantial biological responses that remain. Some genes are induced in both wild-type and Stat1-null cells and thus are truly Stat1-independent. Others are subject to more complex regulation in response to IFN-γ, repressed by Stat1 in wild-type cells and activated in Stat1-null cells. Many genes induced by IFN-γ in Stat1-null fibroblasts also are induced by platelet-derived growth factor in wild-type cells and thus are likely to be involved in cell proliferation. In mouse cells expressing the docking site mutant Y440F of human IFN-γ receptor subunit 1, the mouse Stat1 is not phosphorylated in response to human IFN-γ, but c-myc and c-jun are still induced, showing that the Stat1 docking site is not required for Stat1-independent signaling. PMID:11390994

  6. Cell-Specific PEAR1 Methylation Studies Reveal a Locus that Coordinates Expression of Multiple Genes

    Directory of Open Access Journals (Sweden)

    Benedetta Izzi

    2018-04-01

    Full Text Available Chromosomal interactions connect distant enhancers and promoters on the same chromosome, activating or repressing gene expression. PEAR1 encodes the Platelet-Endothelial Aggregation Receptor 1, a contact receptor involved in platelet function and megakaryocyte and endothelial cell proliferation. PEAR1 expression during megakaryocyte differentiation is controlled by DNA methylation at its first CpG island. We identified a PEAR1 cell-specific methylation sensitive region in endothelial cells and megakaryocytes that showed strong chromosomal interactions with ISGL20L2, RRNAD1, MRLP24, HDGF and PRCC, using available promoter capture Hi-C datasets. These genes are involved in ribosome processing, protein synthesis, cell cycle and cell proliferation. We next studied the methylation and expression profile of these five genes in Human Umbilical Vein Endothelial Cells (HUVECs and megakaryocyte precursors. While cell-specific PEAR1 methylation corresponded to variability in expression for four out of five genes, no methylation change was observed in their promoter regions across cell types. Our data suggest that PEAR1 cell-type specific methylation changes may control long distance interactions with other genes. Further studies are needed to show whether such interaction data might be relevant for the genome-wide association data that showed a role for non-coding PEAR1 variants in the same region and platelet function, platelet count and cardiovascular risk.

  7. Heme oxygenase-1 and abscisic acid effects MAPK´s gene expression in soybean seeds

    International Nuclear Information System (INIS)

    Giacometti, R.; Santa Cruz, D.; Noriega, G.; Balestrasse, K.

    2012-01-01

    In soybean previous studies enabled the identification of MAPK3 and 6 whose activity is enhanced within the signaling pathway leading to defense reactions. In this study the effects of different compounds related to hemeoxygenase (HO-1) biosynthesis on mitogen-activated protein kinase (MAPK’s) genes expression in soybean seeds were tested. To this end, 20μM hemine, 22μM ZnPPIX, 0.5mM furidine or 100μM 8-bromoguanosine 3',5'-cyclic monophosphate (8Br) were added to pre-hydrated seeds for 5 days. MAPK’s genes expression was enhanced in seeds treated with hemine. This result indicates that heme catabolism could be involved in the signaling mediated by this cascade pathway. To confirm this hypothesis experiments were carried out in the precsence of ZnPPIX, a potent irreversible HO-1 inhibitor. In this case, no gene induction was observed. On the other hand, 8Br, a cGMP analog, induced HO-1 gene expression but did not modulate MAPK’s, indicating that this effect could not be mediated by cGMP. When the action of furidine, an abscisic acid inhibitor, was tested a diminution of HO-1 gene expression was observed. In this regard, MAPK’s showed a different response, being MAPK6 the only transcript that showed a diminished respect to controls, while MAPK3 mRNA as well as MAPKK1 was enhanced. These results were confirmed by western blotting and activity determinations. (authors)

  8. ERK1/2 mediates glucose-regulated POMC gene expression in hypothalamic neurons.

    Science.gov (United States)

    Zhang, Juan; Zhou, Yunting; Chen, Cheng; Yu, Feiyuan; Wang, Yun; Gu, Jiang; Ma, Lian; Ho, Guyu

    2015-04-01

    Hypothalamic glucose-sensing neurons regulate the expression of genes encoding feeding-related neuropetides POMC, AgRP, and NPY - the key components governing metabolic homeostasis. AMP-activated protein kinase (AMPK) is postulated to be the molecular mediator relaying glucose signals to regulate the expression of these neuropeptides. Whether other signaling mediator(s) plays a role is not clear. In this study, we investigated the role of ERK1/2 using primary hypothalamic neurons as the model system. The primary neurons were differentiated from hypothalamic progenitor cells. The differentiated neurons possessed the characteristic neuronal cell morphology and expressed neuronal post-mitotic markers as well as leptin-regulated orexigenic POMC and anorexigenic AgRP/NPY genes. Treatment of cells with glucose dose-dependently increased POMC and decreased AgRP/NPY expression with a concurrent suppression of AMPK phosphorylation. In addition, glucose treatment dose-dependently increased the ERK1/2 phosphorylation. Blockade of ERK1/2 activity with its specific inhibitor PD98059 partially (approximately 50%) abolished glucose-induced POMC expression, but had little effect on AgRP/NPY expression. Conversely, blockade of AMPK activity with its specific inhibitor produced a partial (approximately 50%) reversion of low-glucose-suppressed POMC expression, but almost completely blunted the low-glucose-induced AgRP/NPY expression. The results indicate that ERK1/2 mediated POMC but not AgRP/NPY expression. Confirming the in vitro findings, i.c.v. administration of PD98059 in rats similarly attenuated glucose-induced POMC expression in the hypothalamus, but again had little effect on AgRP/NPY expression. The results are indicative of a novel role of ERK1/2 in glucose-regulated POMC expression and offer new mechanistic insights into hypothalamic glucose sensing. © 2015 Society for Endocrinology.

  9. Effects of the addition of nanoparticulate calcium carbonate on setting time, dimensional change, compressive strength, solubility and pH of MTA.

    Science.gov (United States)

    Bernardi, A; Bortoluzzi, E A; Felippe, W T; Felippe, M C S; Wan, W S; Teixeira, C S

    2017-01-01

    To evaluate nanoparticulate calcium carbonate (NPCC) using transmission electron microscopy and the effects of NPCC addition to MTA in regard to the setting time, dimensional change, compressive strength, solubility and pH. The experimental groups were G1 (MTA), G2 (MTA with 5% NPCC) and G3 (MTA with 10% NPCC). The tests followed ISO and ADA standards. The specimens in the dimensional change and compressive strength tests were measured immediately after setting, after 24 h and after 30 days. In the solubility test, rings filled with cement were weighed after setting and after 30 days. The pH was measured after 24 h and 30 days. The data were analysed with the ANOVA, Tukey's and Kruskal-Wallis tests (α = 5%). The setting time was reduced (P  G2 > G3). The solubility test revealed a difference amongst the groups when the specimens were hydrated: G2 > G1 > G3 and dehydrated: G3 > G2 > G1. The pH of the groups was similar at 24 h with higher values in each group after 30 days (P calcium carbonate had a cubic morphology with few impurities. The addition of nanoparticulate calcium carbonate to MTA accelerated the setting time, decreased compressive strength and, after 30 days, resulted in lower dimensional change (G2), higher solubility and a higher pH. © 2015 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  10. TRL1 gene expression in tomato (Solanum lycopersicum) floral organs after γ-irradiation

    International Nuclear Information System (INIS)

    Bondarenco, V.S.; Barbacar, N.I.

    2009-01-01

    The article describes the expression patterns of a novel RAD16-like TRL1 (tomato RAD16-like 1) gene in the floral organs of tomato during anther meiosis and mature flower stages. The data on the induction of the TRL1 expression as a result of γ-irradiation is discussed. (authors)

  11. Effect of Twist, Snail and YB-1 gene expression in cervical cancer tissue on cell invasion and epithelial-mesenchymal transition

    Directory of Open Access Journals (Sweden)

    Xin-Qin Kang1

    2017-05-01

    Full Text Available Objective: To study the effect of Twist, Snail and YB-1 gene expression in cervical cancer tissue on cell invasion and epithelial-mesenchymal transition. Methods: Cervical cancer tissue samples and tissue samples adjacent to carcinoma were collected from 138 patients with radical operation for cervical cancer, fluorescence quantitative PCR method was used to detect the mRNA expression of Twist, Snail and YB-1 genes, cell invasion-related genes and epithelial-mesenchymal transition marker genes, the Pearson test was used to analyze the correlation of Twist, Snail and YB-1 gene mRNA expression in cervical cancer tissue with cell invasion and epithelial-mesenchymal transition. Results: Twist, Snail and YB-1 gene mRNA expression in cervical cancer tissue were higher than those in tissue adjacent to carcinoma, the invasion genes STAT3, YAP1, TUG1, FoxM1 and Rab11 mRNA expression were higher than those in tissue adjacent to carcinoma, and the epithelial-mesenchymal transition markers E-cadherin and β-catenin gene mRNA expression were lower than those in tissue adjacent to carcinoma while vimentin gene mRNA expression was higher than that in tissue adjacent to carcinoma. Pearson test showed that Twist, Snail and YB-1 gene mRNA expression in cervical cancer tissue were directly correlated with cell invasion and epithelial-mesenchymal transition. Conclusion: Twist, Snail and YB-1 genes are highly expressed in cervical cancer tissue, and their abnormal expression directly leads to the increased tumor cell invasion activity and the aggravated epithelial-mesenchymal transition.

  12. Mapping of gene expression reveals CYP27A1 as a susceptibility gene for sporadic ALS.

    Directory of Open Access Journals (Sweden)

    Frank P Diekstra

    Full Text Available Amyotrophic lateral sclerosis (ALS is a progressive, neurodegenerative disease characterized by loss of upper and lower motor neurons. ALS is considered to be a complex trait and genome-wide association studies (GWAS have implicated a few susceptibility loci. However, many more causal loci remain to be discovered. Since it has been shown that genetic variants associated with complex traits are more likely to be eQTLs than frequency-matched variants from GWAS platforms, we conducted a two-stage genome-wide screening for eQTLs associated with ALS. In addition, we applied an eQTL analysis to finemap association loci. Expression profiles using peripheral blood of 323 sporadic ALS patients and 413 controls were mapped to genome-wide genotyping data. Subsequently, data from a two-stage GWAS (3,568 patients and 10,163 controls were used to prioritize eQTLs identified in the first stage (162 ALS, 207 controls. These prioritized eQTLs were carried forward to the second sample with both gene-expression and genotyping data (161 ALS, 206 controls. Replicated eQTL SNPs were then tested for association in the second-stage GWAS data to find SNPs associated with disease, that survived correction for multiple testing. We thus identified twelve cis eQTLs with nominally significant associations in the second-stage GWAS data. Eight SNP-transcript pairs of highest significance (lowest p = 1.27 × 10(-51 withstood multiple-testing correction in the second stage and modulated CYP27A1 gene expression. Additionally, we show that C9orf72 appears to be the only gene in the 9p21.2 locus that is regulated in cis, showing the potential of this approach in identifying causative genes in association loci in ALS. This study has identified candidate genes for sporadic ALS, most notably CYP27A1. Mutations in CYP27A1 are causal to cerebrotendinous xanthomatosis which can present as a clinical mimic of ALS with progressive upper motor neuron loss, making it a plausible

  13. Thy-1 attenuates TNF-alpha-activated gene expression in mouse embryonic fibroblasts via Src family kinase.

    Directory of Open Access Journals (Sweden)

    Bin Shan

    Full Text Available Heterogeneous surface expression of Thy-1 in fibroblasts modulates inflammation and may thereby modulate injury and repair. As a paradigm, patients with idiopathic pulmonary fibrosis, a disease with pathologic features of chronic inflammation, demonstrate an absence of Thy-1 immunoreactivity within areas of fibrotic activity (fibroblast foci in contrast to the predominant Thy-1 expressing fibroblasts in the normal lung. Likewise, Thy-1 deficient mice display more severe lung fibrosis in response to an inflammatory injury than wildtype littermates. We investigated the role of Thy-1 in the response of fibroblasts to the pro-inflammatory cytokine TNF-alpha. Our study demonstrates distinct profiles of TNF-alpha-activated gene expression in Thy-1 positive (Thy-1+ and negative (Thy-1- subsets of mouse embryonic fibroblasts (MEF. TNF-alpha induced a robust activation of MMP-9, ICAM-1, and the IL-8 promoter driven reporter in Thy-1- MEFs, in contrast to only a modest increase in Thy-1+ counterparts. Consistently, ectopic expression of Thy-1 in Thy-1- MEFs significantly attenuated TNF-alpha-activated gene expression. Mechanistically, TNF-alpha activated Src family kinase (SFK only in Thy-1- MEFs. Blockade of SFK activation abrogated TNF-alpha-activated gene expression in Thy-1- MEFs, whereas restoration of SFK activation rescued the TNF-alpha response in Thy-1+ MEFs. Our findings suggest that Thy-1 down-regulates TNF-alpha-activated gene expression via interfering with SFK- and NF-kappaB-mediated transactivation. The current study provides a novel mechanistic insight to the distinct roles of fibroblast Thy-1 subsets in inflammation.

  14. Studies of Wilms’ Tumor (WT1 Gene Expression in Adult Acute Leukemias in Singapore

    Directory of Open Access Journals (Sweden)

    Che Kang Lim

    2007-01-01

    Full Text Available Biomarkers provide certain values for diagnosis, monitor treatment effi cacy, or for the development of novel therapeutic approach for particular diseases. Thus, the identifi cation of specifi c of biomarkers for specifi c medical problems, including malignant diseases may be valuable in medical practice. In the study, we have used the Wilms’ tumor gene (WT1 as a biomarker to evaluate its expression in local adult patients with newly diagnosed acute leukemia, including both acute myeloid and lymphoid leukemias (AML and ALL.Aim: To investigate WT1 gene expression in adult patients with acute leukemia at diagnosis.Methods: Eighteen patients with acute leukemia diagnosed at Singapore General Hospital, Singapore, between September, 2004 and July, 2005 were included in this study. There were fifteen AML and three ALL cases aged from 18 to 71 years old. Total RNA and DNA was extracted from peripheral blood mononuclear cells (PBMCs. Expression of WT1 was detected by nested reverse-transcription polymerase chain reaction (Nested RT-PCR. K562, and 3T3 cells were used as positive- and negative-controls. The results were revalidated using real-time PCR. HLA-A genotyping was performed using sequence specific oligonucleotide polymorphism (SSOP analysis.Results: WT1 gene was exclusively expressed in all eighteen, including three ALL and fi fteen AML, patients. In contrast with WT1 gene, the HLA-A genotyping was remarkably heterogeneous in these patients.Conclusions: WT1 gene expression was observed in local patients with acute leukemia at diagnosis. It may be used as a potential molecular marker for diagnosis, clinical progression of the diseases or monitoring the response to treatment, as well as a target for the development of novel therapeutic approaches.

  15. Polycistronic gene expression in Aspergillus niger.

    Science.gov (United States)

    Schuetze, Tabea; Meyer, Vera

    2017-09-25

    Genome mining approaches predict dozens of biosynthetic gene clusters in each of the filamentous fungal genomes sequenced so far. However, the majority of these gene clusters still remain cryptic because they are not expressed in their natural host. Simultaneous expression of all genes belonging to a biosynthetic pathway in a heterologous host is one approach to activate biosynthetic gene clusters and to screen the metabolites produced for bioactivities. Polycistronic expression of all pathway genes under control of a single and tunable promoter would be the method of choice, as this does not only simplify cloning procedures, but also offers control on timing and strength of expression. However, polycistronic gene expression is a feature not commonly found in eukaryotic host systems, such as Aspergillus niger. In this study, we tested the suitability of the viral P2A peptide for co-expression of three genes in A. niger. Two genes descend from Fusarium oxysporum and are essential to produce the secondary metabolite enniatin (esyn1, ekivR). The third gene (luc) encodes the reporter luciferase which was included to study position effects. Expression of the polycistronic gene cassette was put under control of the Tet-On system to ensure tunable gene expression in A. niger. In total, three polycistronic expression cassettes which differed in the position of luc were constructed and targeted to the pyrG locus in A. niger. This allowed direct comparison of the luciferase activity based on the position of the luciferase gene. Doxycycline-mediated induction of the Tet-On expression cassettes resulted in the production of one long polycistronic mRNA as proven by Northern analyses, and ensured comparable production of enniatin in all three strains. Notably, gene position within the polycistronic expression cassette matters, as, luciferase activity was lowest at position one and had a comparable activity at positions two and three. The P2A peptide can be used to express at

  16. A systems biology analysis of the changes in gene expression via silencing of HPV-18 E1 expression in HeLa cells.

    Science.gov (United States)

    Castillo, Andres; Wang, Lu; Koriyama, Chihaya; Eizuru, Yoshito; Jordan, King; Akiba, Suminori

    2014-10-01

    Previous studies have reported the detection of a truncated E1 mRNA generated from HPV-18 in HeLa cells. Although it is unclear whether a truncated E1 protein could function as a replicative helicase for viral replication, it would still retain binding sites for potential interactions with different host cell proteins. Furthermore, in this study, we found evidence in support of expression of full-length HPV-18 E1 mRNA in HeLa cells. To determine whether interactions between E1 and cellular proteins play an important role in cellular processes other than viral replication, genome-wide expression profiles of HPV-18 positive HeLa cells were compared before and after the siRNA knockdown of E1 expression. Differential expression and gene set enrichment analysis uncovered four functionally related sets of genes implicated in host defence mechanisms against viral infection. These included the toll-like receptor, interferon and apoptosis pathways, along with the antiviral interferon-stimulated gene set. In addition, we found that the transcriptional coactivator E1A-binding protein p300 (EP300) was downregulated, which is interesting given that EP300 is thought to be required for the transcription of HPV-18 genes in HeLa cells. The observed changes in gene expression produced via the silencing of HPV-18 E1 expression in HeLa cells indicate that in addition to its well-known role in viral replication, the E1 protein may also play an important role in mitigating the host's ability to defend against viral infection.

  17. The Chromatin Protein DUET/MMD1 Controls Expression of the Meiotic Gene TDM1 during Male Meiosis in Arabidopsis.

    Science.gov (United States)

    Andreuzza, Sébastien; Nishal, Bindu; Singh, Aparna; Siddiqi, Imran

    2015-09-01

    Meiosis produces haploid cells essential for sexual reproduction. In yeast, entry into meiosis activates transcription factors which trigger a transcriptional cascade that results in sequential co-expression of early, middle and late meiotic genes. However, these factors are not conserved, and the factors and regulatory mechanisms that ensure proper meiotic gene expression in multicellular eukaryotes are poorly understood. Here, we report that DUET/MMD1, a PHD finger protein essential for Arabidopsis male meiosis, functions as a transcriptional regulator in plant meiosis. We find that DUET-PHD binds H3K4me2 in vitro, and show that this interaction is critical for function during meiosis. We also show that DUET is required for proper microtubule organization during meiosis II, independently of its function in meiosis I. Remarkably, DUET protein shows stage-specific expression, confined to diplotene. We identify two genes TDM1 and JAS with critical functions in cell cycle transitions and spindle organization in male meiosis, as DUET targets, with TDM1 being a direct target. Thus, DUET is required to regulate microtubule organization and cell cycle transitions during male meiosis, and functions as a direct transcription activator of the meiotic gene TDM1. Expression profiling showed reduced expression of a subset comprising about 12% of a known set of meiosis preferred genes in the duet mutant. Our results reveal the action of DUET as a transcriptional regulator during male meiosis in plants, and suggest that transcription of meiotic genes is under stagewise control in plants as in yeast.

  18. Assays for noninvasive imaging of reporter gene expression

    International Nuclear Information System (INIS)

    Gambhir, S.S.; Barrio, J.R.; Herschman, H.R.; Phelps, M.E.

    1999-01-01

    Repeated, noninvasive imaging of reporter gene expression is emerging as a valuable tool for monitoring the expression of genes in animals and humans. Monitoring of organ/cell transplantation in living animals and humans, and the assessment of environmental, behavioral, and pharmacologic modulation of gene expression in transgenic animals should soon be possible. The earliest clinical application is likely to be monitoring human gene therapy in tumors transduced with the herpes simplex virus type 1 thymidine kinase (HSV1-tk) suicide gene. Several candidate assays for imaging reporter gene expression have been studied, utilizing cytosine deaminase (CD), HSV1-tk, and dopamine 2 receptor (D2R) as reporter genes. For the HSV1-tk reporter gene, both uracil nucleoside derivatives (e.g., 5-iodo-2'-fluoro-2'-deoxy-1-β-D-arabinofuranosyl-5-iodouracil [FIAU] labeled with 124 I, 131 I ) and acycloguanosine derivatives {e.g., 8-[ 18 F]fluoro-9-[[2-hydroxy-1-(hydroxymethyl)ethoxy]methyl]guanine (8-[ 18 F]-fluoroganciclovir) ([ 18 F]FGCV), 9-[(3-[ 18 F]fluoro-1-hydroxy-2-propoxy)methyl]guanine ([ 18 F]FHPG)} have been investigated as reporter probes. For the D2R reporter gene, a derivative of spiperone {3-(2'-[ 18 F]-Fluoroethyl)spiperone ([ 18 F]FESP)} has been used with positron emission tomography (PET) imaging. In this review, the principles and specific assays for imaging reporter gene expression are presented and discussed. Specific examples utilizing adenoviral-mediated delivery of a reporter gene as well as tumors expressing reporter genes are discussed

  19. Analysis of gene expression of myo1c and inpp5k genes involved in endometrial adenocarcinoma

    International Nuclear Information System (INIS)

    Koul, A.M.; Nadeem, A.; Baryalai, P.

    2012-01-01

    Abstract: Inpp5k gene encodes a protein which plays a very vital role in a number of metabolic pathways. It is very significant in the glucose metabolism where it regulates the signalling of the insulin pathway. But the full molecular details of the pathways regulated by Inpp5k encoded protein are not known. It is speculated that Inpp5k gene expression is altered in case of endometrial adenocarcinoma. Myolc gene encodes for a protein called Myosin-lc which acts an actin-based molecular motor in the cells. II has been studied that this gene down-regulates during endometrial adenocarcinoma and colorectal cancers. In this study the expression analysis of these two was carried out using multiplex PCR. An endogenous control was used for this PCR. ACTS gene served as the endogenous control because of it being a house keeping gene. It thus shows a universal expression in all cells. Thus in this study the gene expression of Inpp5k and Myulc genes was comparatively analysed with ACTS gene. The results that came out of this study showed an over-expression of Inpp5k gene and down-regulation of myolc gene with respect to ACTS gene in cancer cell lines as was indicated by the previous studies with these genes. Expression of both genes i.e. Inpp5k and Myolc was statistically compared between normal and cancerous cell lines and was found statistically significant at a value of P< O.O I in most of the cases. (author)

  20. Comparison between visual assessment of MTA and hippocampal volumes in an elderly, non-demented population

    International Nuclear Information System (INIS)

    Cavallin, Lena; Axelsson, Rimma; Bronge, Lena; Zhang, Yi; Oeksengaard, Anne-Rita; Wahlund, Lars-Olof; Fratiglioni, Laura

    2012-01-01

    Background: It is important to have a replicable easy method for monitoring atrophy progression in Alzheimer's disease. Volumetric methods for calculating hippocampal volume are time-consuming and commonly used in research. Visual assessments of medial temporal lobe atrophy (vaMTA) is a rapid method for clinical use. This method has not been tested in a large non-demented population in comparison with volumetry measurements. Since hippocampal volume decreases with time even in normal aging there is also a need to study the normal age differences of medial temporal lobe atrophy. Purpose: To compare visual assessment of medial temporal lobe atrophy (vaMTA) with hippocampal volume in a healthy, non-demented elderly population. To describe normal ageing using vaMTA. Material and Methods: Non-demented individuals aged 60, 66, 72, 78, 81, 84, and ≥87 years old were recruited from the Swedish National study on Ageing and Care in Kungsholmen (SNAC-K), Sweden. Standard magnetic resonance imaging (MRI) scans, vaMTA, and calculations of hippocampal volumes were performed in 544 subjects. Results: Significant correlation (rs = -0.32, P 80-year-old individuals

  1. Imaging Herpes Simplex Virus Type 1 Amplicon Vector–Mediated Gene Expression in Human Glioma Spheroids

    OpenAIRE

    Christine Kaestle; Alexandra Winkeler; Raphaela Richter; Heinrich Sauer; Jürgen Hescheler; Cornel Fraefel; Maria Wartenberg; Andreas H. Jacobs

    2011-01-01

    Vectors derived from herpes simplex virus type 1 (HSV-1) have great potential for transducing therapeutic genes into the central nervous system; however, inefficient distribution of vector particles in vivo may limit their therapeutic potential in patients with gliomas. This study was performed to investigate the extent of HSV-1 amplicon vector–mediated gene expression in a three-dimensional glioma model of multicellular spheroids by imaging highly infectious HSV-1 virions expressing green fl...

  2. Determining Physical Mechanisms of Gene Expression Regulation from Single Cell Gene Expression Data

    OpenAIRE

    Ezer, Daphne; Moignard, Victoria; G?ttgens, Berthold; Adryan, Boris

    2016-01-01

    Many genes are expressed in bursts, which can contribute to cell-to-cell heterogeneity. It is now possible to measure this heterogeneity with high throughput single cell gene expression assays (single cell qPCR and RNA-seq). These experimental approaches generate gene expression distributions which can be used to estimate the kinetic parameters of gene expression bursting, namely the rate that genes turn on, the rate that genes turn off, and the rate of transcription. We construct a complete ...

  3. Sterol regulatory element binding protein-1 (SREBP1) gene expression is similarly increased in polycystic ovary syndrome and endometrial cancer.

    Science.gov (United States)

    Shafiee, Mohamad N; Mongan, Nigel; Seedhouse, Claire; Chapman, Caroline; Deen, Suha; Abu, Jafaru; Atiomo, William

    2017-05-01

    Women with polycystic ovary syndrome have a three-fold higher risk of endometrial cancer. Insulin resistance and hyperlipidemia may be pertinent factors in the pathogenesis of both conditions. The aim of this study was to investigate endometrial sterol regulatory element binding protein-1 gene expression in polycystic ovary syndrome and endometrial cancer endometrium, and to correlate endometrial sterol regulatory element binding protein-1 gene expression with serum lipid profiles. A cross-sectional study was performed at Nottingham University Hospital, UK. A total of 102 women (polycystic ovary syndrome, endometrial cancer and controls; 34 participants in each group) were recruited. Clinical and biochemical assessments were performed before endometrial biopsies were obtained from all participants. Taqman real-time polymerase chain reaction for endometrial sterol regulatory element binding protein-1 gene and its systemic protein expression were analyzed. The body mass indices of women with polycystic ovary syndrome (29.28 ± 2.91 kg/m 2 ) and controls (28.58 ± 2.62 kg/m 2 ) were not significantly different. Women with endometrial cancer had a higher mean body mass index (32.22 ± 5.70 kg/m 2 ). Sterol regulatory element binding protein-1 gene expression was significantly increased in polycystic ovary syndrome and endometrial cancer endometrium compared with controls (p ovary syndrome, but this was not statistically significant. Similarly, statistically insignificant positive correlations were found between endometrial sterol regulatory element binding protein-1 gene expression and body mass index in endometrial cancer (r = 0.643, p = 0.06) and waist-hip ratio (r = 0.096, p = 0.073). Sterol regulatory element binding protein-1 gene expression was significantly positively correlated with triglyceride in both polycystic ovary syndrome and endometrial cancer (p = 0.028 and p = 0.027, respectively). Quantitative serum sterol regulatory element

  4. Human Sterol Regulatory Element-Binding Protein 1a Contributes Significantly to Hepatic Lipogenic Gene Expression

    Directory of Open Access Journals (Sweden)

    Andreas Bitter

    2015-01-01

    Full Text Available Background/Aims: Sterol regulatory element-binding protein (SREBP 1, the master regulator of lipogenesis, was shown to be associated with non-alcoholic fatty liver disease, which is attributed to its major isoform SREBP1c. Based on studies in mice, the minor isoform SREBP1a is regarded as negligible for hepatic lipogenesis. This study aims to elucidate the expression and functional role of SREBP1a in human liver. Methods: mRNA expression of both isoforms was quantified in cohorts of human livers and primary human hepatocytes. Hepatocytes were treated with PF-429242 to inhibit the proteolytic activation of SREBP precursor protein. SREBP1a-specifc and pan-SREBP1 knock-down were performed by transfection of respective siRNAs. Lipogenic SREBP-target gene expression was analyzed by real-time RT-PCR. Results: In human liver, SREBP1a accounts for up to half of the total SREBP1 pool. Treatment with PF-429242 indicated SREBP-dependent auto-regulation of SREBP1a, which however was much weaker than of SREBP1c. SREBP1a-specifc knock-down also reduced significantly the expression of SREBP1c and of SREBP-target genes. Regarding most SREBP-target genes, simultaneous knock-down of both isoforms resulted in effects of only similar extent as SREBP1a-specific knock-down. Conclusion: We here showed that SREBP1a is significantly contributing to the human hepatic SREBP1 pool and has a share in human hepatic lipogenic gene expression.

  5. Expression of human mag-1 gene in E. coli and preparation of its antibody

    International Nuclear Information System (INIS)

    Lin Huiyun; Xu Yuanji; Wang Yan; Chen Huihua; Du Zhiyan; Tan Xiaogang; Lu Yinglin

    2006-01-01

    Objective: To further investigate the new metastasis associated gene, mag-1 expressed in E. coli and its anti-body was prepared in rabbit. Methods: mag-1 was amplified by PCR from pcDNA3-mag-1 and directly cloned into pET-28a vector. The fusion protein was expressed in BL21 and identified by Western blot using anti-His monoclonal antibody. Rabbit was immunized with partially purified fusion protein subcutaneously. Results: Sequence analysis revealed identity of the sequence obtained to the previous report. The recombinant His-mag-1 could be expressed in E. coli as a fusion protein of 18 x 10 3 . The recombinant protein was mostly expressed in the inclusion bodies on the induction by 0.1 mmol/L IPTG at 37 degree C for 6 hours. Western blot analysis showed that the recombinant protein could be recognized by His monoclonal anti-body. The titer of polyclonal antibody against mag-1 was 1:160000. Conclusion: The mag-1 gene is expressed in E. coli highly and its antibody is prepared successfully. (authors)

  6. Cloning of radiation-induced new gene RS1 expressed in mouse intestinal epithelium by enhanced RACE

    International Nuclear Information System (INIS)

    Wang Fengchao; Wang Junping; Su Yongping; Gao Jinsheng; Lou Shufen; Liu Xiaohong; Ren Jiong; Zhang Bo

    2003-01-01

    Objective: To obtain full-length cDNA of radiation-induced new gene RS1 expressed in mouse intestinal epithelium. Methods: The tissue expression profile of RS1 was analyzed by semi-quantitative RT-PCR to find the target tissue which highly expresses RS1. The total RNA extracted from the corresponding tissue was taken as the template for reverse-transcription. Enhanced RACE PCR was used to clone the full-length cDNA of RS1, including enrichment of the target gene through biotin-labeled probe for magnetic bead purification and nested PCR. Results: About a 2 kb long 3' end was successfully cloned and cloning of the 5' end proceeded well. Conclusion: The result is consistent with our experiment design. The set of combined techniques has been identified with the cloning of full-length cDNA from EST sequence especially when the optimal gene-specific primers are not available or the expression level of target gene is low

  7. Studies on the metabolism and toxicological detection of the designer drug 4-methylthioamphetamine (4-MTA) in human urine using gas chromatography-mass spectrometry.

    Science.gov (United States)

    Ewald, Andreas H; Peters, Frank T; Weise, Magdalene; Maurer, Hans H

    2005-09-25

    4-Methylthioamphetamine (4-MTA) is a scheduled designer drug that has appeared on the illicit drug market and led to several non-fatal or even fatal poisonings. Only few data are available on its metabolism. The first aim of this study was to identify the 4-MTA metabolites in human urine and then to study whether the authors' STA procedure is suitable for screening for and identification of 4-MTA and/or its metabolites in urine. After enzymatic cleavage of conjugates, solid-phase extraction (SPE) and acetylation the following metabolites could be identified by full-scan gas chromatography-mass spectrometry (GC-MS): deamino-oxo 4-MTA, deamino-hydroxy 4-MTA, ring hydroxy and beta-hydroxy 4-MTA. 4-MTA sulfoxide could be identified as possible artifact. In urine samples after enzymatic hydrolysis, acidic extraction, and methylation, 4-methylthiobenzoic acid could be identified. The authors' systematical toxicological analysis (STA) procedure using full-scan GC-MS after acid hydrolysis, liquid-liquid extraction (LLE) and acetylation allowed detection of 4-MTA as target analyte plus all the above-mentioned metabolites with the exception of 4-methylthiobenzoic acid. The extraction efficiency of 4-MTA was approximately 70% and the limit of detection (LOD) was 30 ng/ml (S/N 3).

  8. Multiple Sclerosis Risk Variant HLA-DRB1*1501 Associates with High Expression of DRB1 Gene in Different Human Populations

    Science.gov (United States)

    Abad-Grau, María del Mar; Fedetz, María; Izquierdo, Guillermo; Lucas, Miguel; Fernández, Óscar; Ndagire, Dorothy; Catalá-Rabasa, Antonio; Ruiz, Agustín; Gayán, Javier; Delgado, Concepción; Arnal, Carmen

    2012-01-01

    The human leukocyte antigen (HLA) DRB1*1501 has been consistently associated with multiple sclerosis (MS) in nearly all populations tested. This points to a specific antigen presentation as the pathogenic mechanism though this does not fully explain the disease association. The identification of expression quantitative trait loci (eQTL) for genes in the HLA locus poses the question of the role of gene expression in MS susceptibility. We analyzed the eQTLs in the HLA region with respect to MS-associated HLA-variants obtained from genome-wide association studies (GWAS). We found that the Tag of DRB1*1501, rs3135388 A allele, correlated with high expression of DRB1, DRB5 and DQB1 genes in a Caucasian population. In quantitative terms, the MS-risk AA genotype carriers of rs3135388 were associated with 15.7-, 5.2- and 8.3-fold higher expression of DQB1, DRB5 and DRB1, respectively, than the non-risk GG carriers. The haplotype analysis of expression-associated variants in a Spanish MS cohort revealed that high expression of DRB1 and DQB1 alone did not contribute to the disease. However, in Caucasian, Asian and African American populations, the DRB1*1501 allele was always highly expressed. In other immune related diseases such as type 1 diabetes, inflammatory bowel disease, ulcerative colitis, asthma and IgA deficiency, the best GWAS-associated HLA SNPs were also eQTLs for different HLA Class II genes. Our data suggest that the DR/DQ expression levels, together with specific structural properties of alleles, seem to be the causal effect in MS and in other immunopathologies rather than specific antigen presentation alone. PMID:22253788

  9. Temporal profile of estrogen-dependent gene expression in LHRH-producing GT1-7 cells.

    Science.gov (United States)

    Varju, Patricia; Chang, Ken C; Hrabovszky, Erik; Merchenthaler, István; Liposits, Zsolt

    2009-02-01

    The long-term cellular effects of estrogens are mediated by nuclear estrogen receptors which act as transcription factors to regulate gene expression. Hypothalamic targets of estrogen action include luteinizing hormone-releasing hormone-secreting neurons controlling reproduction in vertebrates. Microarray analysis and qRT-PCR studies were performed on GT1-7, immortalized LHRH neurons after 17beta-estradiol treatment to reveal the nature of estrogen-regulated genes and the time course of changes in their expression profile. More than 1000 transcripts showed robust responses to estrogen treatment and the majority of responding genes were up-regulated. Early-responding genes showed altered expression 0.5-2h after estrogen exposure, whereas late-responding genes changed after 24-48h treatment. Up-regulated genes encoded transcription factors, molecules involved in cellular movement, cell death, immune response, neurotransmitter and neuropeptide receptors, ion channels and transporters. The 17beta-estradiol modulation of 12 genes - representing characteristic gene clusters - has been confirmed by qRT-PCR. Our studies highlighted diverse gene networks, cell regulatory mechanisms and metabolic pathways through which estrogen may alter gene expression in immortalized LHRH neurons. The findings also support the notion that genomic effects of estrogen targeting in vivo directly the LHRH neuronal network of mammals play an important role in the central feedback regulation of the reproductive axis by estrogen.

  10. Combination of Mineral Trioxide Aggregate and Platelet-rich Fibrin Promotes the Odontoblastic Differentiation and Mineralization of Human Dental Pulp Cells via BMP/Smad Signaling Pathway.

    Science.gov (United States)

    Woo, Su-Mi; Kim, Won-Jae; Lim, Hae-Soon; Choi, Nam-Ki; Kim, Sun-Hun; Kim, Seon-Mi; Jung, Ji-Yeon

    2016-01-01

    Recent reports have shown that the combined use of platelet-rich fibrin (PRF), an autologous fibrin matrix, and mineral trioxide aggregate (MTA) as root filling material is beneficial for the endodontic management of an open apex. However, the potential of the combination of MTA and PRF as an odontogenic inducer in human dental pulp cells (HDPCs) in vitro has not yet been studied. The purpose of this study was to evaluate the effect of the combination of MTA and PRF on odontoblastic maturation in HDPCs. HDPCs extracted from third molars were directly cultured with MTA and PRF extract (PRFe). Odontoblastic differentiation of HDPCs was evaluated by measuring the alkaline phosphatase (ALP) activity, and the expression of odontogenesis-related genes was detected using reverse-transcription polymerase chain reaction or Western blot. Mineralization formation was assessed by alizarin red staining. HDPCs treated with MTA and PRFe significantly up-regulated the expression of dentin sialoprotein and dentin matrix protein-1 and enhanced ALP activity and mineralization compared with those with MTA or PRFe treatment alone. In addition, the combination of MTA and PRFe induced the activation of bone morphogenic proteins (BMP)/Smad, whereas LDN193189, the bone morphogenic protein inhibitor, attenuated dentin sialophosphoprotein and dentin matrix protein-1 expression, ALP activity, and mineralization enhanced by MTA and PRFe treatment. This study shows that the combination of MTA and PRF has a synergistic effect on the stimulation of odontoblastic differentiation of HDPCs via the modulation of the BMP/Smad signaling pathway. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  11. Evaluation of the tissue reaction to fast endodontic cement (CER) and Angelus MTA.

    Science.gov (United States)

    Gomes-Filho, João Eduardo; Rodrigues, Guilherme; Watanabe, Simone; Estrada Bernabé, Pedro Felício; Lodi, Carolina Simonett; Gomes, Alessandra Cristina; Faria, Max Doulgas; Domingos Dos Santos, Alailson; Silos Moraes, João Carlos

    2009-10-01

    A new cement (CER; Cimento Endodôntico Rápido or fast endodontic cement) has been developed to improve handling properties. It is a formulation that has Portland cement in gel. However, there had not yet been any study evaluating its biologic properties. The purpose of this study was to evaluate the rat subcutaneous tissue response to CER and Angelus MTA. The materials were placed in polyethylene tubes and implanted into dorsal connective tissue of Wistar rats for 7, 30, and 60 days. The specimens were prepared to be stained with hematoxylin-eosin or von Kossa or not stained for polarized light. The presence of inflammation, predominant cell type, calcification, and thickness of fibrous connective tissue were recorded. Scores were defined as follows: 0, none or few inflammatory cells, no reaction; 1, 125 cells, severe reaction. Fibrous capsule was categorized as thin when thickness was 150 mum. Necrosis and formation of calcification were both recorded. Both materials Angelus MTA and CER caused moderate reactions at 7 days, which decreased with time. The response was similar to the control at 30 and 60 days with Angelus MTA and CER, characterized by organized connective tissue and presence of some chronic inflammatory cells. Mineralization and granulations birefringent to polarized light were observed with both materials. It was possible to conclude that CER was biocompatible and stimulated mineralization.

  12. Changes in the topology of gene expression networks by human immunodeficiency virus type 1 (HIV-1) integration in macrophages.

    Science.gov (United States)

    Soto-Girón, María Juliana; García-Vallejo, Felipe

    2012-01-01

    One key step of human immunodeficiency virus type 1 (HIV-1) infection is the integration of its viral cDNA. This process is mediated through complex networks of host-virus interactions that alter several normal cell functions of the host. To study the complexity of disturbances in cell gene expression networks by HIV-1 integration, we constructed a network of human macrophage genes located close to chromatin regions rich in proviruses. To perform the network analysis, we selected 28 genes previously identified as the target of cDNA integration and their transcriptional profiles were obtained from GEO Profiles (NCBI). A total of 2770 interactions among the 28 genes located around the HIV-1 proviruses in human macrophages formed a highly dense main network connected to five sub-networks. The overall network was significantly enriched by genes associated with signal transduction, cellular communication and regulatory processes. To simulate the effects of HIV-1 integration in infected macrophages, five genes with the most number of interaction in the normal network were turned off by putting in zero the correspondent expression values. The HIV-1 infected network showed changes in its topology and alteration in the macrophage functions reflected in a re-programming of biosynthetic and general metabolic process. Understanding the complex virus-host interactions that occur during HIV-1 integration, may provided valuable genomic information to develop new antiviral treatments focusing on the management of some specific gene expression networks associated with viral integration. This is the first gene network which describes the human macrophages genes interactions related with HIV-1 integration. Copyright © 2011 Elsevier B.V. All rights reserved.

  13. Expression of the BRCA1 gene in a breast tumor: Correlation with the effect of neoadjuvant chemotherapy

    Science.gov (United States)

    Tsyganov, M. M.; Ibragimova, M. K.; Deryusheva, I. V.; Slonimskaya, E. M.; Litviakov, N. V.

    2017-09-01

    Most current research is limited by only germinal mutations of the BRCA1 gene (more often 5382insC) and the number of studies, which characterize various somatic alterations of the BRCA1 gene in a tumor, namely the expression of this gene and its correlation with the efficiency of chemotherapy, which is scarce. Taking into account the data on the connection between the genetic mutation of BRCA1 with the high efficiency of the platinum medication one may suggest that the expression of the BRCA1 gene is also associated with the high sensitivity of the tumor to the platinum medication. Aim: to evaluate the correlation between the expression of the BRCA1 gene in a breast tumor with the neoadjuvant chemotherapy (NACT) efficiency. The research included 86 patients with BC. We evaluated the expression of BRCA1 in the tumor material before and after NACT. We established that objective response to NACT is connected with a high level of BRCA1 in the general group of patients (p = 0.01) and in case of docetaxel monotherapy (p < 0.05).

  14. Molecular cloning, sequence identification and expression profile of domestic guinea pig (Cavia porcellus UGT1A1 gene

    Directory of Open Access Journals (Sweden)

    Yang Deming

    2016-01-01

    Full Text Available Domestic guinea pig is a model animal for human disease research. Uridine diphosphate glucuronosyltransferase 1 family, polypeptide A1 (UGT1A1 is an important human disease-related gene. In this study, the complete coding sequence of domestic guinea pig gene UGT1A1 was amplified by reverse transcription-polymerase chain reaction. The open reading frame of the domestic guinea pig UGT1A1 gene is 1602 bp in length and was found to encode a protein of 533 amino acids. Sequence analysis revealed that the UGT1A1 protein of domestic guinea pig shared high homology with the UGT1A1 proteins of degu (84%, damara mole-rat (84%, human (80%, northern white-cheeked gibbon (80%, Colobus angolensis palliatus (80% and golden snub-nosed monkey (79%. This gene contains five exons and four introns, as revealed by the computer-assisted analysis. The results also showed that the domestic guinea pig UGT1A1 gene had a close genetic relationship with the UGT1A1 gene of degu. The prediction of transmembrane helices showed that domestic guinea pig UGT1A1 might be a transmembrane protein. Expression profile analysis indicated that the domestic guinea pig UGT1A1 gene was differentially expressed in detected domestic guinea pig tissues. Our experiment laid a primary foundation for using the domestic guinea pig as a model animal to study the UGT1A1-related human diseases.

  15. Relative IGF-1 and IGF-2 gene expression in maternal and fetal tissues from diabetic swine

    International Nuclear Information System (INIS)

    Wolverton, C.K.; Leaman, D.W.; White, M.E.; Ramsay, T.G.

    1990-01-01

    Fourteen pregnant, crossbred gilts were utilized in this study. Seven gilts were injected with alloxan (50 mg/kg) at day 75 of gestation to induce diabetes. Gilts underwent caesarean section on day 105 of gestation. Samples were collected from maternal skeletal muscle, adipose tissue, uterus and endometrium; and from fetal skeletal muscle, adipose tissue, placenta, liver, lung, kidney, heart, brain and spleen. Tissues were frozen in liquid nitrogen for later analysis of IGF-1 and IGF-2 gene expression. Samples were pooled and total RNA was isolated using the guanidine isothiocynate method. Total mRNA was analyzed by dot blot hybridization. Blots were probed with 32 P-cDNA for porcine IGF-1 and rat IGF-2. IGF-1 gene expression in maternal tissues was unaffected by diabetes. Maternal diabetes increased IGF-2 mRNA in maternal adipose tissue but exhibited no effect in muscle or uterus. Expression of IGF-2 by maternal endometrium was decreased by diabetes. Maternal diabetes induced an increase in IGF-1 gene expression in muscle and placenta while causing an increase in IGF-2 expression in fetal liver and placenta. IGF-2 mRNA was lower in lung from fetuses of diabetic mothers than in controls. These results suggest that maternal diabetes alters IGF-1 and IGF-2 gene expression in specific tissues and differential regulation of these genes appears to exist in the mother and developing fetus

  16. Poly(ADP-ribose) polymerase inhibitors suppress UV-induced human immunodeficiency virus type 1 gene expression at the posttranscriptional level

    International Nuclear Information System (INIS)

    Yamagoe, S.; Kohda, T.; Oishi, M.

    1991-01-01

    Gene expression of human immunodeficiency virus type 1 (HIV-1) is induced not only by trans activation mediated through a gene product (tat) encoded by the virus but also by treatment of virus-carrying cells with DNA-damaging agents such as UV light. Employing an artificially constructed DNA in which the chloramphenicol acetyltransferase gene was placed under the control of the HIV-1 long terminal repeat, we analyzed the induction process in HeLa cells and found that inhibitors of poly(ADP-ribose) polymerase suppressed UV-induced HIV-1 gene expression but not tat-mediated expression. We also found that suppression occurs at the posttranscriptional level. These results indicate that HIV-1 gene expression is activated by at least two different mechanisms, one of which involves poly-ADP ribosylation. A possible new role of poly-ADP ribosylation in the regulation of specific gene expression is also discussed

  17. Expression profile of Lgi1 gene in mouse brain during development.

    Science.gov (United States)

    Ribeiro, Patrícia A O; Sbragia, Lourenço; Gilioli, Rovilson; Langone, Francesco; Conte, Fábio F; Lopes-Cendes, Iscia

    2008-07-01

    Mutations in LGI1 were described in patients with autosomal dominant partial epilepsy with auditory features (ADPEAF), and recent clinical findings have implicated LGI1 in human brain development. However, the precise role of LGI1 in epileptogenesis remains largely unknown. The objective of this study was to determine the expression pattern of Lgi1 in mice brain during development and in adult animals. Real-time polymerase chain reaction (PCR) quantification and Western blot experiments showed a relative low expression during intrauterine stages, increasing until adulthood. In addition, we did not find significant differences between left and right hemispheres. The hippocampus presented higher levels of Lgi1 expression when compared to the neocortex and the cerebellum of adult animals; however, these results did not reach statistical significance. This study was the first to determine a specific profile of Lgi1 gene expression during central nervous system development, which suggests a possible inhibitory function in latter stages of development. In addition, we did not find differences in hemispheric expression that could explain the predominance of left-sided abnormalities in patients with ADPEAF.

  18. Identifying key genes in rheumatoid arthritis by weighted gene co-expression network analysis.

    Science.gov (United States)

    Ma, Chunhui; Lv, Qi; Teng, Songsong; Yu, Yinxian; Niu, Kerun; Yi, Chengqin

    2017-08-01

    This study aimed to identify rheumatoid arthritis (RA) related genes based on microarray data using the WGCNA (weighted gene co-expression network analysis) method. Two gene expression profile datasets GSE55235 (10 RA samples and 10 healthy controls) and GSE77298 (16 RA samples and seven healthy controls) were downloaded from Gene Expression Omnibus database. Characteristic genes were identified using metaDE package. WGCNA was used to find disease-related networks based on gene expression correlation coefficients, and module significance was defined as the average gene significance of all genes used to assess the correlation between the module and RA status. Genes in the disease-related gene co-expression network were subject to functional annotation and pathway enrichment analysis using Database for Annotation Visualization and Integrated Discovery. Characteristic genes were also mapped to the Connectivity Map to screen small molecules. A total of 599 characteristic genes were identified. For each dataset, characteristic genes in the green, red and turquoise modules were most closely associated with RA, with gene numbers of 54, 43 and 79, respectively. These genes were enriched in totally enriched in 17 Gene Ontology terms, mainly related to immune response (CD97, FYB, CXCL1, IKBKE, CCR1, etc.), inflammatory response (CD97, CXCL1, C3AR1, CCR1, LYZ, etc.) and homeostasis (C3AR1, CCR1, PLN, CCL19, PPT1, etc.). Two small-molecule drugs sanguinarine and papaverine were predicted to have a therapeutic effect against RA. Genes related to immune response, inflammatory response and homeostasis presumably have critical roles in RA pathogenesis. Sanguinarine and papaverine have a potential therapeutic effect against RA. © 2017 Asia Pacific League of Associations for Rheumatology and John Wiley & Sons Australia, Ltd.

  19. Profiling Gene Expression in Germinating Brassica Roots.

    Science.gov (United States)

    Park, Myoung Ryoul; Wang, Yi-Hong; Hasenstein, Karl H

    2014-01-01

    Based on previously developed solid-phase gene extraction (SPGE) we examined the mRNA profile in primary roots of Brassica rapa seedlings for highly expressed genes like ACT7 (actin7), TUB (tubulin1), UBQ (ubiquitin), and low expressed GLK (glucokinase) during the first day post-germination. The assessment was based on the mRNA load of the SPGE probe of about 2.1 ng. The number of copies of the investigated genes changed spatially along the length of primary roots. The expression level of all genes differed significantly at each sample position. Among the examined genes ACT7 expression was most even along the root. UBQ was highest at the tip and root-shoot junction (RS). TUB and GLK showed a basipetal gradient. The temporal expression of UBQ was highest in the MZ 9 h after primary root emergence and higher than at any other sample position. Expressions of GLK in EZ and RS increased gradually over time. SPGE extraction is the result of oligo-dT and oligo-dA hybridization and the results illustrate that SPGE can be used for gene expression profiling at high spatial and temporal resolution. SPGE needles can be used within two weeks when stored at 4 °C. Our data indicate that gene expression studies that are based on the entire root miss important differences in gene expression that SPGE is able to resolve for example growth adjustments during gravitropism.

  20. Transgenic Citrus Expressing an Arabidopsis NPR1 Gene Exhibit Enhanced Resistance against Huanglongbing (HLB; Citrus Greening).

    Science.gov (United States)

    Dutt, Manjul; Barthe, Gary; Irey, Michael; Grosser, Jude

    2015-01-01

    Commercial sweet orange cultivars lack resistance to Huanglongbing (HLB), a serious phloem limited bacterial disease that is usually fatal. In order to develop sustained disease resistance to HLB, transgenic sweet orange cultivars 'Hamlin' and 'Valencia' expressing an Arabidopsis thaliana NPR1 gene under the control of a constitutive CaMV 35S promoter or a phloem specific Arabidopsis SUC2 (AtSUC2) promoter were produced. Overexpression of AtNPR1 resulted in trees with normal phenotypes that exhibited enhanced resistance to HLB. Phloem specific expression of NPR1 was equally effective for enhancing disease resistance. Transgenic trees exhibited reduced diseased severity and a few lines remained disease-free even after 36 months of planting in a high-disease pressure field site. Expression of the NPR1 gene induced expression of several native genes involved in the plant defense signaling pathways. The AtNPR1 gene being plant derived can serve as a component for the development of an all plant T-DNA derived consumer friendly GM tree.

  1. Transgenic Citrus Expressing an Arabidopsis NPR1 Gene Exhibit Enhanced Resistance against Huanglongbing (HLB; Citrus Greening.

    Directory of Open Access Journals (Sweden)

    Manjul Dutt

    Full Text Available Commercial sweet orange cultivars lack resistance to Huanglongbing (HLB, a serious phloem limited bacterial disease that is usually fatal. In order to develop sustained disease resistance to HLB, transgenic sweet orange cultivars 'Hamlin' and 'Valencia' expressing an Arabidopsis thaliana NPR1 gene under the control of a constitutive CaMV 35S promoter or a phloem specific Arabidopsis SUC2 (AtSUC2 promoter were produced. Overexpression of AtNPR1 resulted in trees with normal phenotypes that exhibited enhanced resistance to HLB. Phloem specific expression of NPR1 was equally effective for enhancing disease resistance. Transgenic trees exhibited reduced diseased severity and a few lines remained disease-free even after 36 months of planting in a high-disease pressure field site. Expression of the NPR1 gene induced expression of several native genes involved in the plant defense signaling pathways. The AtNPR1 gene being plant derived can serve as a component for the development of an all plant T-DNA derived consumer friendly GM tree.

  2. Increased MCP-1 gene expression in monocytes of severe OSA patients and under intermittent hypoxia.

    Science.gov (United States)

    Chuang, Li-Pang; Chen, Ning-Hung; Lin, Yuling; Ko, Wen-Shan; Pang, Jong-Hwei S

    2016-03-01

    Obstructive sleep apnea (OSA) is known to be a risk factor of coronary artery disease. Monocyte chemoattractant protein-1 (MCP-1), as a critical factor for monocyte infiltration, is known to play a role in the development of atherosclerosis. This study aimed to investigate the effect of intermittent hypoxia, the hallmark of OSA, on the MCP-1 expression of monocytes. Peripheral blood was sampled from 61 adults enrolled for suspected OSA. RNA was prepared from the isolated monocytes for the analysis of MCP-1. The effect of in vitro intermittent hypoxia on the regulation and function of MCP-1 was investigated on THP-1 monocytic cells and human monocytes. The mRNA and secreted protein levels were investigated by RT/real-time PCR and enzyme-linked immunosorbent assay, respectively. Monocytic MCP-1 gene expression was found to be increased significantly in severe OSA patients. In vitro intermittent hypoxia was demonstrated to increase the mRNA and protein expression levels of MCP-1 dose- and time-dependently in THP-1 monocytic cells. The MCP-1 mRNA expression in monocytes isolated from OSA patient was induced to a much higher level compared to that from normal control. Pre-treatment with inhibitor for p42/44 MAPK or p38 MAPK suppressed the activation of MCP-1 expression by intermittent hypoxia. This is the first study to demonstrate the increase of MCP-1 gene expression in monocytes of severe OSA patients. In addition, monocytic MCP-1 gene expression can be induced under intermittent hypoxia.

  3. Sirtuin 1 stimulates the proliferation and the expression of glycolysis genes in pancreatic neoplastic lesions.

    Science.gov (United States)

    Pinho, Andreia V; Mawson, Amanda; Gill, Anthony; Arshi, Mehreen; Warmerdam, Max; Giry-Laterriere, Marc; Eling, Nils; Lie, Triyana; Kuster, Evelyne; Camargo, Simone; Biankin, Andrew V; Wu, Jianmin; Rooman, Ilse

    2016-11-15

    Metabolic reprogramming is a feature of neoplasia and tumor growth. Sirtuin 1 (SIRT1) is a lysine deacetylase of multiple targets including metabolic regulators such as p53. SIRT1 regulates metaplasia in the pancreas. Nevertheless, it is unclear if SIRT1 affects the development of neoplastic lesions and whether metabolic gene expression is altered.To assess neoplastic lesion development, mice with a pancreas-specific loss of Sirt1 (Pdx1-Cre;Sirt1-lox) were bred into a KrasG12D mutant background (KC) that predisposes to the development of pancreatic intra-epithelial neoplasia (PanIN) and ductal adenocarcinoma (PDAC). Similar grade PanIN lesions developed in KC and KC;Sirt1-lox mice but specifically early mucinous PanINs occupied 40% less area in the KC;Sirt1-lox line, attributed to reduced proliferation. This was accompanied by reduced expression of proteins in the glycolysis pathway, such as GLUT1 and GAPDH.The stimulatory effect of SIRT1 on proliferation and glycolysis gene expression was confirmed in a human PDAC cell line. In resected PDAC samples, higher proliferation and expression of glycolysis genes correlated with poor patient survival. SIRT1 expression per se was not prognostic but low expression of Cell Cycle and Apoptosis Regulator 2 (CCAR2), a reported SIRT1 inhibitor, corresponded to poor patient survival.These findings open perspectives for novel targeted therapies in pancreatic cancer.

  4. Sirtuin 1 stimulates the proliferation and the expression of glycolysis genes in pancreatic neoplastic lesions

    Science.gov (United States)

    Pinho, Andreia V.; Mawson, Amanda; Gill, Anthony; Arshi, Mehreen; Warmerdam, Max; Giry-Laterriere, Marc; Eling, Nils; Lie, Triyana; Kuster, Evelyne; Camargo, Simone; Biankin, Andrew V.; Wu, Jianmin; Rooman, Ilse

    2016-01-01

    Metabolic reprogramming is a feature of neoplasia and tumor growth. Sirtuin 1 (SIRT1) is a lysine deacetylase of multiple targets including metabolic regulators such as p53. SIRT1 regulates metaplasia in the pancreas. Nevertheless, it is unclear if SIRT1 affects the development of neoplastic lesions and whether metabolic gene expression is altered. To assess neoplastic lesion development, mice with a pancreas-specific loss of Sirt1 (Pdx1-Cre;Sirt1-lox) were bred into a KrasG12D mutant background (KC) that predisposes to the development of pancreatic intra-epithelial neoplasia (PanIN) and ductal adenocarcinoma (PDAC). Similar grade PanIN lesions developed in KC and KC;Sirt1-lox mice but specifically early mucinous PanINs occupied 40% less area in the KC;Sirt1-lox line, attributed to reduced proliferation. This was accompanied by reduced expression of proteins in the glycolysis pathway, such as GLUT1 and GAPDH. The stimulatory effect of SIRT1 on proliferation and glycolysis gene expression was confirmed in a human PDAC cell line. In resected PDAC samples, higher proliferation and expression of glycolysis genes correlated with poor patient survival. SIRT1 expression per se was not prognostic but low expression of Cell Cycle and Apoptosis Regulator 2 (CCAR2), a reported SIRT1 inhibitor, corresponded to poor patient survival. These findings open perspectives for novel targeted therapies in pancreatic cancer. PMID:27494892

  5. Time-Course Gene Set Analysis for Longitudinal Gene Expression Data.

    Directory of Open Access Journals (Sweden)

    Boris P Hejblum

    2015-06-01

    Full Text Available Gene set analysis methods, which consider predefined groups of genes in the analysis of genomic data, have been successfully applied for analyzing gene expression data in cross-sectional studies. The time-course gene set analysis (TcGSA introduced here is an extension of gene set analysis to longitudinal data. The proposed method relies on random effects modeling with maximum likelihood estimates. It allows to use all available repeated measurements while dealing with unbalanced data due to missing at random (MAR measurements. TcGSA is a hypothesis driven method that identifies a priori defined gene sets with significant expression variations over time, taking into account the potential heterogeneity of expression within gene sets. When biological conditions are compared, the method indicates if the time patterns of gene sets significantly differ according to these conditions. The interest of the method is illustrated by its application to two real life datasets: an HIV therapeutic vaccine trial (DALIA-1 trial, and data from a recent study on influenza and pneumococcal vaccines. In the DALIA-1 trial TcGSA revealed a significant change in gene expression over time within 69 gene sets during vaccination, while a standard univariate individual gene analysis corrected for multiple testing as well as a standard a Gene Set Enrichment Analysis (GSEA for time series both failed to detect any significant pattern change over time. When applied to the second illustrative data set, TcGSA allowed the identification of 4 gene sets finally found to be linked with the influenza vaccine too although they were found to be associated to the pneumococcal vaccine only in previous analyses. In our simulation study TcGSA exhibits good statistical properties, and an increased power compared to other approaches for analyzing time-course expression patterns of gene sets. The method is made available for the community through an R package.

  6. TEAD1-dependent expression of the FoxO3a gene in mouse skeletal muscle

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    Xu Xuewen

    2011-01-01

    Full Text Available Abstract Background TEAD1 (TEA domain family member 1 is constitutively expressed in cardiac and skeletal muscles. It acts as a key molecule of muscle development, and trans-activates multiple target genes involved in cell proliferation and differentiation pathways. However, its target genes in skeletal muscles, regulatory mechanisms and networks are unknown. Results In this paper, we have identified 136 target genes regulated directly by TEAD1 in skeletal muscle using integrated analyses of ChIP-on-chip. Most of the targets take part in the cell process, physiology process, biological regulation metabolism and development process. The targets also play an important role in MAPK, mTOR, T cell receptor, JAK-STAT, calcineurin and insulin signaling pathways. TEAD1 regulates foxo3a transcription through binding to the M-CAT element in foxo3a promoter, demonstrated with independent ChIP-PCR, EMSA and luciferase reporter system assay. In addition, results of over-expression and inhibition experiments suggest that foxo3a is positively regulated by TEAD1. Conclusions Our present data suggests that TEAD1 plays an important role in the regulation of gene expression and different signaling pathways may co-operate with each other mediated by TEAD1. We have preliminarily concluded that TEAD1 may regulate FoxO3a expression through calcineurin/MEF2/NFAT and IGF-1/PI3K/AKT signaling pathways in skeletal muscles. These findings provide important clues for further analysis of the role of FoxO3a gene in the formation and transformation of skeletal muscle fiber types.

  7. The expression of propionicin PLG-1 gene (plg-1) by lactic starters.

    Science.gov (United States)

    Mohamed, Sameh E; Tahoun, Mahmoud K

    2015-05-01

    Propionicin PLG-1 is a bacteriocin produced by Propionibacterium thoenii P127. Such bacteriocin inhibits wide range of food-borne pathogens such as pathogenic Escherichia coli, Pseudomonas aeruginosa, Vibrio parahaemolyticus, Yersinia enterocolitica and a strain of Corynebacterium sp. In the present study, plg-1 gene expressing propionicin PLG-1 was isolated, sequenced for the first time and the resulting sequence was analysed using several web-based bioinformatics programs. The PCR product containing plg-1 gene was transferred to different lactic acid bacterial (LAB) strains using pLEB590 as a cloning vector to give the modified vector pLEBPLG-1. LAB transformants showed an antimicrobial activity against Esch. coli DH5α (most affected strain), Listeria monocytogenes 18116, and Salmonella enterica 25566 as model pathogenic strains. Such LAB transformants can be used in dairy industry to control the food-borne pathogens that are largely distributed worldwide and to feed schoolchildren in the poor countries where dangerous epidemic diseases and diarrhoea prevail.

  8. Genomic organization, expression, and chromosome localization of a third aurora-related kinase gene, Aie1.

    Science.gov (United States)

    Hu, H M; Chuang, C K; Lee, M J; Tseng, T C; Tang, T K

    2000-11-01

    We previously reported two novel testis-specific serine/threonine kinases, Aie1 (mouse) and AIE2 (human), that share high amino acid identities with the kinase domains of fly aurora and yeast Ipl1. Here, we report the entire intron-exon organization of the Aie1 gene and analyze the expression patterns of Aie1 mRNA during testis development. The mouse Aie1 gene spans approximately 14 kb and contains seven exons. The sequences of the exon-intron boundaries of the Aie1 gene conform to the consensus sequences (GT/AG) of the splicing donor and acceptor sites of most eukaryotic genes. Comparative genomic sequencing revealed that the gene structure is highly conserved between mouse Aie1 and human AIE2. However, much less homology was found in the sequence outside the kinase-coding domains. The Aie1 locus was mapped to mouse chromosome 7A2-A3 by fluorescent in situ hybridization. Northern blot analysis indicates that Aie1 mRNA likely is expressed at a low level on day 14 and reaches its plateau on day 21 in the developing postnatal testis. RNA in situ hybridization indicated that the expression of the Aie1 transcript was restricted to meiotically active germ cells, with the highest levels detected in spermatocytes at the late pachytene stage. These findings suggest that Aie1 plays a role in spermatogenesis.

  9. Comprehensive analysis of gene expression patterns of hedgehog-related genes

    Directory of Open Access Journals (Sweden)

    Baillie David

    2006-10-01

    Full Text Available Abstract Background The Caenorhabditis elegans genome encodes ten proteins that share sequence similarity with the Hedgehog signaling molecule through their C-terminal autoprocessing Hint/Hog domain. These proteins contain novel N-terminal domains, and C. elegans encodes dozens of additional proteins containing only these N-terminal domains. These gene families are called warthog, groundhog, ground-like and quahog, collectively called hedgehog (hh-related genes. Previously, the expression pattern of seventeen genes was examined, which showed that they are primarily expressed in the ectoderm. Results With the completion of the C. elegans genome sequence in November 2002, we reexamined and identified 61 hh-related ORFs. Further, we identified 49 hh-related ORFs in C. briggsae. ORF analysis revealed that 30% of the genes still had errors in their predictions and we improved these predictions here. We performed a comprehensive expression analysis using GFP fusions of the putative intergenic regulatory sequence with one or two transgenic lines for most genes. The hh-related genes are expressed in one or a few of the following tissues: hypodermis, seam cells, excretory duct and pore cells, vulval epithelial cells, rectal epithelial cells, pharyngeal muscle or marginal cells, arcade cells, support cells of sensory organs, and neuronal cells. Using time-lapse recordings, we discovered that some hh-related genes are expressed in a cyclical fashion in phase with molting during larval development. We also generated several translational GFP fusions, but they did not show any subcellular localization. In addition, we also studied the expression patterns of two genes with similarity to Drosophila frizzled, T23D8.1 and F27E11.3A, and the ortholog of the Drosophila gene dally-like, gpn-1, which is a heparan sulfate proteoglycan. The two frizzled homologs are expressed in a few neurons in the head, and gpn-1 is expressed in the pharynx. Finally, we compare the

  10. Gene Expression Analyses of HER-2/neu and ESR1 in Patients with Breast Cancer

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    Omid Kheyri Nadergoli

    2017-10-01

    Full Text Available ABSTRACT Background: Her-2 and ESR1 genes, that interact in the cell signaling pathway, are the most important molecular markers of breast cancer, which have been amplified or overexpressed in 30% and 70%, respectively. This study was performed to evaluate the gene expression levels of Her-2 and ESR1 genes in tumor cells and its adjacent normal tissue of breast cancer patients and compared them whit clinical-pathological features. Methods: In total, 80 tissue specimens from 40 patients, with an average age of 48.47 years, were examined by Real-time PCR technique, and ultimately evaluated the expression level of Her-2 and ESR1genes. The data were analyzed by REST 2009 V2.0.13 statistical software. Results: HER2 and ESR1 overexpression was identified in 19 (48% and 12 (30% of 40 patients respectively, which was higher and lower than that recorded in international statistics, respectively. ESR1 overexpression was associated with Stage 3A and lymph node involvement 2 (N2 (P = 0.04 and P = 0.047, respectively. No significant correlation was observed between the expression of HER2 and ESR1 and other clinical-pathological features, however, the relative differences were identified in the expression levels of genes between main group and groups that were classified according to the clinical-pathological features and age. Conclusions: Overexpression of Her-2 and ESR1 genes in the patients of our study are higher and lower than international statistics, respectively, indicating the differences in genetic, environmental and ethnic factors that involved in the developing of breast cancer.

  11. Multiple sclerosis risk variant HLA-DRB1*1501 associates with high expression of DRB1 gene in different human populations.

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    Antonio Alcina

    Full Text Available The human leukocyte antigen (HLA DRB1*1501 has been consistently associated with multiple sclerosis (MS in nearly all populations tested. This points to a specific antigen presentation as the pathogenic mechanism though this does not fully explain the disease association. The identification of expression quantitative trait loci (eQTL for genes in the HLA locus poses the question of the role of gene expression in MS susceptibility. We analyzed the eQTLs in the HLA region with respect to MS-associated HLA-variants obtained from genome-wide association studies (GWAS. We found that the Tag of DRB1*1501, rs3135388 A allele, correlated with high expression of DRB1, DRB5 and DQB1 genes in a Caucasian population. In quantitative terms, the MS-risk AA genotype carriers of rs3135388 were associated with 15.7-, 5.2- and 8.3-fold higher expression of DQB1, DRB5 and DRB1, respectively, than the non-risk GG carriers. The haplotype analysis of expression-associated variants in a Spanish MS cohort revealed that high expression of DRB1 and DQB1 alone did not contribute to the disease. However, in Caucasian, Asian and African American populations, the DRB1*1501 allele was always highly expressed. In other immune related diseases such as type 1 diabetes, inflammatory bowel disease, ulcerative colitis, asthma and IgA deficiency, the best GWAS-associated HLA SNPs were also eQTLs for different HLA Class II genes. Our data suggest that the DR/DQ expression levels, together with specific structural properties of alleles, seem to be the causal effect in MS and in other immunopathologies rather than specific antigen presentation alone.

  12. Soybean DREB1/CBF-type transcription factors function in heat and drought as well as cold stress-responsive gene expression.

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    Kidokoro, Satoshi; Watanabe, Keitaro; Ohori, Teppei; Moriwaki, Takashi; Maruyama, Kyonoshin; Mizoi, Junya; Myint Phyu Sin Htwe, Nang; Fujita, Yasunari; Sekita, Sachiko; Shinozaki, Kazuo; Yamaguchi-Shinozaki, Kazuko

    2015-02-01

    Soybean (Glycine max) is a globally important crop, and its growth and yield are severely reduced by abiotic stresses, such as drought, heat, and cold. The cis-acting element DRE (dehydration-responsive element)/CRT plays an important role in activating gene expression in response to these stresses. The Arabidopsis DREB1/CBF genes that encode DRE-binding proteins function as transcriptional activators in the cold stress responsive gene expression. In this study, we identified 14 DREB1-type transcription factors (GmDREB1s) from a soybean genome database. The expression of most GmDREB1 genes in soybean was strongly induced by a variety of abiotic stresses, such as cold, drought, high salt, and heat. The GmDREB1 proteins activated transcription via DREs (dehydration-responsive element) in Arabidopsis and soybean protoplasts. Transcriptome analyses using transgenic Arabidopsis plants overexpressing GmDREB1s indicated that many of the downstream genes are cold-inducible and overlap with those of Arabidopsis DREB1A. We then comprehensively analyzed the downstream genes of GmDREB1B;1, which is closely related to DREB1A, using a transient expression system in soybean protoplasts. The expression of numerous genes induced by various abiotic stresses were increased by overexpressing GmDREB1B;1 in soybean, and DREs were the most conserved element in the promoters of these genes. The downstream genes of GmDREB1B;1 included numerous soybean-specific stress-inducible genes that encode an ABA receptor family protein, GmPYL21, and translation-related genes, such as ribosomal proteins. We confirmed that GmDREB1B;1 directly activates GmPYL21 expression and enhances ABRE-mediated gene expression in an ABA-independent manner. These results suggest that GmDREB1 proteins activate the expression of numerous soybean-specific stress-responsive genes under diverse abiotic stress conditions. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

  13. The functional landscape of mouse gene expression

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    Zhang Wen

    2004-12-01

    Full Text Available Abstract Background Large-scale quantitative analysis of transcriptional co-expression has been used to dissect regulatory networks and to predict the functions of new genes discovered by genome sequencing in model organisms such as yeast. Although the idea that tissue-specific expression is indicative of gene function in mammals is widely accepted, it has not been objectively tested nor compared with the related but distinct strategy of correlating gene co-expression as a means to predict gene function. Results We generated microarray expression data for nearly 40,000 known and predicted mRNAs in 55 mouse tissues, using custom-built oligonucleotide arrays. We show that quantitative transcriptional co-expression is a powerful predictor of gene function. Hundreds of functional categories, as defined by Gene Ontology 'Biological Processes', are associated with characteristic expression patterns across all tissues, including categories that bear no overt relationship to the tissue of origin. In contrast, simple tissue-specific restriction of expression is a poor predictor of which genes are in which functional categories. As an example, the highly conserved mouse gene PWP1 is widely expressed across different tissues but is co-expressed with many RNA-processing genes; we show that the uncharacterized yeast homolog of PWP1 is required for rRNA biogenesis. Conclusions We conclude that 'functional genomics' strategies based on quantitative transcriptional co-expression will be as fruitful in mammals as they have been in simpler organisms, and that transcriptional control of mammalian physiology is more modular than is generally appreciated. Our data and analyses provide a public resource for mammalian functional genomics.

  14. Kaempferol stimulates gene expression of low-density lipoprotein receptor through activation of Sp1 in cultured hepatocytes

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    Ochiai, Ayasa; Miyata, Shingo; Iwase, Masamori; Shimizu, Makoto; Inoue, Jun; Sato, Ryuichiro

    2016-01-01

    A high level of plasma low-density lipoprotein (LDL) cholesterol is considered a risk factor for atherosclerosis. Because the hepatic LDL receptor (LDLR) is essential for clearing plasma LDL cholesterol, activation of LDLR is a promising therapeutic target for patients with atherosclerotic disease. Here we demonstrated how the flavonoid kaempferol stimulated the gene expression and activity of LDLR in HepG2 cells. The kaempferol-mediated stimulation of LDLR gene expression was completely inhibited by knockdown of Sp1 gene expression. Treatment of HepG2 cells with kaempferol stimulated the recruitment of Sp1 to the promoter region of the LDLR gene, as well as the phosphorylation of Sp1 on Thr-453 and Thr-739. Moreover, these kaempferol-mediated processes were inhibited in the presence of U0126, an ERK pathway inhibitor. These results suggest that kaempferol may increase the activity of Sp1 through stimulation of Sp1 phosphorylation by ERK1/2 and subsequent induction of LDLR expression and activity. PMID:27109240

  15. Evaluation of suitable reference genes for gene expression studies in bovine muscular tissue

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    Dunner Susana

    2008-09-01

    Full Text Available Abstract Background Real-time reverse transcriptase quantitative polymerase chain reaction (real-time RTqPCR is a technique used to measure mRNA species copy number as a way to determine key genes involved in different biological processes. However, the expression level of these key genes may vary among tissues or cells not only as a consequence of differential expression but also due to different factors, including choice of reference genes to normalize the expression levels of the target genes; thus the selection of reference genes is critical for expression studies. For this purpose, ten candidate reference genes were investigated in bovine muscular tissue. Results The value of stability of ten candidate reference genes included in three groups was estimated: the so called 'classical housekeeping' genes (18S, GAPDH and ACTB, a second set of genes used in expression studies conducted on other tissues (B2M, RPII, UBC and HMBS and a third set of novel genes (SF3A1, EEF1A2 and CASC3. Three different statistical algorithms were used to rank the genes by their stability measures as produced by geNorm, NormFinder and Bestkeeper. The three methods tend to agree on the most stably expressed genes and the least in muscular tissue. EEF1A2 and HMBS followed by SF3A1, ACTB, and CASC3 can be considered as stable reference genes, and B2M, RPII, UBC and GAPDH would not be appropriate. Although the rRNA-18S stability measure seems to be within the range of acceptance, its use is not recommended because its synthesis regulation is not representative of mRNA levels. Conclusion Based on geNorm algorithm, we propose the use of three genes SF3A1, EEF1A2 and HMBS as references for normalization of real-time RTqPCR in muscle expression studies.

  16. A Single Dose of LSD Does Not Alter Gene Expression of the Serotonin 2A Receptor Gene (HTR2A) or Early Growth Response Genes (EGR1-3) in Healthy Subjects

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    Dolder, Patrick C.; Grünblatt, Edna; Müller, Felix; Borgwardt, Stefan J.; Liechti, Matthias E.

    2017-01-01

    Rationale: Renewed interest has been seen in the use of lysergic acid diethylamide (LSD) in psychiatric research and practice. The repeated use of LSD leads to tolerance that is believed to result from serotonin (5-HT) 5-HT2A receptor downregulation. In rats, daily LSD administration for 4 days decreased frontal cortex 5-HT2A receptor binding. Additionally, a single dose of LSD acutely increased expression of the early growth response genes EGR1 and EGR2 in rat and mouse brains through 5-HT2A receptor stimulation. No human data on the effects of LSD on gene expression has been reported. Therefore, we investigated the effects of single-dose LSD administration on the expression of the 5-HT2A receptor gene (HTR2A) and EGR1-3 genes. Methods: mRNA expression levels were analyzed in whole blood as a peripheral biomarker in 15 healthy subjects before and 1.5 and 24 h after the administration of LSD (100 μg) and placebo in a randomized, double-blind, placebo-controlled, cross-over study. Results: LSD did not alter the expression of the HTR2A or EGR1-3 genes 1.5 and 24 h after administration compared with placebo. Conclusion: No changes were observed in the gene expression of LSD’s primary target receptor gene or genes that are implicated in its downstream effects. Remaining unclear is whether chronic LSD administration alters gene expression in humans. PMID:28701958

  17. Validation of suitable reference genes for quantitative gene expression analysis in Panax ginseng

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    Meizhen eWang

    2016-01-01

    Full Text Available Reverse transcription-qPCR (RT-qPCR has become a popular method for gene expression studies. Its results require data normalization by housekeeping genes. No single gene is proved to be stably expressed under all experimental conditions. Therefore, systematic evaluation of reference genes is necessary. With the aim to identify optimum reference genes for RT-qPCR analysis of gene expression in different tissues of Panax ginseng and the seedlings grown under heat stress, we investigated the expression stability of eight candidate reference genes, including elongation factor 1-beta (EF1-β, elongation factor 1-gamma (EF1-γ, eukaryotic translation initiation factor 3G (IF3G, eukaryotic translation initiation factor 3B (IF3B, actin (ACT, actin11 (ACT11, glyceraldehyde-3-phosphate dehydrogenase (GAPDH and cyclophilin ABH-like protein (CYC, using four widely used computational programs: geNorm, Normfinder, BestKeeper, and the comparative ΔCt method. The results were then integrated using the web-based tool RefFinder. As a result, EF1-γ, IF3G and EF1-β were the three most stable genes in different tissues of P. ginseng, while IF3G, ACT11 and GAPDH were the top three-ranked genes in seedlings treated with heat. Using three better reference genes alone or in combination as internal control, we examined the expression profiles of MAR, a multiple function-associated mRNA-like non-coding RNA (mlncRNA in P. ginseng. Taken together, we recommended EF1-γ/IF3G and IF3G/ACT11 as the suitable pair of reference genes for RT-qPCR analysis of gene expression in different tissues of P. ginseng and the seedlings grown under heat stress, respectively. The results serve as a foundation for future studies on P. ginseng functional genomics.

  18. Expression study of CYP19A1 gene in a cohort of Iranian leiomyoma patients

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    Leila Emrahi

    2018-07-01

    Full Text Available Background: CYP19A1 gene encodes aromatase, a microsomal key enzyme that catalyzes the synthesis of estrogens from androgens. Accumulating evidence has revealed that aromatase plays an important role in the pathogenesis of leiomyoma through increasing local concentration of estrogens. In this study, we examined the levels of CYP19A1 mRNA to determine the impact of aromatase overexpression in uterine leiomyoma growth. Subjects and methods: Tissues were obtained via myomectomy or hysterectomy from 30 patients. Total RNA was extracted and cDNA was synthesized from each frozen sample. Using SYBR Green dye, Real-time PCR assay was performed by sequence-specific primers. Relative mRNA expression was normalized to the mean of the Ct values determined for HPRT1. Gene expression ratio in each sample was determined relative to the mean ΔCt value of tumor-free margin samples. Results: PCR efficiencies for amplification reactions of HPRT1, and CYP19A genes were calculated as 0.93 and 0.96, respectively. Regression coefficients (R for standard curves were above 0.90. The obtained data revealed that the mean fold increase of CYP19A1 gene expression in leiomyoma samples relative to normal samples was 3.551 (95% CI: 0.04–6.64, S.E., 0.29–5.35. Conclusions: Our results were in accordance with previous studies and imply that up-regulation of CYP19A1 is correlated with the pathogenesis of leiomyoma tumors. We also observed that expression level of CYP19A1 was not linked to the tumor size or localization. It can be concluded that; up-regulation of aromatase is a key factor in the initiation of tumor development as well as tumor growth. Keywords: Leiomyoma, CYP19A1, Real-time PCR, Gene expression study

  19. Investigation of Fasciculation and Elongation Protein ζ-1 (FEZ1 in Peripheral Blood Reveals Differences in Gene Expression in Patients with Schizophrenia

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    Vachev T.I.

    2015-06-01

    Full Text Available Schizophrenia (SZ is a chronic neuropsychiatric disorder characterized by affective, neuromorphological and cognitive impairment, deteriorated social functioning and psychosis with underlying molecular abnormalities, including gene expression changes. Observations have suggested that fasciculation and elongation protein ζ-1 (FEZ1 may be implicated in the pathogenesis of schizophrenia. Nevertheless, our current knowledge of the expression of FEZ1 in peripheral blood of schizophrenia patients remains unclear. The purpose of this study was to identify the characteristic gene expression patterns of FEZ1 in peripheral blood samples from schizophrenia patients. We performed quantitative reverse-transcriptase (qRT-PCR analysis using peripheral blood from drug-free schizophrenia patients (n = 29 and age and gender-matched general population controls (n = 24. For the identification of FEZ1 gene expression patterns, we applied a comparative threshold cycle (CT method. A statistically significant difference of FEZ1 mRNA level was revealed in schizophrenia subjects compared to healthy controls (p = 0.0034. To the best of our knowledge, this study is the first describing a down-regulation of FEZ1 gene expression in peripheral blood of patients with schizophrenia. Our results suggested a possible functional role of FEZ1 in the pathogenesis of schizophrenia and confirmed the utility of peripheral blood samples for molecular profiling of psychiatric disorders including schizophrenia. The current study describes FEZ1 gene expression changes in peripheral blood of patients with schizophrenia with significantly down-regulation of FEZ1 mRNA. Thus, our results provide support for a model of SZ pathogenesis that includes the effects of FEZ1 expression.

  20. Validation of Tuba1a as Appropriate Internal Control for Normalization of Gene Expression Analysis during Mouse Lung Development

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    Aditi Mehta

    2015-02-01

    Full Text Available The expression ratio between the analysed gene and an internal control gene is the most widely used normalization method for quantitative RT-PCR (qRT-PCR expression analysis. The ideal reference gene for a specific experiment is the one whose expression is not affected by the different experimental conditions tested. In this study, we validate the applicability of five commonly used reference genes during different stages of mouse lung development. The stability of expression of five different reference genes (Tuba1a, Actb Gapdh, Rn18S and Hist4h4 was calculated within five experimental groups using the statistical algorithm of geNorm software. Overall, Tuba1a showed the least variability in expression among the different stages of lung development, while Hist4h4 and Rn18S showed the maximum variability in their expression. Expression analysis of two lung specific markers, surfactant protein C (SftpC and Clara cell-specific 10 kDA protein (Scgb1a1, normalized to each of the five reference genes tested here, confirmed our results and showed that incorrect reference gene choice can lead to artefacts. Moreover, a combination of two internal controls for normalization of expression analysis during lung development will increase the accuracy and reliability of results.

  1. Molecular Cloning and Expression Analysis of a Hexokinase Gene, MdHXK1 in Apple

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    Jin Zhao

    2016-03-01

    Full Text Available A hexokinase gene named MdHXK1 (MDP0000309677 was cloned from ‘Gala’ apple (Malus × domestica Borkh.. Sequence analysis showed that the MdHXK1 gene was 1 497 bp long and encoded 499 amino acids. The predicted molecular mass of this protein was 54.05 kD, and the pI was 5.76. A phylogenetic tree indicated apple MdHXK1 exhibited the highest sequence similarity to Pyrus bretschneideri PbHXK1. Analysis of the functional domain showed that the MdHXK1 protein included two conserved kinase domains. The prediction of subcellular localization suggested that the MdHXK1 protein was mainly localized in the cytoplasm. There was an indication that MdHXK1 existed as one copy in the apple genome by Southern blotting. Silico analysis suggested that the promoter sequence contained several typical cis-acting elements, including defense, sugar signaling and phytohormone responsive elements. Quantitative real-time PCR analysis demonstrated that the MdHXK1 gene was mainly expressed in stem and flower tissues. During the development of apple fruits, the expression of the MdHXK1 gene initially increased and then decreased. The changes on Glc phosphorylation relative activity and glucose concentration showed the same trend. In addition, the expression of this gene was induced by salt stress, low temperature, and abscisic acid (ABA. Finally, we obtained and purified the fused MdHXK1 protein by recombinant prokaryotic expression. Studies have demonstrated that MdHXK1 may participate in sugar metabolism in apple fruits. Enzyme encoded by MdHXK1 is a key factor in the mediation of sugar accumulation. Recently, researchers on hexokinase at home and abroad mainly focused on model plants, such as Arabidopsis, tobacco and rice, but orchard fruit like apple were underresearched. Our research established the foundation for the further study of the functions of MdHXK1.

  2. Expression of Sox genes in tooth development.

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    Kawasaki, Katsushige; Kawasaki, Maiko; Watanabe, Momoko; Idrus, Erik; Nagai, Takahiro; Oommen, Shelly; Maeda, Takeyasu; Hagiwara, Nobuko; Que, Jianwen; Sharpe, Paul T; Ohazama, Atsushi

    2015-01-01

    Members of the Sox gene family play roles in many biological processes including organogenesis. We carried out comparative in situ hybridization analysis of seventeen sox genes (Sox1-14, 17, 18, 21) during murine odontogenesis from the epithelial thickening to the cytodifferentiation stages. Localized expression of five Sox genes (Sox6, 9, 13, 14 and 21) was observed in tooth bud epithelium. Sox13 showed restricted expression in the primary enamel knots. At the early bell stage, three Sox genes (Sox8, 11, 17 and 21) were expressed in pre-ameloblasts, whereas two others (Sox5 and 18) showed expression in odontoblasts. Sox genes thus showed a dynamic spatio-temporal expression during tooth development.

  3. Genome-Wide Gene Expression Disturbance by Single A1/C1 Chromosome Substitution in Brassica rapa Restituted From Natural B. napus

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    Bin Zhu

    2018-03-01

    Full Text Available Alien chromosome substitution (CS lines are treated as vital germplasms for breeding and genetic mapping. Previously, a whole set of nine Brassica rapa-oleracea monosonic alien addition lines (MAALs, C1-C9 was established in the background of natural B. napus genotype “Oro,” after the restituted B. rapa (RBR for Oro was realized. Herein, a monosomic substitution line with one alien C1 chromosome (Cs1 in the RBR complement was selected in the progenies of MAAL C1 and RBR, by the PCR amplification of specific gene markers and fluorescence in situ hybridization. Cs1 exhibited the whole plant morphology similar to RBR except for the defective stamens without fertile pollen grains, but it produced some seeds and progeny plants carrying the C1 chromosome at high rate besides those without the alien chromosome after pollinated by RBR. The viability of the substitution and its progeny for the RBR diploid further elucidated the functional compensation between the chromosome pairs with high homoeology. To reveal the impact of such aneuploidy on genome-wide gene expression, the transcriptomes of MAAL C1, Cs1 and euploid RBR were analyzed. Compared to RBR, Cs1 had sharply reduced gene expression level across chromosome A1, demonstrating the loss of one copy of A1 chromosome. Both additional chromosome C1 in MAAL and substitutional chromosome C1 in Cs1 caused not only cis-effect but also prevalent trans-effect differentially expressed genes. A dominant gene dosage effects prevailed among low expressed genes across chromosome A1 in Cs1, and moreover, dosage effects for some genes potentially contributed to the phenotype deviations. Our results provided novel insights into the transcriptomic perturbation and gene dosage effects on phenotype in CS related to one naturally evolved allopolyploid.

  4. Physical properties and biological effects of mineral trioxide aggregate mixed with methylcellulose and calcium chloride.

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    Lee, Bin-Na; Chun, Soo-Ji; Chang, Hoon-Sang; Hwang, Yun-Chan; Hwang, In-Nam; Oh, Won-Mann

    2017-01-01

    Methylcellulose (MC) is a chemical compound derived from cellulose. MTA mixed with MC reduces setting time and increases plasticity. This study assessed the influence of MC as an anti-washout ingredient and CaCl2 as a setting time accelerator on the physical and biological properties of MTA. Test materials were divided into 3 groups; Group 1(control): distilled water; Group 2: 1% MC/CaCl2; Group 3: 2% MC/CaCl2. Compressive strength, pH, flowability and cell viability were tested. The gene expression of bone sialoprotein (BSP) was detected by RT-PCR and real- time PCR. The expression of alkaline phosphatase (ALP) and mineralization behavior were evaluated using an ALP staining and an alizarin red staining. Compressive strength, pH, and cell viability of MTA mixed with MC/CaCl2 were not significantly different compared to the control group. The flowability of MTA with MC/CaCI2 has decreased significantly when compared to the control (pphysical and biological effect of MTA. It suggests that these cements may be useful as a root-end filling material.

  5. Aging-dependent DNA hypermethylation and gene expression of GSTM1 involved in T cell differentiation.

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    Yeh, Shu-Hui; Liu, Cheng-Ling; Chang, Ren-Chieh; Wu, Chih-Chiang; Lin, Chia-Hsueh; Yang, Kuender D

    2017-07-25

    This study investigated whether aging was associated with epigenetic changes of DNA hypermethylation on immune gene expression and lymphocyte differentiation. We screened CG sites of methylation in blood leukocytes from different age populations, picked up genes with age-related increase of CG methylation content more than 15%, and validated immune related genes with CG hypermethylation involved in lymphocyte differentiation in the aged population. We found that 12 genes (EXHX1、 IL-10、 TSP50、 GSTM1、SLC5A5、SPI1、F2R、LMO2、PTPN6、FGFR2、MMP9、MET) were associated with promoter or exon one DNA hypermethylation in the aged group. Two immune related genes, GSTM1 and LMO2, were chosen to validate its aging-related CG hypermethylation in different leukocytes. We are the first to validate that GSTM1_P266 and LMO2_E128 CG methylation contents in T lymphocytes but not polymorphonuclear cells (PMNs) or mononuclear cells (MNCs) were significantly increased in the aged population. The GSTM1 mRNA expression in T lymphocytes but not PMNs or MNCs was inversely associated with the GSTM1 CG hypermethylation levels in the aged population studied. Further studies showed that lower GSTM1 CG methylation content led to the higher GSTM1 mRNA expression in T cells and knockdown of GSTM1 mRNA expression decreased type 1 T helper cell (Th1) differentiation in Jurkat T cells and normal adult CD4 T cells. The GSTM1_P266 hypermethylation in the aged population associated with lower GSTM1 mRNA expression was involved in Th1 differentiation, highlighting that modulation of aging-associated GSTM1 methylation may be able to enhance T helper cell immunity in the elders.

  6. BRCA1 gene expression in relation to prognostic parameters of breast cancer

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    Manal Kamal

    2011-09-01

    Full Text Available The tumor suppressor gene, BRCA1 has been conferred to increase the susceptibility to breast cancer in younger women. This work studied the expression of BRCA1 (mRNA in women with breast cancer in relation to other prognostic parameters such as histological type and grade of cancer, hormone receptor status, human epidermal growth factor receptor 2 (HER2/neu and CA15-3. Thirty patients with positive family history of breast cancer and a control group of 20 healthy subjects were also included for the study. Ribonucleic acid (RNA extraction from breast cancer tissues was done (considered suitable for RNA extraction if 70% or more of the tissue section contained tumor and was followed by real-time reverse transcription polymerase chain reaction. BRCA1 expression was assessed and correlated with age, histological type and grade of breast cancer, estrogen and progesterone receptor (ER, PR status, HER2/neu expression and CA15-3 levels. The mean age of patients was 54.8 ± 10.49 years. Of the 30 breast cancer cases studied, the majority (77% was of high histological grade and the most common histological type was infiltrating ductal carcinoma (20 cases. ER expression was positive in 53.3% of breast cancers, while PR expression was positive in 50% of cancers. BRCA1 mRNA was found in 6 patient samples (20% of the breast cancer patients while the remaining 24 patients (80% showed negative BRCA1 mRNA expression as well as the control group. A positive significant relationship was demonstrated between BRCA1 (mRNA expression and high histological grade, negative estrogen and progesterone receptor status, and high levels of serum CA15-3. A significant negative correlation was found between BRCA1 mRNA expression and age (r = −0.683; p < 0.01. The study demonstrated lack of BRCA1 gene expression (mRNA in the majority of breast cancer cases and confirmed the relationship between BRCA1 expression and parameters that determine poor prognosis in breast cancer. The

  7. Taurine Transporter Gene Expression in Mononuclear Blood Cells of Type 1 Diabetes Patients.

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    Napoli, Zaleida; Seghieri, Giuseppe; Bianchi, Loria; Anichini, Roberto; De Bellis, Alessandra; Campesi, Ilaria; Carru, Ciriaco; Occhioni, Stefano; Zinellu, Angelo; Franconi, Flavia

    2016-01-01

    Taurine transporter gene expression (RNA-TauT) has a role in retinal cell function and is modulated in vitro and in vivo by hyperglycemia and/or oxidative stress. This study was aimed at testing whether RNA-TauT gene expression is modified in blood mononuclear peripheral cells (MPCs) of type 1 diabetic patients, is related to plasma markers of oxidative stress or endothelial dysfunction, or, finally, is related to presence of retinopathy. RNA-TauT was measured in MPCs by real-time PCR-analysis in 35 type 1 diabetic patients and in 33 age- and sex-matched controls, additionally measuring plasma and cell taurine and markers of oxidative stress and endothelial dysfunction. RNA-TauT, expressed as 2(-ΔΔCt), was significantly higher in MPCs of type 1 diabetic patients than in controls [median (interquartile range): 1.32(0.31) versus 1.00(0.15); P = 0.01]. In diabetic patients RNA-TauT was related to HbA1c (r = 0.42; P = 0.01) and inversely to plasma homocysteine (r = -0.39; P = 0.02) being additionally significantly higher in MPCs of patients without retinopathy [(n = 22); 1.36(0.34)] compared to those with retinopathy [(n = 13); 1.16(0.20)], independently from HbA1c or diabetes duration. RNA-TauT gene expression is significantly upregulated in MPCs of type 1 diabetes patients and is related to HbA1c levels and inversely to plasma homocysteine. Finally, in diabetes patients, RNA-TauT upregulation seems to be blunted in patients with retinopathy independently of their metabolic control or longer diabetes duration.

  8. Mechanism of Gene Expression of Arabidopsis Glutathione S-Transferase, AtGST1, and AtGST11 in Response to Aluminum Stress1

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    Ezaki, Bunichi; Suzuki, Masakatsu; Motoda, Hirotoshi; Kawamura, Masako; Nakashima, Susumu; Matsumoto, Hideaki

    2004-01-01

    The gene expression of two Al-induced Arabidopsis glutathione S-transferase genes, AtGST1 and AtGST11, was analyzed to investigate the mechanism underlying the response to Al stress. An approximately 1-kb DNA fragment of the 5′-upstream region of each gene was fused to a β-glucuronidase (GUS) reporter gene (pAtGST1::GUS and pAtGST11::GUS) and introduced into Arabidopsis ecotype Landsberg erecta. The constructed transgenic lines showed a time-dependent gene expression to a different degree in the root and/or leaf by Al stress. The pAtGST1::GUS gene was induced after a short Al treatment (maximum expression after a 2-h exposure), while the pAtGST11::GUS gene was induced by a longer Al treatment (approximately 8 h for maximum expression). Since the gene expression was observed in the leaf when only the root was exposed to Al stress, a signaling system between the root and shoot was suggested in Al stress. A GUS staining experiment using an adult transgenic line carrying the pAtGST11::GUS gene supported this suggestion. Furthermore, Al treatment simultaneously with various Ca depleted conditions in root region enhanced the gene expression of the pAtGST11::GUS in the shoot region. This result suggested that the degree of Al toxicity in the root reflects the gene response of pAtGST11::GUS in the shoot via the deduced signaling system. Both transgenic lines also showed an increase of GUS activity after cold stress, heat stress, metal toxicity, and oxidative damages, suggesting a common induction mechanism in response to the tested stresses including Al stress. PMID:15047894

  9. Expression of nm23-H1 gene product in esophageal squamous cell carcinoma and its association with vessel invasion and survival

    International Nuclear Information System (INIS)

    Tomita, Masaki; Ayabe, Takanori; Matsuzaki, Yasunori; Edagawa, Masao; Maeda, Masayuki; Shimizu, Tetsuya; Hara, Masaki; Onitsuka, Toshio

    2001-01-01

    We assessed the nm23-H1 gene product expression and its relationship with lymphatic and blood vessel invasion in patients with esophageal squamous cell carcinoma. Formalin-fixed and paraffin-embedded tissue sections from 45 patients who were treated surgically were used in this study. Pathologists graded lymphatic and blood vessel invasion in each of the tissue samples. Expression of nm23-Hl gene product was determined using a specific monoclonal antibody. Expression of nm23-H1 gene product was present in 17 (37.8%) cases. We found an inverse correlation between nm23-H1 gene product expression and lymphatic vessel invasion, whereas no correlation between nm23-H1 gene product expression and blood vessel invasion. Overall survival rate was not different between nm23-H1 gene product positive and negative patients (p = 0.21). However, reduced expression of nm23-H1 gene product was associated with shorter overall survival in patients with involved lymph nodes (p < 0.05), but not in patients without involved lymph nodes (p = 0.87). In patients with esophageal squamous cell carcinoma, there appears to be an inverse relationship between nm23-H1 gene product expression and lymphatic vessel invasion. Furthermore, nm23-H1 gene product expression might be a prognostic marker in patients with involved lymph nodes. Our data does not demonstrate any correlation between nm23-H1 gene product expression and blood vessel invasion

  10. Two human homeobox genes, c1 and c8: structure analysis and expression in embryonic development.

    Science.gov (United States)

    Simeone, A; Mavilio, F; Acampora, D; Giampaolo, A; Faiella, A; Zappavigna, V; D'Esposito, M; Pannese, M; Russo, G; Boncinelli, E

    1987-07-01

    Two human cDNA clones (HHO.c1.95 and HHO.c8.5111) containing a homeobox region have been characterized, and the respective genomic regions have been partially analyzed. Expression of the corresponding genes, termed c1 and c8, was evaluated in different organs and body parts during human embryonic/fetal development. HHO.c1.95 apparently encodes a 217-amino acid protein containing a class I homeodomain that shares 60 out of 61 amino acid residues with the Antennapedia homeodomain of Drosophila melanogaster. HHO.c8.5111 encodes a 153-amino acid protein containing a homeodomain identical to that of the frog AC1 gene. Clones HHO.c1 and HHO.c8 detect by blot-hydridization one and two specific polyadenylylated transcripts, respectively. These are differentially expressed in spinal cord, backbone rudiments, limb buds (or limbs), heart, and skin of human embryos and early fetuses in the 5- to 9-week postfertilization period, thus suggesting that the c1 and c8 genes play a key role in a variety of developmental processes. Together, the results of the embryonic/fetal expression of c1 and c8 and those of two previously analyzed genes (c10 and c13) indicate a coherent pattern of expression of these genes in early human ontogeny.

  11. Two human homeobox genes, c1 and c8: structure analysis and expression in embryonic development

    International Nuclear Information System (INIS)

    Simeone, A.; Mavilio, F.; Acampora, D.

    1987-01-01

    Two human cDNA clones (HHO.c1.95 and HHO.c8.5111) containing a homeobox region have been characterized, and the respective genomic regions have been partially analyzed. Expression of the corresponding genes, termed c1 and c8, was evaluated in different organs and body parts during human embryonic/fetal development. HHO.c1.95 apparently encodes a 217-amino acid protein containing a class I homeodomain that shares 60 out of 61 amino acid residues with the Antennapedia homeodomain of Drosophila melanogaster. HHO.c8.5111 encodes a 153-amino acid protein containing a homeodomains identical to that of the frog AC1 gene. Clones HHO.c1 and HHO.c8 detect by blot-hybridization one and two specific polyadenylylated transcripts, respectively. These are differentially expressed in spinal cord, backbone rudiments, limb buds (or limbs), heart, and skin of human embryos and early fetuses in the 5- to 9-week postfertilization period, thus suggesting that the c1 and c8 genes play a key role in a variety of developmental processes. Together, the results of the embryonic/fetal expression of c1 and c8 and those of two previously analyzed genes (c10 and c13) indicate a coherent pattern of expression of these genes in early human ontogeny

  12. Identification of suitable reference genes for gene expression studies of shoulder instability.

    Directory of Open Access Journals (Sweden)

    Mariana Ferreira Leal

    Full Text Available Shoulder instability is a common shoulder injury, and patients present with plastic deformation of the glenohumeral capsule. Gene expression analysis may be a useful tool for increasing the general understanding of capsule deformation, and reverse-transcription quantitative polymerase chain reaction (RT-qPCR has become an effective method for such studies. Although RT-qPCR is highly sensitive and specific, it requires the use of suitable reference genes for data normalization to guarantee meaningful and reproducible results. In the present study, we evaluated the suitability of a set of reference genes using samples from the glenohumeral capsules of individuals with and without shoulder instability. We analyzed the expression of six commonly used reference genes (ACTB, B2M, GAPDH, HPRT1, TBP and TFRC in the antero-inferior, antero-superior and posterior portions of the glenohumeral capsules of cases and controls. The stability of the candidate reference gene expression was determined using four software packages: NormFinder, geNorm, BestKeeper and DataAssist. Overall, HPRT1 was the best single reference gene, and HPRT1 and B2M composed the best pair of reference genes from different analysis groups, including simultaneous analysis of all tissue samples. GenEx software was used to identify the optimal number of reference genes to be used for normalization and demonstrated that the accumulated standard deviation resulting from the use of 2 reference genes was similar to that resulting from the use of 3 or more reference genes. To identify the optimal combination of reference genes, we evaluated the expression of COL1A1. Although the use of different reference gene combinations yielded variable normalized quantities, the relative quantities within sample groups were similar and confirmed that no obvious differences were observed when using 2, 3 or 4 reference genes. Consequently, the use of 2 stable reference genes for normalization, especially

  13. Control of Secreted Protein Gene Expression and the Mammalian Secretome by the Metabolic Regulator PGC-1α.

    Science.gov (United States)

    Minsky, Neri; Roeder, Robert G

    2017-01-06

    Secreted proteins serve pivotal roles in the development of multicellular organisms, acting as structural matrix, extracellular enzymes, and signal molecules. However, how the secretome is regulated remains incompletely understood. Here we demonstrate, unexpectedly, that peroxisome proliferator-activated receptor γ coactivator 1-α (PGC-1α), a critical transcriptional co-activator of metabolic gene expression, functions to down-regulate the expression of diverse genes encoding secreted molecules and extracellular matrix components to modulate the secretome. Using cell lines, primary cells, and mice, we show that both endogenous and exogenous PGC-1α down-regulate the expression of numerous genes encoding secreted molecules. Mechanistically, results obtained using mRNA stability measurements as well as intronic RNA expression analysis are consistent with a transcriptional effect of PGC-1α on the expression of genes encoding secreted proteins. Interestingly, PGC-1α requires the central heat shock response regulator heat shock factor protein 1 (HSF1) to affect some of its targets, and both factors co-reside on several target genes encoding secreted molecules in cells. Finally, using a mass spectrometric analysis of secreted proteins, we demonstrate that PGC-1α modulates the secretome of mouse embryonic fibroblasts. Our results define a link between a key pathway controlling metabolic regulation and the regulation of the mammalian secretome. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  14. Loss of the NKX3.1 tumorsuppressor promotes the TMPRSS2-ERG fusion gene expression in prostate cancer

    International Nuclear Information System (INIS)

    Thangapazham, Rajesh; Saenz, Francisco; Katta, Shilpa; Mohamed, Ahmed A; Tan, Shyh-Han; Petrovics, Gyorgy; Srivastava, Shiv; Dobi, Albert

    2014-01-01

    In normal prostate epithelium the TMPRSS2 gene encoding a type II serine protease is directly regulated by male hormones through the androgen receptor. In prostate cancer ERG protooncogene frequently gains hormonal control by seizing gene regulatory elements of TMPRSS2 through genomic fusion events. Although, the androgenic activation of TMPRSS2 gene has been established, little is known about other elements that may interact with TMPRSS2 promoter sequences to modulate ERG expression in TMPRSS2-ERG gene fusion context. Comparative genomic analyses of the TMPRSS2 promoter upstream sequences and pathway analyses were performed by the Genomatix Software. NKX3.1 and ERG genes expressions were evaluated by immunoblot or by quantitative Real-Time PCR (qRT-PCR) assays in response to siRNA knockdown or heterologous expression. QRT-PCR assay was used for monitoring the gene expression levels of NKX3.1-regulated genes. Transcriptional regulatory function of NKX3.1 was assessed by luciferase assay. Recruitment of NKX3.1 to its cognate elements was monitored by Chromatin Immunoprecipitation assay. Comparative analysis of the TMPRSS2 promoter upstream sequences among different species revealed the conservation of binding sites for the androgen inducible NKX3.1 tumor suppressor. Defects of NKX3.1, such as, allelic loss, haploinsufficiency, attenuated expression or decreased protein stability represent established pathways in prostate tumorigenesis. We found that NKX3.1 directly binds to TMPRSS2 upstream sequences and negatively regulates the expression of the ERG protooncogene through the TMPRSS2-ERG gene fusion. These observations imply that the frequently noted loss-of-function of NKX3.1 cooperates with the activation of TMPRSS2-ERG fusions in prostate tumorigenesis

  15. Involvement of histone methyltransferase GLP in HIV-1 latency through catalysis of H3K9 dimethylation

    International Nuclear Information System (INIS)

    Ding, Donglin; Qu, Xiying; Li, Lin; Zhou, Xin; Liu, Sijie; Lin, Shiguan; Wang, Pengfei; Liu, Shaohui; Kong, Chuijin; Wang, Xiaohui; Liu, Lin; Zhu, Huanzhang

    2013-01-01

    Understanding the mechanism of HIV-1 latency is crucial to eradication of the viral reservoir in HIV-1-infected individuals. However, the role of histone methyltransferase (HMT) G9a-like protein (GLP) in HIV-1 latency is still unclear. In the present work, we established four clonal cell lines containing HIV-1 vector. We found that the integration sites of most clonal cell lines favored active gene regions. However, we also observed hypomethylation of CpG of HIV 5′LTR in all four clonal cell lines. Additionally, 5′-deoxy-5′-methylthioadenosine (MTA), a broad-spectrum histone methyltransferase inhibitor, was used to examine the role of histone methylation in HIV-1 latency. MTA was found to decrease the level of H3K9 dimethylation, causing reactivation of latent HIV-1 in C11 cells. GLP knockdown by small interfering RNA clearly induced HIV-1 LTR expression. Results suggest that GLP may play a significant role in the maintenance of HIV-1 latency by catalyzing dimethylation of H3K9. - Highlights: ► We have established an in vitro model of HIV-1 latency. ► The integration sites of most clonal cell lines favor in active gene regions. ► Hypomethylation occurs in CpG islands of HIV 5′LTR in all four clonal cell lines. ► MTA can reactivate latent HIV-1 by decreasing the level of H3K9 me2 in C11 cells. ► HMT GLP may play a significant role in the maintenance of HIV-1 latency

  16. Porcine EEF1A1 and EEF1A2 genes: genomic structure, polymorphism, mapping and expression

    Czech Academy of Sciences Publication Activity Database

    Svobodová, K.; Horák, Pavel; Stratil, Antonín; Bartenschlager, H.; Van Poucke, M.; Chalupová, P.; Dvořáková, Věra; Knorr, Ch.; Stupka, R.; Čítek, J.; Šprysl, M.; Palánová, Anna; Peelman, L. J.; Geldermann, H.; Knoll, A.

    2015-01-01

    Roč. 42, č. 8 (2015), s. 1257-1264 ISSN 0301-4851 R&D Projects: GA ČR(CZ) GA523/06/1302; GA ČR GA523/09/0844 Institutional support: RVO:67985904 Keywords : EEF1A1 * EEF1A2 * gene expression Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.698, year: 2015

  17. Clock Genes Influence Gene Expression in Growth Plate and Endochondral Ossification in Mice*

    Science.gov (United States)

    Takarada, Takeshi; Kodama, Ayumi; Hotta, Shogo; Mieda, Michihiro; Shimba, Shigeki; Hinoi, Eiichi; Yoneda, Yukio

    2012-01-01

    We have previously shown transient promotion by parathyroid hormone of Period-1 (Per1) expression in cultured chondrocytes. Here we show the modulation by clock genes of chondrogenic differentiation through gene transactivation of the master regulator of chondrogenesis Indian hedgehog (IHH) in chondrocytes of the growth plate. Several clock genes were expressed with oscillatory rhythmicity in cultured chondrocytes and rib growth plate in mice, whereas chondrogenesis was markedly inhibited in stable transfectants of Per1 in chondrocytic ATDC5 cells and in rib growth plate chondrocytes from mice deficient of brain and muscle aryl hydrocarbon receptor nuclear translocator-like (BMAL1). Ihh promoter activity was regulated by different clock gene products, with clear circadian rhythmicity in expression profiles of Ihh in the growth plate. In BMAL1-null mice, a predominant decrease was seen in Ihh expression in the growth plate with a smaller body size than in wild-type mice. BMAL1 deficit led to disruption of the rhythmic expression profiles of both Per1 and Ihh in the growth plate. A clear rhythmicity was seen with Ihh expression in ATDC5 cells exposed to dexamethasone. In young mice defective of BMAL1 exclusively in chondrocytes, similar abnormalities were found in bone growth and Ihh expression. These results suggest that endochondral ossification is under the regulation of particular clock gene products expressed in chondrocytes during postnatal skeletogenesis through a mechanism relevant to the rhythmic Ihh expression. PMID:22936800

  18. Hypomethylation and Aberrant Expression of the Glioma Pathogenesis-Related 1 Gene in Wilms Tumors

    Directory of Open Access Journals (Sweden)

    Laxmi Chilukamarri

    2007-11-01

    Full Text Available Wilms tumors (WTs have a complex etiology, displaying genetic and epigenetic changes, including loss of imprinting (LOI and tumor suppressor gene silencing. To identify new regions of epigenetic perturbation in WTs, we screened kidney and tumor DNA using CpG island (CGI tags associated with cancer-specific DNA methylation changes. One such tag corresponded to a paralog of the glioma pathogenesis-related 1/related to testis-specific, vespid, and pathogenesis proteins 1 (GLIPR1/RTVP-1 gene, previously reported to be a tumor-suppressor gene silenced by hypermethylation in prostate cancer. Here we report methylation analysis of the GLIPR1/RTVP-1 gene in WTs and normal fetal and pediatric kidneys. Hypomethylation of the GLIPR1/RTVP-1 5'-region in WTs relative to normal tissue is observed in 21/24 (87.5% of WTs analyzed. Quantitative analysis of GLIPR1/RTVP-1 expression in 24 WTs showed elevated transcript levels in 16/24 WTs (67%, with 12 WTs displaying in excess of 20-fold overexpression relative to fetal kidney (FK control samples. Immunohistochemical analysis of FK and WT corroborates the RNA expression data and reveals high GLIPR1/RTVP-1 in WT blastemal cells together with variable levels in stromal and epithelial components. Hypomethylation is also evident in the WT precursor lesions and nephrogenic rests (NRs, supporting a role for GLIPR1/RTVP-1 deregulation early in Wilms tumorigenesis. Our data show that, in addition to gene dosage changes arising from LOI and hypermethylation-induced gene silencing, gene activation resulting from hypomethylation is also prevalent in WTs.

  19. Expression profiling analyses of porcine MuRF1 gene and its ...

    African Journals Online (AJOL)

    Using the INRA radiation hybrid panel (IMpRH) technique, the MuRF1 gene was assigned to SSC6q21-26, closely linked to microsatellite markers SW1823 and SW709. The tissue distribution patterns revealed that MuRF1 mRNA was exclusively expressed in cardiac and skeletal muscle tissues. Real-time quantitative ...

  20. Thylakoid redox signals are integrated into organellar-gene-expression-dependent retrograde signalling in the prors1-1 mutant

    Directory of Open Access Journals (Sweden)

    Luca eTadini

    2012-12-01

    Full Text Available Perturbations in organellar gene expression (OGE and the thylakoid redox state (TRS activate retrograde signalling pathways that adaptively modify nuclear gene expression (NGE, according to developmental and metabolic needs. The prors1-1 mutation in Arabidopsis down-regulates the expression of the nuclear gene Prolyl-tRNA Synthetase1 (PRORS1 which acts in both plastids and mitochondria, thereby impairing protein synthesis in both organelles and triggering OGE-dependent retrograde signalling. Because the mutation also affects thylakoid electron transport, TRS-dependent signals may likewise have an impact on the changes in NGE observed in this genotype. In this study, we have investigated whether signals related to TRS are actually integrated into the OGE-dependent retrograde signalling pathway. To this end, the chaos mutation (for chlorophyll a/b binding protein harvesting-organelle specific, which shows a partial loss of PSII antennae proteins and thus a reduction in PSII light absorption capability, was introduced into the prors1-1 mutant background. The resulting double mutant displayed a prors1-1-like reduction in plastid translation rate and a chaos-like decrease in PSII antenna size, whereas the hyper-reduction of the thylakoid electron transport chain, caused by the prors1-1 mutation, was alleviated, as determined by monitoring chlorophyll (Chl fluorescence and thylakoid phosphorylation. Interestingly, a substantial fraction of the nucleus-encoded photosynthesis genes down-regulated in the prors1-1 mutant are expressed at nearly wild-type rates in prors1-1 chaos leaves, and this recovery is reflected in the steady-state levels of their protein products in the chloroplast. We therefore conclude that signals related to photosynthetic electron transport and TRS, and indirectly to carbohydrate metabolism and energy balance, are indeed fed into the OGE-dependent retrograde pathway to modulate NGE and adjust the abundance of chloroplast proteins.

  1. Extracting gene expression patterns and identifying co-expressed genes from microarray data reveals biologically responsive processes

    Directory of Open Access Journals (Sweden)

    Paules Richard S

    2007-11-01

    Full Text Available Abstract Background A common observation in the analysis of gene expression data is that many genes display similarity in their expression patterns and therefore appear to be co-regulated. However, the variation associated with microarray data and the complexity of the experimental designs make the acquisition of co-expressed genes a challenge. We developed a novel method for Extracting microarray gene expression Patterns and Identifying co-expressed Genes, designated as EPIG. The approach utilizes the underlying structure of gene expression data to extract patterns and identify co-expressed genes that are responsive to experimental conditions. Results Through evaluation of the correlations among profiles, the magnitude of variation in gene expression profiles, and profile signal-to-noise ratio's, EPIG extracts a set of patterns representing co-expressed genes. The method is shown to work well with a simulated data set and microarray data obtained from time-series studies of dauer recovery and L1 starvation in C. elegans and after ultraviolet (UV or ionizing radiation (IR-induced DNA damage in diploid human fibroblasts. With the simulated data set, EPIG extracted the appropriate number of patterns which were more stable and homogeneous than the set of patterns that were determined using the CLICK or CAST clustering algorithms. However, CLICK performed better than EPIG and CAST with respect to the average correlation between clusters/patterns of the simulated data. With real biological data, EPIG extracted more dauer-specific patterns than CLICK. Furthermore, analysis of the IR/UV data revealed 18 unique patterns and 2661 genes out of approximately 17,000 that were identified as significantly expressed and categorized to the patterns by EPIG. The time-dependent patterns displayed similar and dissimilar responses between IR and UV treatments. Gene Ontology analysis applied to each pattern-related subset of co-expressed genes revealed underlying

  2. Comparative gene expression of intestinal metabolizing enzymes.

    Science.gov (United States)

    Shin, Ho-Chul; Kim, Hye-Ryoung; Cho, Hee-Jung; Yi, Hee; Cho, Soo-Min; Lee, Dong-Goo; Abd El-Aty, A M; Kim, Jin-Suk; Sun, Duxin; Amidon, Gordon L

    2009-11-01

    The purpose of this study was to compare the expression profiles of drug-metabolizing enzymes in the intestine of mouse, rat and human. Total RNA was isolated from the duodenum and the mRNA expression was measured using Affymetrix GeneChip oligonucleotide arrays. Detected genes from the intestine of mouse, rat and human were ca. 60% of 22690 sequences, 40% of 8739 and 47% of 12559, respectively. Total genes of metabolizing enzymes subjected in this study were 95, 33 and 68 genes in mouse, rat and human, respectively. Of phase I enzymes, the mouse exhibited abundant gene expressions for Cyp3a25, Cyp4v3, Cyp2d26, followed by Cyp2b20, Cyp2c65 and Cyp4f14, whereas, the rat showed higher expression profiles of Cyp3a9, Cyp2b19, Cyp4f1, Cyp17a1, Cyp2d18, Cyp27a1 and Cyp4f6. However, the highly expressed P450 enzymes were CYP3A4, CYP3A5, CYP4F3, CYP2C18, CYP2C9, CYP2D6, CYP3A7, CYP11B1 and CYP2B6 in the human. For phase II enzymes, glucuronosyltransferase Ugt1a6, glutathione S-transferases Gstp1, Gstm3 and Gsta2, sulfotransferase Sult1b1 and acyltransferase Dgat1 were highly expressed in the mouse. The rat revealed predominant expression of glucuronosyltransferases Ugt1a1 and Ugt1a7, sulfotransferase Sult1b1, acetyltransferase Dlat and acyltransferase Dgat1. On the other hand, in human, glucuronosyltransferases UGT2B15 and UGT2B17, glutathione S-transferases MGST3, GSTP1, GSTA2 and GSTM4, sulfotransferases ST1A3 and SULT1A2, acetyltransferases SAT1 and CRAT, and acyltransferase AGPAT2 were dominantly detected. Therefore, current data indicated substantial interspecies differences in the pattern of intestinal gene expression both for P450 enzymes and phase II drug-metabolizing enzymes. This genomic database is expected to improve our understanding of interspecies variations in estimating intestinal prehepatic clearance of oral drugs.

  3. Reference Gene Screening for Analyzing Gene Expression Across Goat Tissue

    Directory of Open Access Journals (Sweden)

    Yu Zhang

    2013-12-01

    Full Text Available Real-time quantitative PCR (qRT-PCR is one of the important methods for investigating the changes in mRNA expression levels in cells and tissues. Selection of the proper reference genes is very important when calibrating the results of real-time quantitative PCR. Studies on the selection of reference genes in goat tissues are limited, despite the economic importance of their meat and dairy products. We used real-time quantitative PCR to detect the expression levels of eight reference gene candidates (18S, TBP, HMBS, YWHAZ, ACTB, HPRT1, GAPDH and EEF1A2 in ten tissues types sourced from Boer goats. The optimal reference gene combination was selected according to the results determined by geNorm, NormFinder and Bestkeeper software packages. The analyses showed that tissue is an important variability factor in genes expression stability. When all tissues were considered, 18S, TBP and HMBS is the optimal reference combination for calibrating quantitative PCR analysis of gene expression from goat tissues. Dividing data set by tissues, ACTB was the most stable in stomach, small intestine and ovary, 18S in heart and spleen, HMBS in uterus and lung, TBP in liver, HPRT1 in kidney and GAPDH in muscle. Overall, this study provided valuable information about the goat reference genes that can be used in order to perform a proper normalisation when relative quantification by qRT-PCR studies is undertaken.

  4. Genome wide gene expression regulation by HIP1 Protein Interactor, HIPPI: Prediction and validation

    Directory of Open Access Journals (Sweden)

    Lahiri Ansuman

    2011-09-01

    Full Text Available Abstract Background HIP1 Protein Interactor (HIPPI is a pro-apoptotic protein that induces Caspase8 mediated apoptosis in cell. We have shown earlier that HIPPI could interact with a specific 9 bp sequence motif, defined as the HIPPI binding site (HBS, present in the upstream promoter of Caspase1 gene and regulate its expression. We also have shown that HIPPI, without any known nuclear localization signal, could be transported to the nucleus by HIP1, a NLS containing nucleo-cytoplasmic shuttling protein. Thus our present work aims at the investigation of the role of HIPPI as a global transcription regulator. Results We carried out genome wide search for the presence of HBS in the upstream sequences of genes. Our result suggests that HBS was predominantly located within 2 Kb upstream from transcription start site. Transcription factors like CREBP1, TBP, OCT1, EVI1 and P53 half site were significantly enriched in the 100 bp vicinity of HBS indicating that they might co-operate with HIPPI for transcription regulation. To illustrate the role of HIPPI on transcriptome, we performed gene expression profiling by microarray. Exogenous expression of HIPPI in HeLa cells resulted in up-regulation of 580 genes (p HIP1 was knocked down. HIPPI-P53 interaction was necessary for HIPPI mediated up-regulation of Caspase1 gene. Finally, we analyzed published microarray data obtained with post mortem brains of Huntington's disease (HD patients to investigate the possible involvement of HIPPI in HD pathogenesis. We observed that along with the transcription factors like CREB, P300, SREBP1, Sp1 etc. which are already known to be involved in HD, HIPPI binding site was also significantly over-represented in the upstream sequences of genes altered in HD. Conclusions Taken together, the results suggest that HIPPI could act as an important transcription regulator in cell regulating a vast array of genes, particularly transcription factors and at least, in part, play a

  5. Leucine zipper, down regulated in cancer-1 gene expression in prostate cancer

    Science.gov (United States)

    Salemi, Michele; Barone, Nunziata; La Vignera, Sandro; Condorelli, Rosita A.; Recupero, Domenico; Galia, Antonio; Fraggetta, Filippo; Aiello, Anna Maria; Pepe, Pietro; Castiglione, Roberto; Vicari, Enzo; Calogero, Aldo E.

    2016-01-01

    Numerous genetic alterations have been implicated in the development of prostate cancer (PCa). DNA and protein microarrays have enabled the identification of genes associated with apoptosis, which is important in PCa development. Despite the molecular mechanisms are not entirely understood, inhibition of apoptosis is a critical pathophysiological factor that contributes to the onset and progression of PCa. Leucine zipper, down-regulated in cancer 1 (LDOC-1) is a known regulator of the nuclear factor (NF)-mediated pathway of apoptosis through the inhibition of NF-κB. The present study investigated the expression of the LDOC-1 gene in LNCaP, PC-3, PNT1A and PNT2 prostate cell lines by reverse transcription-quantitative polymerase chain reaction. In addition LDOC-1 protein expression in normal prostate tissues and PCa was studied by immunohistochemistry. LDOC-1 messenger RNA resulted overexpressed in LNCaP and PC-3 PCa cell lines compared with the two normal prostate cell lines PNT1A and PNT2. The results of immunohistochemistry demonstrated a positive cytoplasmic LDOC-1 staining in all PCa and normal prostate samples, whereas no nuclear staining was observed in any sample. Furthermore, a more intense signal was evidenced in PCa samples. LDOC-1 gene overexpression in PCa suggests an activity of LDOC-1 in PCa cell lines. PMID:27698860

  6. Validation of commonly used reference genes for sleep-related gene expression studies

    Directory of Open Access Journals (Sweden)

    Castro Rosa MRPS

    2009-05-01

    Full Text Available Abstract Background Sleep is a restorative process and is essential for maintenance of mental and physical health. In an attempt to understand the complexity of sleep, multidisciplinary strategies, including genetic approaches, have been applied to sleep research. Although quantitative real time PCR has been used in previous sleep-related gene expression studies, proper validation of reference genes is currently lacking. Thus, we examined the effect of total or paradoxical sleep deprivation (TSD or PSD on the expression stability of the following frequently used reference genes in brain and blood: beta-actin (b-actin, beta-2-microglobulin (B2M, glyceraldehyde-3-phosphate dehydrogenase (GAPDH, and hypoxanthine guanine phosphoribosyl transferase (HPRT. Results Neither TSD nor PSD affected the expression stability of all tested genes in both tissues indicating that b-actin, B2M, GAPDH and HPRT are appropriate reference genes for the sleep-related gene expression studies. In order to further verify these results, the relative expression of brain derived neurotrophic factor (BDNF and glycerol-3-phosphate dehydrogenase1 (GPD1 was evaluated in brain and blood, respectively. The normalization with each of four reference genes produced similar pattern of expression in control and sleep deprived rats, but subtle differences in the magnitude of expression fold change were observed which might affect the statistical significance. Conclusion This study demonstrated that sleep deprivation does not alter the expression stability of commonly used reference genes in brain and blood. Nonetheless, the use of multiple reference genes in quantitative RT-PCR is required for the accurate results.

  7. TOPAZ1, a novel germ cell-specific expressed gene conserved during evolution across vertebrates.

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    Adrienne Baillet

    Full Text Available BACKGROUND: We had previously reported that the Suppression Subtractive Hybridization (SSH approach was relevant for the isolation of new mammalian genes involved in oogenesis and early follicle development. Some of these transcripts might be potential new oocyte and granulosa cell markers. We have now characterized one of them, named TOPAZ1 for the Testis and Ovary-specific PAZ domain gene. PRINCIPAL FINDINGS: Sheep and mouse TOPAZ1 mRNA have 4,803 bp and 4,962 bp open reading frames (20 exons, respectively, and encode putative TOPAZ1 proteins containing 1,600 and 1653 amino acids. They possess PAZ and CCCH domains. In sheep, TOPAZ1 mRNA is preferentially expressed in females during fetal life with a peak during prophase I of meiosis, and in males during adulthood. In the mouse, Topaz1 is a germ cell-specific gene. TOPAZ1 protein is highly conserved in vertebrates and specifically expressed in mouse and sheep gonads. It is localized in the cytoplasm of germ cells from the sheep fetal ovary and mouse adult testis. CONCLUSIONS: We have identified a novel PAZ-domain protein that is abundantly expressed in the gonads during germ cell meiosis. The expression pattern of TOPAZ1, and its high degree of conservation, suggests that it may play an important role in germ cell development. Further characterization of TOPAZ1 may elucidate the mechanisms involved in gametogenesis, and particularly in the RNA silencing process in the germ line.

  8. Twist and YB-1 gene expression in cervical cancer and precancerous tissue and their correlation with cell invasion

    Directory of Open Access Journals (Sweden)

    Qin Tian

    2017-04-01

    Full Text Available Objective: To study the correlation of Twist and YB-1 gene expression in cervical cancer and precancerous tissue with cell invasion. Methods: Cervical cancer tissue, precancerous tissue and normal cervical tissue surgically removed in our hospital between May 2013 and April 2015 were collected; immunohistochemical staining kits were used to detect the positive protein expression rate of Twist and YB-1 gene; fluorescence quantitative PCR kits were used to detect Twist, YB-1 and invasion gene mRNA expression. Results: Twist and YB-1 mRNA expression and positive protein expression rate as well as USP22, Rab11, Rac1 and ANXA5 mRNA expression in cervical cancer tissue and precancerous tissue were significantly higher than those in normal cervical tissue, Twist and YB-1 mRNA expression and positive protein expression rate as well as USP22, Rab11, Rac1 and ANXA5 mRNA expression in cervical cancer tissue were significantly higher than those in precancerous tissue; USP22, Rab11, Rac1 and ANXA5 mRNA expression in cervical cancer tissue and precancerous tissue with positive Twist and YB-1 expression were significantly higher than those in cervical cancer tissue and precancerous tissue with negative Twist and YB-1 expression. Conclusion: Highly expressed Twist and YB-1 in cervical cancer and precancerous tissue can promote cell invasion.

  9. The gsdf gene locus harbors evolutionary conserved and clustered genes preferentially expressed in fish previtellogenic oocytes.

    Science.gov (United States)

    Gautier, Aude; Le Gac, Florence; Lareyre, Jean-Jacques

    2011-02-01

    The gonadal soma-derived factor (GSDF) belongs to the transforming growth factor-β superfamily and is conserved in teleostean fish species. Gsdf is specifically expressed in the gonads, and gene expression is restricted to the granulosa and Sertoli cells in trout and medaka. The gsdf gene expression is correlated to early testis differentiation in medaka and was shown to stimulate primordial germ cell and spermatogonia proliferation in trout. In the present study, we show that the gsdf gene localizes to a syntenic chromosomal fragment conserved among vertebrates although no gsdf-related gene is detected on the corresponding genomic region in tetrapods. We demonstrate using quantitative RT-PCR that most of the genes localized in the synteny are specifically expressed in medaka gonads. Gsdf is the only gene of the synteny with a much higher expression in the testis compared to the ovary. In contrast, gene expression pattern analysis of the gsdf surrounding genes (nup54, aff1, klhl8, sdad1, and ptpn13) indicates that these genes are preferentially expressed in the female gonads. The tissue distribution of these genes is highly similar in medaka and zebrafish, two teleostean species that have diverged more than 110 million years ago. The cellular localization of these genes was determined in medaka gonads using the whole-mount in situ hybridization technique. We confirm that gsdf gene expression is restricted to Sertoli and granulosa cells in contact with the premeiotic and meiotic cells. The nup54 gene is expressed in spermatocytes and previtellogenic oocytes. Transcripts corresponding to the ovary-specific genes (aff1, klhl8, and sdad1) are detected only in previtellogenic oocytes. No expression was detected in the gonocytes in 10 dpf embryos. In conclusion, we show that the gsdf gene localizes to a syntenic chromosomal fragment harboring evolutionary conserved genes in vertebrates. These genes are preferentially expressed in previtelloogenic oocytes, and thus, they

  10. Expression of the metastasis-associated mts1 gene during mouse development

    DEFF Research Database (Denmark)

    Klingelhöfer, Jörg; Ambartsumian, N S; Lukanidin, E M

    1997-01-01

    motility. In order to understand the function of this gene, we studied the expression of the mts1 mRNA and protein in vivo during mouse development. Both mRNA and protein were present in high concentrations from 12.5 to 18.5 days post coitum (dpc) in a variety of developing embryonic tissue of mesodermal....... In developing bone, Mts1 was expressed in invasive mesenchymal cells and in osteoclasts. The results presented here suggest that Mtsl plays an important role in mouse development during differentiation and function of macrophages and might be involved in different processes associated with mesenchymal...

  11. A synthetic cadmium metallothionein gene (PMCd1syn) of Paramecium species: expression, purification and characteristics of metallothionein protein.

    Science.gov (United States)

    Dar, Saira; Shuja, Rukhsana N; Shakoori, Abdul Rauf

    2013-02-01

    Metallothioneins (MTs) are metal binding proteins that are rich in cysteine residues constituting 10-30 % of the total protein, and in which the thiol groups bind to the metal ions. The increasing amount of metal ions in the medium have shown increased production of MTs by different organisms such as bacteria, protozoa and mammals like humans. PMCd1 is the first gene ever discovered in Paramecium, a ciliated protozoan, that could produce this MT in response to cadmium. In this study the PMCd1syn gene has been cloned in pET41a expression vector and expressed in an Escherichia coli BL21-codonplus strain for the first time. Since the gene PMCd1 amplified from Paramecium contained 10 codons, which could act as stop codons during expression in E. coli, this gene of 612 bps was synthesized to substitute these (stop) codons for the Paramecium sp. specific amino acids. For stability of the expressed protein, glutathione-S-transferase gene was fused with PMCd1syn gene and coexpressed. The cells expressing PMCd1syn demonstrated increased accumulation of cadmium. This is the first report of cadmium MT protein expressed from Paramecium species, particularly from synthetic MT gene (PMCd1syn). This fusion protein, the molecular weight of which has been confirmed to be 53.03 kDa with MALDI analysis, is rich in cysteine residues, and has been shown for the first time in this ciliate to bind to and sequester Cd(2+)-ions.

  12. Gene-expression analysis of matrix metalloproteinases 1 and 2 and their tissue inhibitors in chronic periapical inflammatory lesions.

    Science.gov (United States)

    Hadziabdic, Naida; Kurtovic-Kozaric, Amina; Pojskic, Naris; Sulejmanagic, Nedim; Todorovic, Ljubomir

    2016-03-01

    Periapical inflammatory lesions have been investigated previously, but understanding of pathogenesis of these lesions (granulomas and radicular cysts) at the molecular level is still questionable. Matrix metalloproteinases (MMPs) are enzymes involved in the development of periapical pathology, specifically inflammation and tissue destruction. To elucidate pathogenesis of periapical granulomas and radicular cysts, we undertook a detailed analysis of gene expression of MMP-1, MMP-2 and their tissue inhibitors, TIMP-1 and TIMP-2. A total of 149 samples were analyzed using real-time PCR (59 radicular cysts, 50 periapical granulomas and 40 healthy gingiva samples as controls) for expression of MMP-1, MMP-2, TIMP-1 and TIMP-2 genes. The determination of best reference gene for expression analysis of periapical lesions was done using a panel of 12 genes. We have shown that β-actin and GAPDH are not the most stable reference controls for gene expression analysis of inflammatory periapical tissues and healthy gingiva. The most suitable reference gene was determined to be SDHA (a succinate dehydrogenase complex, subunit A, flavoprotein [Fp]). We found that granulomas (n = 50) and radicular cysts (n = 59) exhibited significantly higher expression of all four examined genes, MMP-1, MMP-2, TIMP-1, and TIMP-2, when compared to healthy gingiva (n = 40; P periapical inflammatory lesions. Since the abovementioned markers were not differentially expressed in periapical granulomas and radicular cysts, the challenge of finding the genetic differences between the two lesions still remains. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  13. Reduction in WT1 gene expression during early treatment predicts the outcome in patients with acute myeloid leukemia.

    Science.gov (United States)

    Andersson, Charlotta; Li, Xingru; Lorenz, Fryderyk; Golovleva, Irina; Wahlin, Anders; Li, Aihong

    2012-12-01

    Wilms tumor gene 1 (WT1) expression has been suggested as an applicable minimal residual disease marker in acute myeloid leukemia (AML). We evaluated the use of this marker in 43 adult AML patients. Quantitative assessment of WT1 gene transcripts was performed using real-time quantitative-polymerase chain reaction assay. Samples from both the peripheral blood and the bone marrow were analyzed at diagnosis and during follow-up. A strong correlation was observed between WT1 normalized with 2 different control genes (β-actin and ABL1, P0.05). A≥1-log reduction in WT1 expression in bone marrow samples taken freedom from relapse (P=0.010) when β-actin was used as control gene. Furthermore, a reduction in WT1 expression by ≥2 logs in peripheral blood samples taken at a later time point significantly correlated with a better outcome for overall survival (P=0.004) and freedom from relapse (P=0.012). This result was achieved when normalizing against both β-actin and ABL1. These results therefore suggest that WT1 gene expression can provide useful information for minimal residual disease detection in adult AML patients and that combined use of control genes can give more informative results.

  14. Glucocorticoid-induced reversal of interleukin-1β-stimulated inflammatory gene expression in human oviductal cells.

    Directory of Open Access Journals (Sweden)

    Stéphanie Backman

    Full Text Available Studies indicate that high-grade serous ovarian carcinoma (HGSOC, the most common epithelial ovarian carcinoma histotype, originates from the fallopian tube epithelium (FTE. Risk factors for this cancer include reproductive parameters associated with lifetime ovulatory events. Ovulation is an acute inflammatory process during which the FTE is exposed to follicular fluid containing both pro- and anti-inflammatory molecules, such as interleukin-1 (IL1, tumor necrosis factor (TNF, and cortisol. Repeated exposure to inflammatory cytokines may contribute to transforming events in the FTE, with glucocorticoids exerting a protective effect. The global response of FTE cells to inflammatory cytokines or glucocorticoids has not been investigated. To examine the response of FTE cells and the ability of glucocorticoids to oppose this response, an immortalized human FTE cell line, OE-E6/E7, was treated with IL1β, dexamethasone (DEX, IL1β and DEX, or vehicle and genome-wide gene expression profiling was performed. IL1β altered the expression of 47 genes of which 17 were reversed by DEX. DEX treatment alone altered the expression of 590 genes, whereas combined DEX and IL1β treatment altered the expression of 784 genes. Network and pathway enrichment analysis indicated that many genes altered by DEX are involved in cytokine, chemokine, and cell cycle signaling, including NFκΒ target genes and interacting proteins. Quantitative real time RT-PCR studies validated the gene array data for IL8, IL23A, PI3 and TACC2 in OE-E6/E7 cells. Consistent with the array data, Western blot analysis showed increased levels of PTGS2 protein induced by IL1β that was blocked by DEX. A parallel experiment using primary cultured human FTE cells indicated similar effects on PTGS2, IL8, IL23A, PI3 and TACC2 transcripts. These findings support the hypothesis that pro-inflammatory signaling is induced in FTE cells by inflammatory mediators and raises the possibility that

  15. Differential gene expression by 1,25(OH)2D3 in an endometriosis stromal cell line.

    Science.gov (United States)

    Ingles, Sue Ann; Wu, Liang; Liu, Benjamin T; Chen, Yibu; Wang, Chun-Yeh; Templeman, Claire; Brueggmann, Doerthe

    2017-10-01

    Endometriosis is a common female reproductive disease characterized by invasion of endometrial cells into other organs, frequently causing pelvic pain and infertility. Alterations of the vitamin D system have been linked to endometriosis incidence and severity. To shed light on the potential mechanism for these associations, we examined the effects of 1,25(OH) 2 D 3 on gene expression in endometriosis cells. Stromal cell lines derived from endometriosis tissue were treated with 1,25(OH) 2 D 3 , and RNA-seq was used to identify genes differentially expressed between treated and untreated cells. Gene ontology and pathway analyses were carried out using Partek Flow and Ingenuity software suites, respectively. We identified 1627 genes that were differentially expressed (886 down-regulated and 741 up-regulated) by 1,25(OH) 2 D 3 . Only one gene, CYP24A1, was strongly up-regulated (369-fold). Many genes were strongly down-regulated. 1,25(OH) 2 D 3 treatment down-regulated several genetic pathways related to neuroangiogenesis, cellular motility, and invasion, including pathways for axonal guidance, Rho GDP signaling, and matrix metalloprotease inhibition. These findings support a role for vitamin D in the pathophysiology of endometriosis, and provide new targets for investigation into possible causes and treatments. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. Identification of reference genes in human myelomonocytic cells for gene expression studies in altered gravity.

    Science.gov (United States)

    Thiel, Cora S; Hauschild, Swantje; Tauber, Svantje; Paulsen, Katrin; Raig, Christiane; Raem, Arnold; Biskup, Josefine; Gutewort, Annett; Hürlimann, Eva; Unverdorben, Felix; Buttron, Isabell; Lauber, Beatrice; Philpot, Claudia; Lier, Hartwin; Engelmann, Frank; Layer, Liliana E; Ullrich, Oliver

    2015-01-01

    Gene expression studies are indispensable for investigation and elucidation of molecular mechanisms. For the process of normalization, reference genes ("housekeeping genes") are essential to verify gene expression analysis. Thus, it is assumed that these reference genes demonstrate similar expression levels over all experimental conditions. However, common recommendations about reference genes were established during 1 g conditions and therefore their applicability in studies with altered gravity has not been demonstrated yet. The microarray technology is frequently used to generate expression profiles under defined conditions and to determine the relative difference in expression levels between two or more different states. In our study, we searched for potential reference genes with stable expression during different gravitational conditions (microgravity, normogravity, and hypergravity) which are additionally not altered in different hardware systems. We were able to identify eight genes (ALB, B4GALT6, GAPDH, HMBS, YWHAZ, ABCA5, ABCA9, and ABCC1) which demonstrated no altered gene expression levels in all tested conditions and therefore represent good candidates for the standardization of gene expression studies in altered gravity.

  17. In vivo oestrogenic modulation of Egr1 and Pitx1 gene expression in female rat pituitary gland.

    Science.gov (United States)

    Gajewska, Alina; Herman, Andrzej P; Wolińska-Witort, Ewa; Kochman, Kazimierz; Zwierzchowski, Lech

    2014-12-01

    EGR1 and PITX1 are transcription factors required for gonadotroph cell Lhb promoter activation. To determine changes in Egr1 and Pitx1 mRNA levels in central and peripheral pituitary stimulations, an in vivo model based on i.c.v. pulsatile (1 pulse/0.5 h over 2 h) GnRH agonist (1.5 nM buserelin) or antagonist (2 nM antide) microinjections was used. The microinjections were given to ovariectomised and 17β-oestradiol (E2) (3×20 μg), ERA (ESR1) agonist propyl pyrazole triol (PPT) (3×0.5 mg), ERB (ESR2) agonist diarylpropionitrile (DPN) (3×0.5 mg) s.c. pre-treated rats 30 min after last pulse anterior pituitaries were excised. Relative mRNA expression was determined by quantitative RT-PCR (qRT-PCR). Results revealed a gene-specific response for GnRH and/or oestrogenic stimulations in vivo. Buserelin pulses enhanced Egr1 expression by 66% in ovariectomised rats, whereas the oestradiol-supplemented+i.c.v. NaCl-microinjected group showed a 50% increase in Egr1 mRNA expression. The oestrogenic signal was transmitted via ERA (ESR1) and ERB (ESR2) activation as administration of PPT and DPN resulted in 97 and 62%, respectively, elevation in Egr1 mRNA expression. A synergistic action of GnRH agonist and 17β-oestradiol (E2) stimulation of the Egr1 gene transcription in vivo were found. GnRHR activity did not affect Pitx1 mRNA expression; regardless of NaCl, buserelin or antide i.c.v. pulses, s.c. oestrogenic supplementation (with E2, PPT or DPN) consistently decreased (by -46, -48 and -41% respectively) the Pitx1 mRNA in the anterior pituitary gland. Orchestrated Egr1 and Pitx1 activities depending on specific central and peripheral regulatory inputs could be responsible for physiologically variable Lhb gene promoter activation in vivo. © 2014 Society for Endocrinology.

  18. Identification and validation of suitable endogenous reference genes for gene expression studies in human peripheral blood

    Directory of Open Access Journals (Sweden)

    Turner Renee J

    2009-08-01

    Full Text Available Abstract Background Gene expression studies require appropriate normalization methods. One such method uses stably expressed reference genes. Since suitable reference genes appear to be unique for each tissue, we have identified an optimal set of the most stably expressed genes in human blood that can be used for normalization. Methods Whole-genome Affymetrix Human 2.0 Plus arrays were examined from 526 samples of males and females ages 2 to 78, including control subjects and patients with Tourette syndrome, stroke, migraine, muscular dystrophy, and autism. The top 100 most stably expressed genes with a broad range of expression levels were identified. To validate the best candidate genes, we performed quantitative RT-PCR on a subset of 10 genes (TRAP1, DECR1, FPGS, FARP1, MAPRE2, PEX16, GINS2, CRY2, CSNK1G2 and A4GALT, 4 commonly employed reference genes (GAPDH, ACTB, B2M and HMBS and PPIB, previously reported to be stably expressed in blood. Expression stability and ranking analysis were performed using GeNorm and NormFinder algorithms. Results Reference genes were ranked based on their expression stability and the minimum number of genes needed for nomalization as calculated using GeNorm showed that the fewest, most stably expressed genes needed for acurate normalization in RNA expression studies of human whole blood is a combination of TRAP1, FPGS, DECR1 and PPIB. We confirmed the ranking of the best candidate control genes by using an alternative algorithm (NormFinder. Conclusion The reference genes identified in this study are stably expressed in whole blood of humans of both genders with multiple disease conditions and ages 2 to 78. Importantly, they also have different functions within cells and thus should be expressed independently of each other. These genes should be useful as normalization genes for microarray and RT-PCR whole blood studies of human physiology, metabolism and disease.

  19. Promotion of Metastasis-associated Gene Expression in Survived PANC-1 Cells Following Trichostatin A Treatment.

    Science.gov (United States)

    Chen, Zongjing; Yang, Yunxiu; Liu, Biao; Wang, Benquan; Sun, Meng; Zhang, Ling; Chen, Bicheng; You, Heyi; Zhou, Mengtao

    2015-01-01

    Histone deacetylase inhibitors represent a promising class of potential anticancer agents for the treatment of human malignancies. In this study, the effects of trichostatin A (TSA) on apoptosis, metastasis-associated gene expression, and activation of the Notch pathway in human pancreatic cancer cell lines were investigated. After treatment with TSA, cell viability and apoptosis were evaluated using the MTT [3-(4,5-dimethylthia-zol-2-yl)-2,5-diphenyltetrazolium bromide] assay, Hoechst 33258 staining, and flow cytometry. Moreover, RT-PCR and western blot analyses were performed to measure the expression levels of apoptosis-associated genes (Bcl-2, Bax, and caspase-3), metastasis-associated genes (E-cadherin, vimentin, and matrix metalloproteinases), and Notch pathway activation (Notch intracellular domain, NICD). The levels of matrix metalloproteinase 2 and NICD were also semi-quantified by immunoassay. Following treatment with TSA for 24 h, PANC-1, SW1990, and MIATACA-2 cells exhibited cell death. The MTT assay revealed that TSA significantly decreased cell viability in a dose-dependent manner in PANC-1 cells. The Hoechst 33258 staining and flow cytometry results evidenced a significant increase in PANC-1 cell apoptosis following TSA treatment. The expression levels of Bax and caspase-3 were increased significantly, whereas Bcl-2 was down-regulated after TSA treatment. In the PANC-1 cells that survived after TSA treatment, the expression levels of vimentin, E-cadherin, and MMP genes were altered by the promotion of potential metastasis and increased expression of NICD. TSA can induce apoptosis of pancreatic cancer cells. In addition, the up-regulation of metastasis-related genes and the activation of the Notch pathway in the survived PANC-1 cells may be associated with a too-low level of TSA or resistance to TSA.

  20. Positive selection on gene expression in the human brain

    DEFF Research Database (Denmark)

    Khaitovich, Philipp; Tang, Kun; Franz, Henriette

    2006-01-01

    Recent work has shown that the expression levels of genes transcribed in the brains of humans and chimpanzees have changed less than those of genes transcribed in other tissues [1] . However, when gene expression changes are mapped onto the evolutionary lineage in which they occurred, the brain...... shows more changes than other tissues in the human lineage compared to the chimpanzee lineage [1] , [2] and [3] . There are two possible explanations for this: either positive selection drove more gene expression changes to fixation in the human brain than in the chimpanzee brain, or genes expressed...... in the brain experienced less purifying selection in humans than in chimpanzees, i.e. gene expression in the human brain is functionally less constrained. The first scenario would be supported if genes that changed their expression in the brain in the human lineage showed more selective sweeps than other genes...

  1. Protein kinase D1 signaling in angiogenic gene expression and VEGF-mediated angiogenesis

    Directory of Open Access Journals (Sweden)

    Bin eRen MD, Phd, FAHA

    2016-05-01

    Full Text Available Protein kinase D 1 (PKD-1 is a signaling kinase important in fundamental cell functions including migration, proliferation and differentiation. PKD-1 is also a key regulator of gene expression and angiogenesis that is essential for cardiovascular development and tumor progression. Further understanding molecular aspects of PKD-1 signaling in the regulation of angiogenesis may have translational implications in obesity, cardiovascular disease and cancer. The author will summarize and provide the insights into molecular mechanisms by which PKD-1 regulates transcriptional expression of angiogenic genes, focusing on the transcriptional regulation of CD36 by PKD-1-FoxO1 signaling axis along with the potential implications of this axis in arterial differentiation and morphogenesis. He will also discuss a new concept of dynamic balance between proangiogenic and antiangiogenic signaling in determining angiogenic switch, and stress how PKD-1 signaling regulates VEGF signaling-mediated angiogenesis.

  2. The Schizophrenia-Associated BRD1 Gene Regulates Behavior, Neurotransmission, and Expression of Schizophrenia Risk Enriched Gene Sets in Mice.

    Science.gov (United States)

    Qvist, Per; Christensen, Jane Hvarregaard; Vardya, Irina; Rajkumar, Anto Praveen; Mørk, Arne; Paternoster, Veerle; Füchtbauer, Ernst-Martin; Pallesen, Jonatan; Fryland, Tue; Dyrvig, Mads; Hauberg, Mads Engel; Lundsberg, Birgitte; Fejgin, Kim; Nyegaard, Mette; Jensen, Kimmo; Nyengaard, Jens Randel; Mors, Ole; Didriksen, Michael; Børglum, Anders Dupont

    2017-07-01

    The schizophrenia-associated BRD1 gene encodes a transcriptional regulator whose comprehensive chromatin interactome is enriched with schizophrenia risk genes. However, the biology underlying the disease association of BRD1 remains speculative. This study assessed the transcriptional drive of a schizophrenia-associated BRD1 risk variant in vitro. Accordingly, to examine the effects of reduced Brd1 expression, we generated a genetically modified Brd1 +/- mouse and subjected it to behavioral, electrophysiological, molecular, and integrative genomic analyses with focus on schizophrenia-relevant parameters. Brd1 +/- mice displayed cerebral histone H3K14 hypoacetylation and a broad range of behavioral changes with translational relevance to schizophrenia. These behaviors were accompanied by striatal dopamine/serotonin abnormalities and cortical excitation-inhibition imbalances involving loss of parvalbumin immunoreactive interneurons. RNA-sequencing analyses of cortical and striatal micropunches from Brd1 +/- and wild-type mice revealed differential expression of genes enriched for schizophrenia risk, including several schizophrenia genome-wide association study risk genes (e.g., calcium channel subunits [Cacna1c and Cacnb2], cholinergic muscarinic receptor 4 [Chrm4)], dopamine receptor D 2 [Drd2], and transcription factor 4 [Tcf4]). Integrative analyses further found differentially expressed genes to cluster in functional networks and canonical pathways associated with mental illness and molecular signaling processes (e.g., glutamatergic, monoaminergic, calcium, cyclic adenosine monophosphate [cAMP], dopamine- and cAMP-regulated neuronal phosphoprotein 32 kDa [DARPP-32], and cAMP responsive element binding protein signaling [CREB]). Our study bridges the gap between genetic association and pathogenic effects and yields novel insights into the unfolding molecular changes in the brain of a new schizophrenia model that incorporates genetic risk at three levels: allelic

  3. Efectividad del M.T.A en el manejo no quirúrgico de perforaciones apicales

    Directory of Open Access Journals (Sweden)

    José Elias Flórez Ariza

    2013-11-01

    Full Text Available Title: MTA effectiveness in not surgical treatment of apical perforations.ResumenLas perforaciones endodónticas son aperturas artificiales que resultan en la comunicación del sistema de conductos radiculares con los tejidos perirradicularres o con la cavidad bucal. Estas son complicaciones indeseables que pueden ocurrir durante cualquier etapa del tratamiento de conductos, o durante la preparación del espacio para perno. Aún cuando también pueden ser producto de caries o procesos resortivos, la mayoría de las perforaciones son realizadas de forma iatrogénica. El manejo terapéutico de perforaciones radiculares a lo largo de la historia es un punto crítico y centro de investigaciones constantes en busca del desarrollo de los materiales idóneos para el sellado de las mismas, pero es el Mineral Trióxido Agregado (MTA el material que por sus propiedades físicas y bioquímicas, tales como biocompatibilidad, y regenerativo de hueso, cemento y de ligamento periodontal, el mayormente utilizado como garantía de éxito en dichos procedimientos. El presente caso muestra el éxito no quirúrgico obtenido a partir del sellado de una perforación apical contaminada, utilizando MTA en la zona de la afección y el relleno complementario con gutapercha termoplástica en un diente antero superior. (DUAZARY 2013 No. 1, 56 - 61.AbstractThe Endodontic perforations are artificial openings that result in the communication of the root canal system with the perirradicular tissues or oral cavity. These are undesirable complications that can occur during any stage of root canal treatment, or during the preparation of the space for the bolt. Although they can be produced by cavities or resorptive processes, the majority of the perforations are made iatrogenically. The therapeutic management of root perforations throughout history has been a critical and ongoing investigation topic in search of the development of suitable materials for their sealing, but the

  4. ALS1 and ALS3 gene expression and biofilm formation in Candida albicans isolated from vulvovaginal candidiasis

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    Shahla Roudbarmohammadi

    2016-01-01

    Conclusion: The results attained indicated that there is an association between the expression of ALS1 and ALS3 genes and fluconazole resistance in C. albicans. A considerable percent of the isolates expressing the ALS1 and ALS3 genes may have contributed to their adherence to vagina and biofilm formation.

  5. Altered gene expression in pulmonary tissue of tryptophan hydroxylase-1 knockout mice: implications for pulmonary arterial hypertension.

    Directory of Open Access Journals (Sweden)

    Richard B Rothman

    Full Text Available The use of fenfluramines can increase the risk of developing pulmonary arterial hypertension (PAH in humans, but the mechanisms responsible are unresolved. A recent study reported that female mice lacking the gene for tryptophan hydroxylase-1 (Tph1(-/- mice were protected from PAH caused by chronic dexfenfluramine, suggesting a pivotal role for peripheral serotonin (5-HT in the disease process. Here we tested two alternative hypotheses which might explain the lack of dexfenfluramine-induced PAH in Tph1(-/- mice. We postulated that: 1 Tph1(-/- mice express lower levels of pulmonary 5-HT transporter (SERT when compared to wild-type controls, and 2 Tph1(-/- mice display adaptive changes in the expression of non-serotonergic pulmonary genes which are implicated in PAH. SERT was measured using radioligand binding methods, whereas gene expression was measured using microarrays followed by quantitative real time PCR (qRT-PCR. Contrary to our first hypothesis, the number of pulmonary SERT sites was modestly up-regulated in female Tph1(-/- mice. The expression of 51 distinct genes was significantly altered in the lungs of female Tph1(-/- mice. Consistent with our second hypothesis, qRT-PCR confirmed that at least three genes implicated in the pathogenesis of PAH were markedly up-regulated: Has2, Hapln3 and Retlna. The finding that female Tph1(-/- mice are protected from dexfenfluramine-induced PAH could be related to compensatory changes in pulmonary gene expression, in addition to reductions in peripheral 5-HT. These observations emphasize the intrinsic limitation of interpreting data from studies conducted in transgenic mice that are not fully characterized.

  6. Structure-related clustering of gene expression fingerprints of thp-1 cells exposed to smaller polycyclic aromatic hydrocarbons.

    Science.gov (United States)

    Wan, B; Yarbrough, J W; Schultz, T W

    2008-01-01

    This study was undertaken to test the hypothesis that structurally similar PAHs induce similar gene expression profiles. THP-1 cells were exposed to a series of 12 selected PAHs at 50 microM for 24 hours and gene expressions profiles were analyzed using both unsupervised and supervised methods. Clustering analysis of gene expression profiles revealed that the 12 tested chemicals were grouped into five clusters. Within each cluster, the gene expression profiles are more similar to each other than to the ones outside the cluster. One-methylanthracene and 1-methylfluorene were found to have the most similar profiles; dibenzothiophene and dibenzofuran were found to share common profiles with fluorine. As expression pattern comparisons were expanded, similarity in genomic fingerprint dropped off dramatically. Prediction analysis of microarrays (PAM) based on the clustering pattern generated 49 predictor genes that can be used for sample discrimination. Moreover, a significant analysis of Microarrays (SAM) identified 598 genes being modulated by tested chemicals with a variety of biological processes, such as cell cycle, metabolism, and protein binding and KEGG pathways being significantly (p < 0.05) affected. It is feasible to distinguish structurally different PAHs based on their genomic fingerprints, which are mechanism based.

  7. Trichloroethylene-induced gene expression and DNA methylation changes in B6C3F1 mouse liver.

    Directory of Open Access Journals (Sweden)

    Yan Jiang

    Full Text Available Trichloroethylene (TCE, widely used as an organic solvent in the industry, is a common contaminant in air, soil, and water. Chronic TCE exposure induced hepatocellular carcinoma in mice, and occupational exposure in humans was suggested to be associated with liver cancer. To understand the role of non-genotoxic mechanism(s for TCE action, we examined the gene expression and DNA methylation changes in the liver of B6C3F1 mice orally administered with TCE (0, 100, 500 and 1000 mg/kg b.w. per day for 5 days. After 5 days TCE treatment at a dose level of 1000 mg/kg b.w., a total of 431 differentially expressed genes were identified in mouse liver by microarray, of which 291 were up-regulated and 140 down-regulated. The expression changed genes were involved in key signal pathways including PPAR, proliferation, apoptosis and homologous recombination. Notably, the expression level of a number of vital genes involved in the regulation of DNA methylation, such as Utrf1, Tet2, DNMT1, DNMT3a and DNMT3b, were dysregulated. Although global DNA methylation change was not detected in the liver of mice exposed to TCE, the promoter regions of Cdkn1a and Ihh were found to be hypo- and hypermethylated respectively, which correlated negatively with their mRNA expression changes. Furthermore, the gene expression and DNA methylation changes induced by TCE were dose dependent. The overall data indicate that TCE exposure leads to aberrant DNA methylation changes, which might alter the expression of genes involved in the TCE-induced liver tumorgenesis.

  8. The targeting expression of the vascular endothelial growth factor gene in endothelial cells regulated by HRE.ppET-1.

    Science.gov (United States)

    Zheng, Xiangrong; Zhang, Shangshang; Yang, Yujia; Wang, Xia; Zhong, Le; Yu, Xiaohe

    2008-11-01

    The success of gene therapy depends largely on the efficacy of gene delivery vector systems that can deliver genes to target organs or cells selectively and efficiently with minimal toxicity. Here, we show that by using the HRE.ppET-1 regulatory element, we were able to restrict expression of the transgene of vascular endothelial growth factor (VEGF) to endothelial cells exclusively in hypoxic conditions. Eukaryotic expression vectors such as pEGFP-HRE.ppET-1, pcDNA3.1-VEGF+Pa, pcDNA3.1-ppET-1+ EGF+Pa, and pcDNA3.1-HRE.ppET-1+VEGF+Pa were constructed by using a series of nuclear molecule handling methods like PCR, enzyme digestion. The recombinant vectors were transfected into HUVEC cells and HL7702 cells by the lipofectin method. GFP expression was observed with a fluorescence microscope to validate the specificity of expression in endothelial cells under the regulation of HRE.ppET-1 element. Cobalt chloride (final concentration 100 mumol/L) was added to the medium to mimic hypoxia in vitro. After transfection of vectors, the expression of VEGF mRNA was detected by RT-PCR, and the expression of VEGF was detected by Western blotting and ELISA methods under normoxia and hypoxia, respectively. The cell proliferation rate was detected by the MTT test. The expression of GFP revealed that the exterior gene was transcripted effectively in endothelial cells regulated by the HRE.ppET-1 element, while the expression of GFP was very weak in nonendothelial cells. The results of RT-PCR, Western blotting and ELISA showed that VEGF gene expression in the pcDNA3.1-HRE.ppET-1+VEGF+Pa group and in the pcDNA3.1-ppET-1+VEGF+Pa group was higher in hypoxia than it was in normoxia (PHRE.ppET-1 element was expressed specifically in endothelial cells, and can increase the expression of VEGF in hypoxia and stimulate proliferation of endothelial cells. Taking advantage of these facts could greatly improve the efficiency of gene therapy. The vector would be valuable for various gene transfer

  9. Sealing ability of MTA, CPM, and MBPc as root-end filling materials: a bacterial leakage study.

    Science.gov (United States)

    Medeiros, Paulo Leal; Bernardineli, Norberti; Cavenago, Bruno Cavalini; Torres, Sérgio Aparecido; Duarte, Marco Antonio Hungaro; Bramante, Clovis Monteiro; Marciano, Marina Angélica

    2016-04-01

    Objectives To evaluate the sealing ability of three root-end filling materials (white MTA, CPM, and MBPc) using an Enterococcus faecalis leakage model. Material and Methods Seventy single-root extracted human teeth were instrumented and root-ends were resected to prepare 3 mm depth cavities. Root-end preparations were filled with white MTA, CPM, and MBPc cements. Enterococcus faecalis was coronally introduced and the apical portion was immersed in BHI culture medium with phenol red indicator. The bacterial leakage was monitored every 24 h for 4 weeks. The statistical analysis was performed using the Wilcoxon-Gehan test (pCPM and the other groups. Conclusions The epoxy resin-based cement MBPc had lower bacterial leakage compared with the calcium silicate-based cements MTA and CPM.

  10. Expression and molecular evolution of two DREB1 genes in black poplar (Populus nigra.

    Directory of Open Access Journals (Sweden)

    Yanguang Chu

    Full Text Available Environmental stresses such as low temperature, drought, and high salinity significantly affect plant growth and yield. As selective forces, these adverse factors play essential roles in shaping phenotypic variation in plant populations. Black poplar (Populus nigra is an economically and ecologically important forest tree species with widely distributed populations and is thus suitable for experiments detecting evolutionary footprints left by stress. Here, we performed expression and evolutionary analysis of two duplicated DREB A1-subgroup (DREB1 genes, PnDREB68 and PnDREB69, encoding transcription factors that are involved in stress responses. The two genes showed partially overlapping but distinct expression patterns in response to stresses. These genes were strongly and rapidly induced by cold stress in leaves, stems, and roots. In leaf tissue, dehydration stress induced the expression of PnDREB68 but not PnDREB69. PnDREB69 displayed more rapid responses and longer expression durations than PnDREB68 under salt and ABA stress, respectively. Based on single nucleotide polymorphism (SNP analysis, we found significant population genetic differentiation, with a greater FST value (0.09189 for PnDREB69 than for PnDREB68 (0.07743. Nucleotide diversity analysis revealed a two-fold higher πT for PnDREB68 than for PnDREB69 (0.00563 vs. 0.00243, reflecting strong purifying selection acting on the former. The results suggest that positive selection acted on PnDREB69, as evidenced by neutral testing using Tajima's D statistic. The distinct selective forces to which each of the genes was subjected may be associated with expression divergence. Linkage disequilibrium (LD was low for the sequenced region, with a higher level for PnDREB68 than for PnDREB69. Additionally, analysis of the relationship among carbon isotope ratios, SNP classes and gene expression, together with motif and domain analysis, suggested that 14 polymorphisms within the two genes may be

  11. Gene Expression of Glucose Transporter 1 (GLUT1, Hexokinase 1 and Hexokinase 2 in Gastroenteropancreatic Neuroendocrine Tumors: Correlation with F-18-fluorodeoxyglucose Positron Emission Tomography and Cellular Proliferation

    Directory of Open Access Journals (Sweden)

    Andreas Kjaer

    2013-10-01

    Full Text Available Neoplastic tissue exhibits high glucose utilization and over-expression of glucose transporters (GLUTs and hexokinases (HKs, which can be imaged by 18F-Fluorodeoxyglucose-positron emission tomography (FDG-PET. The aim of the present study was to investigate the expression of glycolysis-associated genes and to compare this with FDG-PET imaging as well as with the cellular proliferation index in two cancer entities with different malignant potential. Using real-time PCR, gene expression of GLUT1, HK1 and HK2 were studied in 34 neuroendocrine tumors (NETs in comparison with 14 colorectal adenocarcinomas (CRAs. The Ki67 proliferation index and, when available, FDG-PET imaging was compared with gene expression. Overexpression of GLUT1 gene expression was less frequent in NETs (38% compared to CRAs (86%, P = 0.004. HK1 was overexpressed in 41% and 71% of NETs and CRAs, respectively (P = 0.111 and HK2 was overexpressed in 50% and 64% of NETs and CRAs, respectively (P = 0.53. There was a significant correlation between the Ki67 proliferation index and GLUT1 gene expression for the NETs (R = 0.34, P = 0.047, but no correlation with the hexokinases. FDG-PET identified foci in significantly fewer NETs (36% than CRAs (86%, (P = 0.04. The gene expression results, with less frequent GLUT1 and HK1 upregulation in NETs, confirmed the lower metabolic activity of NETs compared to the more aggressive CRAs. In accordance with this, fewer NETs were FDG-PET positive compared to CRA tumors and FDG uptake correlated with GLUT1 gene expression.

  12. Cloning and expression analysis of FaPR-1 gene in strawberry

    Science.gov (United States)

    Mo, Fan; Luo, Ya; Ge, Cong; Mo, Qin; Ling, Yajie; Luo, Shu; Tang, Haoru

    2018-04-01

    The FaPR-1 gene was cloned by RT-PCR from `Benihoppe' strawberry and its bioinformatics analysis was conducted. The results showed that the open reading frame was 483 bp encoding encoding l60 amino acids which protein molecular weight and theoretical isoelectricity were 17854.17 and 8.72 respectively. Subcellular localization prediction shows that this gene is located extracellularly. By comparing strawberry FaPR-l and other plant Pathogenesis-related protein, homology and phylogenetic tree construction showed that the homology with grapes, peach is relatively close. In the treatments of ABA, sucrose and the mixture of the two, the expression of FaPR-1 in strawberry fruit were significantly increased.

  13. Gene-expression profiling after exposure to C-ion beams

    International Nuclear Information System (INIS)

    Saegusa, Kumiko; Furuno, Aki; Ishikawa, Kenichi; Ishikawa, Atsuko; Ohtsuka, Yoshimi; Kawai, Seiko; Imai, Takashi; Nojima, Kumie

    2005-01-01

    It is recognized that carbon-ion beam kills cancer cells more efficiently than X-ray. In this study we have compared cellular gene expression response after carbon-ion beam exposure with that after X-ray exposure. Gene expression profiles of cultured neonatal human dermal fibroblasts (NHDF) at 0, 1, 3, 6, 12, 18, and 24 hr after exposure to 0.1, 2 and 5 Gy of X-ray or carbon-ion beam were obtained using 22K oligonucleotide microarray. N-way ANOVA analysis of whole gene expression data sets selected 960 genes for carbon-ion beam and 977 genes for X-ray, respectively. Interestingly, majority of these genes (91% for carbon-ion beam and 88% for X-ray, respectively) were down regulated. The selected genes were further classified by their dose-dependence or time-dependence of gene expression change (fold change>1.5). It was revealed that genes involved in cell proliferation had tendency to show time-dependent up regulation by carbon-ion beam. Another N-way ANOVA analysis was performed to select 510 genes, and further selection was made to find 70 genes that showed radiation species-dependent gene expression change (fold change>1.25). These genes were then categorized by the K-Mean clustering method into 4 clusters. Each cluster showed tendency to contain genes involved in cell cycle regulation, cell death, responses to stress and metabolisms, respectively. (author)

  14. Expression profiling identifies genes involved in emphysema severity

    Directory of Open Access Journals (Sweden)

    Bowman Rayleen V

    2009-09-01

    Full Text Available Abstract Chronic obstructive pulmonary disease (COPD is a major public health problem. The aim of this study was to identify genes involved in emphysema severity in COPD patients. Gene expression profiling was performed on total RNA extracted from non-tumor lung tissue from 30 smokers with emphysema. Class comparison analysis based on gas transfer measurement was performed to identify differentially expressed genes. Genes were then selected for technical validation by quantitative reverse transcriptase-PCR (qRT-PCR if also represented on microarray platforms used in previously published emphysema studies. Genes technically validated advanced to tests of biological replication by qRT-PCR using an independent test set of 62 lung samples. Class comparison identified 98 differentially expressed genes (p p Gene expression profiling of lung from emphysema patients identified seven candidate genes associated with emphysema severity including COL6A3, SERPINF1, ZNHIT6, NEDD4, CDKN2A, NRN1 and GSTM3.

  15. Alteration in CatSper1 and 2 genes expression, sperm parameters and testis histology in varicocelized rats.

    Science.gov (United States)

    Soleimani, Maryam Zohour; Jalali Mashayekhi, Farideh; Mousavi Hasanzade, Morteza; Baazm, Maryam

    2018-03-01

    CatSper gene, a member of cation channel sperm family, has an essential role in sperm motility and male fertility. Following varicocele, sperm parameters especially sperm movement decreases. For this reason, we hypothesized that CatSper gene expression might be reduced after varicocele induction in an animal model. The aim of this study was to evaluate the expression of CatSper 1 and 2 genes, sperm parameters and testis histology following varicocele induction . A total of 30 Wistar male rats were randomly divided into three following groups (n=10/ each): control, sham, and varicocele group. Experimental varicocele was induced by partial ligation of the left renal vein. The epididymal sperm parameters, CatSper 1 and 2 genes expression, and testes histology were studied two months after varicocele induction. Our results revealed that motility (32.73±16.14%), morphology (48.80±17%) and viability (31.23±9.82%) of sperms significantly reduced following varicocele induction. In addition, we showed a significant decrease in the number of spermatogonia (43.63±5.31) and seminiferous tubules diameters (190.51±19.23 mm) in experimental varicocele rats. The level of CatSper 1 and 2 genes expression evaluated using real-time polymerase chain reaction was significantly downregulated 2 months after varicocele induction. Our data indicated that experimental varicocele has deleterious effects on sperm parameters, testis structure as well as the expression of CatSper 1 and 2 genes.

  16. Regulation of Gene Expression in Shewanella oneidensis MR-1 during Electron Acceptor Limitation and Bacterial Nanowire Formation

    Science.gov (United States)

    Barchinger, Sarah E.; Pirbadian, Sahand; Baker, Carol S.; Leung, Kar Man; Burroughs, Nigel J.; El-Naggar, Mohamed Y.

    2016-01-01

    ABSTRACT In limiting oxygen as an electron acceptor, the dissimilatory metal-reducing bacterium Shewanella oneidensis MR-1 rapidly forms nanowires, extensions of its outer membrane containing the cytochromes MtrC and OmcA needed for extracellular electron transfer. RNA sequencing (RNA-Seq) analysis was employed to determine differential gene expression over time from triplicate chemostat cultures that were limited for oxygen. We identified 465 genes with decreased expression and 677 genes with increased expression. The coordinated increased expression of heme biosynthesis, cytochrome maturation, and transport pathways indicates that S. oneidensis MR-1 increases cytochrome production, including the transcription of genes encoding MtrA, MtrC, and OmcA, and transports these decaheme cytochromes across the cytoplasmic membrane during electron acceptor limitation and nanowire formation. In contrast, the expression of the mtrA and mtrC homologs mtrF and mtrD either remains unaffected or decreases under these conditions. The ompW gene, encoding a small outer membrane porin, has 40-fold higher expression during oxygen limitation, and it is proposed that OmpW plays a role in cation transport to maintain electrical neutrality during electron transfer. The genes encoding the anaerobic respiration regulator cyclic AMP receptor protein (CRP) and the extracytoplasmic function sigma factor RpoE are among the transcription factor genes with increased expression. RpoE might function by signaling the initial response to oxygen limitation. Our results show that RpoE activates transcription from promoters upstream of mtrC and omcA. The transcriptome and mutant analyses of S. oneidensis MR-1 nanowire production are consistent with independent regulatory mechanisms for extending the outer membrane into tubular structures and for ensuring the electron transfer function of the nanowires. IMPORTANCE Shewanella oneidensis MR-1 has the capacity to transfer electrons to its external surface

  17. [Blue-light induced expression of S-adenosy-L-homocysteine hydrolase-like gene in Mucor amphibiorum RCS1].

    Science.gov (United States)

    Gao, Ya; Wang, Shu; Fu, Mingjia; Zhong, Guolin

    2013-09-04

    To determine blue-light induced expression of S-adenosyl-L-homocysteine hydrolase-like (sahhl) gene in fungus Mucor amphibiorum RCS1. In the random process of PCR, a sequence of 555 bp was obtained from M. amphibiorum RCS1. The 555 bp sequence was labeled with digoxin to prepare the probe for northern hybridization. By northern hybridization, the transcription of sahhl gene was analyzed in M. amphibiorum RCS1 mycelia culture process from darkness to blue light to darkness. Simultaneously real-time PCR method was used to the sahhl gene expression analysis. Compared with the sequence of sahh gene from Homo sapiens, Mus musculus and some fungi species, a high homology of the 555 bp sequence was confirmed. Therefore, the preliminary confirmation has supported that the 555 bp sequence should be sahhl gene from M. amphibiorum RCS1. Under the dark pre-culture in 24 h, a large amounts of transcript of sahhl gene in the mycelia can be detected by northern hybridization and real-time PCR in the condition of 24 h blue light. But a large amounts of transcript of sahhl gene were not found in other detection for the dark pre-culture of 48 h, even though M. amphibiorum RCS1 mycelia were induced by blue light. Blue light can induce the expression of sahhl gene in the vigorous growth of M. amphibiorum RCS1 mycelia.

  18. Expression of SSX-1 and SSX-5 genes in the peripheral blood of ...

    African Journals Online (AJOL)

    Amal Fawzy

    2013-12-07

    Dec 7, 2013 ... Aim: This study aims to evaluate the SSX-1 and SSX-5 mRNA expression in tumor cells circu- lating in the peripheral blood (PB) of a cohort of Egyptian patients with HCC and to find out any possible associations between these genes expression and different clinical/laboratory parameters. Subjects and ...

  19. Selection of reference genes for quantitative gene expression normalization in flax (Linum usitatissimum L.

    Directory of Open Access Journals (Sweden)

    Neutelings Godfrey

    2010-04-01

    Full Text Available Abstract Background Quantitative real-time PCR (qRT-PCR is currently the most accurate method for detecting differential gene expression. Such an approach depends on the identification of uniformly expressed 'housekeeping genes' (HKGs. Extensive transcriptomic data mining and experimental validation in different model plants have shown that the reliability of these endogenous controls can be influenced by the plant species, growth conditions and organs/tissues examined. It is therefore important to identify the best reference genes to use in each biological system before using qRT-PCR to investigate differential gene expression. In this paper we evaluate different candidate HKGs for developmental transcriptomic studies in the economically-important flax fiber- and oil-crop (Linum usitatissimum L. Results Specific primers were designed in order to quantify the expression levels of 20 different potential housekeeping genes in flax roots, internal- and external-stem tissues, leaves and flowers at different developmental stages. After calculations of PCR efficiencies, 13 HKGs were retained and their expression stabilities evaluated by the computer algorithms geNorm and NormFinder. According to geNorm, 2 Transcriptional Elongation Factors (TEFs and 1 Ubiquitin gene are necessary for normalizing gene expression when all studied samples are considered. However, only 2 TEFs are required for normalizing expression in stem tissues. In contrast, NormFinder identified glyceraldehyde-3-phosphate dehydrogenase (GADPH as the most stably expressed gene when all samples were grouped together, as well as when samples were classed into different sub-groups. qRT-PCR was then used to investigate the relative expression levels of two splice variants of the flax LuMYB1 gene (homologue of AtMYB59. LuMYB1-1 and LuMYB1-2 were highly expressed in the internal stem tissues as compared to outer stem tissues and other samples. This result was confirmed with both ge

  20. Selection of reference genes for quantitative gene expression normalization in flax (Linum usitatissimum L.).

    Science.gov (United States)

    Huis, Rudy; Hawkins, Simon; Neutelings, Godfrey

    2010-04-19

    Quantitative real-time PCR (qRT-PCR) is currently the most accurate method for detecting differential gene expression. Such an approach depends on the identification of uniformly expressed 'housekeeping genes' (HKGs). Extensive transcriptomic data mining and experimental validation in different model plants have shown that the reliability of these endogenous controls can be influenced by the plant species, growth conditions and organs/tissues examined. It is therefore important to identify the best reference genes to use in each biological system before using qRT-PCR to investigate differential gene expression. In this paper we evaluate different candidate HKGs for developmental transcriptomic studies in the economically-important flax fiber- and oil-crop (Linum usitatissimum L). Specific primers were designed in order to quantify the expression levels of 20 different potential housekeeping genes in flax roots, internal- and external-stem tissues, leaves and flowers at different developmental stages. After calculations of PCR efficiencies, 13 HKGs were retained and their expression stabilities evaluated by the computer algorithms geNorm and NormFinder. According to geNorm, 2 Transcriptional Elongation Factors (TEFs) and 1 Ubiquitin gene are necessary for normalizing gene expression when all studied samples are considered. However, only 2 TEFs are required for normalizing expression in stem tissues. In contrast, NormFinder identified glyceraldehyde-3-phosphate dehydrogenase (GADPH) as the most stably expressed gene when all samples were grouped together, as well as when samples were classed into different sub-groups.qRT-PCR was then used to investigate the relative expression levels of two splice variants of the flax LuMYB1 gene (homologue of AtMYB59). LuMYB1-1 and LuMYB1-2 were highly expressed in the internal stem tissues as compared to outer stem tissues and other samples. This result was confirmed with both geNorm-designated- and Norm

  1. The expression of Hedgehog genes (Ihh, Dhh) and Hedgehog target genes (Ptc1, Gli1, Coup-TfII) is affected by estrogenic stimuli in the uterus of immature female rats

    International Nuclear Information System (INIS)

    Katayama, Seiichi; Ashizawa, Koji; Gohma, Hiroshi; Fukuhara, Tadahiro; Narumi, Kazunori; Tsuzuki, Yasuhiro; Tatemoto, Hideki; Nakada, Tadashi; Nagai, Kenji

    2006-01-01

    The objective of this study was to investigate the effects of estrogen receptor (ER) agonists and an ER antagonist on the expression of Hedgehog genes (Indian hedgehog: Ihh; Desert hedgehog: Dhh) and Hedgehog target genes (Patched 1: Ptc1; glioma-associated oncogene homolog 1: Gli1; chicken ovalbumin upstream promoter transcription factor II: Coup-TfII) in the rat uterus. Immature female rats were administered once with 17α-ethynyl estradiol (EE, an ER agonist), propyl pyrazole triole (PPT, an ERα-selective agonist), diarylpropionitrile (DPN, an ERβ-selective agonist), or ICI 182,780 (an ER antagonist). Expression of mRNA for Ihh, Dhh, and Ptc1 was dose-dependently downregulated by EE in the uterus of immature rats, mediated by ER as confirmed by coadministration of ICI 182,780. The mRNA expression levels of Ptc1, Gli1, and Coup-TfII were simultaneously downregulated during the period in which the mRNA expression levels of Ihh and Dhh were downregulated in the uterus after administration of EE. PPT downregulated the transcription of Ihh, Dhh, Ptc1, Gli1, and Coup-TfII, indicating that expression of these genes was regulated by the ERα-dependent pathway. DPN also downregulated the transcription of Ihh and Dhh, although the effect was weaker than that of PPT, indicating that the regulation of uterine Ihh and Dhh transcription was also affected by the ERβ-dependent pathway. These results suggest that the expression of Hedgehog genes (Ihh, Dhh) and Hedgehog target genes (Ptc1, Gli1, Coup-TfII) is affected by estrogenic stimuli in the uterus of immature female rats

  2. The expression of Hedgehog genes (Ihh, Dhh) and Hedgehog target genes (Ptc1, Gli1, Coup-TfII) is affected by estrogenic stimuli in the uterus of immature female rats.

    Science.gov (United States)

    Katayama, Seiichi; Ashizawa, Koji; Gohma, Hiroshi; Fukuhara, Tadahiro; Narumi, Kazunori; Tsuzuki, Yasuhiro; Tatemoto, Hideki; Nakada, Tadashi; Nagai, Kenji

    2006-12-15

    The objective of this study was to investigate the effects of estrogen receptor (ER) agonists and an ER antagonist on the expression of Hedgehog genes (Indian hedgehog: Ihh; Desert hedgehog: Dhh) and Hedgehog target genes (Patched 1: Ptc1; glioma-associated oncogene homolog 1: Gli1; chicken ovalbumin upstream promoter transcription factor II: Coup-TfII) in the rat uterus. Immature female rats were administered once with 17alpha-ethynyl estradiol (EE, an ER agonist), propyl pyrazole triole (PPT, an ERalpha-selective agonist), diarylpropionitrile (DPN, an ERbeta-selective agonist), or ICI 182,780 (an ER antagonist). Expression of mRNA for Ihh, Dhh, and Ptc1 was dose-dependently downregulated by EE in the uterus of immature rats, mediated by ER as confirmed by coadministration of ICI 182,780. The mRNA expression levels of Ptc1, Gli1, and Coup-TfII were simultaneously downregulated during the period in which the mRNA expression levels of Ihh and Dhh were downregulated in the uterus after administration of EE. PPT downregulated the transcription of Ihh, Dhh, Ptc1, Gli1, and Coup-TfII, indicating that expression of these genes was regulated by the ERalpha-dependent pathway. DPN also downregulated the transcription of Ihh and Dhh, although the effect was weaker than that of PPT, indicating that the regulation of uterine Ihh and Dhh transcription was also affected by the ERbeta-dependent pathway. These results suggest that the expression of Hedgehog genes (Ihh, Dhh) and Hedgehog target genes (Ptc1, Gli1, Coup-TfII) is affected by estrogenic stimuli in the uterus of immature female rats.

  3. Expression of porcine myosin heavy chain 1 gene in Berkshire loins ...

    African Journals Online (AJOL)

    Expression of porcine myosin heavy chain 1 gene in Berkshire loins with a high pH24 value. Jin Hun Kang, Woo Young Bang, Eun Jung Kwon, Yong Hwa Lee, Da Hye Park, Eun Seok Cho, Min Ji Kim, Jong-Soon Choi, Hwa Chun Park, Beom Young Park, Chul Wook Kim ...

  4. Sustained expression of a neuron-specific isoform of the Taf1 gene in development stages and aging in mice

    International Nuclear Information System (INIS)

    Jambaldorj, Jamiyansuren; Makino, Satoshi; Munkhbat, Batmunkh; Tamiya, Gen

    2012-01-01

    Highlights: ► We identified the mouse homologue of neuron-specific TAF1 (N-Taf1). ► Taf1 mRNA was expressed in most tissues and cell lines. ► N-Taf1 mRNA was expressed in the brain and Neuroblastoma N2a cell lines. ► Taf1 and N-Taf1 showed different expression profile in development stage and aging. -- Abstract: TATA-box binding protein associated factor 1 (TAF1) protein is the largest and the essential component of the TFIID complex in the pathway of RNA polymerase II–mediated gene transcription, and it regulates transcription of a large number of genes related to cell division. The neuron-specific isoform of the TAF1 gene (N-TAF1), which we reported previously, may have an essential role in neurons through transcriptional regulation of many neuron-specific genes. In the present study, we cloned the full-length cDNA that encodes the mouse homologue of N-TAF1 (N-Taf1) protein. By carrying out of real time RT-PCR, we investigated the expression analysis of the N-Taf1 mRNA in mouse tissues and cell lines. As well as the human N-TAF1, the N-Taf1 showed limited expression in the brain and neuroblastoma, whereas Taf1 expressed elsewhere. Furthermore, in mouse embryo head or mouse brain, mRNA expression of TAF1 changes dramatically during development but N-Taf1 showed sustained expression. Our result suggests that the N-Taf1 gene has an important role in non-dividing neuronal cell rather than in cell division and proliferation during neurogenesis.

  5. Sustained expression of a neuron-specific isoform of the Taf1 gene in development stages and aging in mice

    Energy Technology Data Exchange (ETDEWEB)

    Jambaldorj, Jamiyansuren [Department of Pharmacology, Institute of Health Biosciences, Graduate School, The University of Tokushima, Tokushima 770-8503 (Japan); Advanced Molecular Epidemiology Research Institute, Yamagata University Faculty of Medicine, Yamagata 990-9585 (Japan); Central Scientific Research Laboratory, Institute of Medical Sciences, Ulaanbaatar (Mongolia); Makino, Satoshi, E-mail: smakino@genetix-h.com [Molecular Neuroscience Research Center, Shiga University of Medical Science, Otsu 520-2192 (Japan); Munkhbat, Batmunkh [Central Scientific Research Laboratory, Institute of Medical Sciences, Ulaanbaatar (Mongolia); Tamiya, Gen [Advanced Molecular Epidemiology Research Institute, Yamagata University Faculty of Medicine, Yamagata 990-9585 (Japan)

    2012-08-24

    Highlights: Black-Right-Pointing-Pointer We identified the mouse homologue of neuron-specific TAF1 (N-Taf1). Black-Right-Pointing-Pointer Taf1 mRNA was expressed in most tissues and cell lines. Black-Right-Pointing-Pointer N-Taf1 mRNA was expressed in the brain and Neuroblastoma N2a cell lines. Black-Right-Pointing-Pointer Taf1 and N-Taf1 showed different expression profile in development stage and aging. -- Abstract: TATA-box binding protein associated factor 1 (TAF1) protein is the largest and the essential component of the TFIID complex in the pathway of RNA polymerase II-mediated gene transcription, and it regulates transcription of a large number of genes related to cell division. The neuron-specific isoform of the TAF1 gene (N-TAF1), which we reported previously, may have an essential role in neurons through transcriptional regulation of many neuron-specific genes. In the present study, we cloned the full-length cDNA that encodes the mouse homologue of N-TAF1 (N-Taf1) protein. By carrying out of real time RT-PCR, we investigated the expression analysis of the N-Taf1 mRNA in mouse tissues and cell lines. As well as the human N-TAF1, the N-Taf1 showed limited expression in the brain and neuroblastoma, whereas Taf1 expressed elsewhere. Furthermore, in mouse embryo head or mouse brain, mRNA expression of TAF1 changes dramatically during development but N-Taf1 showed sustained expression. Our result suggests that the N-Taf1 gene has an important role in non-dividing neuronal cell rather than in cell division and proliferation during neurogenesis.

  6. [Gene clone and expression of Barx1 in different tooth of the mini-pig at embryonic day 40].

    Science.gov (United States)

    Zhang, Ying; Yin, Ji-rong; Yang, Kai

    2012-10-01

    To partially clone and compare the quantitative expression of tooth development-related gene Barx1 in different teeth of the mini-pig embryo at embryonic day 40, and to investigate the relationship between Barx1 spatial quantitative expression and tooth morphogenesis. The mini-pig Barx1 genes was partially cloned and the mRNA sequences of human Barx1 genes was aligned with expressed sequence tags (EST) of pig by basic local alignment search tool (BLAST), which were assembled with DNAman v5.2.2. With designed primers, Barx1 was partially cloned in use of reverse transcription polymerase chain reaction (PCR), and tested by BLAST with all the species in NCBI database and confirmed as one part of target gene. Laser capture microdissection was used to collect tooth samples from frozen sections which were prepared before in -80°C freezer. Real-time PCR was carried out to analyze quantitative expression in different teeth. Partial mini-pig Barx1 gene of 698 bp was cloned. Real-time PCR showed that, glyceraldehyde-3-phosphate dehydrogenase used as loading control, the figures of 2(-ΔCT) of lower deciduous incisor, canine, the third premolar and molar were 0.000 249, 0.000 715, 0.026 096 and 0.112 656, respectively. There was a trend of increasing expression from anterior to posterior teeth. Barx1 gene could be related to the number or differentiation of tooth cusps.

  7. Expression of the NRF2 Target Gene NQO1 Is Enhanced in Mononuclear Cells in Human Chronic Kidney Disease

    DEFF Research Database (Denmark)

    Shen, Jianlin; Rasmussen, Marianne; Dong, Qi-Rong

    2017-01-01

    increase in NQO1 gene expression in CKD 1-5 (n = 29; 3.5 for NQO1/ribosomal protein L41; p disease prevalence was higher in CKD 1-5 patients with higher compared to those with lower NQO1 gene expression (p = 0...

  8. A longitudinal study of gene expression in healthy individuals

    Directory of Open Access Journals (Sweden)

    Tessier Michel

    2009-06-01

    Full Text Available Abstract Background The use of gene expression in venous blood either as a pharmacodynamic marker in clinical trials of drugs or as a diagnostic test requires knowledge of the variability in expression over time in healthy volunteers. Here we defined a normal range of gene expression over 6 months in the blood of four cohorts of healthy men and women who were stratified by age (22–55 years and > 55 years and gender. Methods Eleven immunomodulatory genes likely to play important roles in inflammatory conditions such as rheumatoid arthritis and infection in addition to four genes typically used as reference genes were examined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR, as well as the full genome as represented by Affymetrix HG U133 Plus 2.0 microarrays. Results Gene expression levels as assessed by qRT-PCR and microarray were relatively stable over time with ~2% of genes as measured by microarray showing intra-subject differences over time periods longer than one month. Fifteen genes varied by gender. The eleven genes examined by qRT-PCR remained within a limited dynamic range for all individuals. Specifically, for the seven most stably expressed genes (CXCL1, HMOX1, IL1RN, IL1B, IL6R, PTGS2, and TNF, 95% of all samples profiled fell within 1.5–2.5 Ct, the equivalent of a 4- to 6-fold dynamic range. Two subjects who experienced severe adverse events of cancer and anemia, had microarray gene expression profiles that were distinct from normal while subjects who experienced an infection had only slightly elevated levels of inflammatory markers. Conclusion This study defines the range and variability of gene expression in healthy men and women over a six-month period. These parameters can be used to estimate the number of subjects needed to observe significant differences from normal gene expression in clinical studies. A set of genes that varied by gender was also identified as were a set of genes with elevated

  9. Differential neutrophil gene expression in early bovine pregnancy

    Directory of Open Access Journals (Sweden)

    Kizaki Keiichiro

    2013-02-01

    Full Text Available Abstract Background In food production animals, especially cattle, the diagnosis of gestation is important because the timing of gestation directly affects the running of farms. Various methods have been used to detect gestation, but none of them are ideal because of problems with the timing of detection or the accuracy, simplicity, or cost of the method. A new method for detecting gestation, which involves assessing interferon-tau (IFNT-stimulated gene expression in peripheral blood leukocytes (PBL, was recently proposed. PBL fractionation methods were used to examine whether the expression profiles of various PBL populations could be used as reliable diagnostic markers of bovine gestation. Methods PBL were collected on days 0 (just before artificial insemination, 7, 14, 17, 21, and 28 of gestation. The gene expression levels of the PBL were assessed with microarray analysis and/or quantitative real-time reverse transcription (q PCR. PBL fractions were collected by flow cytometry or density gradient cell separation using Histopaque 1083 or Ficoll-Conray solutions. The expression levels of four IFNT-stimulated genes, interferon-stimulated protein 15 kDa (ISG15, myxovirus-resistance (MX 1 and 2, and 2′-5′-oligoadenylate synthetase (OAS1, were then analyzed in each fraction through day 28 of gestation using qPCR. Results Microarray analysis detected 72 and 28 genes in whole PBL that were significantly higher on days 14 and 21 of gestation, respectively, than on day 0. The upregulated genes included IFNT-stimulated genes. The expression levels of these genes increased with the progression of gestation until day 21. In flow cytometry experiments, on day 14 the expression levels of all of the genes were significantly higher in the granulocyte fraction than in the other fractions. Their expression gradually decreased through day 28 of gestation. Strong correlations were observed between the expression levels of the four genes in the granulocyte

  10. LINE FUSION GENES: a database of LINE expression in human genes

    Directory of Open Access Journals (Sweden)

    Park Hong-Seog

    2006-06-01

    Full Text Available Abstract Background Long Interspersed Nuclear Elements (LINEs are the most abundant retrotransposons in humans. About 79% of human genes are estimated to contain at least one segment of LINE per transcription unit. Recent studies have shown that LINE elements can affect protein sequences, splicing patterns and expression of human genes. Description We have developed a database, LINE FUSION GENES, for elucidating LINE expression throughout the human gene database. We searched the 28,171 genes listed in the NCBI database for LINE elements and analyzed their structures and expression patterns. The results show that the mRNA sequences of 1,329 genes were affected by LINE expression. The LINE expression types were classified on the basis of LINEs in the 5' UTR, exon or 3' UTR sequences of the mRNAs. Our database provides further information, such as the tissue distribution and chromosomal location of the genes, and the domain structure that is changed by LINE integration. We have linked all the accession numbers to the NCBI data bank to provide mRNA sequences for subsequent users. Conclusion We believe that our work will interest genome scientists and might help them to gain insight into the implications of LINE expression for human evolution and disease. Availability http://www.primate.or.kr/line

  11. LPL gene expression is associated with poor prognosis in CLL and closely related to NOTCH1 mutations

    DEFF Research Database (Denmark)

    Kristensen, Louise; Kielsgaard Kristensen, Thomas; Abildgaard, Niels

    2016-01-01

    these markers. AIM: To evaluate LPL gene expression together with the well-established prognostic markers of CLL and investigate correlations with more recently identified prognostic markers, NOTCH1 and TP53 mutations. METHODS: On 149 patients LPL gene expression was analyzed by real-time RT-PCR. Exon 34...... of NOTCH1 was PCR amplified and directly sequenced. RESULTS: LPL gene expression could be measured as a categorical variable (LPL+/LPL-) and was associated with time to treatment (p... and new prognostic markers, 3 out of 4 patients, who received treatment within 24 months after diagnosis, were LPL+ (p=0.03). There was a strong correlation between NOTCH1 mutation and LPL+ (p=0.005). The unfavorable prognosis of LPL+ was maintained in CLL with wild-type NOTCH1. CONCLUSIONS: NOTCH1...

  12. Alteration of BRCA1 expression affects alcohol-induced transcription of RNA Pol III-dependent genes.

    Science.gov (United States)

    Zhong, Qian; Shi, Ganggang; Zhang, Yanmei; Lu, Lei; Levy, Daniel; Zhong, Shuping

    2015-02-01

    Emerging evidence has indicated that alcohol consumption is an established risk factor for breast cancer. Deregulation of RNA polymerase III (Pol III) transcription enhances cellular Pol III gene production, leading to an increase in translational capacity to promote cell transformation and tumor formation. We have reported that alcohol intake increases Pol III gene transcription to promote cell transformation and tumor formation in vitro and in vivo. Studies revealed that tumor suppressors, pRb, p53, PTEN and Maf1 repress the transcription of Pol III genes. BRCA1 is a tumor suppressor and its mutation is tightly related to breast cancer development. However, it is not clear whether BRCA1 expression affects alcohol-induced transcription of Pol III genes. At the present studies, we report that restoring BRCA1 in HCC 1937 cells, which is a BRCA1 deficient cell line, represses Pol III gene transcription. Expressing mutant or truncated BRCA1 in these cells does not affect the ability of repression on Pol III genes. Our analysis has demonstrated that alcohol induces Pol III gene transcription. More importantly, overexpression of BRCA1 in estrogen receptor positive (ER+) breast cancer cells (MCF-7) decreases the induction of tRNA(Leu) and 5S rRNA genes by alcohol, whereas reduction of BRCA1 by its siRNA slightly increases the transcription of the class of genes. This suggests that BRCA1 is associated with alcohol-induced deregulation of Pol III genes. These studies for the first time demonstrate the role of BRCA1 in induction of Pol III genes by alcohol and uncover a novel mechanism of alcohol-associated breast cancer. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. The analysis of correlation between IL-1B gene expression and genotyping in multiple sclerosis patients.

    Science.gov (United States)

    Heidary, Masoumeh; Rakhshi, Nahid; Pahlevan Kakhki, Majid; Behmanesh, Mehrdad; Sanati, Mohammad Hossein; Sanadgol, Nima; Kamaladini, Hossein; Nikravesh, Abbas

    2014-08-15

    IL-1B is released by monocytes, astrocytes and brain endothelial cells and seems to be involved in inflammatory reactions of the central nervous system (CNS) in multiple sclerosis (MS). This study aims to evaluate the expression level of IL-1B mRNA in peripheral blood mononuclear cells (PBMCs), genotype the rs16944 SNP and find out the role of this SNP on the expression level of IL-1B in MS patients. We found that the expression level of IL-1B in MS patients increased 3.336 times more than controls in PBMCs but the rs16944 SNP in the promoter region of IL-1B did not affect the expression level of this gene and there was not association of this SNP with MS in the examined population. Also, our data did not reveal any correlation between normalized expressions of IL-1B gene with age of participants, age of onset, and disease duration. Copyright © 2014 Elsevier B.V. All rights reserved.

  14. Effects and mechanism of integrin-β1 gene expression inhibited by shRNA in invasion of pancreatic carcinoma PANC-1 cells.

    Science.gov (United States)

    Yu, Feng; Li, Hua; Bu, Xuefeng; Zhang, Yongjun

    2012-01-01

    To investigate the effects of integrin-β1 gene expression inhibited by shRNA on invasion of pancreatic carcinoma PANC-1 cells in vitro. The eukaryotic expression plasmid of short hairpin RNA (shRNA) targeting integrin-β1 gene (integrin-β1-shRNA) was constructed and transfected into PANC-1 cells. The expressions of integrin-β1 mRNA and protein were detected by real-time quantitative polymerase chain reaction (PCR) and western blot assay, respectively. The invasive ability of PANC-1 cells was observed with a transwell cell culture chamber and the expressions of MMP-2 and MMP-9 were assayed. Compared to the untransfected group, recombinant expression plasmid integrin-β1-shRNA resulted in reduction of integrin-β1 mRNA and protein by 78.58%±7.24% and 92.88%±3.18%, respectively and the average number of invading PANC-1 cells were decreased from 52±5 to 21±4 (pPANC-1 cells in vitro significantly.

  15. The porcine lymphotropic herpesvirus 1 encodes functional regulators of gene expression

    International Nuclear Information System (INIS)

    Lindner, I.; Ehlers, B.; Noack, S.; Dural, G.; Yasmum, N.; Bauer, C.; Goltz, M.

    2007-01-01

    The porcine lymphotropic herpesviruses (PLHV) are discussed as possible risk factors in xenotransplantation because of the high prevalence of PLHV-1, PLHV-2 and PLHV-3 in pig populations world-wide and the fact that PLHV-1 has been found to be associated with porcine post-transplant lymphoproliferative disease. To provide structural and functional knowledge on the PLHV immediate-early (IE) transactivator genes, the central regions of the PLHV genomes were characterized by genome walking, sequence and splicing analysis. Three spliced genes were identified (ORF50, ORFA6/BZLF1 h , ORF57) encoding putative IE transactivators, homologous to (i) ORF50 and BRLF1/Rta (ii) K8/K-bZIP and BZLF1/Zta and (iii) ORF57 and BMLF1 of HHV-8 and EBV, respectively. Expressed as myc-tag or HA-tag fusion proteins, they were located to the cellular nucleus. In reporter gene assays, several PLHV-promoters were mainly activated by PLHV-1 ORF50, to a lower level by PLHV-1 ORFA6/BZLF1 h and not by PLHV-1 ORF57. However, the ORF57-encoded protein acted synergistically on ORF50-mediated activation

  16. Low-dose Norfloxacin-treated leptospires induce less IL-1β release in J774A.1 cells following discrepant leptospiral gene expression.

    Science.gov (United States)

    Cao, Yongguo; Xie, Xufeng; Zhang, Wenlong; Wu, Dianjun; Tu, Changchun

    2018-06-01

    Currently, accumulating evidence is challenging subtherapeutic therapy. Low-dose Norfloxacin (Nor) has been reported to suppress the immune response and worsen leptospirosis. In this study, we investigated the influence of low-dose Nor (0.03 μg/ml, 0.06 μg/ml, 0.125 μg/ml) on leptospiral gene expression and analyzed the immunomodulatory effects of low-dose Nor-treated leptospires in J774A.1 cells. To study the expression profiles of low-dose Nor-treated leptospires, we chose LipL71/LipL21 as reference genes determined by the geNorm applet in this experiment. The results showed that low-dose Nor up-regulated the expression of FlaB and inhibited the expression of 16S rRNA, LipL32, LipL41, Loa22, KdpA, and KdpB compared with the untreated leptospires. These results indicated that low-dose Nor could regulate leptospiral gene expression. Using RT-PCR, the gene expression of IL-1β and TNF-α in J774A.1 cells was detected. Nor-treated leptospires induced higher expression levels of both IL-1β and TNF-α. However, when analyzed by ELISA, the release of mature IL-1β was reduced compared with that observed in cells induced with no Nor-treated leptospires, although the TNF-α protein level showed no significant change. Our study indicated that the gene expression of leptospires could be modulated by low-dose Nor, which induced less IL-1β release in J774A.1 cells. Copyright © 2018 Elsevier Ltd. All rights reserved.

  17. Inhibition of HIV-1 Integrase gene expression by 10-23 DNAzyme

    Indian Academy of Sciences (India)

    We have designed three novel DNAzymes, DIN54, DIN116, and DIN152, against HIV-1 Integrase gene using Mfold software and evaluated them for target site cleavage activity on the in vitro transcribed mRNA. All DNAzymes were tested for its inhibition of expression of HIV Integrase protein in the transiently transfected cell ...

  18. Expression profile and distribution of Efhc1 gene transcript during rodent brain development.

    Science.gov (United States)

    Conte, Fábio F; Ribeiro, Patrícia A O; Marchesini, Rafael B; Pascoal, Vinícius D B; Silva, Joelcimar M; Oliveira, Amanda R; Gilioli, Rovílson; Sbragia, Lourenço; Bittencourt, Jackson C; Lopes-Cendes, Iscia

    2009-09-01

    One of the putative causative genes for juvenile myoclonic epilepsy (JME) is EFHC1. We report here the expression profile and distribution of Efhc1 messenger RNA (mRNA) during mouse and rat brain development. Real-time polymerase chain reaction revealed that there is no difference in the expression of Efhc1 mRNA between right and left hemispheres in both species. In addition, the highest levels of Efhc1 mRNA were found at intra-uterine stages in mouse and in adulthood in rat. In common, there was a progressive decrease in Efhc1 expression from 1-day-old neonates to 14-day-old animals in both species. In situ hybridization studies showed that rat and mouse Efhc1 mRNAs are expressed in ependymal cells of ventricle walls. Our findings suggest that Efhc1 expression is more important during initial phases of brain development and that at this stage it could be involved in key developmental mechanisms underlying JME.

  19. Clock-controlled output gene Dbp is a regulator of Arnt/Hif-1β gene expression in pancreatic islet β-cells

    International Nuclear Information System (INIS)

    Nakabayashi, Hiroko; Ohta, Yasuharu; Yamamoto, Masayoshi; Susuki, Yosuke; Taguchi, Akihiko; Tanabe, Katsuya; Kondo, Manabu; Hatanaka, Masayuki; Nagao, Yuko; Tanizawa, Yukio

    2013-01-01

    Highlights: •Arnt mRNA expressed in a circadian manner in mouse pancreatic islets. •Expressions of Dbp and Arnt damped in the islets of a diabetic model mouse. •DBP and E4BP4 regulate Arnt promoter activity by direct binding. •Arnt may have a role in connecting circadian rhythm and metabolism. -- Abstract: Aryl hydrocarbon receptor nuclear translocator (ARNT)/hypoxia inducible factor-1β (HIF-1β) has emerged as a potential determinant of pancreatic β-cell dysfunction and type 2 diabetes in humans. An 82% reduction in Arnt expression was observed in islets from type 2 diabetic donors as compared to non-diabetic donors. However, few regulators of Arnt expression have been identified. Meanwhile, disruption of the clock components CLOCK and BMAL1 is known to result in hypoinsulinemia and diabetes, but the molecular details remain unclear. In this study, we identified a novel molecular connection between Arnt and two clock-controlled output genes, albumin D-element binding protein (Dbp) and E4 binding protein 4 (E4bp4). By conducting gene expression studies using the islets of Wfs1 −/− A y /a mice that develop severe diabetes due to β-cell apoptosis, we demonstrated clock-related gene expressions to be altered in the diabetic mice. Dbp mRNA decreased by 50%, E4bp4 mRNA increased by 50%, and Arnt mRNA decreased by 30% at Zeitgever Time (ZT) 12. Mouse pancreatic islets exhibited oscillations of clock gene expressions. E4BP4, a D-box negative regulator, oscillated anti-phase to DBP, a D-box positive regulator. We also found low-amplitude circadian expression of Arnt mRNA, which peaked at ZT4. Over-expression of DBP raised both mRNA and protein levels of ARNT in HEK293 and MIN6 cell lines. Arnt promoter-driven luciferase reporter assay in MIN6 cells revealed that DBP increased Arnt promoter activity by 2.5-fold and that E4BP4 competitively inhibited its activation. In addition, on ChIP assay, DBP and E4BP4 directly bound to D-box elements within the Arnt

  20. Clock-controlled output gene Dbp is a regulator of Arnt/Hif-1β gene expression in pancreatic islet β-cells

    Energy Technology Data Exchange (ETDEWEB)

    Nakabayashi, Hiroko; Ohta, Yasuharu, E-mail: yohta@yamaguchi-u.ac.jp; Yamamoto, Masayoshi; Susuki, Yosuke; Taguchi, Akihiko; Tanabe, Katsuya; Kondo, Manabu; Hatanaka, Masayuki; Nagao, Yuko; Tanizawa, Yukio, E-mail: tanizawa@yamaguchi-u.ac.jp

    2013-05-03

    Highlights: •Arnt mRNA expressed in a circadian manner in mouse pancreatic islets. •Expressions of Dbp and Arnt damped in the islets of a diabetic model mouse. •DBP and E4BP4 regulate Arnt promoter activity by direct binding. •Arnt may have a role in connecting circadian rhythm and metabolism. -- Abstract: Aryl hydrocarbon receptor nuclear translocator (ARNT)/hypoxia inducible factor-1β (HIF-1β) has emerged as a potential determinant of pancreatic β-cell dysfunction and type 2 diabetes in humans. An 82% reduction in Arnt expression was observed in islets from type 2 diabetic donors as compared to non-diabetic donors. However, few regulators of Arnt expression have been identified. Meanwhile, disruption of the clock components CLOCK and BMAL1 is known to result in hypoinsulinemia and diabetes, but the molecular details remain unclear. In this study, we identified a novel molecular connection between Arnt and two clock-controlled output genes, albumin D-element binding protein (Dbp) and E4 binding protein 4 (E4bp4). By conducting gene expression studies using the islets of Wfs1{sup −/−} A{sup y}/a mice that develop severe diabetes due to β-cell apoptosis, we demonstrated clock-related gene expressions to be altered in the diabetic mice. Dbp mRNA decreased by 50%, E4bp4 mRNA increased by 50%, and Arnt mRNA decreased by 30% at Zeitgever Time (ZT) 12. Mouse pancreatic islets exhibited oscillations of clock gene expressions. E4BP4, a D-box negative regulator, oscillated anti-phase to DBP, a D-box positive regulator. We also found low-amplitude circadian expression of Arnt mRNA, which peaked at ZT4. Over-expression of DBP raised both mRNA and protein levels of ARNT in HEK293 and MIN6 cell lines. Arnt promoter-driven luciferase reporter assay in MIN6 cells revealed that DBP increased Arnt promoter activity by 2.5-fold and that E4BP4 competitively inhibited its activation. In addition, on ChIP assay, DBP and E4BP4 directly bound to D-box elements within the

  1. Small-molecule screen identifies modulators of EWS/FLI1 target gene expression and cell survival in Ewing's sarcoma.

    Science.gov (United States)

    Boro, Aleksandar; Prêtre, Kathya; Rechfeld, Florian; Thalhammer, Verena; Oesch, Susanne; Wachtel, Marco; Schäfer, Beat W; Niggli, Felix K

    2012-11-01

    Ewing's sarcoma family of tumors (EFT) is characterized by the presence of chromosomal translocations leading to the expression of oncogenic transcription factors such as, in the majority of cases, EWS/FLI1. Because of its key role in Ewing's sarcoma development and maintenance, EWS/FLI1 represents an attractive therapeutic target. Here, we characterize PHLDA1 as a novel direct target gene whose expression is repressed by EWS/FLI1. Using this gene and additional specific well-characterized target genes such as NROB1, NKX2.2 and CAV1, all activated by EWS/FLI1, as a read-out system, we screened a small-molecule compound library enriched for FDA-approved drugs that modulated the expression of EWS/FLI1 target genes. Among a hit-list of nine well-known drugs such as camptothecin, fenretinide, etoposide and doxorubicin, we also identified the kinase inhibitor midostaurin (PKC412). Subsequent experiments demonstrated that midostaurin is able to induce apoptosis in a panel of six Ewing's sarcoma cell lines in vitro and can significantly suppress xenograft tumor growth in vivo. These results suggest that midostaurin might be a novel drug that is active against Ewing's cells, which might act by modulating the expression of EWS/FLI1 target genes. Copyright © 2012 UICC.

  2. Gene expression profiling following NRF2 and KEAP1 siRNA knockdown in human lung fibroblasts identifies CCL11/Eotaxin-1 as a novel NRF2 regulated gene

    Science.gov (United States)

    2012-01-01

    Background Oxidative Stress contributes to the pathogenesis of many diseases. The NRF2/KEAP1 axis is a key transcriptional regulator of the anti-oxidant response in cells. Nrf2 knockout mice have implicated this pathway in regulating inflammatory airway diseases such as asthma and COPD. To better understand the role the NRF2 pathway has on respiratory disease we have taken a novel approach to define NRF2 dependent gene expression in a relevant lung system. Methods Normal human lung fibroblasts were transfected with siRNA specific for NRF2 or KEAP1. Gene expression changes were measured at 30 and 48 hours using a custom Affymetrix Gene array. Changes in Eotaxin-1 gene expression and protein secretion were further measured under various inflammatory conditions with siRNAs and pharmacological tools. Results An anti-correlated gene set (inversely regulated by NRF2 and KEAP1 RNAi) that reflects specific NRF2 regulated genes was identified. Gene annotations show that NRF2-mediated oxidative stress response is the most significantly regulated pathway, followed by heme metabolism, metabolism of xenobiotics by Cytochrome P450 and O-glycan biosynthesis. Unexpectedly the key eosinophil chemokine Eotaxin-1/CCL11 was found to be up-regulated when NRF2 was inhibited and down-regulated when KEAP1 was inhibited. This transcriptional regulation leads to modulation of Eotaxin-1 secretion from human lung fibroblasts under basal and inflammatory conditions, and is specific to Eotaxin-1 as NRF2 or KEAP1 knockdown had no effect on the secretion of a set of other chemokines and cytokines. Furthermore, the known NRF2 small molecule activators CDDO and Sulphoraphane can also dose dependently inhibit Eotaxin-1 release from human lung fibroblasts. Conclusions These data uncover a previously unknown role for NRF2 in regulating Eotaxin-1 expression and further the mechanistic understanding of this pathway in modulating inflammatory lung disease. PMID:23061798

  3. The hepatic Raldh1 expression is elevated in Zucker fatty rats and its over-expression introduced the retinal-induced Srebp-1c expression in INS-1 cells.

    Directory of Open Access Journals (Sweden)

    Yang Li

    Full Text Available The roles of vitamin A (VA in the development of metabolic diseases remain unanswered. We have reported that retinoids synergized with insulin to induce the expression of sterol-regulatory element-binding protein 1c gene (Srebp-1c expression in primary rat hepatocytes. Additionally, the hepatic Srebp-1c expression is elevated in Zucker fatty (ZF rats, and reduced in those fed a VA deficient diet. VA is metabolized to retinoic acid (RA for regulating gene expression. We hypothesized that the expression of RA production enzymes contributes to the regulation of the hepatic Srebp-1c expression. Therefore, we analyzed their expression levels in Zucker lean (ZL and ZF rats. The mRNA levels of retinaldehyde dehydrogenase family 1 gene (Raldh1 were found to be higher in the isolated and cultured primary hepatocytes from ZF rats than that from ZL rats. The RALDH1 protein level was elevated in the liver of ZF rats. Retinol and retinal dose- and time-dependently induced the expression of RA responsive Cyp26a1 gene in hepatocytes and hepatoma cells. INS-1 cells were identified as an ideal tool to study the effects of RA production on the regulation of gene expression because only RA, but not retinal, induced Srebp-1c mRNA expression in them. Recombinant adenovirus containing rat Raldh1 cDNA was made and used to infect INS-1 cells. The over-expression of RALDH1 introduced the retinal-mediated induction of Srebp-1c expression in INS-1 cells. We conclude that the expression levels of the enzymes for RA production may contribute to the regulation of RA responsive genes, and determine the responses of the cells to retinoid treatments. The elevated hepatic expression of Raldh1 in ZF rats may cause the excessive RA production from retinol, and in turn, result in higher Srebp-1c expression. This excessive RA production may be one of the factors contributing to the elevated lipogenesis in the liver of ZF rats.

  4. SATB1 packages densely-looped, transciptionally-active chromatinfor coordinated expression of cytokine genes

    Energy Technology Data Exchange (ETDEWEB)

    Cai, Shutao; Lee, Charles C.; Kohwi-Shigematsu, Terumi

    2006-05-23

    SATB1 is an important regulator of nuclear architecture that anchors specialized DNA sequences onto its cage-like network and recruits chromatin remodeling/modifying factors to control gene transcription. We studied the role of SATB1 in regulating the coordinated expression of Il5, Il4, and Il13 from the 200kb cytokine gene cluster region of mouse chromosome 11 during T-helper 2 (Th2)-cell activation. We show that upon cell activation, SATB1 is rapidly induced to form a unique transcriptionally-active chromatin structure that includes the cytokine gene region. Chromatin is folded into numerous small loops all anchored by SATB1, is histone H3 acetylated at lysine 9/14, and associated with Th2-specific factors, GATA3, STAT6, c-Maf, the chromatin-remodeling enzyme Brg-1, and RNA polymerase II across the 200kb region. Before activation, the chromatin displays some of these features, such as association with GATA3 and STAT6, but these were insufficient for cytokine gene expression. Using RNA interference (RNAi), we show that upon cell activation, SATB1 is not only required for chromatin folding into dense loops, but also for c-Maf induction and subsequently for Il4, Il5, and Il13 transcription. Our results show that SATB1 is an important determinant for chromatin architecture that constitutes a novel higher-order, transcriptionally-active chromatin structure upon Th2-cell activation.

  5. Porcine Knock-in Fibroblasts Expressing hDAF on α-1,3-Galactosyltransferase (GGTA1) Gene Locus.

    Science.gov (United States)

    Kim, Ji Woo; Kim, Hye-Min; Lee, Sang Mi; Kang, Man-Jong

    2012-10-01

    The Galactose-α1,3-galactose (α1,3Gal) epitope is responsible for hyperacute rejection in pig-to-human xenotransplantation. Human decay-accelerating factor (hDAF) is a cell surface regulatory protein that serves as a complement inhibitor to protect self cells from complement attack. The generation of α1,3-galactosyltransferase (GGTA1) knock-out pigs expressing DAF is a necessary step for their use as organ donors for humans. In this study, we established GGTA1 knock-out cell lines expressing DAF from pig ear fibroblasts for somatic cell nuclear transfer. hDAF expression was detected in hDAF knock-in heterozygous cells, but not in normal pig cells. Expression of the GGTA1 gene was lower in the knock-in heterozygous cell line compared to the normal pig cell. Knock-in heterozygous cells afforded more effective protection against cytotoxicity with human serum than with GGTA1 knock-out heterozygous and control cells. These cell lines may be used in the production of GGTA1 knock-out and DAF expression pigs for xenotransplantation.

  6. FARO server: Meta-analysis of gene expression by matching gene expression signatures to a compendium of public gene expression data

    DEFF Research Database (Denmark)

    Manijak, Mieszko P.; Nielsen, Henrik Bjørn

    2011-01-01

    circumvented by instead matching gene expression signatures to signatures of other experiments. FINDINGS: To facilitate this we present the Functional Association Response by Overlap (FARO) server, that match input signatures to a compendium of 242 gene expression signatures, extracted from more than 1700...... Arabidopsis microarray experiments. CONCLUSIONS: Hereby we present a publicly available tool for robust characterization of Arabidopsis gene expression experiments which can point to similar experimental factors in other experiments. The server is available at http://www.cbs.dtu.dk/services/faro/....

  7. Validation of reference genes for quantitative RT-PCR studies of gene expression in perennial ryegrass (Lolium perenne L.

    Directory of Open Access Journals (Sweden)

    Thrush Anthony

    2010-01-01

    Full Text Available Abstract Background Perennial ryegrass (Lolium perenne L. is an important pasture and turf crop. Biotechniques such as gene expression studies are being employed to improve traits in this temperate grass. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR is among the best methods available for determining changes in gene expression. Before analysis of target gene expression, it is essential to select an appropriate normalisation strategy to control for non-specific variation between samples. Reference genes that have stable expression at different biological and physiological states can be effectively used for normalisation; however, their expression stability must be validated before use. Results Existing Serial Analysis of Gene Expression data were queried to identify six moderately expressed genes that had relatively stable gene expression throughout the year. These six candidate reference genes (eukaryotic elongation factor 1 alpha, eEF1A; TAT-binding protein homolog 1, TBP-1; eukaryotic translation initiation factor 4 alpha, eIF4A; YT521-B-like protein family protein, YT521-B; histone 3, H3; ubiquitin-conjugating enzyme, E2 were validated for qRT-PCR normalisation in 442 diverse perennial ryegrass (Lolium perenne L. samples sourced from field- and laboratory-grown plants under a wide range of experimental conditions. Eukaryotic EF1A is encoded by members of a multigene family exhibiting differential expression and necessitated the expression analysis of different eEF1A encoding genes; a highly expressed eEF1A (h, a moderately, but stably expressed eEF1A (s, and combined expression of multigene eEF1A (m. NormFinder identified eEF1A (s and YT521-B as the best combination of two genes for normalisation of gene expression data in perennial ryegrass following different defoliation management in the field. Conclusions This study is unique in the magnitude of samples tested with the inclusion of numerous field-grown samples

  8. Repair genes expression profile of MLH1, MSH2 and ATM in the normal oral mucosa of chronic smokers.

    Science.gov (United States)

    Alves, Mônica Ghislaine Oliveira; Carta, Celina Faig Lima; de Barros, Patrícia Pimentel; Issa, Jaqueline Scholz; Nunes, Fábio Daumas; Almeida, Janete Dias

    2017-01-01

    The aim of this study was to evaluate the effect of chronic smoking on the expression profile of the repair genes MLH1, MSH2 and ATM in the normal oral mucosa of chronic smokers and never smokers. The sample consisted of thirty exfoliative cytology smears per group obtained from Smokers and Never Smokers. Total RNA was extracted and expression of the MLH1, MSH2 and ATM genes were evaluated by quantitative real-time and immunocytochemistry. The gene and protein expression data were correlated to the clinical data. Gene expression was analyzed statistically using the Student t-test and Pearson's correlation coefficient, with pMLH1, MSH2 and ATM genes were downregulated in the smoking group compared to the control with significant values for MLH1 (p=0.006), MSH2 (p=0.0001) and ATM (p=0.0001). Immunocytochemical staining for anti-MLH1, anti-MSH2 and anti-ATM was negative in Never Smokers; in Smokers it was rarely positive. No significant correlation was observed among the expression of MLH1, MSH2, ATM and age, number of cigarettes consumed per day, time of smoking during life, smoking history or levels of CO in expired air. The expression of genes and proteins related to DNA repair mechanism MLH1, MSH2 and ATM in the normal oral mucosa of chronic smokers was reduced. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Bcmimp1, a Botrytis cinerea gene transiently expressed in planta, encodes a mitochondrial protein

    Directory of Open Access Journals (Sweden)

    David eBenito-Pescador

    2016-02-01

    Full Text Available Botrytis cinerea is a widespread necrotrophic fungus which infects more than 200 plant species. In an attempt to characterize the physiological status of the fungus in planta and to identify genetic factors contributing to its ability to infect the host cells, a differential gene expression analysis during the interaction B. cinerea-tomato was carried out. Gene Bcmimp1 codes for a mRNA detected by differential display in the course of this analysis. During the interaction with the host, it shows a transient expression pattern with maximal expression levels during the colonization and maceration of the infected tissues. Bioinformatic analysis suggested that BCMIMP1 is an integral membrane protein located in the mitochondrial inner membrane. Co-localization experiments with a BCMIMP1-GFP fusion protein confirmed that the protein is targeted to the mitochondria. ΔBcmimp1 mutants do not show obvious phenotypic differences during saprophytic growth and their infection ability was unaltered as compared to the wild-type. Interestingly, the mutants produced increased levels of ROS, likely as a consequence of disturbed mitochondrial function. Although Bcmimp1 expression is enhanced in planta it cannot be considered a pathogenicity factor.

  10. Regulation of Nuclear Receptor Interacting Protein 1 (NRIP1) Gene Expression in Response to Weight Loss and Exercise in Humans

    DEFF Research Database (Denmark)

    De Marinis, Yang Z; Sun, Jiangming; Bompada, Pradeep

    2017-01-01

    Objective: Nuclear receptor interacting protein 1 (NRIP1) is an important energy regulator, but few studies have addressed its role in humans. This study investigated adipose tissue and skeletal muscle NRIP1 gene expression and serum levels in response to weight loss and exercise in humans. Methods...... network/module. Conclusions: NRIP1 gene expression and serum levels are strongly associated with metabolic states such as obesity, weight loss, different types of exercise, and peripheral tissue insulin resistance, potentially as a mediator of sedentary effects.......: In patients with obesity, adipose tissue NRIP1 mRNA expression increased during weight loss and weight maintenance and showed strong associations with metabolic markers and anthropometric parameters. Serum NRIP1 protein levels also increased after weight loss. In skeletal muscle, imposed rest increased NRIP1...

  11. Altered expression of hypoxia-inducible factor-1α (HIF-1α and its regulatory genes in gastric cancer tissues.

    Directory of Open Access Journals (Sweden)

    Jihan Wang

    Full Text Available Tissue hypoxia induces reprogramming of cell metabolism and may result in normal cell transformation and cancer progression. Hypoxia-inducible factor 1-alpha (HIF-1α, the key transcription factor, plays an important role in gastric cancer development and progression. This study aimed to investigate the underlying regulatory signaling pathway in gastric cancer using gastric cancer tissue specimens. The integration of gene expression profile and transcriptional regulatory element database (TRED was pursued to identify HIF-1α ↔ NFκB1 → BRCA1 → STAT3 ← STAT1 gene pathways and their regulated genes. The data showed that there were 82 differentially expressed genes that could be regulated by these five transcription factors in gastric cancer tissues and these genes formed 95 regulation modes, among which seven genes (MMP1, TIMP1, TLR2, FCGR3A, IRF1, FAS, and TFF3 were hub molecules that are regulated at least by two of these five transcription factors simultaneously and were associated with hypoxia, inflammation, and immune disorder. Real-Time PCR and western blot showed increasing of HIF-1α in mRNA and protein levels as well as TIMP1, TFF3 in mRNA levels in gastric cancer tissues. The data are the first study to demonstrate HIF-1α-regulated transcription factors and their corresponding network genes in gastric cancer. Further study with a larger sample size and more functional experiments is needed to confirm these data and then translate into clinical biomarker discovery and treatment strategy for gastric cancer.

  12. Pterostilbene acts through metastasis-associated protein 1 to inhibit tumor growth, progression and metastasis in prostate cancer.

    Directory of Open Access Journals (Sweden)

    Kun Li

    Full Text Available The development of natural product agents with targeted strategies holds promise for enhanced anticancer therapy with reduced drug-associated side effects. Resveratrol found in red wine, has anticancer activity in various tumor types. We reported earlier on a new molecular target of resveratrol, the metastasis-associated protein 1 (MTA1, which is a part of nucleosome remodeling and deacetylation (NuRD co-repressor complex that mediates gene silencing. We identified resveratrol as a regulator of MTA1/NuRD complex and re-activator of p53 acetylation in prostate cancer (PCa. In the current study, we addressed whether resveratrol analogues also possess the ability to inhibit MTA1 and to reverse p53 deacetylation. We demonstrated that pterostilbene (PTER, found in blueberries, had greater increase in MTA1-mediated p53 acetylation, confirming superior potency over resveratrol as dietary epigenetic agent. In orthotopic PCa xenografts, resveratrol and PTER significantly inhibited tumor growth, progression, local invasion and spontaneous metastasis. Furthermore, MTA1-knockdown sensitized cells to these agents resulting in additional reduction of tumor progression and metastasis. The reduction was dependent on MTA1 signaling showing increased p53 acetylation, higher apoptotic index and less angiogenesis in vivo in all xenografts treated with the compounds, and particularly with PTER. Altogether, our results indicate MTA1 as a major contributor in prostate tumor malignant progression, and support the use of strategies targeting MTA1. Our strong pre-clinical data indicate PTER as a potent, selective and pharmacologically safe natural product that may be tested in advanced PCa.

  13. Antisense imaging of epidermal growth factor-induced p21{sup WAF-1/CIP-1} gene expression in MDA-MB-468 human breast cancer xenografts

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Judy; Chen, Paul; Mrkobrada, Marko [Leslie Dan Faculty of Pharmacy, University of Toronto, 19 Russell Street, M5S 2S2, Toronto, Ontario (Canada); Hu, Meiduo [Leslie Dan Faculty of Pharmacy, University of Toronto, 19 Russell Street, M5S 2S2, Toronto, Ontario (Canada); Department of Pharmaceutical Sciences, University of Toronto, Toronto, Ontario (Canada); Vallis, Katherine A. [Department of Radiation Oncology, Princess Margaret Hospital, University Health Network, 610 University Avenue, Toronto, Ontario (Canada); Department of Radiation Oncology, University of Toronto, Toronto, Ontario (Canada); Department of Medical Biophysics, University of Toronto, Toronto, Ontario (Canada); Reilly, Raymond M. [Department of Medical Imaging, University of Toronto, Toronto, Ontario (Canada)

    2003-09-01

    Molecular imaging of the expression of key genes which determine the response to DNA damage following cancer treatment may predict the effectiveness of a particular treatment strategy. A prominent early response gene for DNA damage is the gene encoding p21{sup WAF-1/CIP-1}, a cyclin-dependent kinase inhibitor that regulates progression through the cell cycle. In this study, we explored the feasibility of imaging p21{sup WAF-1/CIP-1} gene expression at the mRNA level using an 18-mer phosphorothioated antisense oligodeoxynucleotide (ODN) labeled with {sup 111}In. The known induction of the p21{sup WAF-1/CIP-1} gene in MDA-MB-468 human breast cancer cells following exposure to epidermal growth factor (EGF) was used as an experimental tool. Treatment of MDA-MB-468 cells in vitro with EGF (20 nM) increased the ratio of p21{sup WAF-1/CIP-1} mRNA/{beta}-actin mRNA threefold within 2 h as measured by the reverse transcription polymerase chain reaction (RT-PCR). A concentration-dependent inhibition of EGF-induced p21{sup WAF-1/CIP-1} protein expression was achieved in MDA-MB-468 cells by treatment with antisense ODNs with up to a tenfold decrease observed at 1 {mu}M. There was a fourfold lower inhibition of p21{sup WAF-1/CIP-1} protein expression by control sense or random sequence ODNs. Intratumoral injections of EGF (15 {mu}g/day x 3 days) were employed to induce p21{sup WAF-1/CIP-1} gene expression in MDA-MB-468 xenografts implanted subcutaneously into athymic mice. RT-PCR of explanted tumors showed a threefold increased level of p21{sup WAF-1/CIP-1} mRNA compared with normal saline-treated tumors. Successful imaging of EGF-induced p21{sup WAF-1/CIP-1} gene expression in MDA-MB-468 xenografts was achieved at 48 h post injection of {sup 111}In-labeled antisense ODNs (3.7 MBq; 2 {mu}g). Tumors displaying basal levels of p21{sup WAF-1/CIP-1} gene expression in the absence of EGF treatment could not be visualized. Biodistribution studies showed a significantly higher tumor

  14. Gene expression in tumor cells and stroma in dsRed 4T1 tumors in eGFP-expressing mice with and without enhanced oxygenation

    International Nuclear Information System (INIS)

    Moen, Ingrid; Øyan, Anne M; Stuhr, Linda EB; Jevne, Charlotte; Wang, Jian; Kalland, Karl-Henning; Chekenya, Martha; Akslen, Lars A; Sleire, Linda; Enger, Per Ø; Reed, Rolf K

    2012-01-01

    The tumor microenvironment is pivotal in tumor progression. Thus, we aimed to develop a mammary tumor model to elucidate molecular characteristics in the stroma versus the tumor cell compartment by global gene expression. Secondly, since tumor hypoxia influences several aspects of tumor pathophysiology, we hypothesized that hyperoxia might have an inhibitory effect on tumor growth per se. Finally, we aimed to identify differences in gene expression and key molecular mechanisms, both in the native state and following treatment. 4T1 dsRed breast cancer cells were injected into eGFP expressing NOD/SCID mice. Group 1 was exposed to 3 intermittent HBO treatments (Day 1, 4 and 7), Group 2 to 7 daily HBO treatments (both 2.5bar, 100% O 2 , à 90 min), whereas the controls were exposed to a normal atmosphere. Tumor growth, histology, vascularisation, cell proliferation, cell death and metastasis were assessed. Fluorescence-activated cell sorting was used to separate tumor cells from stromal cells prior to gene expression analysis. The purity of sorted cells was verified by fluorescence microscopy. Gene expression profiling demonstrated that highly expressed genes in the untreated tumor stroma included constituents of the extracellular matrix and matrix metalloproteinases. Tumor growth was significantly inhibited by HBO, and the MAPK pathway was found to be significantly reduced. Immunohistochemistry indicated a significantly reduced microvessel density after intermittent HBO, whereas daily HBO did not show a similar effect. The anti-angiogenic response was reflected in the expression trends of angiogenic factors. The present in vivo mammary tumor model enabled us to separate tumor and stromal cells, and demonstrated that the two compartments are characterized by distinct gene expressions, both in the native state and following HBO treatments. Furthermore, hyperoxia induced a significant tumor growth-inhibitory effect, with significant down-regulation of the MAPK pathway

  15. Understanding gene expression in coronary artery disease through ...

    Indian Academy of Sciences (India)

    Understanding gene expression in coronary artery disease through global profiling, network analysis and independent validation of key candidate genes. Prathima ... Table 2. Differentially expressed genes in CAD compared to age and gender matched controls. .... Regulation of nuclear pre-mRNA domain containing 1A.

  16. Radiopacity evaluation of root canal sealers containing calcium hydroxide and MTA

    Directory of Open Access Journals (Sweden)

    Juliane Maria Guerreiro- Tanomaru

    2009-06-01

    Full Text Available The purpose of this study was to evaluate the radiopacity of root canal sealers containing calcium hydroxide and MTA (Acroseal, Sealer 26, Sealapex, Endo CPM Sealer, Epiphany and Intrafill. Five disc-shaped specimens (10 x 1 mm were fabricated from each material, according to the ISO 6876/2001 standard. After setting of the materials, radiographs were taken using occlusal film and a graduated aluminum stepwedge varying from 2 to 16 mm in thickness. The dental X-ray unit (GE1000 was set at 50 kVp, 10 mA, 18 pulses/s and distance of 33.5 cm. The radiographs were digitized and the radiopacity compared to that of the aluminum stepwedge using VIXWIN-2000 software (Gendex. The data (mmAl were analyzed statistically by ANOVA and Tukey's test at the 5% significance level. Epiphany and Intrafill presented the highest radiopacity values (8.3 mmAl and 7.5 mmAl respectively, p < 0.05 followed by Sealer 26 (6.3 mmAl, Sealapex (6.1 mmAl and Endo CPM Sealer (6 mmAl. Acroseal was the least radiopaque material (4 mmAl, p < 0.05. In conclusion, the calcium hydroxide- and MTA-containing root canal sealers had different radiopacities. However, all materials presented radiopacity values above the minimum recommended by the ISO standard.

  17. The ordering of expression among a few genes can provide simple cancer biomarkers and signal BRCA1 mutations

    Directory of Open Access Journals (Sweden)

    Parmigiani Giovanni

    2009-08-01

    Full Text Available Abstract Background A major challenge in computational biology is to extract knowledge about the genetic nature of disease from high-throughput data. However, an important obstacle to both biological understanding and clinical applications is the "black box" nature of the decision rules provided by most machine learning approaches, which usually involve many genes combined in a highly complex fashion. Achieving biologically relevant results argues for a different strategy. A promising alternative is to base prediction entirely upon the relative expression ordering of a small number of genes. Results We present a three-gene version of "relative expression analysis" (RXA, a rigorous and systematic comparison with earlier approaches in a variety of cancer studies, a clinically relevant application to predicting germline BRCA1 mutations in breast cancer and a cross-study validation for predicting ER status. In the BRCA1 study, RXA yields high accuracy with a simple decision rule: in tumors carrying mutations, the expression of a "reference gene" falls between the expression of two differentially expressed genes, PPP1CB and RNF14. An analysis of the protein-protein interactions among the triplet of genes and BRCA1 suggests that the classifier has a biological foundation. Conclusion RXA has the potential to identify genomic "marker interactions" with plausible biological interpretation and direct clinical applicability. It provides a general framework for understanding the roles of the genes involved in decision rules, as illustrated for the difficult and clinically relevant problem of identifying BRCA1 mutation carriers.

  18. Real-time PCR expression profiling of genes encoding potential virulence factors in Candida albicans biofilms: identification of model-dependent and -independent gene expression

    Directory of Open Access Journals (Sweden)

    Řičicová Markéta

    2010-04-01

    Full Text Available Abstract Background Candida albicans infections are often associated with biofilm formation. Previous work demonstrated that the expression of HWP1 (hyphal wall protein and of genes belonging to the ALS (agglutinin-like sequence, SAP (secreted aspartyl protease, PLB (phospholipase B and LIP (lipase gene families is associated with biofilm growth on mucosal surfaces. We investigated using real-time PCR whether genes encoding potential virulence factors are also highly expressed in biofilms associated with abiotic surfaces. For this, C. albicans biofilms were grown on silicone in microtiter plates (MTP or in the Centres for Disease Control (CDC reactor, on polyurethane in an in vivo subcutaneous catheter rat (SCR model, and on mucosal surfaces in the reconstituted human epithelium (RHE model. Results HWP1 and genes belonging to the ALS, SAP, PLB and LIP gene families were constitutively expressed in C. albicans biofilms. ALS1-5 were upregulated in all model systems, while ALS9 was mostly downregulated. ALS6 and HWP1 were overexpressed in all models except in the RHE and MTP, respectively. The expression levels of SAP1 were more pronounced in both in vitro models, while those of SAP2, SAP4 and SAP6 were higher in the in vivo model. Furthermore, SAP5 was highly upregulated in the in vivo and RHE models. For SAP9 and SAP10 similar gene expression levels were observed in all model systems. PLB genes were not considerably upregulated in biofilms, while LIP1-3, LIP5-7 and LIP9-10 were highly overexpressed in both in vitro models. Furthermore, an elevated lipase activity was detected in supernatans of biofilms grown in the MTP and RHE model. Conclusions Our findings show that HWP1 and most of the genes belonging to the ALS, SAP and LIP gene families are upregulated in C. albicans biofilms. Comparison of the fold expression between the various model systems revealed similar expression levels for some genes, while for others model-dependent expression

  19. Analyzing Plasmodium falciparum erythrocyte membrane protein 1 gene expression by a next generation sequencing based method

    DEFF Research Database (Denmark)

    Jespersen, Jakob S.; Petersen, Bent; Seguin-Orlando, Andaine

    2013-01-01

    at identifying PfEMP1 features associated with high virulence. Here we present the first effective method for sequence analysis of var genes expressed in field samples: a sequential PCR and next generation sequencing based technique applied on expressed var sequence tags and subsequently on long range PCR......, encoded by ~60 highly variable 'var' genes per haploid genome. PfEMP1 is exported to the surface of infected erythrocytes and is thought to be fundamental to immune evasion by adhesion to host and parasite factors. The highly variable nature has constituted a roadblock in var expression studies aimed...

  20. Over-expression of Eph and ephrin genes in advanced ovarian cancer: ephrin gene expression correlates with shortened survival

    Directory of Open Access Journals (Sweden)

    Lincoln Douglas

    2006-06-01

    Full Text Available Abstract Background Increased expression of Eph receptor tyrosine kinases and their ephrin ligands has been implicated in tumor progression in a number of malignancies. This report describes aberrant expression of these genes in ovarian cancer, the commonest cause of death amongst gynaecological malignancies. Methods Eph and ephrin expression was determined using quantitative real time RT-PCR. Correlation of gene expression was measured using Spearman's rho statistic. Survival was analysed using log-rank analysis and (was visualised by Kaplan-Meier survival curves. Results Greater than 10 fold over-expression of EphA1 and a more modest over-expression of EphA2 were observed in partially overlapping subsets of tumors. Over-expression of EphA1 strongly correlated (r = 0.801; p Conclusion These data imply that increased levels of ephrins A1 and A5 in the presence of high expression of Ephs A1 and A2 lead to a more aggressive tumor phenotype. The known functions of Eph/ephrin signalling in cell de-adhesion and movement may explain the observed correlation of ephrin expression with poor prognosis.

  1. Adrenomedullin Regulates IL-1β Gene Expression in F4/80+ Macrophages during Synovial Inflammation

    Science.gov (United States)

    Takano, Shotaro; Miyagi, Masayuki; Inoue, Gen; Aikawa, Jun; Iwabuchi, Kazuya; Takaso, Masashi

    2017-01-01

    Adrenomedullin (AM) plays an important role in the regulation of inflammatory processes; however, the role and expression of AM in synovial inflammation have not been determined. To investigate the expression and role of AM in inflamed synovial tissue (ST), the gene expression profiles of AM in the ST, including synovial macrophages and fibroblasts, of a murine patellar surgical dislocation model were characterized. In addition, the effects of interleukin- (IL-) 1β and AM in cultured synovial cells were also examined. CD11c+ macrophages were found to be elevated in ST of the surgically dislocated patella. Higher gene expression of CD11c, IL-1β, AM, receptor activity-modifying proteins 2 (RAMP2), and 3 (RAMP3) was also observed in ST obtained from the dislocated side. AM expression was also significantly increased in synovial fibroblasts and macrophages in response to IL-1β treatment. Synovial macrophages also highly expressed RAMP3 compared to fibroblasts and this expression was further stimulated by exogenously added IL-1β. Further, the treatment of the F4/80-positive cell fraction obtained from ST with AM inhibited IL-1β expression. Taken together, these findings demonstrated that AM was produced by synovial fibroblasts and macrophages in inflamed ST and that increased levels of AM may exert anti-inflammatory effects on synovial macrophages. PMID:28299347

  2. Three gene expression vector sets for concurrently expressing multiple genes in Saccharomyces cerevisiae.

    Science.gov (United States)

    Ishii, Jun; Kondo, Takashi; Makino, Harumi; Ogura, Akira; Matsuda, Fumio; Kondo, Akihiko

    2014-05-01

    Yeast has the potential to be used in bulk-scale fermentative production of fuels and chemicals due to its tolerance for low pH and robustness for autolysis. However, expression of multiple external genes in one host yeast strain is considerably labor-intensive due to the lack of polycistronic transcription. To promote the metabolic engineering of yeast, we generated systematic and convenient genetic engineering tools to express multiple genes in Saccharomyces cerevisiae. We constructed a series of multi-copy and integration vector sets for concurrently expressing two or three genes in S. cerevisiae by embedding three classical promoters. The comparative expression capabilities of the constructed vectors were monitored with green fluorescent protein, and the concurrent expression of genes was monitored with three different fluorescent proteins. Our multiple gene expression tool will be helpful to the advanced construction of genetically engineered yeast strains in a variety of research fields other than metabolic engineering. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  3. Transgenic rice plants expressing synthetic cry2AX1 gene exhibits resistance to rice leaffolder (Cnaphalocrosis medinalis).

    Science.gov (United States)

    Manikandan, R; Balakrishnan, N; Sudhakar, D; Udayasuriyan, V

    2016-06-01

    Bacillus thuringiensis is a major source of insecticidal genes imparting insect resistance in transgenic plants. Level of expression of transgenes in transgenic plants is important to achieve desirable level of resistance against target insects. In order to achieve desirable level of expression, rice chloroplast transit peptide sequence was fused with synthetic cry2AX1 gene to target its protein in chloroplasts. Sixteen PCR positive lines of rice were generated by Agrobacterium mediated transformation using immature embryos. Southern blot hybridization analysis of T 0 transgenic plants confirmed the integration of cry2AX1 gene in two to five locations of rice genome and ELISA demonstrated its expression. Concentration of Cry2AX1 in transgenic rice events ranged 5.0-120 ng/g of fresh leaf tissue. Insect bioassay of T 0 transgenic rice plants against neonate larvae of rice leaffolder showed larval mortality ranging between 20 and 80 % in comparison to control plant. Stable inheritance and expression of cry2AX1 gene was demonstrated in T 1 progenies through Southern and ELISA. In T 1 progenies, the highest concentration of Cry2AX1 and mortality of rice leaffolder larvae were recorded as 150 ng/g of fresh leaf tissue and 80 %, respectively. The Cry2AX1 expression even at a very low concentration (120-150 ng/g) in transgenic rice plants was found effective against rice leaffolder larvae.

  4. Development of the foremost light-curable calcium-silicate MTA cement as root-end in oral surgery. Chemical-physical properties, bioactivity and biological behavior.

    Science.gov (United States)

    Gandolfi, Maria Giovanna; Taddei, Paola; Siboni, Francesco; Modena, Enrico; Ciapetti, Gabriela; Prati, Carlo

    2011-07-01

    An innovative light-curable calcium-silicate cement containing a HEMA-TEGDMA-based resin (lc-MTA) was designed to obtain a bioactive fast setting root-end filling and root repair material. lc-MTA was tested for setting time, solubility, water absorption, calcium release, alkalinizing activity (pH of soaking water), bioactivity (apatite-forming ability) and cell growth-proliferation. The apatite-forming ability was investigated by micro-Raman, ATR-FTIR and ESEM/EDX after immersion at 37°C for 1-28 days in DPBS or DMEM+FBS. The marginal adaptation of cement in root-end cavities of extracted teeth was assessed by ESEM/EDX, and the viability of Saos-2 cell on cements was evaluated. lc-MTA demonstrated a rapid setting time (2min), low solubility, high calcium release (150-200ppm) and alkalinizing power (pH 10-12). lc-MTA proved the formation of bone-like apatite spherulites just after 1 day. Apatite precipitates completely filled the interface porosities and created a perfect marginal adaptation. lc-MTA allowed Saos-2 cell viability and growth and no compromising toxicity was exerted. HEMA-TEGDMA creates a polymeric network able to stabilize the outer surface of the cement and a hydrophilic matrix permeable enough to allow water absorption. SiO(-)/Si-OH groups from the mineral particles induce heterogeneous nucleation of apatite by sorption of calcium and phosphate ions. Oxygen-containing groups from poly-HEMA-TEGDMA provide additional apatite nucleating sites through the formation of calcium chelates. The strong novelty was that the combination of a hydraulic calcium-silicate powder and a poly-HEMA-TEGDMA hydrophilic resin creates the conditions (calcium release and functional groups able to chelate Ca ions) for a bioactive fast setting light-curable material for clinical applications in dental and maxillofacial surgery. The first and unique/exclusive light-curable calcium-silicate MTA cement for endodontics and root-end application was created, with a potential

  5. Clinical and radiographic evaluation of indirect pulp treatment with MTA and calcium hydroxide in primary teeth (in-vivo study

    Directory of Open Access Journals (Sweden)

    Vimi George

    2015-01-01

    Full Text Available Objectives: Clinical and radiographic effects of mineral trioxide aggregate (white MTA and calcium hydroxide (Dycal in indirect pulp treatment (IPT of primary teeth over a period of 6 months. Materials and Methods: A clinical trial with sample size of 40 primary molars between the age group of 5-9 years, of which, 20 teeth were considered, each for MTA and Dycal. Measurements on the digitized radiographs were performed at baseline, third and sixth month, increase in dentin was then measured using Corel Draw software. Result: Independent t-test had indicated that at the end of 3 months and 6 months, a statistically significant increase in dentin thickness with both MTA and Dycal (P-value ≤ 0.001 was found. Within the MTA group, the thickness of dentin formed was 0.089 mm ± 0.031 mm at first 3 months and 0.055 ± 0.022 mm at the second 3 months, (P ≤ 0.001 evaluated using paired t-test. In the Dycal group, increment in dentin deposited was 0.068 mm at the first 3 months and second 3 months, it was 0.030 mm (P-value ≤ 0.001. Conclusion: Clinically and radiographically, MTA is superior to Dycal as a good IPT medicament in primary teeth.

  6. Clinical and radiographic evaluation of indirect pulp treatment with MTA and calcium hydroxide in primary teeth (in-vivo study).

    Science.gov (United States)

    George, Vimi; Janardhanan, Suresh Kumar; Varma, Balagopal; Kumaran, Parvathy; Xavier, Arun Mamachan

    2015-01-01

    Clinical and radiographic effects of mineral trioxide aggregate (white MTA) and calcium hydroxide (Dycal) in indirect pulp treatment (IPT) of primary teeth over a period of 6 months. A clinical trial with sample size of 40 primary molars between the age group of 5-9 years, of which, 20 teeth were considered, each for MTA and Dycal. Measurements on the digitized radiographs were performed at baseline, third and sixth month, increase in dentin was then measured using Corel Draw software. Independent t-test had indicated that at the end of 3 months and 6 months, a statistically significant increase in dentin thickness with both MTA and Dycal (P-value ≤ 0.001) was found. Within the MTA group, the thickness of dentin formed was 0.089 mm ± 0.031 mm at first 3 months and 0.055 ± 0.022 mm at the second 3 months, (P ≤ 0.001) evaluated using paired t-test. In the Dycal group, increment in dentin deposited was 0.068 mm at the first 3 months and second 3 months, it was 0.030 mm (P-value ≤ 0.001). Clinically and radiographically, MTA is superior to Dycal as a good IPT medicament in primary teeth.

  7. Candidate genes expressed in human islets and their role in the pathogenesis of type 1 diabetes

    DEFF Research Database (Denmark)

    Storling, Joachim; Brorsson, Caroline Anna

    2013-01-01

    In type 1 diabetes (T1D), the insulin-producing β cells are destroyed by an immune-mediated process leading to complete insulin deficiency. There is a strong genetic component in T1D. Genes located in the human leukocyte antigen (HLA) region are the most important genetic determinants of disease......, but more than 40 additional loci are known to significantly affect T1D risk. Since most of the currently known genetic candidates have annotated immune cell functions, it is generally considered that most of the genetic susceptibility in T1D is caused by variation in genes affecting immune cell function....... Recent studies, however, indicate that most T1D candidate genes are expressed in human islets suggesting that the functions of the genes are not restricted to immune cells, but also play roles in the islets and possibly the β cells. Several candidates change expression levels within the islets following...

  8. A comparative study of BioAggregate and ProRoot MTA on adhesion, migration, and attachment of human dental pulp cells.

    Science.gov (United States)

    Zhu, Lingxin; Yang, Jingwen; Zhang, Jie; Peng, Bin

    2014-08-01

    The aim of the present study was to evaluate the effects of a novel bioceramic nanoparticular cement, BioAggregate (Innovative Bioceramix, Vancouver, BC, Canada), on the adhesion, migration, and attachment of human dental pulp cells (HDPCs) and to compare its performance with that of ProRoot mineral trioxide aggregate (MTA) (Dentsply, Tulsa, OK). Primary cultured HDPCs were treated with various dilutions of BioAggregate and MTA extracts to assess the cell viability using the Cell Counting Kit-8 (Dojindo, Kumamoto, Japan). Cell adhesion assay was performed using type I collagen-coated plates. An in vitro scratch wound healing model was used to determine cell migration. Focal adhesion formation and cytoskeleton organization were further examined by double immunofluorescence labeling for vinculin and fibrous actin. To assess cell attachment, HDPCs were directly seeded onto the material surfaces and observed by scanning electron microscopy. HDPCs exposed to BioAggregate extracts showed the highest viabilities at all extract concentrations at 24 and 48 hours, whereas cells exposed to original MTA extracts displayed suppressed viabilities at 72 hours compared with the control. Treatment with BioAggregate extracts enhanced cellular adhesion and migration of HDPCs in a concentration-dependent manner, which was superior to the effects induced by MTA extracts. Immunofluorescence staining indicated that both BioAggregate and MTA optimized focal adhesion formation and stress fiber assembly. Furthermore, scanning electron microscopic analysis revealed that HDPCs attached onto BioAggregate were more flattened and exhibited better spreading than cells on MTA. BioAggregate is able to promote cellular adhesion, migration, and attachment of HDPCs, indicating its excellent cytocompatibility. Therefore, BioAggregate appears to be a possible alternative to MTA for pulp capping. Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

  9. The MRC1/CD68 ratio is positively associated with adipose tissue lipogenesis and with muscle mitochondrial gene expression in humans.

    Directory of Open Access Journals (Sweden)

    José María Moreno-Navarrete

    Full Text Available BACKGROUND: Alternative macrophages (M2 express the cluster differentiation (CD 206 (MCR1 at high levels. Decreased M2 in adipose tissue is known to be associated with obesity and inflammation-related metabolic disturbances. Here we aimed to investigate MCR1 relative to CD68 (total macrophages gene expression in association with adipogenic and mitochondrial genes, which were measured in human visceral [VWAT, n = 147] and subcutaneous adipose tissue [SWAT, n = 76] and in rectus abdominis muscle (n = 23. The effects of surgery-induced weight loss were also longitudinally evaluated (n = 6. RESULTS: MCR1 and CD68 gene expression levels were similar in VWAT and SWAT. A higher proportion of CD206 relative to total CD68 was present in subjects with less body fat and lower fasting glucose concentrations. The ratio MCR1/CD68was positively associated with IRS1gene expression and with the expression of lipogenic genes such as ACACA, FASN and THRSP, even after adjusting for BMI. The ratio MCR1/CD68 in SWAT increased significantly after the surgery-induced weight loss (+44.7%; p = 0.005 in parallel to the expression of adipogenic genes. In addition, SWAT MCR1/CD68ratio was significantly associated with muscle mitochondrial gene expression (PPARGC1A, TFAM and MT-CO3. AT CD206 was confirmed by immunohistochemistry to be specific of macrophages, especially abundant in crown-like structures. CONCLUSION: A decreased ratio MCR1/CD68 is linked to adipose tissue and muscle mitochondrial dysfunction at least at the level of expression of adipogenic and mitochondrial genes.

  10. The SKP1-like gene family of Arabidopsis exhibits a high degree of differential gene expression and gene product interaction during development.

    Directory of Open Access Journals (Sweden)

    Mohammad H Dezfulian

    Full Text Available The Arabidopsis thaliana genome encodes several families of polypeptides that are known or predicted to participate in the formation of the SCF-class of E3-ubiquitin ligase complexes. One such gene family encodes the Skp1-like class of polypeptide subunits, where 21 genes have been identified and are known to be expressed in Arabidopsis. Phylogenetic analysis based on deduced polypeptide sequence organizes the family of ASK proteins into 7 clades. The complexity of the ASK gene family, together with the close structural similarity among its members raises the prospect of significant functional redundancy among select paralogs. We have assessed the potential for functional redundancy within the ASK gene family by analyzing an expanded set of criteria that define redundancy with higher resolution. The criteria used include quantitative expression of locus-specific transcripts using qRT-PCR, assessment of the sub-cellular localization of individual ASK:YFP auto-fluorescent fusion proteins expressed in vivo as well as the in planta assessment of individual ASK-F-Box protein interactions using bimolecular fluorescent complementation techniques in combination with confocal imagery in live cells. The results indicate significant functional divergence of steady state transcript abundance and protein-protein interaction specificity involving ASK proteins in a pattern that is poorly predicted by sequence-based phylogeny. The information emerging from this and related studies will prove important for defining the functional intersection of expression, localization and gene product interaction that better predicts the formation of discrete SCF complexes, as a prelude to investigating their molecular mode of action.

  11. Vascular Gene Expression in Nonneoplastic and Malignant Brain

    Science.gov (United States)

    Madden, Stephen L.; Cook, Brian P.; Nacht, Mariana; Weber, William D.; Callahan, Michelle R.; Jiang, Yide; Dufault, Michael R.; Zhang, Xiaoming; Zhang, Wen; Walter-Yohrling, Jennifer; Rouleau, Cecile; Akmaev, Viatcheslav R.; Wang, Clarence J.; Cao, Xiaohong; St. Martin, Thia B.; Roberts, Bruce L.; Teicher, Beverly A.; Klinger, Katherine W.; Stan, Radu-Virgil; Lucey, Brenden; Carson-Walter, Eleanor B.; Laterra, John; Walter, Kevin A.

    2004-01-01

    Malignant gliomas are uniformly lethal tumors whose morbidity is mediated in large part by the angiogenic response of the brain to the invading tumor. This profound angiogenic response leads to aggressive tumor invasion and destruction of surrounding brain tissue as well as blood-brain barrier breakdown and life-threatening cerebral edema. To investigate the molecular mechanisms governing the proliferation of abnormal microvasculature in malignant brain tumor patients, we have undertaken a cell-specific transcriptome analysis from surgically harvested nonneoplastic and tumor-associated endothelial cells. SAGE-derived endothelial cell gene expression patterns from glioma and nonneoplastic brain tissue reveal distinct gene expression patterns and consistent up-regulation of certain glioma endothelial marker genes across patient samples. We define the G-protein-coupled receptor RDC1 as a tumor endothelial marker whose expression is distinctly induced in tumor endothelial cells of both brain and peripheral vasculature. Further, we demonstrate that the glioma-induced gene, PV1, shows expression both restricted to endothelial cells and coincident with endothelial cell tube formation. As PV1 provides a framework for endothelial cell caveolar diaphragms, this protein may serve to enhance glioma-induced disruption of the blood-brain barrier and transendothelial exchange. Additional characterization of this extensive brain endothelial cell gene expression database will provide unique molecular insights into vascular gene expression. PMID:15277233

  12. Gene expression profile data for mouse facial development

    Directory of Open Access Journals (Sweden)

    Sonia M. Leach

    2017-08-01

    Full Text Available This article contains data related to the research articles "Spatial and Temporal Analysis of Gene Expression during Growth and Fusion of the Mouse Facial Prominences" (Feng et al., 2009 [1] and “Systems Biology of facial development: contributions of ectoderm and mesenchyme” (Hooper et al., 2017 In press [2]. Embryonic mammalian craniofacial development is a complex process involving the growth, morphogenesis, and fusion of distinct facial prominences into a functional whole. Aberrant gene regulation during this process can lead to severe craniofacial birth defects, including orofacial clefting. As a means to understand the genes involved in facial development, we had previously dissected the embryonic mouse face into distinct prominences: the mandibular, maxillary or nasal between E10.5 and E12.5. The prominences were then processed intact, or separated into ectoderm and mesenchyme layers, prior analysis of RNA expression using microarrays (Feng et al., 2009, Hooper et al., 2017 in press [1,2]. Here, individual gene expression profiles have been built from these datasets that illustrate the timing of gene expression in whole prominences or in the separated tissue layers. The data profiles are presented as an indexed and clickable list of the genes each linked to a graphical image of that gene׳s expression profile in the ectoderm, mesenchyme, or intact prominence. These data files will enable investigators to obtain a rapid assessment of the relative expression level of any gene on the array with respect to time, tissue, prominence, and expression trajectory.

  13. Large scale gene expression meta-analysis reveals tissue-specific, sex-biased gene expression in humans

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    Benjamin Mayne

    2016-10-01

    Full Text Available The severity and prevalence of many diseases are known to differ between the sexes. Organ specific sex-biased gene expression may underpin these and other sexually dimorphic traits. To further our understanding of sex differences in transcriptional regulation, we performed meta-analyses of sex biased gene expression in multiple human tissues. We analysed 22 publicly available human gene expression microarray data sets including over 2500 samples from 15 different tissues and 9 different organs. Briefly, by using an inverse-variance method we determined the effect size difference of gene expression between males and females. We found the greatest sex differences in gene expression in the brain, specifically in the anterior cingulate cortex, (1818 genes, followed by the heart (375 genes, kidney (224 genes, colon (218 genes and thyroid (163 genes. More interestingly, we found different parts of the brain with varying numbers and identity of sex-biased genes, indicating that specific cortical regions may influence sexually dimorphic traits. The majority of sex-biased genes in other tissues such as the bladder, liver, lungs and pancreas were on the sex chromosomes or involved in sex hormone production. On average in each tissue, 32% of autosomal genes that were expressed in a sex-biased fashion contained androgen or estrogen hormone response elements. Interestingly, across all tissues, we found approximately two-thirds of autosomal genes that were sex-biased were not under direct influence of sex hormones. To our knowledge this is the largest analysis of sex-biased gene expression in human tissues to date. We identified many sex-biased genes that were not under the direct influence of sex chromosome genes or sex hormones. These may provide targets for future development of sex-specific treatments for diseases.

  14. Crosstalk between thyroid hormone receptor and liver X receptor in the regulation of selective Alzheimer's disease indicator-1 gene expression.

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    Emi Ishida

    Full Text Available Selective Alzheimer's disease (AD indicator 1 (Seladin-1 has been identified as a gene down-regulated in the degenerated lesions of AD brain. Up-regulation of Seladin-1 reduces the accumulation of β-amyloid and neuronal death. Thyroid hormone (TH exerts an important effect on the development and maintenance of central nervous systems. In the current study, we demonstrated that Seladin-1 gene and protein expression in the forebrain was increased in thyrotoxic mice compared with that of euthyroid mice. However, unexpectedly, no significant decrease in the gene and protein expression was observed in hypothyroid mice. Interestingly, an agonist of liver X receptor (LXR, TO901317 (TO administration in vivo increased Seladin-1 gene and protein expression in the mouse forebrain only in a hypothyroid state and in the presence of mutant TR-β, suggesting that LXR-α would compensate for TR-β function to maintain Seladin-1 gene expression in hypothyroidism and resistance to TH. TH activated the mouse Seladin-1 gene promoter (-1936/+21 bp and site 2 including canonical TH response element (TRE half-site in the region between -159 and -154 bp is responsible for the positive regulation. RXR-α/TR-β heterodimerization was identified on site 2 by gel-shift assay, and chromatin immunoprecipitation assay revealed the recruitment of TR-β to site 2 and the recruitment was increased upon TH administration. On the other hand, LXR-α utilizes a distinct region from site 2 (-120 to -102 bp to activate the mouse Seladin-1 gene promoter. Taking these findings together, we concluded that TH up-regulates Seladin-1 gene expression at the transcriptional level and LXR-α maintains the gene expression.

  15. Prediction of the contact sensitizing potential of chemicals using analysis of gene expression changes in human THP-1 monocytes.

    Science.gov (United States)

    Arkusz, Joanna; Stępnik, Maciej; Sobala, Wojciech; Dastych, Jarosław

    2010-11-10

    The aim of this study was to find differentially regulated genes in THP-1 monocytic cells exposed to sensitizers and nonsensitizers and to investigate if such genes could be reliable markers for an in vitro predictive method for the identification of skin sensitizing chemicals. Changes in expression of 35 genes in the THP-1 cell line following treatment with chemicals of different sensitizing potential (from nonsensitizers to extreme sensitizers) were assessed using real-time PCR. Verification of 13 candidate genes by testing a large number of chemicals (an additional 22 sensitizers and 8 nonsensitizers) revealed that prediction of contact sensitization potential was possible based on evaluation of changes in three genes: IL8, HMOX1 and PAIMP1. In total, changes in expression of these genes allowed correct detection of sensitization potential of 21 out of 27 (78%) test sensitizers. The gene expression levels inside potency groups varied and did not allow estimation of sensitization potency of test chemicals. Results of this study indicate that evaluation of changes in expression of proposed biomarkers in THP-1 cells could be a valuable model for preliminary screening of chemicals to discriminate an appreciable majority of sensitizers from nonsensitizers. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

  16. Chromosome 15 structural abnormalities: effect on IGF1R gene expression and function

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    Rossella Cannarella

    2017-09-01

    Full Text Available Insulin-like growth factor 1 receptor (IGF1R, mapping on the 15q26.3 chromosome, is required for normal embryonic and postnatal growth. The aim of the present study was to evaluate the IGF1R gene expression and function in three unrelated patients with chromosome 15 structural abnormalities. We report two male patients with the smallest 15q26.3 chromosome duplication described so far, and a female patient with ring chromosome 15 syndrome. Patient one, with a 568 kb pure duplication, had overgrowth, developmental delay, mental and psychomotor retardation, obesity, cryptorchidism, borderline low testis volume, severe oligoasthenoteratozoospermia and gynecomastia. We found a 1.8-fold increase in the IGF1R mRNA and a 1.3-fold increase in the IGF1R protein expression (P < 0.05. Patient two, with a 650 kb impure duplication, showed overgrowth, developmental delay, mild mental retardation, precocious puberty, low testicular volume and severe oligoasthenoteratozoospermia. The IGF1R mRNA and protein expression was similar to that of the control. Patient three, with a 46,XX r(15 (p10q26.2 karyotype, displayed intrauterine growth retardation, developmental delay, mental and psychomotor retardation. We found a <0.5-fold decrease in the IGF1R mRNA expression and an undetectable IGF1R activity. After reviewing the previously 96 published cases of chromosome 15q duplication, we found that neurological disorders, congenital cardiac defects, typical facial traits and gonadal abnormalities are the prominent features in patients with chromosome 15q duplication. Interestingly, patients with 15q deletion syndrome display similar features. We speculate that both the increased and decreased IGF1R gene expression may play a role in the etiology of neurological and gonadal disorders.

  17. Involvement of aberrant DNA methylation on reduced expression of lysophosphatidic acid receptor-1 gene in rat tumor cell lines

    International Nuclear Information System (INIS)

    Tsujiuchi, Toshifumi; Shimizu, Kyoko; Onishi, Mariko; Sugata, Eriko; Fujii, Hiromasa; Mori, Toshio; Honoki, Kanya; Fukushima, Nobuyuki

    2006-01-01

    Lysophosphatidic acid (LPA) is a bioactive phospholipid that stimulates cell proliferation, migration, and protects cells from apoptosis. It interacts with specific G protein-coupled transmembrane receptors. Recently, it has been reported that alterations of LPA receptor expression might be important in the malignant transformation of tumor cells. Therefore, to assess an involvement of DNA methylation in reduced expression of the LPA receptor-1 (lpa1) gene, we investigated the expression of the lpa1 gene and its DNA methylation patterns in rat tumor cell lines. Both rat brain-derived neuroblastoma B103 and liver-derived hepatoma RH7777 cells used in this study indicated no expression of lpa1. For the analysis of methylation status, bisulfite sequencing was performed with B103 and RH7777 cells, comparing with other lpa1 expressed cells and normal tissues of brain and liver. The lpa1 expressed cells and tissues were all unmethylated in this region of lpa1. In contrast, both B103 and RH7777 cells were highly methylated, correlating with reduced expression of the lpa1. Treatment with 5-aza 2'-deoxycytidine induced expression of lpa1 gene in B103 and RH7777 cells after 24 h. In RH7777 cells treated with 5-aza 2'-deoxycytidine, stress fiber formation was also observed in response to LPA in RH7777 cells, but not in untreated RH7777 cells. These results suggest that aberrant DNA methylation of the lpa1 gene may be involved in its reduced expression in rat tumor cells

  18. Evaluation of Appropriate Reference Genes for Gene Expression Normalization during Watermelon Fruit Development.

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    Qiusheng Kong

    Full Text Available Gene expression analysis in watermelon (Citrullus lanatus fruit has drawn considerable attention with the availability of genome sequences to understand the regulatory mechanism of fruit development and to improve its quality. Real-time quantitative reverse-transcription PCR (qRT-PCR is a routine technique for gene expression analysis. However, appropriate reference genes for transcript normalization in watermelon fruits have not been well characterized. The aim of this study was to evaluate the appropriateness of 12 genes for their potential use as reference genes in watermelon fruits. Expression variations of these genes were measured in 48 samples obtained from 12 successive developmental stages of parthenocarpic and fertilized fruits of two watermelon genotypes by using qRT-PCR analysis. Considering the effects of genotype, fruit setting method, and developmental stage, geNorm determined clathrin adaptor complex subunit (ClCAC, β-actin (ClACT, and alpha tubulin 5 (ClTUA5 as the multiple reference genes in watermelon fruit. Furthermore, ClCAC alone or together with SAND family protein (ClSAND was ranked as the single or two best reference genes by NormFinder. By using the top-ranked reference genes to normalize the transcript abundance of phytoene synthase (ClPSY1, a good correlation between lycopene accumulation and ClPSY1 expression pattern was observed in ripening watermelon fruit. These validated reference genes will facilitate the accurate measurement of gene expression in the studies on watermelon fruit biology.

  19. Evaluation of Appropriate Reference Genes for Gene Expression Normalization during Watermelon Fruit Development.

    Science.gov (United States)

    Kong, Qiusheng; Yuan, Jingxian; Gao, Lingyun; Zhao, Liqiang; Cheng, Fei; Huang, Yuan; Bie, Zhilong

    2015-01-01

    Gene expression analysis in watermelon (Citrullus lanatus) fruit has drawn considerable attention with the availability of genome sequences to understand the regulatory mechanism of fruit development and to improve its quality. Real-time quantitative reverse-transcription PCR (qRT-PCR) is a routine technique for gene expression analysis. However, appropriate reference genes for transcript normalization in watermelon fruits have not been well characterized. The aim of this study was to evaluate the appropriateness of 12 genes for their potential use as reference genes in watermelon fruits. Expression variations of these genes were measured in 48 samples obtained from 12 successive developmental stages of parthenocarpic and fertilized fruits of two watermelon genotypes by using qRT-PCR analysis. Considering the effects of genotype, fruit setting method, and developmental stage, geNorm determined clathrin adaptor complex subunit (ClCAC), β-actin (ClACT), and alpha tubulin 5 (ClTUA5) as the multiple reference genes in watermelon fruit. Furthermore, ClCAC alone or together with SAND family protein (ClSAND) was ranked as the single or two best reference genes by NormFinder. By using the top-ranked reference genes to normalize the transcript abundance of phytoene synthase (ClPSY1), a good correlation between lycopene accumulation and ClPSY1 expression pattern was observed in ripening watermelon fruit. These validated reference genes will facilitate the accurate measurement of gene expression in the studies on watermelon fruit biology.

  20. Cloning and heterologous expression of a novel insecticidal gene (tccC1) from Xenorhabdus nematophilus strain

    International Nuclear Information System (INIS)

    Joo Lee, Pom; Ahn, Ji-Young; Kim, Yang-Hoon; Wook Kim, Seung; Kim, Ji-Yeon; Park, Jae-Sung; Lee, Jeewon

    2004-01-01

    We have identified and cloned a novel toxin gene (tccC1/xptB1) from Xenorhabdus nematophilus strain isolated from Korea-specific entomophagous nematode Steinernema glaseri MK. The DNA sequence of cloned toxin gene (3048 bp) has an open reading frame encoding 1016 amino acids with a predicted molecular mass of 111058 Da. The toxin sequence shares 50-96% identical amino acid residues with the previously reported tccC1 cloned from X. nematophilus (AJ308438), Photorhabdus luminescens W14 (AF346499) P. luminescens TTO1 (BX571873), and Yersinia pestis CO92 (NC 0 03143). The toxin gene was successfully expressed in Escherichia coli, and the recombinant toxin protein caused a rapid cessation in mortality of Galleria mellonella larvae (80% death of larvae within 2 days). Conclusively, the heterologous expression of the novel gene tccC1 cloned into E. coli plasmid vector produced recombinant toxin with high insecticidal activity

  1. A Functional Role for the Epigenetic Regulator ING1 in Activity-induced Gene Expression in Primary Cortical Neurons.

    Science.gov (United States)

    Leighton, Laura J; Zhao, Qiongyi; Li, Xiang; Dai, Chuanyang; Marshall, Paul R; Liu, Sha; Wang, Yi; Zajaczkowski, Esmi L; Khandelwal, Nitin; Kumar, Arvind; Bredy, Timothy W; Wei, Wei

    2018-01-15

    Epigenetic regulation of activity-induced gene expression involves multiple levels of molecular interaction, including histone and DNA modifications, as well as mechanisms of DNA repair. Here we demonstrate that the genome-wide deposition of inhibitor of growth family member 1 (ING1), which is a central epigenetic regulatory protein, is dynamically regulated in response to activity in primary cortical neurons. ING1 knockdown leads to decreased expression of genes related to synaptic plasticity, including the regulatory subunit of calcineurin, Ppp3r1. In addition, ING1 binding at a site upstream of the transcription start site (TSS) of Ppp3r1 depends on yet another group of neuroepigenetic regulatory proteins, the Piwi-like family, which are also involved in DNA repair. These findings provide new insight into a novel mode of activity-induced gene expression, which involves the interaction between different epigenetic regulatory mechanisms traditionally associated with gene repression and DNA repair. Copyright © 2017 IBRO. Published by Elsevier Ltd. All rights reserved.

  2. The roles of Tenascin C and Fibronectin 1 in adhesive capsulitis: a pilot gene expression study

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    Carina Cohen

    Full Text Available OBJECTIVES: We evaluated mRNA expression levels of genes that encode TGF-β1; the TGF-β1 receptor; the collagen-modifying enzymes LOX, PLOD1, and PLOD2; and the extracellular matrix proteins COMP, FN1, TNC and TNXB in synovial/capsule specimens from patients with idiopathic adhesive capsulitis. Possible associations between the measured mRNA levels and clinical parameters were also investigated. METHODS: We obtained glenohumeral joint synovium/capsule specimens from 9 patients with idiopathic adhesive capsulitis who had not shown improvement in symptoms after 5 months of physiotherapy. Adhesive capsulitis was confirmed in all patients by magnetic resonance imaging. We also obtained specimens from 8 control patients who had underwent surgery for acute acromioclavicular joint dislocation and who had radiological indication of glenohumeral capsule alteration based on arthroscopic evaluation. mRNA expression in the synovium/capsule specimens was analyzed by quantitative reverse transcription PCR. The B2M and HPRT1 genes were used as references to normalize target gene expression in the shoulder tissue samples. RESULTS: The synovium/capsule samples from the patients with adhesive capsulitis had significantly higher TNC and FN1 expression than those from the controls. Additionally, symptom duration directly correlated with expression of TGFβ1 receptor I. CONCLUSION: Elevated levels of TNC and FN1 expression may be a marker of capsule injury. Upregulation of TGFβ1 receptor I seems to be dependent on symptom duration; therefore, TGFβ signaling may be involved in adhesive capsulitis. As such, TNC, FN1 and TGFβ1 receptor I may also play roles in adhesive capsulitis by contributing to capsule inflammation and fibrosis.

  3. An incoherent feedforward loop facilitates adaptive tuning of gene expression.

    Science.gov (United States)

    Hong, Jungeui; Brandt, Nathan; Abdul-Rahman, Farah; Yang, Ally; Hughes, Tim; Gresham, David

    2018-04-05

    We studied adaptive evolution of gene expression using long-term experimental evolution of Saccharomyces cerevisiae in ammonium-limited chemostats. We found repeated selection for non-synonymous variation in the DNA binding domain of the transcriptional activator, GAT1, which functions with the repressor, DAL80 in an incoherent type-1 feedforward loop (I1-FFL) to control expression of the high affinity ammonium transporter gene, MEP2. Missense mutations in the DNA binding domain of GAT1 reduce its binding to the GATAA consensus sequence. However, we show experimentally, and using mathematical modeling, that decreases in GAT1 binding result in increased expression of MEP2 as a consequence of properties of I1-FFLs. Our results show that I1-FFLs, one of the most commonly occurring network motifs in transcriptional networks, can facilitate adaptive tuning of gene expression through modulation of transcription factor binding affinities. Our findings highlight the importance of gene regulatory architectures in the evolution of gene expression. © 2018, Hong et al.

  4. Placental triglyceride accumulation in maternal type 1 diabetes is associated with increased lipase gene expression

    DEFF Research Database (Denmark)

    Lindegaard, Marie Louise Skakkebæk; Damm, Peter; Mathiesen, Elisabeth R

    2006-01-01

    Maternal diabetes can cause fetal macrosomia and increased risk of obesity, diabetes, and cardiovascular disease in adulthood of the offspring. Although increased transplacental lipid transport could be involved, the impact of maternal type 1 diabetes on molecular mechanisms for lipid transport...... in placenta is largely unknown. To examine whether maternal type 1 diabetes affects placental lipid metabolism, we measured lipids and mRNA expression of lipase-encoding genes in placentas from women with type 1 diabetes (n = 27) and a control group (n = 21). The placental triglyceride (TG) concentration....... These results suggest that maternal type 1 diabetes is associated with TG accumulation and increased EL and HSL gene expression in placenta and that optimal metabolic control reduces these effects....

  5. Insulin-Like Growth Factor-1 Inscribes a Gene Expression Profile for Angiogenic Factors and Cancer Progression in Breast Epithelial Cells

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    J.S. Oh

    2002-01-01

    Full Text Available Activation of the insulin-like growth factor-1 receptor (IGF-11R by IGF-1 is associated with the risk and progression of many types of cancer, although despite this it remains unclear how activated IGF-1 R contributes to cancer progression. In this study, gene expression changes elicited by IGF-1 were profiled in breast epithelial cells. We noted that many genes are functionally linked to cancer progression and angiogenesis. To validate some of the changes observed, the RNA and/or protein was confirmed for c-fos, cytochrome P4501Al, cytochrome P450 1131, interleukin-1 beta, fas ligand, vascular endothelial growth factor, and urokinase plasminogen activator. Nuclear proteins were also temporally monitored to address how gene expression changes were regulated. We found that IGF-1 stimulated the nuclear translocation of phosphorylated AKT, hypoxic-inducible factor-1 alpha, and phosphorylated cAMP-responsive element-binding protein, which correlated with temporal changes in gene expression. Next, the promoter regions of IGF-1-regulated genes were searched in silico. The promoters of genes that clustered together had similar regulatory regions. In summary, IGF-1 inscribes a gene expression profile relevant to cancer progression, and this study provides insight into the mechanism(s whereby some of these changes occur.

  6. Gene expression profiles reveal key genes for early diagnosis and treatment of adamantinomatous craniopharyngioma.

    Science.gov (United States)

    Yang, Jun; Hou, Ziming; Wang, Changjiang; Wang, Hao; Zhang, Hongbing

    2018-04-23

    Adamantinomatous craniopharyngioma (ACP) is an aggressive brain tumor that occurs predominantly in the pediatric population. Conventional diagnosis method and standard therapy cannot treat ACPs effectively. In this paper, we aimed to identify key genes for ACP early diagnosis and treatment. Datasets GSE94349 and GSE68015 were obtained from Gene Expression Omnibus database. Consensus clustering was applied to discover the gene clusters in the expression data of GSE94349 and functional enrichment analysis was performed on gene set in each cluster. The protein-protein interaction (PPI) network was built by the Search Tool for the Retrieval of Interacting Genes, and hubs were selected. Support vector machine (SVM) model was built based on the signature genes identified from enrichment analysis and PPI network. Dataset GSE94349 was used for training and testing, and GSE68015 was used for validation. Besides, RT-qPCR analysis was performed to analyze the expression of signature genes in ACP samples compared with normal controls. Seven gene clusters were discovered in the differentially expressed genes identified from GSE94349 dataset. Enrichment analysis of each cluster identified 25 pathways that highly associated with ACP. PPI network was built and 46 hubs were determined. Twenty-five pathway-related genes that overlapped with the hubs in PPI network were used as signatures to establish the SVM diagnosis model for ACP. The prediction accuracy of SVM model for training, testing, and validation data were 94, 85, and 74%, respectively. The expression of CDH1, CCL2, ITGA2, COL8A1, COL6A2, and COL6A3 were significantly upregulated in ACP tumor samples, while CAMK2A, RIMS1, NEFL, SYT1, and STX1A were significantly downregulated, which were consistent with the differentially expressed gene analysis. SVM model is a promising classification tool for screening and early diagnosis of ACP. The ACP-related pathways and signature genes will advance our knowledge of ACP pathogenesis

  7. Expression of gentisate 1,2-dioxygenase (gdoA) genes involved in aromatic degradation in two haloarchaeal genera.

    Science.gov (United States)

    Fairley, D J; Wang, G; Rensing, C; Pepper, I L; Larkin, M J

    2006-12-01

    Gentisate-1,2-dioxygenase genes (gdoA), with homology to a number of bacterial dioxygenases, and genes encoding a putative coenzyme A (CoA)-synthetase subunit (acdB) and a CoA-thioesterase (tieA) were identified in two haloarchaeal isolates. In Haloarcula sp. D1, gdoA was expressed during growth on 4-hydroxybenzoate but not benzoate, and acdB and tieA were not expressed during growth on any of the aromatic substrates tested. In contrast, gdoA was expressed in Haloferax sp. D1227 during growth on benzoate, 3-hydroxybenzoate, cinnamate and phenylpropionate, and both acdB and tieA were expressed during growth on benzoate, cinnamate and phenylpropionate, but not on 3-hydroxybenzoate. This pattern of induction is consistent with these genes encoding steps in a CoA-mediated benzoate pathway in this strain.

  8. Differential Gene Expression and Aging

    Directory of Open Access Journals (Sweden)

    Laurent Seroude

    2002-01-01

    Full Text Available It has been established that an intricate program of gene expression controls progression through the different stages in development. The equally complex biological phenomenon known as aging is genetically determined and environmentally modulated. This review focuses on the genetic component of aging, with a special emphasis on differential gene expression. At least two genetic pathways regulating organism longevity act by modifying gene expression. Many genes are also subjected to age-dependent transcriptional regulation. Some age-related gene expression changes are prevented by caloric restriction, the most robust intervention that slows down the aging process. Manipulating the expression of some age-regulated genes can extend an organism's life span. Remarkably, the activity of many transcription regulatory elements is linked to physiological age as opposed to chronological age, indicating that orderly and tightly controlled regulatory pathways are active during aging.

  9. Differentially expressed genes in iron-induced prion protein conversion

    International Nuclear Information System (INIS)

    Kim, Minsun; Kim, Eun-hee; Choi, Bo-Ran; Woo, Hee-Jong

    2016-01-01

    The conversion of the cellular prion protein (PrP C ) to the protease-resistant isoform is the key event in chronic neurodegenerative diseases, including transmissible spongiform encephalopathies (TSEs). Increased iron in prion-related disease has been observed due to the prion protein-ferritin complex. Additionally, the accumulation and conversion of recombinant PrP (rPrP) is specifically derived from Fe(III) but not Fe(II). Fe(III)-mediated PK-resistant PrP (PrP res ) conversion occurs within a complex cellular environment rather than via direct contact between rPrP and Fe(III). In this study, differentially expressed genes correlated with prion degeneration by Fe(III) were identified using Affymetrix microarrays. Following Fe(III) treatment, 97 genes were differentially expressed, including 85 upregulated genes and 12 downregulated genes (≥1.5-fold change in expression). However, Fe(II) treatment produced moderate alterations in gene expression without inducing dramatic alterations in gene expression profiles. Moreover, functional grouping of identified genes indicated that the differentially regulated genes were highly associated with cell growth, cell maintenance, and intra- and extracellular transport. These findings showed that Fe(III) may influence the expression of genes involved in PrP folding by redox mechanisms. The identification of genes with altered expression patterns in neural cells may provide insights into PrP conversion mechanisms during the development and progression of prion-related diseases. - Highlights: • Differential genes correlated with prion degeneration by Fe(III) were identified. • Genes were identified in cell proliferation and intra- and extracellular transport. • In PrP degeneration, redox related genes were suggested. • Cbr2, Rsad2, Slc40a1, Amph and Mvd were expressed significantly.

  10. Genetic Variants Contribute to Gene Expression Variability in Humans

    Science.gov (United States)

    Hulse, Amanda M.; Cai, James J.

    2013-01-01

    Expression quantitative trait loci (eQTL) studies have established convincing relationships between genetic variants and gene expression. Most of these studies focused on the mean of gene expression level, but not the variance of gene expression level (i.e., gene expression variability). In the present study, we systematically explore genome-wide association between genetic variants and gene expression variability in humans. We adapt the double generalized linear model (dglm) to simultaneously fit the means and the variances of gene expression among the three possible genotypes of a biallelic SNP. The genomic loci showing significant association between the variances of gene expression and the genotypes are termed expression variability QTL (evQTL). Using a data set of gene expression in lymphoblastoid cell lines (LCLs) derived from 210 HapMap individuals, we identify cis-acting evQTL involving 218 distinct genes, among which 8 genes, ADCY1, CTNNA2, DAAM2, FERMT2, IL6, PLOD2, SNX7, and TNFRSF11B, are cross-validated using an extra expression data set of the same LCLs. We also identify ∼300 trans-acting evQTL between >13,000 common SNPs and 500 randomly selected representative genes. We employ two distinct scenarios, emphasizing single-SNP and multiple-SNP effects on expression variability, to explain the formation of evQTL. We argue that detecting evQTL may represent a novel method for effectively screening for genetic interactions, especially when the multiple-SNP influence on expression variability is implied. The implication of our results for revealing genetic mechanisms of gene expression variability is discussed. PMID:23150607

  11. Identification of adaptive mutations in the influenza A virus non-structural 1 gene that increase cytoplasmic localization and differentially regulate host gene expression.

    Directory of Open Access Journals (Sweden)

    Nicole Forbes

    Full Text Available The NS1 protein of influenza A virus (IAV is a multifunctional virulence factor. We have previously characterized gain-of-function mutations in the NS1 protein arising from the experimental adaptation of the human isolate A/Hong Kong/1/1968(H3N2 (HK to the mouse. The majority of these mouse adapted NS1 mutations were demonstrated to increase virulence, viral fitness, and interferon antagonism, but differ in binding to the post-transcriptional processing factor cleavage and polyadenylation specificity factor 30 (CPSF30. Because nuclear trafficking is a major genetic determinant of influenza virus host adaptation, we assessed subcellular localization and host gene expression of NS1 adaptive mutations. Recombinant HK viruses with adaptive mutations in the NS1 gene were assessed for NS1 protein subcellular localization in mouse and human cells using confocal microscopy and cellular fractionation. In human cells the HK wild-type (HK-wt virus NS1 protein partitioned equivalently between the cytoplasm and nucleus but was defective in cytoplasmic localization in mouse cells. Several adaptive mutations increased the proportion of NS1 in the cytoplasm of mouse cells with the greatest effects for mutations M106I and D125G. The host gene expression profile of the adaptive mutants was determined by microarray analysis of infected mouse cells to show either high or low extents of host-gene regulation (HGR or LGR phenotypes. While host genes were predominantly down regulated for the HGR group of mutants (D2N, V23A, F103L, M106I+L98S, L98S, M106V, and M106V+M124I, the LGR phenotype mutants (D125G, M106I, V180A, V226I, and R227K were characterized by a predominant up regulation of host genes. CPSF30 binding affinity of NS1 mutants did not predict effects on host gene expression. To our knowledge this is the first report of roles of adaptive NS1 mutations that impact intracellular localization and regulation of host gene expression.

  12. NMD and microRNA expression profiling of the HPCX1 locus reveal MAGEC1 as a candidate prostate cancer predisposition gene

    International Nuclear Information System (INIS)

    Mattila, Henna; Schindler, Martin; Isotalo, Jarkko; Ikonen, Tarja; Vihinen, Mauno; Oja, Hannu; Tammela, Teuvo LJ; Wahlfors, Tiina; Schleutker, Johanna

    2011-01-01

    Several predisposition loci for hereditary prostate cancer (HPC) have been suggested, including HPCX1 at Xq27-q28, but due to the complex structure of the region, the susceptibility gene has not yet been identified. In this study, nonsense-mediated mRNA decay (NMD) inhibition was used for the discovery of truncating mutations. Six prostate cancer (PC) patients and their healthy brothers were selected from a group of HPCX1-linked families. Expression analyses were done using Agilent 44 K oligoarrays, and selected genes were screened for mutations by direct sequencing. In addition, microRNA expression levels in the lymphoblastic cells were analyzed to trace variants that might alter miRNA expression and explain partly an inherited genetic predisposion to PC. Seventeen genes were selected for resequencing based on the NMD array, but no truncating mutations were found. The most interesting variant was MAGEC1 p.Met1?. An association was seen between the variant and unselected PC (OR = 2.35, 95% CI = 1.10-5.02) and HPC (OR = 3.38, 95% CI = 1.10-10.40). miRNA analysis revealed altogether 29 miRNAs with altered expression between the PC cases and controls. miRNA target analysis revealed that 12 of them also had possible target sites in the MAGEC1 gene. These miRNAs were selected for validation process including four miRNAs located in the X chromosome. The expressions of 14 miRNAs were validated in families that contributed to the significant signal differences in Agilent arrays. Further functional studies are needed to fully understand the possible contribution of these miRNAs and MAGEC1 start codon variant to PC

  13. Effects of C-reactive protein on adipokines genes expression in 3T3-L1 adipocytes

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    Yuan, Guoyue, E-mail: yuanguoyue@hotmail.com [Department of Endocrinology, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu 212001 (China); Jia, Jue; Di, Liangliang [Department of Endocrinology, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu 212001 (China); Zhou, Libin [Ruijin Hospital, Center of Molecular Medicine, Shanghai Institute of Endocrine and Metabolic Diseases, State Key Laboratory of Medical Genomics, Shanghai Jiaotong University Medical School, 197, Ruijin Road II, Shanghai 200025 (China); Dong, Sijing; Ye, Jingjing; Wang, Dong; Yang, Ling; Wang, Jifang [Department of Endocrinology, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu 212001 (China); Li, Lianxi [Department of Endocrinology and Metabolism, Shanghai Jiao Tong University Affiliated Sixth People' s Hospital, 600, Yishan Road, Shanghai 200233 (China); Yang, Ying [Ruijin Hospital, Center of Molecular Medicine, Shanghai Institute of Endocrine and Metabolic Diseases, State Key Laboratory of Medical Genomics, Shanghai Jiaotong University Medical School, 197, Ruijin Road II, Shanghai 200025 (China); Mao, Chaoming [Department of Endocrinology, The Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu 212001 (China); Chen, Mingdao, E-mail: mingdaochensh@yahoo.com [Ruijin Hospital, Center of Molecular Medicine, Shanghai Institute of Endocrine and Metabolic Diseases, State Key Laboratory of Medical Genomics, Shanghai Jiaotong University Medical School, 197, Ruijin Road II, Shanghai 200025 (China)

    2012-08-03

    Highlights: Black-Right-Pointing-Pointer CRP increases TNF-{alpha} and IL-6 genes expression in matured 3T3-L1 adipocytes. Black-Right-Pointing-Pointer CRP suppresses adiponectin, leptin and PPAR-{gamma} mRNA levels in matured 3T3-L1 cells. Black-Right-Pointing-Pointer Wortmannin reverses effects of CRP on adiponectin, TNF-{alpha} and leptin mRNA levels. Black-Right-Pointing-Pointer CRP may regulate IR, obesity and metabolic syndrome by this mechanism. -- Abstract: Adipose tissue is now recognized to be an important endocrine organ, secreting a variety of adipokines that are involved in the regulation of energy metabolism, insulin resistance and metabolic syndrome. C-reactive protein (CRP) is considered as one of the most sensitive markers of inflammation. A number of studies have shown that elevation of CRP concentrations is an independent predictive parameter of type 2 diabetes mellitus, which is also strongly associated with various components of the metabolic syndrome. The aim of the present study is to investigate the effects of CRP on adipokines genes expression in 3T3-L1 adipocytes. Quantitative real-time PCR analysis revealed that CRP inhibited adiponectin, leptin and peroxisome proliferator-activated receptor-gamma (PPAR-{gamma}) genes expression and raised tumor necrosis factor-{alpha} (TNF-{alpha}) and interleukin-6 (IL-6) mRNA levels in matured 3T3-L1 adipocytes in a dose and time-dependent manner. Pharmacological inhibition of phosphatidylinositol (PI)-3 kinase by wortmannin partially reversed the effects of CRP on adiponectin, TNF-{alpha} and leptin genes expression. These results collectively suggest that CRP regulates adiponectin, TNF-{alpha}, leptin, IL-6 and PPAR-{gamma} genes expression, and that might represent a mechanism by which CRP regulates insulin resistance, obesity and metabolic syndrome.

  14. Effects of C-reactive protein on adipokines genes expression in 3T3-L1 adipocytes

    International Nuclear Information System (INIS)

    Yuan, Guoyue; Jia, Jue; Di, Liangliang; Zhou, Libin; Dong, Sijing; Ye, Jingjing; Wang, Dong; Yang, Ling; Wang, Jifang; Li, Lianxi; Yang, Ying; Mao, Chaoming; Chen, Mingdao

    2012-01-01

    Highlights: ► CRP increases TNF-α and IL-6 genes expression in matured 3T3-L1 adipocytes. ► CRP suppresses adiponectin, leptin and PPAR-γ mRNA levels in matured 3T3-L1 cells. ► Wortmannin reverses effects of CRP on adiponectin, TNF-α and leptin mRNA levels. ► CRP may regulate IR, obesity and metabolic syndrome by this mechanism. -- Abstract: Adipose tissue is now recognized to be an important endocrine organ, secreting a variety of adipokines that are involved in the regulation of energy metabolism, insulin resistance and metabolic syndrome. C-reactive protein (CRP) is considered as one of the most sensitive markers of inflammation. A number of studies have shown that elevation of CRP concentrations is an independent predictive parameter of type 2 diabetes mellitus, which is also strongly associated with various components of the metabolic syndrome. The aim of the present study is to investigate the effects of CRP on adipokines genes expression in 3T3-L1 adipocytes. Quantitative real-time PCR analysis revealed that CRP inhibited adiponectin, leptin and peroxisome proliferator-activated receptor-gamma (PPAR-γ) genes expression and raised tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) mRNA levels in matured 3T3-L1 adipocytes in a dose and time-dependent manner. Pharmacological inhibition of phosphatidylinositol (PI)-3 kinase by wortmannin partially reversed the effects of CRP on adiponectin, TNF-α and leptin genes expression. These results collectively suggest that CRP regulates adiponectin, TNF-α, leptin, IL-6 and PPAR-γ genes expression, and that might represent a mechanism by which CRP regulates insulin resistance, obesity and metabolic syndrome.

  15. The effect of thyroid hormones on the white adipose tissue gene expression of PAI-1 and its serum concentration

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    C. Biz

    2009-12-01

    Full Text Available Metabolic syndrome is associated with an increased risk of developing cardiovascular diseases and Plasminogen activator inhibitor 1 (PAI-1 overexpression may play a significant role in this process. A positive correlation between adipose tissue gene expression of PAI-1 and its serum concentration has been reported. Furthermore, high serum levels of thyroid hormones (T3 and T4 and PAI-1 have been observed in obese children. The present study evaluates the impact of thyroid hormone treatment on white adipose tissue PAI-1 gene expression and its serum concentration. Male Wistar rats (60 days old were treated for three weeks with T4 (50 µg/day, Hyper or with saline (control. Additionally, 3T3-L1 adipocytes were treated for 24 h with T4 (100 nM or T3 (100 nM. PAI-1 gene expression was determined by real-time PCR, while the serum concentration of PAI-1 was measured by ELISA using a commercial kit (Innovative Research, USA. Both the serum concentration of PAI-1 and mRNA levels were similar between groups in retroperitoneal and epididymal white adipose tissue. Using 3T3-L1 adipocytes, in vitro treatment with T4 and T3 increased the gene expression of PAI-1, suggesting non-genomic and genomic effects, respectively. These results demonstrate that thyroid hormones have different effects in vitro and in vivo on PAI-1 gene expression in adipocytes.

  16. Allelic variants of OsSUB1A cause differential expression of transcription factor genes in response to submergence in rice.

    Science.gov (United States)

    Sharma, Niharika; Dang, Trang Minh; Singh, Namrata; Ruzicic, Slobodan; Mueller-Roeber, Bernd; Baumann, Ute; Heuer, Sigrid

    2018-01-08

    Flooding during seasonal monsoons affects millions of hectares of rice-cultivated areas across Asia. Submerged rice plants die within a week due to lack of oxygen, light and excessive elongation growth to escape the water. Submergence tolerance was first reported in an aus-type rice landrace, FR13A, and the ethylene-responsive transcription factor (TF) gene SUB1A-1 was identified as the major tolerance gene. Intolerant rice varieties generally lack the SUB1A gene but some intermediate tolerant varieties, such as IR64, carry the allelic variant SUB1A-2. Differential effects of the two alleles have so far not been addressed. As a first step, we have therefore quantified and compared the expression of nearly 2500 rice TF genes between IR64 and its derived tolerant near isogenic line IR64-Sub1, which carries the SUB1A-1 allele. Gene expression was studied in internodes, where the main difference in expression between the two alleles was previously shown. Nineteen and twenty-six TF genes were identified that responded to submergence in IR64 and IR64-Sub1, respectively. Only one gene was found to be submergence-responsive in both, suggesting different regulatory pathways under submergence in the two genotypes. These differentially expressed genes (DEGs) mainly included MYB, NAC, TIFY and Zn-finger TFs, and most genes were downregulated upon submergence. In IR64, but not in IR64-Sub1, SUB1B and SUB1C, which are also present in the Sub1 locus, were identified as submergence responsive. Four TFs were not submergence responsive but exhibited constitutive, genotype-specific differential expression. Most of the identified submergence responsive DEGs are associated with regulatory hormonal pathways, i.e. gibberellins (GA), abscisic acid (ABA), and jasmonic acid (JA), apart from ethylene. An in-silico promoter analysis of the two genotypes revealed the presence of allele-specific single nucleotide polymorphisms, giving rise to ABRE, DRE/CRT, CARE and Site II cis-elements, which

  17. The Expression of Activating Receptor Gene of Natural Killer Cells (KLRC3 in Patients with Type 1 Diabetes Mellitus (T1DM

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    Dalia Shalaby

    2017-07-01

    Full Text Available Objectives: To identify the possible role of natural killer (NK cells in the pathogenesis of type 1 diabetes mellitus (T1DM through studying the expression of the KLRC3 gene, which encodes the NK cell activating receptor (NKG2E. Methods: This study was conducted at Alexandria University Children’s Hospital from April to October 2015. The study was conducted with 30 newly diagnosed T1DM patients (15 males and 15 females, aged 7–13 years (10.6±1.8 years and 20 non-diabetic subjects served as age- and sex-matched controls. The patients were further sub-divided into two groups; group I included patients who first presented with classical symptoms of DM (polyuria, polydipsia, and polyphagia without diabetes ketoacidosis (DKA and group II included patients who first presented with DKA. The expression of the KLRC3 gene was measured in each group using the real-time polymerase chain reaction. Results: KLRC3 gene expression was significantly downregulated in T1DM cases compared to healthy controls (p = 0.001. Expression was more downregulated in group I patients (p = 0.008. Moreover, there was higher mean value of glycated heamoglobin and lower C-peptide levels in group I than group II. Serum pancreatic amylase showed no significant difference between the two groups. Conclusions: KLRC3 gene expression was downregulated in patients with T1DM compared to healthy controls. Downregulation of expression was greater in DKA patients compared to those who presented with classical symptoms. Expression of KLRC3 in T1DM might play a role in the pathogenesis of T1DM and could be a predictor of its severity.

  18. Lack of co-ordinate expression of the alpha1(I) and alpha1(III) procollagen genes in fibroblast clonal cultures.

    Science.gov (United States)

    Yamaguchi, Y; Crane, S; Zhou, L; Ochoa, S M; Falanga, V

    2000-12-01

    Several extracellular matrix genes, most notably alpha1(I) and alpha1(III) procollagen, are reported to be co-ordinately expressed in cultures of dermal fibroblasts. However, it remains unclear whether the expression of these genes is truly co-ordinate or whether it may be the result of averaging the phenotypic expression of different fibroblast subpopulations present within each culture. Objectives To determine by Northern analysis the correlation between alpha1(I) and alpha1(III) procollagen mRNA levels in clonal populations of human dermal fibroblasts. As previously described, clonal cultures were derived from parent strains of human dermal fibroblasts by a microscopically controlled dilution technique and by stimulation of single cells with low oxygen tension in the early phases of clonal growth. In agreement with previous reports, we found that baseline steady-state levels of alpha1(I) procollagen mRNA were co-ordinately regulated with the alpha1(III) procollagen mRNA in 26 parent strains (r = 0. 9003; P ordinate regulation observed in non-clonal cultures, suggesting that these two genes operate under different sets of regulatory controls. This clonal heterogeneity may provide additional flexibility to the process of tissue repair and fibroblast clonal expansion.

  19. Wound-induced expression of DEFECTIVE IN ANTHER DEHISCENCE1 and DAD1-like lipase genes is mediated by both CORONATINE INSENSITIVE1-dependent and independent pathways in Arabidopsis thaliana.

    Science.gov (United States)

    Ruduś, Izabela; Terai, Haruka; Shimizu, Takafumi; Kojima, Hisae; Hattori, Kazuki; Nishimori, Yuka; Tsukagoshi, Hironaka; Kamiya, Yuji; Seo, Mitsunori; Nakamura, Kenzo; Kępczyński, Jan; Ishiguro, Sumie

    2014-06-01

    Endogenous JA production is not necessary for wound-induced expression of JA-biosynthetic lipase genes such as DAD1 in Arabidopsis. However, the JA-Ile receptor COI1 is often required for their JA-independent induction. Wounding is a serious event in plants that may result from insect feeding and increase the risk of pathogen infection. Wounded plants produce high amounts of jasmonic acid (JA), which triggers the expression of insect and pathogen resistance genes. We focused on the transcriptional regulation of DEFECTIVE IN ANTHER DEHISCENCE1 and six of its homologs including DONGLE (DGL) in Arabidopsis, which encode lipases involved in JA biosynthesis. Plants constitutively expressing DAD1 accumulated a higher amount of JA than control plants after wounding, indicating that the expression of these lipase genes contributes to determining JA levels. We found that the expression of DAD1, DGL, and other DAD1-LIKE LIPASE (DALL) genes is induced upon wounding. Some DALLs were also expressed in unwounded leaves. Further experiments using JA-biosynthetic and JA-response mutants revealed that the wound induction of these genes is regulated by several distinct pathways. DAD1 and most of its homologs other than DALL4 were fully induced without relying on endogenous JA-Ile production and were only partly affected by JA deficiency, indicating that positive feedback by JA is not necessary for induction of these genes. However, DAD1 and DGL required CORONATINE INSENSITIVE1 (COI1) for their expression, suggesting that a molecule other than JA might act as a regulator of COI1. Wound induction of DALL1, DALL2, and DALL3 did not require COI1. This differential regulation of DAD1 and its homologs might explain their functions at different time points after wounding.

  20. Enhancing expression of SSU1 genes in Saccharomyces uvarum leads to an increase in sulfite tolerance and a transcriptome profile change.

    Science.gov (United States)

    Liu, X Z; Sang, M; Zhang, X A; Zhang, T K; Zhang, H Y; He, X; Li, S X; Sun, X D; Zhang, Z M

    2017-05-01

    Saccharomyces uvarum is a good wine yeast species that may have great potential for the future. However, sulfur tolerance of most S. uvarum strains is very poor. In addition there is still little information about the SSU1 gene of S. uvarum, which encodes a putative transporter conferring sulfite tolerance. In order to analyze the function of the SSU1 gene, two expression vectors that contained different SSU1 genes were constructed and transferred into a sulfite-tolerant S. uvarum strain, A9. Then sulfite tolerance, SO2 production, and PCR, sequencing, RT-qPCR and transcriptome analyses were used to access the function of the S. uvarum SSU1 gene. Our results illustrated that enhancing expression of the SSU1 gene can promote sulfite resistance in S. uvarum, and an insertion fragment ahead of the additional SSU1 gene, as seen in some alleles, could affect the expression of other genes and the sulfite tolerance level of S. uvarum. This is the first report on enhancing the expression of the SSU1 gene of S. uvarum. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.